BASIC AND TRANSLATIONAL MEDICINE
BASIC RESEARCH OF CARDIOVASCULAR DISEASE
GW34-e0013
Yang Yang
Northwest University
OBJECTIVES Sepsis is a major health threat and often results in heart failure as a major endpoint. Growth arrest-specific gene 6 (GAS6) participates in immune regulation and inflammation through binding to the TAM receptor family. This study was designed to examine the cardioprotective role of GAS6 in sepsis.
METHODS ELISA and single-cell RNA sequencing, immunohistochemical analysis or qPCR were performed to assess GAS6 levels in cardiomyocytes from patients and mice with sepsis. Mice with GAS6 overexpression/knockout and cardiomyocytes with high/low expression of GAS6 were constructed to discern the possible role of GAS6 in septic myocardial injury. Next, the involvement of NLRP3 in GAS6-induced septic myocardial injury responses, if any, was determined. Finally, the cardioprotective effects of vitamin K1 (VK1), a safe and effective GAS6 agonist screened from FDA library, was evaluated in sepsis.
RESULTS Serum GAS6 levels were increased in septic patients and mice while myocardial GAS6 level was decreased in septic mice and patients. Single-cell RNA sequencing further revealed a decline in GAS6 levels in nearly all cell clusters including cardiomyocytes. GAS6 overexpression overtly improved cardiac dysfunction and injury in CLP-challenged mice, along with alleviated mitochondrial injury, endoplasmic reticulum stress, oxidative stress, and apoptosis. However, GAS6-elicited beneficial effects were partially removed by GAS6 knockout. NLRP3 served as a downstream effector for GAS6-initiated myocardial benefits. GAS6 knockout led to elevated levels of NLRP3 and cytokines, the effect of which was reconciled by GAS6 overexpression. Vitamin K1, a safe and effective GAS6 agonist, was identified from the FDA-approved drug library and possessed anti-sepsis myocardial injury through regulation of GAS6/NLRP3 signaling.
CONCLUSIONS These results revealed the therapeutical potential of targeting GAS6/NLRP3 signaling in the management of heart anomalies in sepsis.
GW34-e0020
Liu Yang1, Deng Jing2, Piao Chunmei1
1Beijing Anzhen Hospital Affiliated to Capital Medical University
2Beijing Institute of Heart Lung and Blood Vessel Diseases
OBJECTIVES Pulmonary hypertension (PH) seriously affects the health of patients. We have found in clinical studies that PH has adverse effects on both maternal and offspring.
METHODS Twenty-four SPF C57 mice aged 7–9 weeks, including 12 females and 12 pregnant mice, were selected and further divided into 4 groups with 6 mice in each group according to random number table method: ① Female mice with normal oxygen; ② Female mice with hypoxia/SU5416; ③ Pregnant mice with normal oxygen; ④ Pregnant mice with hypoxia/SU5416. All mice were fed and water freely, two groups of mice were fed in normal oxygen environment, the other two groups of mice with hypoxia/SU5416 were fed in a low oxygen chamber, and nitrogen was continuously injected into the chamber. The controller sensed the oxygen concentration in the chamber through the probe and adjusted the nitrogen intake in real time to maintain the oxygen concentration at 10%. Avoid mice suffocation. SU5416 was injected subcutaneously twice a week, 20 mg/kg. At the same time, water and substrate were replaced, nitrogen was replaced, oxygen content was maintained at 10%, and weight and death of mice were monitored. Considering that the mice delivered at 21 days, 19 days of gestation before delivery was selected as the experimental end point to detect weight, right ventricular systolic pressure (RVSP), right ventricular hypertrophy index (RVHI).
RESULTS There was no significant difference in RVSP and RVHI between female and pregnant mice under the same condition. Compared with normal oxygen condition, two groups of mice in hypoxia/SU5416 had poor development, RVSP and RVHI were significantly increased, the number of fetal mice was small, hypoplasia, degeneration and even abortion.
CONCLUSIONS The model of mice PH was successfully established. PH affects the development and health of female and pregnant mice, and seriously affects the fetuses.
GW34-e0025
Xiaodong Jing
The Second Affiliated Hospital of Chongqing Medical University
OBJECTIVES Epicardial cells (EpiCs) are a group of progenitor cells with multi-differentiation potential. But this differentiation potential is quiescent in adulthood. Our objective is to investigate the effects of extracellular matrix (ECM) Agrin on epicardial cells activation and proliferation after myocardial infarction (MI) in mice.
METHODS Mouse myocardial infarction models were constructed and primary mouse embryonic epicardial cells were cultured. The dynamic changes of myocardial Agrin was determined during heart development and after myocardial infarction in mice. Then, we determined the role of myocardial injection of recombinant rat Agrin (rrAgrin) on EpiCs activation and proliferation followed by 1, 3, 5, 7 and 14 days after MI in mouse models.
RESULTS Agrin in myocardium declined from embryonic to adult in mice. Agrin in sub-epicardium decreased significantly in the early stage after MI, but then significantly increased. The myocardial injection of rrAgrin significantly upregulated the epicardial activation marker Tbx18 and Wt1, and the proliferation markers Ki67, PH3, Aurora B and Brd U after MI. Agrin significantly thickened the epicardium by promoting EpiCs proliferation, and the increased EpiCs may migrate into myocardium. In addition, Agrin can also promoted the proliferation and migration of EpiCs by binding to its receptor α-dystroglycan (α-DAG) in vitro.
CONCLUSIONS Myocardial Agrin changes dynamically during heart development and after cardiac injury. Agrin may bind to its receptor α-DAG to promote activation, proliferation, and migration of EpiCs after MI.
GW34-e0043
Dingqian Liu
Zhongshan Hospital, Fudan University
OBJECTIVES Cardiomyocytes autophagy plays a key role in prevention of heart failure (HF), identification of key autophagy-related genes is particularly important to further study the pathological features of HF.
METHODS In this study, autophagy-related genes (ARGs) and HF-related datasets (GSE21610 and GSE57345) were extracted from the human autophagy database and gene expression omnibus (GEO) database, respectively. Firstly, the differentially expressed genes (DEGs) between HF and normal control (HC) samples were screened by “limma”. Secondly, the HF-related genes (module genes) were identified by weighted gene co-expression network analysis (WGCNA). Then, the common genes were obtained by intersecting the DEGs, module genes and ARGs. Besides, the function enrichment analyses of these common genes were conducted by “ClusterProfiler”. Then, the key genes were calculated and identified by both training dataset (GSE21610) and validation dateset (GSE57345). Moreover, the gene set enrichment analysis (GSEA) and immune cell infiltration analysis were performed to reveal the partial molecular mechanisms. Furthermore, the targeted drugs of key genes were predicted in Drug-Gene Interaction database.
RESULTS Five common genes associated with autophagy and catabolic process were obtained by intersecting 447 DEGs, 6370 module genes and 222 ARGs. BNIP3L, CDKN1BI and HIF1A were the key genes of HF, which were associated with endocytosis, lysosome, protein processing in endoplasmic reticulum, ubiquitin mediated proteolysis and etc. Besides, HIF1A and BNIP3L were significantly negative associated with the proportion of monocytes. Furthermore, Epoetin alfa and Tretinoin were special drugs synchronously associated with CDKN1B and HIF1A.
CONCLUSIONS This study revealed the potential molecular mechanisms of autophagy-related genes in HF, which could provide novel insights for the clinical diagnosis and treatment of HF.
GW34-e0066
Jiachun Xia1, Yanan Pang1, Chenshan Gao2, Lei Hou1
1Institute of Cardiovascular Diseases, Tongren Hospital, Shanghai Jiao Tong University School of Medicine
2Collaborative Innovation Centre of Regenerative Medicine and Medical BioResource Development and Application Co-constructed by the Province and Ministry, Guangxi Medical University
OBJECTIVES To explore the effects and potential mechanisms of PCSK9 on ventricular remodeling after myocardial infarction.
METHODS Mouse were used to construct a model of acute myocardial infarction. To assess the effect of PCSK9, we used WT mice treated with PCSK9 inhibitor (alirocumab) or PCSK9−/− mice. Regulatory T (Treg) cells in WT mice were depleted by intraperitoneal injection of CD25 antibody. Cardiac function was assessed by cardiac ultrasound on day 1, 14 and 28 after successful construction of the myocardial infarction model; the infarct size was measured by Masson staining of tissue sections, the cross-sectional area of cardiomyocytes was measured by WGA staining and the collagen content was assessed by Sirius Red (PSR) staining 28 days after myocardial infarction. The proportion of Treg cells, proliferative Treg cells, M1 and M2 macrophages in the cardiac tissue was analyzed by flow cytometry on day 7 after myocardial infarction. The induced human iTreg cells were treated with human-derived PCSK9 recombinant protein-treated, and the cellular reactive oxygen species (ROS) level and cell proliferation was analyzed by flow cytometry.
RESULTS Knockdown and inhibition of PCSK9 improved cardiac function, reduced infarct size and cross-sectional area of cardiomyocytes, increased collagen content in the margins of the infarct zone, and increased the proportion of Treg cells and proliferative Treg cells in cardiac tissue. PCSK9 inhibitor promoted M2 polarization of macrophages in cardiac tissue. PCSK9 recombinant protein promoted ROS production and inhibited proliferation of human iTreg cells. After depletion of Treg cells in mice, PCSK9 inhibitor did not improve cardiac function.
CONCLUSIONS Knockdown and inhibition of PCSK9 alleviated ventricular remodeling after myocardial infarction by regulating the proportion of Treg cells in cardiac tissue and promoting proliferation.
GW34-e0068
Liuhua Zhou, Jiateng Sun, Tongtong Yang, Lingfeng Gu, Liansheng Wang
The First Affiliated Hospital of Nanjing Medical University
OBJECTIVES Neonatal mice achieve complete functional and structural repair through endogenous myocardial regeneration after apical resection (AR), but this capacity is rapidly lost 7 days after birth. As one of the DNA chaperone, HMGB2 is widely involved in the regulation of transcription, recombination, repair and genome homeostasis. At present, the role of HMGB2 in cardiac regeneration is unknown. Given that HMGB2 had a significant positive regulation on cell proliferation, targeting cardiomyocytes to promote HMGB2 seems to be a promising attempt at myocardial regeneration therapy. In this research, we investigated whether HMGB2 can promote cardiac regeneration.
METHODS Cardiomyocytes of neonatal and adult mice were detected by proteome TMT mass spectroscopy. And we combined with SnNA-Seq data which was related to dynamic transcriptional responses to regenerative and non-regenerative cardiomyocyte injury to select the target gene HMGB2. Subsequently, the HMGB2 expression was detected by WB, PCR and immunofluorescence staining at the peri myocardial regeneration time. The effect of HMGB2 on the myocardial regeneration potential of cardiomyocytes (CMs) was evaluated using CM-specific HMGB2 overexpression or knockdown adenovirus vector. The models of P1-AR and P7-AR were constructed to verify the effect of HMGB2 on myocardial cell proliferation and myocardial repair. The mechanism of HMGB2 mediated cardiac regeneration was detected by RNA-seq.
RESULTS The results of biogenic analysis showed that HMGB2 was consistent with the spatial and temporal changes of cardiomyocyte regeneration in terms of functional protein and RNA expression. And the expression of HMGB2 was downregulated during postnatal cardiac development. Overexpression of HMGB2 stimulateed the self-proliferating potential of NMCMs, while HMGB2 knockdown had the opposite biological function. In addition, HMGB2 overexpression could prolong the proliferation time window of newborn myocardium. In the real microenvironment in vivo, HMGB2 knockdown inhibited cell cycle activity and cardiac function. However, HMGB2 overexpression promoted cell cycle activity and cardiac function. We transfected NMCMs with Ad5: cTNT-HMGB2 and Ad5: cTNT-Vector and then performed transcriptomic sequencing (RNA-seq) between the two groups. The results of RNA-seq showed that the differential genes were not only enriched in the regulation of cell cycle process, but also in the glycolysis, PI3K/AKT and HIF-1α signaling pathways after HMGB2 overexpression. In terms of mechanism, overexpression of HMGB2 upregulated PI3K/AKT/HIF-1α pathway to activate glycolysis and promote cardiac regeneration after myocardial injury.
CONCLUSIONS In summary, our results suggested that HMGB2 modulated the glucose metabolism via the PI3K/AKT/HIF-1α signaling pathways, which might contribute to the cardiac regeneration after myocardial injury.
GW34-e0072
WeiBin He, Guang Tong, Lin Zeng, WenLong He, XiaoPan Chen, Cien Zhen, PengYuan Chen, Ning Tan, ZhongChan Sun, PengCheng He
Guangdong Cardiovascular Institute
OBJECTIVES Doxorubicin (DOX)-induced cardiotoxicity (DIC) is frequently observed in clinical practice. Autophagy dysregulation is known to be a mechanism of DIC. Mitochondrial-Endoplasmic Reticulum Contact sites (MERCs) is where autophagy initiates and autophagosomes form. However, the role of MERCs in autophagy dysregulation in DIC remains elusive. FUNDC1 is a mitochondria outer membrane protein that serves as a mitophagy receptor as well as a tethering protein of MERCs. The aim of our study was to investigate the effect of DOX on MERCs and explore whether it is involved in the dysregulation of autophagy in DIC.
METHODS We use confocal microscopy and transmission electron microscopy to assess MERCs structure. Overall autophagic flux was analyzed by mCherry-EGFP-LC3B fluorescence assay and western blotting for LC3BII. A total dose of 15 mg/kg of doxorubicin was administrated in mice to construct a DIC model in vivo. Adeno-associated virus (AAV) was used to cardiac-specifically overexpress FUNDC1. Cardiac remodeling was evaluated by Masson’s trichrome staining. Cardiac function (left ventricle ejection fraction, LVEF; left ventricle fraction shortening, LVFS) was measured by echocardiography.
RESULTS DOX treatment led to a significant downregulation of multiple tethering proteins (including FUNDC1) of MERCs and inhibition of MERCs formation, as demonstrated by decreased colocalization coefficients of mitochondria and endoplasmic reticulum in cardiomyocytes. The overexpression of FUNDC1 abolished DOX-induced inhibition on MERCs formation. In addition, DOX treatment blocked the normal autophagic flux, as demonstrated by the decreased autophagosomes synthesis and impaired autophagosomes turnover. Maintaining the MERCs structure by FUNDC1 overexpression was able to restore the blocked autophagic flux mentioned above. Moreover, FUNDC1 overexpression restored the blocked autophagic flux in vivo, as demonstrated by the increased LC3B turnover compared to the DOX treatment group. Importantly, global heart function of FUNDC1 overexpressed mice was significantly improved post DOX treatment compared to the wildtype DOX treatment group, as demonstrated by the increased LVEF and LVFS. In addition, LV fibrosis level was found to be significantly decreased in FUNDC1 overexpressed mice treated with DOX compared to the wildtype DOX treatment group.
CONCLUSIONS We reported for the first time that the impaired MERCs-formation in DOX-treated cardiomyocytes and revealed that restoring the disrupted MERCs structure by FUNDC1 overexpression was able to revert the blocked autophagic flux induced by DOX treatment. Furthermore, cardiac-specific overexpression of FUNDC1 protected DOX-treated mice against adverse cardiac remodeling and improved cardiac function. These findings may shed light on the prevention and treatment of DIC.
GW34-e0092
Chang Cui, Jiuzhou Chen, Huiyuan Qin, Minglong Chen
Division of Cardiology, The First Affiliated Hospital of Nanjing Medical University, Nanjing, China
OBJECTIVES Atrial cardiomyopathy forms the substrate for atrial fibrillation (AF) and underlies the potential for atrial thrombus formation and subsequent stroke. However, generating stable animal models that accurately reflect the whole course of atrial lesions, especially the development of AF is highly challenging. The isoform of CRE-binding protein modulator (CREM-IbΔC-X) has been demonstrated to be involved in the regulation of cardiac development and atrial rhythm. We investigated if cardiac-specific overexpression of CREM-IbΔC-X via CRISPER/Cas9 (CS-CREM) in mice could be a suitable disease model of atrial cardiomyopathy.
METHODS Immunofluorescence staining, immunoblotting, and quantitative real-time PCR were used to verify the expression of CREM-IbΔC-X in patients with AF. CRISPR/Cas9 technology was employed to construct the CS-CREM mice. Subsequently, survival analysis, repeated ECG recordings, echocardiography, histology, proteomics analysis and responses to antiarrhythmic drugs were assessed.
RESULTS CREM-IbΔC-X was highly expressed in atrial biopsies from the patients with AF. The CS-CREM mice well illustrated the development of atrial cardiomyopathy in terms of electrophysiological and structural remodeling over time. Further proteomics analysis of atrial samples identified differentially expressed proteins, which were enriched in extracellular matrix (up-regulated) and metabolic processes (down-regulated) in the CS-CREM mice. Moreover, the correspond responses to anti-arrhythmia drugs, i.e., amiodarone and propafenone, indicated that CS-CREM mice may be an ideal in vivo model for drug testing.
CONCLUSIONS Our study introduced a new model of atrial cardiomyopathy with spontaneous atrial fibrillation by cardiac-specifically overexpressing CREM-IbΔC-X in mice, which would be helpful for investigating the mechanism and therapeutic target of atrial cardiomyopathy.
GW34-e0095
Luxun Tang1, Yu Shi2, Yongjian Yang1, Chunyu Zeng2
1Department of Cardiovascular Medicine, The General Hospital of Western Theater Command PLA
2Department of Cardiology, Daping Hospital, The Third Military Medical University (Army Medical University)
OBJECTIVES The neonatal mammalian heart has a remarkable regenerative capacity, while the adult heart has difficulty regenerating. A metabolic reprogramming from glycolysis to fatty acid oxidation (FAO) occurs along with the loss of cardiomyocyte proliferative capacity shortly after birth. In this study, we sought to determine if and how metabolic reprogramming regulates cardiomyocyte proliferation.
METHODS Cardiac-specific Cpt1a and Cpt1b knockout mice were used to investigate the effect of reversing metabolic reprogramming by CPT1 inhibition on cardiomyocyte proliferation. The proliferation of cardiomyocytes was evaluated by immunofluorescent staining. The infraction area of heart after myocardial infraction was determined by Masson staining. Etomoxir was also used to inhibit CPT1 activity. An in vitro system with time lapse microscopy can quantify adult cardiomyocyte cytokinesis. We used time-lapse microscopy to verify the cytokinesis rate of adult cardiomyocytes. We used time-lapse microscopy to verify the cytokinesis rate of adult cardiomyocytes. Additionally, we used a lineage-tracing system in which we crossed cardiomyocyte-specific Myh6mERcremER mice with Mosaic Analysis with Double Markers (MADM) mice to examine cardiomyocyte division.
RESULTS Reversing metabolic reprogramming by CPT1 inhibition, using cardiac-specific Cpt1a and Cpt1b knockout mice promoted cardiomyocyte proliferation and improved cardiac function post-myocardial infarction (MI). The inhibition of CPT1 is of pharmacological significance because those protective effects were replicated by etomoxir, a CPT1 inhibitor. CPT1 inhibition, by decreasing PARP1 expression, reduced ADP-ribosylation of DUSP1 in cardiomyocytes, leading to decreased p38 MAPK phosphorylation, and stimulation of cardiomyocyte proliferation.
CONCLUSIONS Our present study indicates that reversing metabolic reprogramming is an effective strategy to stimulate adult cardiomyocyte proliferation. CPT1 is a potential therapeutic target for promoting heart regeneration and MI treatment.
GW34-e0100
Jiachun Xia1, Chenshan Gao2, Yanan Pang1, Lei Hou1
1Institute of Cardiovascular Diseases, Tongren Hospital, Shanghai Jiao Tong University School of Medicine
2Collaborative Innovation Centre of Regenerative Medicine and Medical BioResource Development and Application Co-constructed by the Province and Ministry, Guangxi Medical University
OBJECTIVES A series of inflammatory responses triggered by ischemia-reperfusion injury can induce adverse remodeling. In addition to its lipid-lowering effect, proprotein convertase subtilisin/kexin 9 (PCSK9) also plays an important role in atherosclerosis and myocardial infarction inflammation. The protective effect of early PCSK9 inhibitor treatment on myocardial salvage index (MSI) and infarct size remains unclear. This trial aimed to assess the effect of early PCSK9 inhibitor treatment on MSI in patients with anterior ST-segment elevation myocardial infarction (STEMI).
METHODS The PERFECT trial was a prospective, randomized controlled trial. Patients with anterior STEMI who were admitted within 24 h of symptom onset were randomized to receive conventional treatment combined with the PCSK9 inhibitor alirocumab at a dose of 75 mg or conventional treatment alone at a 1:1 ratio. The primary endpoint was MSI, measured by cardiac magnetic resonance imaging (CMR) 5–8 days after PCI. The secondary endpoints were LVEF and the peak time for CK-MB and hs-cTnT.
RESULTS In total, 20 patients met the inclusion criteria and were randomized to the alirocumab group (n=10) and control group (n=10). At 5–8 days, the MSI was comparable between the two groups (56.66±18.3% vs. 48.3±14.34%, P=0.31), albeit the alirocumab group outcome seemed to be better. There were no significant differences in secondary endpoints.
CONCLUSIONS Among patients with anterior STEMI, early application of alirocumab after PCI did not significantly increase the MSI at 5–8 days. Larger trials are required to evaluate the effect of early administration of PCSK9 inhibitors after myocardial infarction.
GW34-e0107
Zhengdong Chen, Naifeng Liu
Department of Cardiology, Zhongda Hospital, School of Medicine, Southeast University 87 Dingjiaqiao, Nanjing 210009, PR China
OBJECTIVES Vascular calcification is the most common pathological change in metabolic syndrome. The abnormal metabolism of iron, a crucial trace element, is involved in the occurrence and development of vascular calcification; DNA damage response inextricably linked with iron homeostasis, is a hallmark of vascular calcification. Moreover, vascular smooth muscle cell is essential for keeping the physiological structure and function of blood vessel. The aim of this experiment is to elucidate whether the genomic instability triggered via iron homeostasis abnormality is the underlying pathogenic mechanism of vascular smooth muscle cell osteoblastic differentiation.
METHODS In vivo, the vascular calcification model was developed in mice by vitamin D3. In vitro, the vascular calcification model, was induced in primary vascular smooth muscle cells of mouse thoracic aortas by calcifying medium. The ferrostatin-1 (Fer-1) or deferoxamine (DFO) was used in anti-calcification studies.
RESULTS In vivo, both the elevated expressions of vascular calcification and DNA damage response markers in mouse calcified thoracic aortas were suppressed by Fer-1 or DFO. In vitro, the high levels of vascular calcification and DNA damage response markers in vascular smooth muscle cells were notably ameliorated by Fer-1 or DFO; additionally, the siRNAs knocking down DNA damage response signalling pathway genes partially abolished the protective effects of Fer-1 or DFO.
CONCLUSIONS Taken together, this study indicated that pharmacological iron restriction is a novel strategy for anti-DNA damage response associated vascular smooth muscle cell calcification.
GW34-e0111
Yongshun Wang
Shenzhen People’s Hospital
OBJECTIVES Coronary slow flow phenomenon (CSFP), characterized by slower contrast agent progression throughout the affected coronary artery under angiography, have been linked to chest pain and cardiac diseases, ranging from arrhythmias to myocardial infarction. Furthermore, CSFP patients exhibit similar patterns in cardiovascular biomarker levels as observed for those aforementioned syndromes. In this study, we investigated whether one such biomarker, the phospholipid lysophosphatidic acid (LPA), was associated with CSFP onset, as well as the possible underlying mechanisms involved.
METHODS Thirty randomly-recruited patients, diagnosed with CSFP under angiography, was age- and gender-matched with 30 non-CSFP, atherosclerosis patients, serving as the control group. Computed tomography myocardial perfusion imaging and coronary computed tomography angiography to construct myocardial blood flow maps, followed by collection of coronary and femoral artery blood and high-performance liquid chromatography/electrospray ionization tandem mass spectrometry to measure LPA levels. To examine the effects of LPA on blood vessels, a mouse aorta organ bath was constructed, followed by application of other vasodilative/vasoconstrictive agents. The effects of LPA on nitric oxide and mitochondrial functioning, particularly in terms of reactive oxygen species (ROS) and glucose metabolism, was then examined using human umbilical vein endothelial cells (HUVECs).
RESULTS CSFP patients, compared to control group, had significantly higher plasma LPA levels, particularly for 18:2 and 20:4 isoforms, which were associated with poorer myocardial perfusion. LPA was able to induce vasoconstriction in mouse thoracic aortas, owing to its suppression of NO and increase in ROS production; LPA-treated HUVECs supported these findings, in which it inhibited eNOS, along with increasing expression of ROS-producing enzymes NADPH oxidase 4 and p22phox, in turn contributing to mitochondrial dysfunction.
CONCLUSIONS LPA contributed to CSFP onset by fostering endothelial dysfunction, via inhibiting NO and increasing ROS, thereby serving as a promising therapeutic target for developing future treatment strategies for CSFP.
GW34-e0118
Dong Zhang1, Hui Wu2
1China Three Gorges University
2Yichang Central People’s Hospital
OBJECTIVES Alleviating myocardial ischemia-reperfusion injury (MIRI) plays a crucial role in improving the prognosis and cardiac function after acute myocardial infarction. Pyroptosis, a novel form of cell death, is involved in the regulation of MIRI. This study aims to investigate the impact of the long form of cFLIP (cFLIPL) on pyroptosis in MIRI and the underlying mechanisms.
METHODS H9C2 cells and SD rats were transfected with a recombinant adenovirus vector carrying cFLIPL, and the transfection was allowed to proceed for 3 days. H9C2 cells underwent hypoxia for 4 hours followed by reoxygenation for 12 hours, simulating ischemia-reperfusion (I/R) injury. SD rats experienced 30 minutes of ischemia followed by 2 hours of reperfusion to establish a MIRI model. Model parameters were evaluated by assessing myocardial cell viability, pyroptosis, and the expression of myocardial injury-related proteins.
RESULTS The expression of cFLIPL is significantly reduced during I/R injury and hypoxia/reoxygenation (H/R) injury. Overexpression of cFLIPL can reduce the infarct size in vivo and increase the viability of H9c2 cells in vitro. I/R and H/R upregulate the expression of ASC, cleaved caspase-1, NLRP3, GSDMD-N, interleukin-1β (IL-1β), and interleukin-18 (IL-18) proteins, inducing pyroptosis. Notably, overexpression of cFLIPL can modulate the related proteins to inhibit myocardial cell pyroptosis.
CONCLUSIONS cFLIPL can inhibit pyroptosis by targeting and binding to caspase-1, suppressing the production of inflammatory factors and blocking cell membrane rupture. Therefore, cFLIPL may serve as a potential intervention target for alleviating MIRI by inhibiting the pyroptotic pathway.
GW34-e0138
Wenjie Chen, Yafeng Zhou
Dushu Lake Hospital Affiliated to Soochow University
OBJECTIVES Hypertension is a major risk factor for cerebrovascular disease. During cardiovascular development and disease, epigenetic mechanisms regulate chromatin structure and spectrum-specific gene expression. However, how epigenetics regulate the development and regression of hypertension remains unknown. This review combines histone deacetylase Sirt3 and hypertension with the aim of elucidating recent advances in the development and regression of hypertension and showing the therapeutic potential of Sirt3 expression for targeting vascular dysfunction and hypertension.
METHODS We have collected and analyzed a large amount of researches to explore the role of Sirt3 in various stages of hypertension development, including oxidative stress, vascular inflammation, endothelial damage, vascular remodeling, cardiac fibrosis, and myocardial hypertrophy. We also analyzed the prospective role of Sirt3 as an intermediate molecule for multiple drugs in the reversal and regression of hypertension through different pathways in treated cells or models.
RESULTS Sirt3 targeted drugs can improve the progression of hypertension in animal models and at the cellular level, and may also play a role in reversing elevated blood pressure in human tissue models.
CONCLUSIONS SIRT3 is an important regulator of hypertensive heart dysfunction, hypertrophy, and fibrosis. Future clinical research could develop new Sirt3 agonists and determine the direction of mitochondrial targeting strategies, highlighting the therapeutic potential of targeting Sirt3 in vascular dysfunction, primary hypertension, and end-organ damage in hypertension.
GW34-e0148
Xi Zhang1,2, Meili Wang1,2, Suli Zhang1,2, Huirong Liu1,2
1Department of Physiology & Pathophysiology, School of Basic Medical Sciences, Capital Medical University, Beijing 100069, China
2Beijing Key Laboratory of Metabolic Disorder Related Cardiovascular Diseases, Capital Medical University, Beijing 100069, China
OBJECTIVES Angiotensin II type 1 receptor (AT1R) is a key mediator of the rennin-angiotensin-aldosterone system (RAAS). Autoantibodies against AT1R (AT1-AA), as an activator of AT1R, was found in abdominal aortic aneurysm (AAA) related diseases, such as hypertension and atherosclerosis. However, the role of AT1-AA in AAA is still unclear.
METHODS We used angiotensin II (AngII)–infused male ApoE−/− mice model of AAA. Ultrasonography measurement, histological assessment, western blot, qRT-PCR and RNA sequencing were used to assess the severity of AAA. Nanoluciferase-based bioluminescence resonance energy transfer (NanoBRET) assay, radio assay of receptors, homogeneous time resolved fluorescence (HTRF) and molecular dynamics (MD) studies were used to measure the activation feature of AT1R and the affinity of AT1R with AT1-AA or AngII.
RESULTS AT1-AA increased the incidence of AAA (vehicle vs. AT1-AA: 47 vs. 81%; P<0.05). Furthermore, AT1-AA greatly increased severity of AAA, due to augmenting the diameter of abdominal aorta and the degradation of elastin by enhancing the expression of MMP2/9 (matrix metalloproteinase) and the accumulation of inflammatory cells. Mechanically, AT1-AA enhanced AngII-induced recruitment of Gq, but not β-arrestin1/2. AT1-AA potentiated the AngII-mediated Gq-dependent inositol phosphate and the activation of ERK1/2. The responses were abolished by AT1R antagonist and Gq inhibitor. And AT1-AA significantly potentiated the binding of AngII with AT1R allosterically (mean KD±SEM: 34.11±0.9765 nM at 0 nM AT1-AA vs. 25.63±1.826 nM at 100 nM AT1-AA; P<0.05), without altering the maximum binding. MD studies showed that AT1-AA increased interaction forces of AngII with AT1R, including Lennard-Jones Short Range (LJ-SR) and Coulombic Short Range (Coul-SR).
CONCLUSIONS AT1-AA, as an allosteric activator of AT1R, promotes AAA through increasing the affinity of AngII with AT1R and activating Gq biasedly.
GW34-e0151
Yang Shen1,2, Hang Zhu1,2, Hao Zhou2,3
1Department of Cardiology, School of Medicine, South China University of Technology, Guangzhou 510006, China
2Department of Cardiology, The Sixth Medical Center of PLA General Hospital of Beijing, Beijing 100048, China
3Department of Vascular Medicine, Peking University Shougang Hospital, Beijing 100144, China
OBJECTIVES Dapagliflozin (DAPA) confers significant protection against heart and kidney diseases. However, whether DAPA can alleviate type 4 cardiorenal syndrome (CRS-4)-related cardiomyopathy remains unclear. Cardiomyocyte resistance to chronic damage is enhanced by both aerobic glycolysis stimulated by pyruvate kinase isozyme M2 (PKM2) and FUN14 domain containing 1 (FUNDC1)-dependent mitophagy. Thus, we tested the hypothesis that DAPA attenuates CRS-4-related myocardial damage through PKM2 induction and FUNDC1-related mitophagy.
METHODS Cardiomyocyte-specific PKM2 knockout (PKM2 CKO ) and FUNDC1 knockout (FUNDC1 CKO ) mice were subjected to subtotal (5/6) nephrectomy to establish a CRS-4 model in vivo. To simulate CRS-4 in vitro, HL-1 cardiomyocytes were treated with uremic sera collected from DAPA-treated mice 12 weeks after nephrectomy. Western blots, co-IP, molecular docking, siRNA knockdown technology, immunofluorescence, and ELISA were performed.
RESULTS Our results demonstrated that DAPA treatment enhanced PKM2 expression and improved myocardial function and structure in vivo, and this effect was abrogated by PKM2 knockdown. Moreover, a significant improvement in mitochondrial function was observed in HL-1 cells exposed to sera from DAPA-treated mice, as featured by increased ATP production, decreased mtROS production, improved mitochondrial membrane potential, preserved mitochondrial complex activity, and reduced mitochondrial apoptosis. DAPA restored FUNDC1-dependent mitophagy through post-transcriptional dephosphorylation in a manner dependent on PKM2 whereas ablation of FUNDC1 abolished the defensive actions of DAPA on myocardium and mitochondria under CRS-4. Co-IP and molecular docking assays indicated that PKM2 directly interacted with protein phosphatase 1 (PP1) and FUNDC1, leading to PP1-mediated FUNDC1 dephosphorylation.
CONCLUSIONS These results suggest that DAPA attenuates CRS-4-related cardiomyopathy through activating the PKM2/PP1/FUNDC1-mitophagy pathway.
GW34-e0189
Siting Hong1, Jiaoyan Li2, Dongxue Liu1, Xin Zhang1, Yingchun Luo1, Zeng Wang1, Yue Zhao1, Zengxiang Dong1, Zhaoguang Liang1
1The First Affiliated Hospital of Harbin Medical University
2Xi’an People’s Hospital (Xi’an Fourth Hospital)
OBJECTIVES The current studies demonstrate that gut microbiota and its metabolites, especially Trimethylamine N-oxide (TMAO), play an important role in the onset and progress of cardiovascular diseases (CVDs). (-)-Epicatechin (EPI), the major polyphenolic compound present in green tea, has shown cardioprotective effect. The purpose of this study was to investigate whether EPI could inhibit TMAO-induced cardiac fibrosis.
METHODS We established cardiac fibrosis mice model with TMAO daily intraperitoneal (i.p.) injection for 14 days. Male C57BL/6 mice were randomly separated into 4 groups. Intragastric administration of EPI or vehicle was conducted in mice for 21 days. The cardiac function was performed by the ultrasonic instrument. Histological analysis of mice’s hearts was accessed by H&E and Masson staining. In vitro, cardiac fibroblasts were treated by TMAO with or without EPI. The protein level of fibrosis markers were quantified by western blot.
RESULTS In vivo, cardiac ultrasound results showed that TMAO could damage the cardiac function of mice and lead to cardiac hypertrophy. Pathological staining showed that TMAO induced myocardial hypertrophy and fibrosis in mice. TMAO upregulated the expression of MMP-2, MMP-9 and α-SMA. EPI treatment significantly improved the cardiac function of mice, inhibited the myocardial fibrosis induced by TMAO and reduced the protein levels of MMP-2, MMP-9 and α-SMA. In vitro, EPI blocked TMAO-induced transformation of cardiac fibroblasts into myofibroblasts. TMAO also elevated MMP-2, MMP-9 and α-SMA expression, but decreased after treatment with EPI.
CONCLUSIONS In summary, EPI suppresses TMAO-induced cardiac fibrosis.
GW34-e0200
Xiang Liu1, Yijia Shao2, Jimei Chen1
1Department of Cardiac Surgery, Guangdong Cardiovascular Institute, Guangdong Provincial People’s Hospital
2Department of Geriatrics, The First Affiliated Hospital, Sun Yat-sen University
OBJECTIVES There is increasing evidence that microbiota has an important effect on heart development. Alternative polyadenylation (APA) is a key mechanism for the regulation of gene expression and has shown to be involved in postnatal heart development. However, it remains unclear whether microbiota can affect postnatal heart development by regulating APA.
METHODS Heart tissues from specific pathogen free (SPF) mice within the first 24 hours after birth (P1), 7-day-old SPF mice and 7-day-old germ-free (GF) mice were subjected to APA sequencing to obtain the genome-wide profiling of APA sites. Functional annotations were performed using Metascape and clusterProfiler package. Protein-protein interactions were constructed using STRING and then visualized on Cytoscape. The landscape of immune cells was assessed by CIBERSORT and the correlations between immune cells and key genes were analyzed via Hmisc package. The switching of 3′ untranslated region (3′ UTR) was verified by qPCR.
RESULTS Heart tissues isolated from specific pathogen free (SPF) mice within the first 24 hours after birth (P1), 7-day-old SPF mice and 7-day-old germ-free (GF) mice were subjected to APA sequencing to obtain the genome-wide profiling of APA sites. Here, we found that GF mice harbored evidently longer 3′ untranslated region (3′ UTR) lengths. And the 3′ UTR lengths of mitochondria-related genes, Rala, Timm13 and Uqcc3, were verified to be significantly altered. Furthermore, GF condition led to an obvious reduction in mitochondrial cristae density and a decreased level of Tomm20 in postnatal hearts. Additionally, Rala was found to be connected to Src, and further related to other differentially expressed genes (DEGs). The DEGs were significantly down-regulated in GF mice except for Thbs1 and Egr1. Finally, GF condition appeared to be associated with a lower infiltration of immune cells, and the levels of resting NK cells, Th17 cells, immature dendritic cells and plasma cells in GF mice were comparable to those in P1 mice. Moreover, these immune cells were shown to be significantly correlated with Rala and the related DEGs.
CONCLUSIONS Our findings suggested that microbiota plays a critical role in postnatal heart development by affecting APA switches and immune cell infiltrations.
GW34-e0211
Ziwei Zhu, Xiaowei Zhang
Lanzhou University Second Hospital
OBJECTIVES Ischemic heart failure (HF) is one of the leading causes of global morbidity and mortality, blocking the apoptotic cascade could help improve adverse outcomes of it. RNA-binding motif protein 25 (RBM25) is an RNA-binding protein related to apoptosis, however, its role remains unknown in ischemic HF. The main purpose of this study is to explore the mechanism of RBM25 in ischemic HF.
METHODS Establishing an ischemic HF model and oxygen-glucose deprivation (OGD) model. ELISA was performed to evaluate the BNP level in the ischemic HF model. Echocardiography and histological analysis were performed to assess cardiac function and infarct size. Proteins were quantitatively and locationally analyzed by western blotting and immunofluorescence. The morphological changes of endoplasmic reticulum (ER) were observed with ER-tracker.
RESULTS Cardiac function and myocardial injury were observed in ischemic HF rats. RBM25 was elevated in cardiomyocytes of hypoxia injury hearts and localized in nucleus both in vitro and in vivo. In addition, cell apoptosis was significantly increased when overexpressed RBM25. Moreover, ER stress stimulated upregulation of RBM25 and promoted cell apoptosis through the CHOP related pathway. Finally, inhibiting the expression of RBM25 could ameliorate the apoptosis and improve cardiac function through blocking the activation of CHOP signaling pathway.
CONCLUSIONS RBM25 is significantly up-regulated in ischemic HF rat heart and OGD model, which leads to apoptosis by modulating the ER stress through CHOP pathway. Knockdown of RBM25 could reverse apoptosis-mediated cardiac dysfunction. RBM25 may be a promising target for the treatment of ischemic HF.
GW34-e0216
Yan Xia, Zhangwei Chen, Juying Qian, Junbo Ge
Zhongshan Hospital, Fudan University
OBJECTIVES Mitochondria are highly abundant in the heart and are the “power houses” of the cell. The maintenance of mitochondrial morphology and structure is of critical importance for cardiac function. Mitochondrial dynamics are closely coordinated with cellular biological and metabolic processes. Mitochondria are double membrane-bound organelles, composed of an inner (IMM) and an outer mitochondrial membrane (OMM). The specific role of IMM protein during the progression of mitochondrial fission remains poorly understood. Mitochondrial fission is considered to play a key role in the pathogenesis of dilated cardiomyopathy (DCM) through mitochondrial quality control. However, the specific regulatory mechanism of mitochondrial fission in the development of doxorubicin (DOX)-induced cardiomyopathy remains unclear. The mitochondrial aspartate-glutamate carrier isoform 1 (AGC1, or aralar) is located in IMM. We report that AGC1 interacts with the fission protein dynamin-related protein 1 (Drp1) and the underlying the functional and molecular mechanism in DOX-induced cardiomyopathy.
METHODS This study examined the role of AGC1 in mitochondrial fission in doxorubicin-induced cardiomyopathy. Left ventricle (LV) samples were obtained from six end-stage DCM patients after heart transplantation in Zhongshan Hospital, Fudan University. We generated DOX-induced mice and Cardiac-specific Drp1 conditional knockout (CKO-Drp1) mice. Stable cell lines were prepared to conduct mechanistic studies.
RESULTS Results of co-immunoprecipitation mass spectrometry (CO-IP MS) anaylsis by based on heart tissue of DCM patients indicated that AGC1 expression was significantly upregulated DCM-induced injury and AGC1 level was closely correlated with mitochondrial morphogenesis and function. We showed that knockdown of AGC1 protected mice from DOX-induced cardiomyopathy by preventing mitochondrial fission, while overexpression of AGC1 in the mouse heart led to impairment of cardiac function. Mechanistically, AGC1 overexpression could upregulate Drp1 expression and contributed to subsequent excessive mitochondrial fission. Specifically, knockdown of AGC1 or use of Drp1-specific inhibitor Mdivi-1 alleviated cardiomyocyte apoptosis and inhibited impairment of mitochondrial function induced by DOX exposure.
CONCLUSIONS In summary, our data illustrate that AGC1, as a novel contributor to DCM, regulates cardiac function via Drp1-mediated mitochondrial fission. Our study indicates that inhibiting AGC1 or mitochondrial fission could be a potential therapeutic strategy and may provide cardioprotection.
GW34-e0232
Fengzhou Liu1,2, Xide Shi3, Chao Liu3, Jiangwei Chen4, Shiqiang Zhou3, Yajuan Li2, Junhui Xue1,2, Fei Li3
1Department of Aviation Medicine, Xijing Hospital, Fourth Military Medical University, Xi’an, China
2Aerospace Clinical Medical Center, School of Aerospace Medicine, Fourth Military Medical University, Xi’an, China
3Department of Cardiology, Xijing Hospital, Fourth Military Medical University, Xi’an, China
4State Key Laboratory of Military Stomatology & National Clinical Research Center for Oral Diseases, Department of Medical Rehabilitation, School of Stomatology, Fourth Military Medical University, Xi’an, China
OBJECTIVES Myocardial microvascular injury is the key event in early diabetic heart disease. The injury of myocardial microvascular endothelial cells (CMECs) is the main cause and trigger of myocardial microvascular disease. Mitochondrial calcium homeostasis plays an important role in maintaining the normal function, survival and death of endothelial cells. Considering that mitochondrial calcium uptake 1 (MICU1) is a key molecule in mitochondrial calcium regulation, this study aimed to investigate the role of MICU1 in CMECs and explore its underlying mechanisms.
METHODS To examine the role of endothelial MICU1 in DCM, we used endothelial-specific MICU1ecKO mice to establish a diabetic mouse model and evaluate the cardiac function. In addition, MICU1 overexpression was conducted by injecting adeno-associated virus 9 carrying MICU1 (AAV9-MICU1). Transcriptome sequencing technology was used to explore underlying molecular mechanisms.
RESULTS Here, we found that MICU1 expression is decreased in CMECs of diabetic mice. Moreover, we demonstrated that endothelial cell MICU1 knockout aggravated the levels of cardiac hypertrophy and interstitial myocardial fibrosis and led to a further reduction in left ventricular function in diabetic mice. Notably, we found that AAV9-MICU1 specifically upregulated the expression of MICU1 in CMECs of diabetic mice, which inhibited nitrification stress, inflammatory reaction, and apoptosis of the CMECs, ameliorated myocardial hypertrophy and fibrosis, and promoted cardiac function. Further mechanistic analysis suggested that MICU1 deficiency result in excessive mitochondrial calcium uptake and homeostasis imbalance which caused nitrification stress-induced endothelial damage and inflammation that disrupted myocardial microvascular endothelial barrier function and ultimately promoted DCM progression.
CONCLUSIONS Our findings demonstrate that MICU1 expression was downregulated in the CMECs of diabetic mice. Overexpression of endothelial MICU1 reduced nitrification stress induced apoptosis and inflammation by inhibiting mitochondrial calcium uptake, which improved myocardial microvascular function and inhibited DCM progression. Our findings suggest that endothelial MICU1 is a molecular intervention target for the potential treatment of DCM.
GW34-e0235
Qi Pan, Cheng Chen, Yuejin Yang, Guihao Chen
State Key Laboratory of Cardiovascular Disease, Department of Cardiology, Fuwai Hospital, National Center for Cardiovascular Diseases, Chinese Academy of Medical Science and Peking Union Medical College
OBJECTIVES The increasing burden of heart failure with preserved ejection (HFpEF) has become a global health problem. HFpEF is characterized by systematic inflammation, cardiac metabolic remodeling and fibrosis. Eosinophils act as an essential, but generally overlooked subgroup of white blood cells, which participant in cardiac fibrosis, as reported in several recent studies. Herein, we explored the role of eosinophils in a “two-hit” preclinical HFpEF model.
METHODS A novel preclinical model of HFpEF induced by normal diet 12-week treatment of high-fat diet (HFD) plus N[w]-nitro-L-arginine methyl ester (L-NAME, 0.5 g/L) and ΔdblGATA1 knock-out (Gata1 KO) C57BL/6N mice, which have complete ablation of the eosinophil lineage, were applied in this study. Blood eosinophil counts were conducted. Mice were divided into 4 groups (n=13–14 per group): wild type (WT) and normal diet (Group I), Gata1 KO and normal diet (Group II), WT and HFpEF model (Group III), and Gata1 KO and HFpEF model (Group IV). Evaluation of heart failure and hypertrophy were performed to examine the association of eosinophils and HFpEF.
RESULTS The level of circulating eosinophil didn’t alter between normal diet and HFpEF mice (normal diet v.s. HFpEF, 11.83±7.02 v.s. 10.87±4.60, P=0.81). Body weight (Group III v.s. IV, 33.46±1.50 v.s. 32.79±1.17 [g], P=0.72), systolic blood pressure (129.3±4.36 v.s. 132.6±4.15 [mmHg], P=0.27), exercise tolerance (running distance, 59.71±21.52 v.s. 63.30±25.35 [m], P=0.98), the ratio of E to e’ (E/e’, 42.34±4.10 v.s. 41.06±5.78, P=0.98), heart weight to tibial length (HW/TL, 6.82±0.18 v.s. 6.77±0.19 [mg/mm], P=0.86) and wet lung weight to dry lung weight (4.48±0.10 v.s. 4.43±0.11, P=0.76) were comparable between Group III and IV. Additionally, no alternation was observed in the levels of B-type natriuretic peptide (3.24±1.05 v.s. 3.57±1.80 [fold change], P=0.99) and transforming growth factor-β (20,691±2949 v.s. 19,049±3660 [pg/g tissue], P=0.81) between Group III and IV.
CONCLUSIONS Deficiency of eosinophils failed to alter the phenotype of HFpEF. The development of HFpEF is independent of eosinophils in terms of the functional, biochemical and histological results.
GW34-e0249
Lang Hu1, Feng Fu2, Yan Li1
1Department of Cardiology, Tangdu Hospital, Airforce Medical University
2Department of Physiology and Pathophysiology, Airforce Medical University
OBJECTIVES The heart derives its energy primarily from fatty acid oxidation, while excessive lipid supply may result in intra-myocardial lipotoxic damage and cardiac dysfunction. Nevertheless, the endogenous mechanism that determines the transition from adaptive lipid utilization to maladaptive lipotoxicity is still not well understood. The present study investigated the role of mitochondria-lipid droplet (LD) membrane contacts (MLC) and its underlying mechanisms in the transition.
METHODS Mice were fed continuous high-fat diet (HFD). Cardiac function and intra-myocardial lipid accumulation were examined. Multiple approaches including fluorescent imaging, lipidomics, fatty acids pulse-chase assay, proximity ligation assay, co-immunoprecipitation, mutagenesis and liquid chromatography tandem mass spectrometry (LC-MS/MS) analysis were used to elucidate the underlying mechanisms.
RESULTS Tight MLC was observed in the hearts of mice fed HFD for 5 weeks, accompanied by adaptive lipid usage. As lipid-overload was prolonged, reduced MLC and increased LD number were detected in the hearts of mice fed HFD for 20 weeks and patients with obesity (BMI≥30 kg/m2), and were associated with maladaptive lipotoxicity. Mechanistically, mitochondria-localized Mitofusin2 (Mfn2) directly binds to LD-localized heat shock cognate protein 70 (Hsc70) via its 649–692 aa domain, tethering mitochondria to LDs and facilitating fatty acids (FAs) transfer from LDs to mitochondria for β-oxidation. Excessive lipid-induced Mfn2 reduction inhibits MLC and impedes FAs transfer, leading to lipid accumulation and cardiac damage. Restoration of Mfn2 reconstituted MLC and relieved myocardial lipotoxicity under lipid overload in-vivo and in-vitro. Hsc70 knockdown largely blocked the above effects of Mfn2. Moreover, Mfn2 inhibition was induced in the ubiquitin-proteasome pathway following Mfn2 acetylation at the K243 site that was triggered by lipid overload.
CONCLUSIONS Mfn2/Hsc70 complex-mediated MLC is a novel regulator of cardiac lipid metabolism. Lipid overload induces Mfn2/Hsc70 complex disassociation, leading to intra-myocardial lipid accumulation and maladaptive lipotoxicity. Therefore, targeting MLC may help in therapeutic interventions in cardiac lipotoxicity.
GW34-e0263
Jing Geng, Xiaoliang Zhang, Dong Guo, Ying Wang, Lang Hu, Yan Li
Department of Cardiology, Tangdu Hospital, Airforce Medical University
OBJECTIVES A high-calories diet (HCD) combined with lack of physical exercise is the major cause of obesity, leading to lipid metabolic disorder and cardiac lipotoxicity. Physical exercise has long been recognized as an effective strategy for preventing diet-induced obesity and improving cardiovascular health. However, it is still unclear whether exercise training would protect against cardiac lipotoxicity in a large number of individuals with HCD.
METHODS Wild-type female C57BL/6J mice were fed ad libitum either a normal diet (ND, 10% calories from fat) or a high-fat, high calories diet (HFCD, 60% calories from fat). Meanwhile, ND and HFCD-fed mice were subjected to sedentary or three different-intensity exercise. Body weight, lipid accumulation, metabolic-related indicators and cardiac function were determined. Lipidomics, proteomics, mitochondrial function analysis was also performed to evaluate cardiac lipid metabolism.
RESULTS HFCD-feeding led to a significant elevation of body weight, blood glucose and serum lipid level. Also, continues HFCD-feeding induced both systolic and diastolic function impairment. As expected, all of the three different-intensity exercise prevented the development of obesity and whole-body metabolic disorders, as indicated by decreased blood glucose, serum lipid, body weight and lipid content in liver and white-adipose tissue. Low-intensity exercise (LIE) showed cardiac protective effect on HFCD-fed mice, as evidenced by improved cardiac diastolic function. However, moderate- or high-intensity exercise (MIE or HIE) failed to protect against HFCD-induced cardiac dysfunction but in contrast deteriorated the development of cardiac dysfunction and pathological hypertrophy. Mechanistically, MIE promoted the activation of lipid intake-related genes (mainly CD36) in HFCD-fed mice, leading to more severe intra-myocardium lipid accumulation. As a result, accumulated toxic lipid further impaired mitochondrial respiratory function and impeded lipid metabolism in heart of HFCD-fed mice.
CONCLUSIONS Our work firstly reported that moderate-intensity exercise, which was previously considered to be beneficial to cardiovascular system, exacerbated HCD-induced cardiac lipotoxicity and dysfunction. The underlying mechanism was associated with excessive cardiac lipid uptake and toxic lipid-induced mitochondrial damage. Our results deepen current insights into the cardiovascular effect of physical exercise on a specified population. Considering the large population with high-calories diet around the world, this study also provided guidance for individuals with HCD to establish rational exercise strategies to avoid the potential negative effects.
GW34-e0268
Weili Li, Yawen Zhang, Guanjing Ling, Nannan Tan, Yan Wei, Jinchi Jiang, Wei Li, Xiaoping Wang, Jing Cao
Beijing University of Chinese Medicine
OBJECTIVES Doxorubicin (DOX), a broad-spectrum chemotherapy drug, has life-threatening cardiotoxicity. Therefore, searching cardioprotective drugs for DOX-induced cardiotoxicity (DIC) is urgently needed. The results of previous high-throughput screening showed that aucubin (AUC) had significant protection on DOX-treated zebrafish. This study aimed to investigate the cardioprotective effects of AUC and the specific mechanism by which AUC alleviates DIC, and to provide an alternative for the treatment of DIC.
METHODS A DIC mice model via tail vein injection with DOX (5 mg⋅kg−1) once weekly for 4 weeks and a DOX-induced H9C2 cell injury model with DOX (1 μmol/L) were established. In vivo, the mice were divided into saline-injected group, DOX-injected group, DOX+low and high dose of AUC-treated group, and DOX+enalapril (ENA)-treated group (Positive control). Echocardiography, histological examination and CK-MB and LDH detection were applied for assessment of protective effects of AUC on cardiac structure and function. TUNEL, DHE, 8-OHdG staining and serum MDA were used to detect myocardial apoptosis and oxidative stress levels. Whereafter, western blot and transmission electron microscope were utilized to evaluate the regulation of autophagy proteins (Beclin-1, LC3 and p62) and autophagy flow by AUC. In vitro, after siRNA of Hipk2 and Nrf2 were combined with AUC, a variety of methods were used, including GFP-mRFP-LC3 double-labeled adenovirus monitoring autophagy flow, DCFH-DA staining to detect ROS, 8-OHdG immunofluorescence staining to quantify DNA damage, Hoechst staining to detect apoptosis rate, and western blot detected autophagy and apoptosis-related proteins, which jointly verified AUC regulating autophagy, oxidative stress and apoptosis through Hipk2 or Nrf2. Finally, siRNA and plasmid overexpression were performed on Hipk2 and Nrf2 respectively, and the protein expression and luciferase intensity of Nrf2 and Hipk2 were detected accordingly, so as to further interpret the cross-talk between Hipk2 and Nrf2.
RESULTS AUC exerted cardioprotection manifested as enhanced cardiac function, and reduced structural damage and myocardial injury markers CK-MB and LDH. In addition, AUC significantly alleviated ROS and MDA production, DNA damage and apoptosis in DIC mice. Mechanically, AUC promoted the expression of Beclin1 and the formation of autophagosomes, and reduced the accumulation of p62 and LC3 proteins in DIC mice. In vitro, AUC increased Beclin-1 expression and reduced DOX-induced accumulation of LC3 and P62 proteins, thus promoting the formation of autophagosomes and reducing the accumulation of autophagolysosomes. In addition, AUC reduced DOX-induced ROS production, DNA oxidative damage, and apoptosis. Interestingly, knockdown of Nrf2 or Hipk2 reversed the above effects on autophagy, oxidative stress and apoptosis. More importantly, there is a cross-talk mechanism between Hipk2 and Nrf2. When Hipk2 was inhibited or overexpressed, the protein expression and luciferase intensity of Nrf2 decreased or increased correspondingly; When Nrf2 was inhibited or overexpressed, the protein expression and luciferase intensity of Hipk2 also decreased or increased correspondingly.
CONCLUSIONS The results partially revealed that AUC exerts cardioprotective effects against DIC through Nrf2-Hipk2 signal crosstalk to activate autophagy and thus alleviate oxidative stress damage and apoptosis induced by DOX.
GW34-e0279
Chengcheng Zhao1,2, Xiangrui Jiang3, Liyuan Peng1,2, Yan Zhang3, Huihui Li1,2, Qiumeng Zhang3, Yinhui Wang1,2, Feipu Yang3, Junfang Wu1,2, Zheng Wen1,2, Jingshan Shen3, Chen Chen1,2, Dao Wen Wang1,2
1Division of Cardiology, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China
2Hubei Key Laboratory of Genetics and Molecular Mechanisms of Cardiological Disorders, Wuhan, China
3Shanghai Institute of Materia Medica, Chinese Academy of Sciences, 555 Zuchongzhi Road, Shanghai 201203, China
OBJECTIVES Epoxyeicosatrienoic acids (EETs), which exert multiple endogenous protective effects, are easily hydrolyzed into less active dihydroxyeicosatrienoic acids (DHETs) by soluble epoxide hydrolase (sEH). However, relative commercial drugs are not in clinic use yet.
METHODS Firstly, the plasma concentration of EETs and DHETs of 316 patients with heart failure (HF) were detected by liquid chromatography-tandem mass spectrometry. Then, transverse aortic constriction (TAC)-induced HF was introduced in cardiomyocyte-specific Ephx2−/− mice. Moreover, Western blot, real-time PCR, luciferase reporter, ChIP assays were employed to explore the underlying mechanism of the reason why sEH was upregulated. Finally, multiple sEH inhibitors were designed, synthesized, and validated in vitro and in vivo.
RESULTS The ratios of DHETs and EETs were increased in the plasma from patients with HF. Meanwhile, the expression of sEH was upregulated in the heart of patients and mice with HF, especially in cardiomyocytes. Cardiomyocyte-specific Ephx2−/− mice ameliorated cardiac dysfunction induced by TAC. Consistently, Ephx2 knockdown protected AngII-treated cardiomyocytes against injury and hypertrophy via increasing EETs in vitro. Mechanistically, AngII could enhance the expression of transcript factor Krüppel-like factor 15 (KLF15), which in turn upregulated sEH. Importantly, glimepiride was identified as a novel sEH inhibitor, which benefited from the elevated EETs during HF.
CONCLUSIONS Glimepiride, as a sEH inhibitor, could attenuate heart failure by increasing EETs concentration.
GW34-e0305
Haizhe Ji, Yinong Jiang
The First Affiliated Hospital of Dalian Medical University
OBJECTIVES Dual specificity phosphatase 1 (DUSP1) is regarded as an anti-inflammatory factor in cardiovascular disorders. Mitophagy removes damaged mitochondria and thus promotes mitochondrial regeneration. We investigated whether DUSP1 could attenuate inflammation-induced cardiomyopathy by improving mitophagy.
METHODS Lipopolysaccharide was used to induce septic cardiomyopathy in wild-type (WT) and DUSP1 transgenic (DUSP1 TG ) mice.
RESULTS Echocardiography revealed that lipopolysaccharide impaired heart function by reducing the cardiac systolic and diastolic capacities of WT mice. Freshly isolated single cardiomyocytes from lipopolysaccharide-treated WT mice also exhibited reduced contractile/relaxation parameters. However, DUSP1 overexpression not only maintained the mechanical properties of cardiomyocytes, but also improved heart performance. Lipopolysaccharide upregulated myocardial inflammatory gene transcription and adhesive factor expression, which increased myocardial neutrophil accumulation and cardiomyocyte apoptosis in WT mice. DUSP1 overexpression inhibited the inflammatory response and therefore promoted cardiomyocyte survival. Lipopolysaccharide disrupted mitochondrial respiration and metabolism in WT cardiomyocytes, but DUSP1 overexpression restored mitochondrial metabolism, maintained the mitochondrial membrane potential and inhibited mitochondrial reactive oxygen species production, possibly by increasing FUN14 domain-containing 1 (FUNDC1)-dependent mitophagy. Silencing of FUNDC1 abolished the protective effects of DUSP1 overexpression on cardiomyocytes and their mitochondria following lipopolysaccharide treatment.
CONCLUSIONS These results demonstrated that DUSP1 is a novel anti-inflammatory factor that protects against septic cardiomyopathy by improving FUNDC1-induced mitophagy.
GW34-e0306
Haizhe Ji, Yinong Jiang
The First Affiliated Hospital of Dalian Medical University
OBJECTIVES Cardiorenal syndrome type-3 (CRS-3) is an abrupt worsening of cardiac function secondary to acute kidney injury. Mitochondrial dysfunction is a key pathological mechanism of CRS-3, and empagliflozin can improve mitochondrial biology by promoting mitophagy. Here, we assessed the effects of empagliflozin on mitochondrial quality surveillance in a mouse model of CRS-3.
METHODS Cardiomyocyte-specific FUNDC1-knockout (FUNDC1 CKO ) mice were subjected to CRS-3 prior to assessment of mitochondrial homeostasis in the presence or absence of empagliflozin.
RESULTS CRS-3 model mice exhibited lower heart function, increased inflammatory responses and exacerbated myocardial oxidative stress than sham-operated controls; however, empagliflozin attenuated these alterations. Empagliflozin stabilized the mitochondrial membrane potential, suppressed mitochondrial reactive oxygen species production, increased mitochondrial respiratory complex activity and restored the oxygen consumption rate in cardiomyocytes from CRS-3 model mice. Empagliflozin also normalized the mitochondrial morphology, mitochondrial dynamics and mitochondrial permeability transition pore opening rate in cardiomyocytes. Cardiomyocyte-specific ablation of FUN14 domain-containing protein 1 (FUNDC1) in mice abolished the protective effects of empagliflozin on mitochondrial homeostasis and myocardial performance. Empagliflozin activated β-catenin and promoted its nuclear retention, thus increasing FUNDC1-induced mitophagy in heart tissues; however, a β-catenin inhibitor reversed these effects.
CONCLUSIONS In summary, empagliflozin activated Wnt/β-catenin to stimulate FUNDC1-dependent mitochondrial quality surveillance, ultimately improving mitochondrial function and cardiac performance during CRS-3. Thus, empagliflozin could be considered for the clinical management of heart function following acute kidney injury.
GW34-e0314
Naixin Wang, Zhonghua Tong, Xiaoqi Wang, Jian Wu, Maomao Zhang
Department of Cardiology, The Second Affiliated Hospital of Harbin Medical University
OBJECTIVES The early activation of reparative signals in monocyte-macrophages is essential for the timely restoration of immune homeostasis and the initiation of the repair process post-MI. However, it remains unknown whether bone marrow (BM) and circulating monocytes are pre-educated to initiate the reparative transcriptional response in the early stage of MI. Furthermore, the mechanisms and molecular cues involved in the initiation of reparative gene activation in peripheral monocytes before cardiac recruitment remain to be fully elucidated. A newly identified epigenetic modification, histone lactylation, induces reparative gene transcription during M1 macrophage polarization to promote immune homeostasis. However, the pattern and involvement of histone lactylation in the early remote activation of reparative response post-MI remain unclear.
METHODS Single-cell RNA sequencing (scRNA-seq) was performed on BM and circulating monocytes to detect the early and remote transcriptional response prior to cardiac recruitment post MI. Western blot and immunofluorescence staining were used to explored the dynamic patterns of histone lactylation levels in monocyte-macrophages post MI. Cleavage under targets and tagmentation (CUT&Tag) and paired RNA sequencing (RNA-seq) were performed to identify potential downstream targets of histone lactylation. CHIP-qPCR and qRT-PCR verified modification and expression level of the target genes. Flow cytometry, EdU labeling, transwell, scratch/wound migration, and tube formation assays assessed the reparative activity of macrophages regulated by histone lactylation. Murine MI models induced by LAD ligation were treated with sodium lactate or a lactate dehydrogenase inhibitor, FX-11, thus manipulate the level of histone lactylation to assess the effects of histone lactylation on the ischemic cardiac immune environment.
RESULTS ScRNA-seq data confirmed the early remote activation of reparative signals in peripheral monocytes post-MI. Meanwhile, we observed increases of histone lactylation levels in peripheral monocytes and cardiac macrophages, including the previously identified histone H3K18 lactylation (H3K18la) early post MI. Reparative genes Lrg1, Vegf-a, and IL-10 were identified as target genes of histone lactylation post-MI. Then we demonstrated that histone lactylation regulates the anti-inflammatory and pro-angiogenic dual activities of monocyte-macrophages by facilitating reparative gene transcription in vitro, confirmed that increased histone lactylation favors a reparative environment and improves cardiac function post-MI in vivo. We also explored the positive role of monocyte histone lactylation in reperfused MI. Mechanistically, we revealed that endogenous glycolysis re-programming in circulating monocytes post-MI and MCT1-mediated extracellular lactate transportation contributed to histone lactylation. Finally, we revealed that GCN5 served as a writer for lactylation, and then confirmed the catalytic effect of IL-1β-dependent GCN5 recruitment on H3K18la and elucidated its potential role as an upstream regulatory element in the regulation of monocyte histone lactylation and downstream reparative gene expression post-MI.
CONCLUSIONS In summary, we demonstrated that histone lactylation favors a reparative environment by promoting reparative gene activation in monocytes early and remotly post-MI, and highlighted that the metabolome-epigenome cascade may provide clues to the endogenous protective effect and underlying mechanism of histone lactylation in MI.
GW34-e0316
Yinhui Wang, Jia Cheng, Ling Zhou, Chunxia Zhao
Department of Internal Medicine, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology
OBJECTIVES Aberrant amino acid metabolism is implicated in cardiac hypertrophy, whereas the involvement of tryptophan metabolism in pathological cardiac hypertrophy remains elusive. Herein, we aimed to investigate the effect and potential mechanism of indoleamine 2,3-dioxygenase (IDO1) and its metabolite kynurenine on pathological cardiac hypertrophy.
METHODS Transverse aortic constriction (TAC) was performed to induce cardiac hypertrophy in IDO1-KO mice and AAV9-cTNT-shIDO1 mice. Liquid chromatography-mass spectrometry (LC-MS) was used to detect the metabolites of tryptophan–kynurenine pathway. Subcutaneous injection was used for delivery of exogenous Kyn and INCB. Chromatin Immunoprecipitation (ChIP) assay and dual luciferase assay were employed to validate the binding of protein and DNA.
RESULTS IDO1 expression was upregulated in both human and murine hypertrophic myocardium, alongside with increased cardiac IDO1 activity and kynurenine (Kyn) content in TAC-induced mice determined by liquid chromatography-mass spectrometry (LC-MS) analysis. Myocardial remodeling and heart function were significantly improved in IDO1-deficient TAC-induced mice, but were greatly exacerbated with subcutaneous kynurenine administration. IDO1 inhibition or kynurenine addition confirmed the alleviation or aggravation of hypertrophy in vitro, respectively, in cardiomyocyte treated with isoprenaline (ISO). Mechanistically, IDO1 and metabolite kynurenine contributed to pathological hypertrophy via the aryl hydrocarbon receptor (AhR)-GATA4 axis.
CONCLUSIONS These findings suggested that IDO1 deficiency and consequent kynurenine insufficiency can protect against pathological cardiac hypertrophy by decreasing GATA4 expression in an aryl hydrocarbon receptor (AhR)-dependent manner, and IDO1 has the potential to be a therapeutic target for pathological cardiac hypertrophy.
GW34-e0318
Shiyu Gong, Ming Zhai, Jiayun Shi, Guanye Yu, Zhijun Lei, Wenhui Peng
Department of Cardiology, Shanghai Tenth People’s Hospital, Tongji University, School of Medicine, Shanghai, China
OBJECTIVES Efferocytosis and metabolic reprogramming of macrophages are necessary for myocardial infarction (MI) reparation. Triggering Receptor Expressed on Myeloid Cells 2 (TREM2) is a transmembrane immune receptor that improves macrophage efferocytosis. However, the potential function of TREM2 and its molecular mechanisms remain unknown in MI. Therefore, we aim to uncover the effect of TREM2+ macrophage and explore the link between efferocytosis and metabolism in MI.
METHODS LysMcre, TREM2flox/flox; LysMcre, B6-G/R; CCR2cre-EGFP, TREM2flox/flox transgenic mice and cardiac macrophages overexpressing TREM2 by adenovirus were used to determine the function of TREM2 in MI. Efferocytosis and metabolic functions were examined using flow cytometry, immunofluorescence, and quantitative polymerase chain reaction. Targeted Metabolomics (LC-MS) was used to detect concentrations of metabolites. RNA-seq was used to screen the potential downstream pathways of TREM2. Protein and molecular docking were predicted with AutoDock Vina software.
RESULTS Macrophage-specific TREM2 knockout mice (Mac-TREM2KO) showed worsened cardiac function and impaired post-MI repair. Deletion of TREM2 induced macrophages to transform into a pro-inflammatory phenotype of CCR2+MHCIIhigh. On the contrary, macrophages expressing TREM2 had decreased SLC25A53 transcription through the SYK-SMAD4 signaling pathway, which impaired NAD+ transport into mitochondria, thereby restricting the tricarboxylic acid cycle and increasing itaconate production in anoxia condition. In vitro, itaconate secreted by TREM2+ macrophage inhibited cardiomyocyte apoptosis and increased fibroblast proliferation.
CONCLUSIONS Our data described a novel role of myeloid-derived macrophage-specific TREM2 in MI, linking efferocytosis to immune metabolism during cardiac repair by transporting NAD+ into mitochondria, mainly through mitochondria protein SLC25A53. Moreover, the metabolism by-product itaconate could alleviate MI injury.
GW34-e0332
Toshihide Hamabe-Horiike1,2,3, Kana Shimizu1,2, Yoichi Sunagawa1,2,3, Yuto Kawase1, Yasufumi Katanasaka1,2,3, Satoshi Shimizu1, Koji Hasegawa1,2, Tatsuya Morimoto1,2,3
1Division of Molecular Medicine, School of Pharmaceutical Sciences, University of Shizuoka
2Division of Translational Research, Clinical Research Institute, Kyoto Medical Center, National Hospital Organization
3Shizuoka General Hospital, Shizuoka
OBJECTIVES Heart failure is a serious public health problem. As the number of people suffering from the disease is expected to continue increasing over the coming decades, heart failure is one of the most significant health problems worldwide. Neurohormonal antagonists, including β blockers and renin-angiotensin system inhibitors, are the established standards for heart failure therapy based on clinical studies demonstrating that these drugs suppress cardiac remodeling and improve prognosis. However, as the prognosis remains poor even with the use of these drugs, new heart failure therapies are urgently needed. Drug repositioning has recently emerged as a strategy for developing new treatments at low cost. In this study, we focused on a drug repositioning strategy for heart failure therapy using a library of approved drugs to screen for compounds that suppress phenylephrine (PE)-induced hypertrophy in primary cultured cardiomyocytes. The antiplatelet drug sarpogrelate, a selective serotonin-2A (5-HT2A) receptor antagonist, was identified as capable of suppressing PE-induced cardiomyocyte hypertrophy. However, it was unclear whether sarpogrelate suppresses cardiac dysfunction in the pressure overload-induced mice model.
METHODS To investigate the effects of sarpogrelate on cardiomyocyte hypertrophy, primary cultured cardiomyocytes prepared from neonatal rats were pretreated with 1 μM sarpogrelate and then stimulated with PE, angiotensin II (Ang II), or endothelin1 (ET-1) for 48 h. Immunofluorescence staining and qPCR analysis were performed. To identify the phosphorylation of ERK and GATA4, Western blot analysis was performed. Next, to investigate the effects of sarpogrelate on systolic function, eight-week-old mice were subjected to transaortic constriction (TAC) surgery. One day after the surgery, mice were randomly assigned to the oral treatment with sarpogrelate (5 mg/kg/day) or vehicle for 8 weeks. Echocardiography, qPCR, and histological qPCR analyses were performed. To evaluate the phosphorylation of ERK and GATA4, Western blot analysis was performed.
RESULTS Immunofluorescence staining showed that sarpogrelate suppressed the cardiomyocyte hypertrophy induced by each of the stimuli. PE-induced increase in hypertrophic-related genes mRNA levels was inhibited by sarpogrelate. Western blotting analysis revealed that sarpogrelate suppressed PE-induced phosphorylation of ERK1/2 and GATA4. Echocardiography showed that 5 mg/kg of sarpogrelate suppressed TAC-induced cardiac hypertrophy and systolic dysfunction. qPCR analysis showed that TAC-induced increase in hypertrophic-related genes mRNA levels was inhibited by sarpogrelate treatment. Wheat germ agglutinin staining showed that sarpogrelate suppressed a TAC-induced increase in the cross-sectional area of the left ventricle, and Masson trichrome staining showed that sarpogrelate also suppressed TAC-induced perivascular fibrosis. Western blotting revealed that sarpogrelate suppressed TAC-induced phosphorylation of ERK1/2 and GATA4.
CONCLUSIONS These results indicate that sarpogrelate suppresses the development of heart failure at least, in part, by inhibiting the ERK1/2–GATA4 signaling pathway. Further study is needed to determine whether the inhibition of ERK1/2, including the inhibition caused by sarpogrelate, may be effective for heart failure therapy.
GW34-e0336
Minghui Li, Zhiwei Chen, Hui Shi, Ruizhen Chen
Department of Cardiology, Zhongshan Hospital, Fudan University
OBJECTIVES The mechanism of bradycardia arrhythmia in viral myocarditis is still unclear. This study aims to investigate this issue.
METHODS Balb/c mice were intraperitoneally injected with CoxsackievirusB3 to establish viral myocarditis model. Survival curves of mice were plotted. Electrocardiograms were recorded to observe changes in heart rhythm. Ten days after infection, the hearts of mice were collected, and some tissues were fixed for histology observation. Some were frozen and stored at −80°C for Western blot analysis of calpain1, MCU, TBX18, and HCN4 protein. In vitro models were established by directly infecting neonate rat cardiomyocytes with CVB3 and observing changes in MCU expression with calpain inhibitor PD150606 treatment.
RESULTS Survival rate of infected mice decreased significantly (P<0.05). Histological examination showed inflammatory cell infiltration in the myocardial tissue of surviving mice at the end of the observation period, while no obvious inflammatory cell infiltration was observed in the myocardium of dead mice during the observation period, suggesting a potential death cause of malignant arrhythmia. Electrocardiogram recordings showed that the heart rate of infected mice decreased significantly (P<0.05). Molecular biology analysis showed that calpain1 expression was upregulated and TBX18 and HCN4 expression were downregulated in infected myocardial tissue (Ps<0.05). In vitro calpain1 inhibition downregulated MCU expression (P<0.05), indicating that calpain1 mediated the MCU upregulation in viral infection and the sinus node autonomic regulation genes alterations.
CONCLUSIONS Calpain1 is activated in infected myocardial tissue, mediating the upregulation of MCU, leading to bradycardia arrhythmia by affecting TBX18 and HCN4 expression.
GW34-e0345
Ayaka Ishima1, Yoichi Sunagawa1,2,3, Masafumi Funamoto1,2, Ayumi Katayama1, Toshihide Hamabe1,2, Yasufumi Katanasaka1,2,3, Ryota Hosomi4, Koji Hasegawa1,2, Tatsuya Morimoto1,2,3
1Division of Molecular Medicine/School of Pharmaceutical Sciences/University of Shizuoka
2Division of Translational Research/Clinical Research Institute/Kyoto Medical Center/National Hospital Organization
3Shizuoka General Hospital
4Laboratory of Food and Nutritional Sciences/Faculty of Chemistry/Materials and Bioengineering/Kansai University
OBJECTIVES Hypertrophic stimuli activate cardiac transcription factors such as SRF, MEF-2, and a zinc finger protein GATA4 and modulate gene expression in cardiac myocytes. These changes finally lead to the development of heart failure in vivo. Acetylation is one of the critical mechanisms that activate these transcriptional factors and is mediated, in part, by intrinsic histone acetyltransferase (HAT) such as a transcriptional coactivator p300. Although many studies have shown a cardioprotective effect of eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), omega-3 unsaturated fatty acids, little is known about the effects of the EPA and DHA on cardiomyocyte hypertrophy and systolic dysfunction. In this study, we investigated the effects of the EPA and DHA on hypertrophic responses in cardiomyocytes and the development of heart failure in rats with myocardial infarction (MI).
METHODS First, Neonatal rat cultured cardiomyocytes were treated with EPA, or DHA. Palmitic acid (PA) or stearic acid (STA) were used as control. These cells were stimulated with saline or 30 μM phenylephrine (PE). After 48 hours, we measured cardiomyocyte surface area, hypertrophic responses gene transcriptions, and the ratio of acetylated histone H3. Next, we performed in vitro HAT assay using recombinant p300-HAT domain to determine the directly effect of EPA and DHA on p300-HAT activity. Finally, Rats with moderate MI (FS<40%) were randomly assigned to 3 groups; vehicle (saline), EPA, and DHA (1 g/kg). Oral administrations were repeated for six weeks from one week after the operation.
RESULTS Treatment with either DHA or EPA significantly inhibited the PE-induced hypertrophic responses such as myofibrillar organization, increase in cell size and mRNA expression of ANF and BNP. Moreover, DHA and EPA repressed the PE-induced acetylation of histone-3 in cardiomyocytes as much. The result of in vitro HAT assay revealed that 100 μM EPA and 100 μM DHA significantly inhibited p300-HAT activity. The echocardiographic analysis demonstrated that both EPA and DHA treatments prevented MI-induced systolic dysfunction and cardiac hypertrophy. Furthermore, EPA and DHA significantly suppressed the MI-induced increase in myocardial cell diameter, perivascular fibrosis, mRNA levels of hypertrophic and fibrotic markers, and acetylation of histone H3K9.
CONCLUSIONS These results suggested that both EPA and DHA suppressed MI-induced development of heart failure by inhibiting p300-HAT activity to the same extent. Thus, omega-3 unsaturated fatty acids EPA and DHA may exert as therapeutic agents to prevent the development of heart failure after MI.
GW34-e0357
Zhen Yao1, Yueli Liu2, Yong Zhang1, Yan Zhang1, Yuanchuan Chen1
1Cardiovascular Center, The Geriatric Hospital Of Hainan Province, Haikou 570000, China
2Pharmacy Institute of Hainan Medical College
OBJECTIVES To study frequency distribution of polymorphism in exon 13 of LDL-R gene in health people of Li and Han nationality in Hainan province and relationship between genotype and blood lipid, in order to understand the abnormal condition of lipid metabolism among Li people.
METHODS Five hundred and eighteen Li people and 592 Han people were recruited during this study. Fasting blood was drawn to evaluate levels of triglyceride (TG), cholesterol (TC), high density lipoprotein (HDL-C), low density of lipoprotein (LDL-C). Polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method was used to determine AvaII Polymorphism of low density lipoprotein receptor gene.
RESULTS The results showed that A−A−, A+A− and A+A+ genotype frequency of low density lipoprotein receptor gene between Li and Han groups are [70.7 (366/518) vs 70.3% (416/592); 24.7 (128/518) vs 27% (160/592); 4.6 (24/518) vs 2.7% (16/592); x2=0.785, P=0.579] respectively, there is no significant difference between them. Comparisons of blood lipid levels in Li and Han group of TG, TC, HDL, and LDL are [(1.32±0.05) mmol/L vs (1.49±0.08) mmol/L, P=0.08]; [(5.03±0.07) mmol/L vs (5.16±0.06) mmol/L, P=0.21]; [(1.57±0.02) mmol/L vs (1.33±0.02) mmol/L, P<0.001]; [(2.72±0.06) mmol/L vs (3.13±0.05) mmol/L, P<0.001] respectively.
CONCLUSIONS In conclusion, Ava II polymorphism of LDL-R gene between Li and Han group is basically identical, and LDL-R gene polymorphism of the healthy of Li and Han group is not associated with blood lipid level. Level of HDL in Li group is significantly higher than that in Han group, and level of LDL in Li group is significantly lower than that in Han group.
GW34-e0361
Liming Chen, Shijun Wang, Junbo Ge, Yunzeng Zou
Department of Cardiology, Shanghai Institute of Cardiovascular Diseases, Zhongshan Hospital and Institute of Biomedical Sciences, Fudan University
OBJECTIVES EphrinB2 (erythropoietin-producing hepatoma interactor B2) is a key member of the Eph/ephrin family, promoting angiogenesis, vasculogenesis, and lymphangiogenesis during embryonic development. However, the role of EphrinB2 in cardiac lymphangiogenesis following myocardial infarction (MI) and the potential molecular mechanism remains to be demonstrated. We sought to investigate whether and how EphrinB2 regulates cardiac lymphangiogenesis in the pathological remodeling process after MI.
METHODS Since homozygous Efnb2 knockout results in embryonic lethality, we utilized heterozygous Efnb2 knockout and adeno-associated virus of serotype 9 (AAV9)-mediated Efnb2 overexpression as loss- and gain-of-function approaches, respectively. We then investigated the effects of Efnb2 and its downstream mediators using RNA-sequencing analysis and in vitro experiments.
RESULTS We found that the expression of EphrinB2, which was rich in endothelium, was downregulated in the ischemic area of the heart two weeks after experimental MI. we subjected global Efnb2+/- mice and lymphatic-specific Efnb2 knockdown mice to either MI or sham operation. After two weeks, we observed that Efnb2 deficiency impaired cardiac lymphangiogenesis and led to adverse cardiac remodeling and ventricular dysfunction. Conversely, AAV9-mediated overexpression of EphrinB2 significantly improved cardiac function after MI. Intriguingly, overexpression of EphrinB2 also accelerated the resolution of inflammation, both macrophages and proinflammatory mediators. Moreover, the beneficial effects of EphrinB2 on inflammation and cardiac function were dependent on Lyve1. Additionally, EphrinB2 overexpression promoted the proliferation and migration of human lymphatic endothelial cells (hLECs) challenged by hypoxia. Our RNA-seq data showed that ISL1 appears to be a key factor in EphrinB2-driven lymphangiogenesis. Knockdown of ISL1 in hLECs greatly reduced the contributions of EphrinB2 in promoting cell proliferation and migration. Eventually, we confirmed the effects of ISL1 on lymphangiogenesis in vivo.
CONCLUSIONS Our findings uncovered a novel mechanism whereby EphrinB2 promotes cardiac lymphangiogenesis to improve myocardial remodeling and function following acute MI.
GW34-e0362
Shijun Wang, Liming Chen
Department of Cardiology, Shanghai Institute of Cardiovascular Diseases, Zhongshan Hospital and Institute of Biomedical Sciences, Fudan University
OBJECTIVES Cardiac dysfunction and pathological remodeling are serious complications of sepsis, which is a leading cause of death in sepsis. RCAN1 is a feedback regulator of cardiac hypertrophy. Here, we aim to investigate the role of RCAN1 in septic cardiomyopathy.
METHODS Mice were randomly divided into control-WT, control-RCAN1−/−, LPS-induced WT, and LPS-induced RCAN1−/− groups, some with Midiv-1 or KN93 treatment. The protein levels of RCAN1, p-ERK1/2, NFAT3, Drp1, p-Drp1, and p-CaMKII in mouse hearts or cultured cardiomyocytes were detected by Western blotting. Cardiac function was assessed by echocardiography. Cardiac hypertrophy and fibrosis were detected by H&E and Masson’s trichrome staining. Mitochondrial morphology was examined by transmission electron microscope. Serum level of LDH was detected by ELISA.
RESULTS RCAN1 was downregulated in septic mouse hearts and LPS-treated cardiomyocytes. RCAN1−/− mice presented severe impairment of cardiac function and elevated myocardial hypertrophy and fibrosis. The protein levels of NFAT3 and p-ERK1/2 were significantly increased in the heart tissues of RCAN1−/− mice. Further, RCAN1 deficiency aggravated sepsis-induced cardiac mitochondrial injury as indicated by increased ROS production, pathological fission, and the loss of mitochondrial membrane potential. Inhibition of fission with Mdivi-1 reversed LPS-induced cardiac hypertrophy, fibrosis, and dysfunction in RCAN1−/− mice. Moreover, RCAN1 depletion promoted mitochondrial translocation of CaMKII, which enhanced fission and septic hypertrophy, while inhibition of CaMKII with KN93 reduced excessive fission, and improved LPS-mediated cardiac remodeling and dysfunction in RCAN1−/− mice.
CONCLUSIONS Our findings demonstrated that RCAN1 deficiency aggravated the mitochondrial injury and septic dysfunction by activating CaMKII. RCAN1 serves as a novel therapeutic target for the treatment of sepsis-related cardiac cardiomyopathy.
GW34-e0374
Sonoka Iwashimizu1, Ono Masaya1, Sunagawa Yoichi1,2,3, Mochizuki Saho1, Inai Kyoko1, Funamoto Masafumi1,2, Shimizu Kana1,2, Katanasaka Yasufumi1,2,3, Hamabe Toshihide1,2,3, Hawke Philip4, Hasegawa Koji1,2, Morimoto Tatsuya1,2,3
1Division of Molecular Medicine, School of Pharmaceutical Sciences, University of Shizuoka
2Division of Translational Research, Clinical Research Institute, Kyoto Medical Center, National Hospital Organization
3Shizuoka General Hospital
4Laboratory of Scientific English, School of Pharmaceutical Sciences, University of Shizuoka
OBJECTIVES The prognosis of patients with malignant neoplasms has been improving due to advances in surgical treatment, radiation therapy, and chemotherapy with anticancer drugs. However, it has recently become clear that this longer period of patient survival has also resulted in increased exposure to the cardiotoxicity of anticancer drugs. The anthracycline anticancer drug doxorubicin (DOX) induces dose-dependent cardiotoxicity, ranging from occult changes in myocardial structure and function to severe cardiomyopathy and congestive heart failure. Since this cardiotoxicity affects the QOL and survival of cancer patients, a solution to this problem is urgently required. Anti-inflammatory activity. However, its effect on DOX-induced cardiotoxicity is still unknown. It has recently been reported that Chrysanthemum morifolium extract (CME) has antioxidant and anti-inflammatory activity. However, its effect on DOX-induced cardiotoxicity is still unknown.
METHODS To investigate whether CME suppressed DOX-induced cytotoxicity, an MTT assay was performed using H9C2 cells and primary cultured cardiomyocytes. Additionally, to examine the effect of CME on the anti-tumor activity of DOX, an MTT assay was performed using various cancer cells. Next, apoptosis was evaluated by TUNEL assay. The expression levels of p53, phosphorylated p53, cleaved-caspase3, and cleaved-caspase9 were also examined by western blotting. Eight-week-old C57BL/6J male mice were randomly assigned to treatment with either vehicle or 400 mg/kg CME orally for 15 days, beginning 2 days before intraperitoneal injection of 20 mg/kg DOX. After this treatment, survival assessments, echocardiography, and a TUNEL assay were performed.
RESULTS An MTT assay revealed that CME reduced DOX-induced cytotoxicity in H9C2 cells and primary cultured cardiomyocytes, but not in the cancer cell lines for which DOX is clinically indicated. A TUNEL assay showed that CME treatment improved DOX-induced apoptosis in H9C2 cells. Moreover, DOX-induced increases in the expression levels of p53, phosphorylated p53, and cleaved caspase-3 and -9 were significantly suppressed by CME treatment. Next, an experiment using a mouse model of acute heart failure showed that CME treatment substantially inhibited the DOX-induced decrease in survival rate. Echocardiographic analysis indicated that CME treatment reduced DOX-induced left ventricular systolic dysfunction. A TUNEL assay showed that CME treatment suppressed DOX-induced apoptosis in the mouse heart.
CONCLUSIONS These results suggest that CME treatment ameliorates DOX-induced cardiotoxicity by suppressing apoptosis. Therefore, CME may have the therapeutic potency to reduce DOX-induced cardiotoxicity in cancer patients. Further clinical study is needed to clarify this effect in humans.
GW34-e0376
Yingwen Lin1,2,3, Dongtu Hu1,2,3, Xiangjie Lin1,2,3, Juncong Li1,2,3, Shuwen Su1,2,3, Dingli Xu1,2,3, Qingchun Zeng1,2,3
1State Key Laboratory of Organ Failure Research, Department of Cardiology, Nanfang Hospital, Southern Medical University
2Guangdong Provincial Key Laboratory of Shock and Microcirculation, Southern Medical University
3Bioland Laboratory (Guangzhou Regenerative Medicine and Health Guangdong Laboratory)
OBJECTIVES Galectin-1, a beta-glycoside binding protein, has been reported to involve a variety of cell functions including proliferation, cell migration and focal adhesion. The role of cell-matrix interaction in calcific aortic valve disease (CAVD) is not fully investigated and whether galectin-1 is involved in this process remains unclear. We hypothesized that galectin-1 mitigates the fibrocalcific response and mechanosensitivity of valve interstitial cells (AVICs) by modulating integrin-mediated cytoskeletal reorganization and focal adhesion.
METHODS We enrolled patients with CAVD and age- and sex-matched controls to detect serum galectin-1.
RESULTS We enrolled patients with CAVD and age- and sex-matched controls to detect serum galectin-1 and found that circulating galectin-1 was lower in CAVD. The expression of galectin-1 decreased in fibrocalcific valves and AVICs cultured in osteogenic medium (OM), as shown by western blot, qPCR, immunohistochemistry (IHC) and immunofluorescence. AVICs cultured in OM also presented enhanced integrin signaling, cytoskeletal reorganization and focal adhesion. Silencing galectin-1 by short-interfering RNA exacerbated the fibrocalcific change of AVICs in vitro while recombinant galectin-1 (rGal-1) prevented these effects. Anti-integrin blocking experiments and co-immunoprecipitation assays demonstrated that the galectin-1 directly interacted with integrin αvβ1 to regulate AVICs adhesion to a fibronectin matrix. Western blot, qPCR and Immunofluorescence assays confirmed that actin cytoskeletal reorganization was induced by αvβ1 activation through Rac1-WRC-Arp2/3 signaling axis. Western blot, alkaline phosphatase (ALP) as well as Alizarin red staining demonstrated that silencing galectin-1 enhanced AVICs sensitivity to piezo1 activation, which is dependent on αvβ1 binding and actin polymerization. In vivo, aortic valve disease was constructed by direct wire injury (DWI), and we showed that overexpression of galectin-1 by adeno-associated virus significantly accelerated the progression of aortic valve lesion induced by DWI in mice.
CONCLUSIONS Galectin-1 interacts with integrin αvβ1 to modulate cytoskeletal reorganization and AVICs adhesion to fibronectin, thus mitigating the fibrocalcific response of AVICs and aortic valve calcification.
GW34-e0391
Dmitry Kashirskikh1, Raisa Surkova1, Igor Sobenin1, Alexander Yakovlev2, Alexander Orekhov1
1Institute of General Pathology and Pathophysiology, Moscow, Russia
2Institute of Higher Nervous Activity and Neurophysiology, Moscow, Russia
OBJECTIVES Previously we found sialidase activity circulating in the blood of atherosclerotic patients. This sialidase is responsible for atherogenic modification of low-density lipoprotein. We have shown that desialylation is an atherogenic modification of LDL that makes the lipoprotein capable of inducing lipid accumulation in arterial wall cells, turning them into foam cells. The aim of this study was to isolate and identify proteins possessing sialidase activity.
METHODS The blood serum of atherosclerotic patients was screened for the presence of significant sialidase activity using commercial kits. Isolation of proteins with sialidase activity was performed using affinity chromatography followed by polyacrylamide gel electrophoresis. In order to identify the isolated proteins, the MALDI-TOF mass spectrometry method was used.
RESULTS About 700 blood serum samples were tested and 84 samples with sialidase activity were selected. Proteins of 65 kDa and 116 kDa were isolated from the eluate after affinity chromatography using polyacrylamide gel electrophoresis. Mass spectrometry revealed a wide range of proteins, but no known human neuraminidase was found among them. During chromatography, exosomes are isolated together with an affinity-binding protein, which explains the wide range of proteins identified by mass spectroscopy and masking the real result of affinity isolation.
CONCLUSIONS The data obtained suggest that an unidentified protein with sialidase activity circulates in the blood, both in free form and non-covalently bound to the extracellular vesicles membrane (exosomes).
This work was supported by the Russian Science Foundation (Grant # 22-65-00005).
GW34-e0392
Dmitry Kashirskikh, Alexandra Melnichenko, Alexander Orekhov
Institute of General Pathology and Pathophysiology, 8 Baltiiskaya Street, Moscow 125315, Russia
OBJECTIVES Electronegative LDL, small dense LDL, and desialylated LDL circulating in the blood of patients were obtained by different methods. Naturally, the question arises what are the similarities and differences between these forms of LDL modification. We believe that multiple modified LDL particle (small, dense, electronegative, desialylated, etc.) occurs in blood. Ex vivo experiments have revealed mechanisms of multiple modification of LDL in the blood.
METHODS Fraction of native LDL was isolated from blood plasma of healthy subjects. Blood serum of patients with assessed atherosclerosis was also obtained. LDL and serum were mixed and incubated for various periods at 37°C.
RESULTS After 1 hour incubation of native LDL with atherosclerotic serum desialylated LDL appears. After 3 hours, LDL becomes able to cause accumulation of cholesterol in cultured cells. After 6 hours, LDL demonstrates reduction of neutral lipids and phospholipids as well as reduction in its size. After 36 hours, an increase in the electronegativity of the lipoprotein particle is detected. After 48–72 hours, loss of α-tocopherol, increase of susceptibility to oxidation, and accumulation of lipid peroxidation products in LDL are observed. Thus, multiple modification of LDL is a cascade of sequential changes in lipoprotein particle, namely: desialylation, loss of lipids, size reduction, increase of electronegative charge, lipid peroxidation in LDL. Desialylation of LDL particle is one of the first or primary events of atherogenic modification. We have established that the reason of LDL desialylation is trans-sialidase. We found trans-sialidase activity in the blood of patients with atherosclerosis and other cardiovascular diseases. We have shown that human neuraminidases 2 and 4 possess trans-sialidase activity. On the other hand, selective inhibitors of viral sialidases suppress trans-sialidase activity in the blood of atherosclerotic patients.
CONCLUSIONS Thus, trans-sialidase causing atherogenic desialylation of LDL may be of both endogenous and exogenous origin.
This work was supported by the Russian Science Foundation (Grant # 23-65-10014).
GW34-e0393
Vasily Sukhorukov, Alexander Orekhov
Institute of General Pathology and Pathophysiology, 8 Baltiiskaya Street, Moscow 125315, Russia
OBJECTIVES Accumulation of lipid-laden (foam) cells in the arterial wall is known to be the earliest step in the pathogenesis of atherosclerosis. There is almost no doubt that atherogenic modified low-density lipoproteins (LDL) are the main sources of accumulating lipids in foam cells. Atherogenic modified LDL are taken up by arterial cells, such as macrophages, pericytes, and smooth muscle cells in an unregulated manner bypassing the LDL receptor. The present study was conducted to reveal possible common mechanisms in the interaction of macrophages with associates of modified LDL and non-lipid latex particles of a similar size.
METHODS To determine regulatory pathways that are potentially responsible for cholesterol accumulation in human macrophages after the exposure to naturally occurring atherogenic or artificially modified LDL, we used transcriptome analysis.
RESULTS Previous studies of our group demonstrated that any type of LDL modification facilitates the self-association of lipoprotein particles. The size of such self-associates hinders their interaction with a specific LDL receptor. As a result, self-associates are taken up by nonspecific phagocytosis bypassing the LDL receptor. That is why we used latex beads as a stimulator of macrophage phagocytotic activity. We revealed at least signaling pathways that were regulated by the interaction of macrophages with the multiple-modified atherogenic naturally occurring LDL and with latex beads in a similar manner. Therefore, modified LDL was shown to stimulate phagocytosis through the upregulation of certain genes. We have identified at least three genes (F2RL1, EIF2AK3, and IL15) encoding inflammatory molecules and associated with signaling pathways that were upregulated in response to the interaction of modified LDL with macrophages. Knockdown of two of these genes, EIF2AK3 and IL15, completely suppressed cholesterol accumulation in macrophages. Correspondingly, the upregulation of EIF2AK3 and IL15 promoted cholesterol accumulation.
CONCLUSIONS These data confirmed our hypothesis of the following chain of events in atherosclerosis: LDL particles undergo atherogenic modification; this is accompanied by the formation of self-associates; large LDL associates stimulate phagocytosis; as a result of phagocytosis stimulation, pro-inflammatory molecules are secreted; these molecules cause or at least contribute to the accumulation of intracellular cholesterol. This chain of events may explain the relationship between cholesterol accumulation and inflammation. The primary sequence of events in this chain is related to inflammatory response rather than cholesterol accumulation.
This work was supported by the Russian Science Foundation (Grant # 23-45-00031).
GW34-e0396
Vasily Sukhorukov, Alexander Orekhov
Institute for Atherosclerosis Research, Osennyaya Street 4-1-207, Moscow 121609, Russia
OBJECTIVES Mitochondrial dysfunction is thought to be one of the causative factors of impaired innate immunity during atherosclerosis development. The presence of heteroplasmic mutations in mitochondrial DNA (mtDNA) can result in the appearance of dysfunctional mitochondria in intimal cells. The aim of this study was to evaluate the effect of the m.15059G>A mitochondrial nonsense mutation on cellular functions related to atherosclerosis, such as lipoidosis, pro-inflammatory response, and mitophagy. It was suggested that heteroplasmic mutations could be responsible for mitochondrial dysfunction, which could lead to a disturbance of the innate immunity response and, ultimately, the chronic inflammation associated with atherosclerosis.
METHODS The human monocytic cell line THP-1 and cytoplasmic hybrid cell line TC-HSMAM1 were used. An original approach based on the CRISPR/Cas9 system was developed and used for the elimination of mtDNA copies carrying the m.15059G>A mutation in the MT-CYB gene. The gene expression levels of genes encoding enzymes related to cholesterol metabolism were analyzed by quantitative RT-PCR. The evaluation of pro-inflammatory cytokine secretion was assessed using ELISA. Mitophagy in cells was detected using confocal microscopy.
RESULTS In contrast to intact TC-HSMAM1 cybrids, in Cas9-TC-HSMAM1 cells, incubation with atherogenic LDL led to a decrease in the expression of the gene encoding fatty acid synthase (FASN). It was found that TC-HSMAM1 cybrids are characterized by defective mitophagy and are also unable to reduce the production of pro-inflammatory cytokines (to form immune tolerance) in response to repeated LPS stimulation. Elimination of mtDNA carrying the m.15059G>A mutation led to the restoration of immune tolerance and activation of mitophagy in the studied cells.
CONCLUSIONS The m.15059G>A mutation was found to be associated with defective mitophagy, immune tolerance, and impaired metabolism of intracellular lipids due to upregulation of the FASN in monocytes and macrophages.
This study was supported by Russian Science Foundation, Grant # 22-15-00064.
GW34-e0397
Victoria Khotina, Alexander Zhuravlev, Vasily Sukhorukov
Institute of General Pathology and Pathophysiology
OBJECTIVES Recent studies have identified mutations in mitochondrial DNA (mtDNA) that are linked with the development of atherosclerosis. Such mutations can cause mitochondrial dysfunction, ultimately leading to the activation of signaling pathways that promote the secretion of pro-inflammatory cytokines. It is possible to facilitate the study of this association using cytoplasmic hybrid cell lines, that can be created by fusion of mitochondria-depleted THP-1 monocytes with enucleated thrombocytes derived from atherosclerotic patients with mtDNA mutations. This study aimed to evaluate the correlation between the m.15059G>A mutation in the mitochondrial MT-CYB gene and pro-inflammatory cytokine secretion, as well as the ability of monocytes to form immune tolerance.
METHODS Human monocyte-like cell line THP-1, cybrid line TC-HSMAM1 with m.15059G>A mutation (MT-CYB), and cybrid cells TC-HSMAM1 with eliminated m.15059G>A mutation (Cas9-TC-HSMAM1) by CRISPR/Cas9 editing were used. The pro-inflammatory response was induced by bacterial lipopolysaccharide (LPS). The secretion of cytokines was assessed by ELISA. To assess immune tolerance, cells were incubated for 16 hours with LPS followed by additional incubation with LPS for 4 h.
RESULTS The study revealed that both TC-HSMAM1 and Cas9-TC-HSMAM1 cybrids showed an increased basal level of IL-1β, IL-6, IL-8, and MCP-1 (CCL2) compared to the control THP-1 cells (P<0.05). Nevertheless, while the basal secretion level of TNFα had no difference between Cas9-TC-HSMAM1 cells and control THP-1 cells, the secretion of this cytokine by TC-HSMAM1 cybrids was significantly increased (P<0.01). Following stimulation of the pro-inflammatory cell phenotype with LPS, there was a significant increase in the secretion of all cytokines in all studied cells. Notably, the secretion of IL-8 and MCP-1 in both TC-HSMAM1 and Cas9-TC-HSMAM1 cybrids did not have any statistically significant differences from control THP-1 cells. Furthermore, the secretion of IL-1β and TNFα by TC-HSMAM1 cybrids was observed to decrease compared to control THP-1 cells after LPS stimulation (P<0.05). On the other hand, the secretion of these cytokines by Cas9-TC-HSMAM1 cells did not differ from the control THP-1 cells. However, IL-6 secretion was significantly increased in TC-HSMAM1 cybrids compared to THP-1 cells (P<0.05), whereas the secretion of this cytokine in Cas9-TC-HSMAM1 exceeded that of both cell lines. The evaluation of the ability to form immune tolerance by cells showed that Cas9-TC-HSMAM1 and THP-1 cells were able to form immune tolerance, which consisted of an increase in TNFα secretion after the first LPS stimulation and a decrease in the secretion of pro-inflammatory cytokines after repeated LPS stimulation. However, TC-HSMAM1 cybrids showed elevated TNFα secretion in response to repeated LPS stimulation demonstrating the inability to form immune tolerance.
CONCLUSIONS The study findings suggest that monocytes with the m.15059G>A mutation in MT-CYB exhibit increased basal secretion and decreased LPS-stimulated secretion of TNFα compared to Cas9-TC-HSMAM1 and THP-1. Additionally, TC-HSMAM1 cells were found to be unable to form immune tolerance in response to repeated stimulation of cells with LPS.
This work was supported by Russian Science Foundation Grant #22-25-00393.
GW34-e0398
Victoria Khotina, Vasily Sukhorukov
Institute of General Pathology and Pathophysiology
OBJECTIVES Recent researches have shown an association between mutations in mtDNA and the development of atherosclerosis. In particular, the m.15059G>A mutation in the mitochondrial MT-CYB gene has been found in lipofibrous plaque cells from patients with atherosclerosis. However, it is still unclear how these mutations affect key processes such as apoptosis regulation, synthetic activity, and proliferation of macrophages, that play a crucial role in atherosclerotic plaque development. To investigate this association, cytoplasmic hybrid cell lines with mutations in mtDNA, created from THP-1 monocyte cells, can be utilized. In this study, we aim to investigate the association between the m.15059G>A mutation in the MT-CYB and the expression of genes related to apoptosis, cell cycle regulation, and protein synthesis in monocytes and macrophages.
METHODS Human monocyte-like cell line THP-1, cybrid line TC-HSMAM1 with m.15059G>A mutation (MT-CYB), and cybrid line TC-HSMAM1 with eliminated m.15059G>A mutation (Cas9-TC-HSMAM1) by CRISPR/Cas9 editing were used. The qPCR was used to assess the expression of apoptotic pathway related genes (BCL2, BAX, CASP3, CASP9, APAF1), the expression of cell cycle regulation related genes (PCNA, CCNB1, CCND1), and the expression of protein synthesis related genes (POLR1A, POLR3A, COL6A1).
RESULTS It was found that TC-HSMAM1 cybrids had an increased expression of BCL2, CASP3 and APAF1, and a decreased expression of BAX and CASP9 compared to THP-1 cells (P<0.001). Elimination of mutated mtDNA from cybrids led to an increase of BAX and CASP9 expression and a decrease of CASP3 expression (P<0.01). However, there were no differences in expression of BCL2 and APAF1 between TC-HSMAM1 and Cas9-TC-HSMAM1 cells (P>0.05). Moreover, BAX/BCL2 ratio in TC-HSMAM1 cells was reduced compared to control THP-1 cells (P<0.001). In contrast, BAX/BCL2 ratio in Cas9-TC-HSMAM1 cells was increased compared to TC-HSMAM1 (P<0.05). It was shown that the expression of PCNA, CCNB1 and CCND1 in TC-HSMAM1 cells was increased compared to THP-1 cells (P<0.001). Increased expression of PCNA and CCND1 was observed in Cas9-TC-HSMAM1 cells compared to the control THP-1 cells (P<0.01), while the expression of the CCNB1 had no difference. No statistically significant difference was found in the expression of the PCNA and CCND1 in cybrid cells before and after the elimination of the m.15059G>A mutation. However, decreased CCNB1 expression was observed in Cas9-TC-HSMAM1 cells compared to TC-HSMAM1 (P<0.01). It was found that the expression of the POLR1A and COL6A1 in the TC-HSMAM1 cybrids was increased compared with THP-1 cells (P<0.001), while the expression of POLR3A was decreased. Increased expression of the POLR3A and COL6A1 was found in Cas9-TC-HSMAM1 cells compared with the control THP cells (P<0.001), while POLR1A expression was significantly decreased (P<0.001). However, POLR1A and COL6A1 expression was decreased in Cas9-TC-HSMAM1 compared with the TC-HSMAM1 cells (P<0.01), while POLR3A expression was increased.
CONCLUSIONS Our findings suggest that the m.15059G>A mutation may be associated with decreased expression of BAX, CASP9 and POLR3A, and the increased expression of CASP3, CCNB1, POLR1A and COL6A1, indicating a possible role of this mutation in regulation of macrophage apoptosis, cell cycle (proliferation activity) and protein synthesis. Furthermore, the presence of this mutation was accompanied by a reduced BAX/BCL2 ratio.
This work was supported by Russian Science Foundation Grant #22-25-00393.
GW34-e0399
Victoria Khotina1, Alexander Zhuravlev1, Vasily Sukhorukov1,2
1Institute of General Pathology and Pathophysiology
2Petrovsky National Research Centre of Surgery
OBJECTIVES A growing body of research demonstrates the role of mutations in mitochondrial DNA (mtDNA) in the development of inflammatory diseases, such as atherosclerosis. These mutations may cause mitochondrial dysfunction, promoting the activation of pro-inflammatory cell response, including the formation of NLRP3 inflammasomes. In turn, NLRP3 inflammasomes produce pro-inflammatory cytokines such as IL-1β. Interestingly, recent findings have discovered the presence of mtDNA mutations in atherosclerotic plaque cells, but little information exists on how these mutations impact the pro-inflammatory response of macrophages. Improved knowledge of the association between mtDNA mutations and the expression of pro-inflammatory cytokines could pave the way for the development of new therapeutic approaches for atherosclerosis treatment. This study aimed to investigate the association between the m.15059G>A mutation in the MT-CYB gene and the expression of NLRP3 inflammasome-related genes.
METHODS Human monocyte-like cell line THP-1, TC-HSMAM1 cybrids with m.15059G>A mutation (MT-CYB), and TC-HSMAM1 cells with eliminated m.15059G>A mutation (Cas9-TC-HSMAM1) by CRISPR/Cas9 editing were used. NLRP3 inflammasome (NLRP3), caspase 1 (CASP1) and interleukin 1β (IL1B) gene expression was measured by qPCR. Pro-inflammatory response was induced by bacterial lipopolysaccharide (LPS). IL-1β secretion was assessed by ELISA.
RESULTS It was shown, that the basal expression of NLRP3 and IL1B genes was increased in TC-HSMAM1 cybrids compared to control THP-1 cells, while the CASP1 expression had no changes. However, elimination of the m.15059G>A mutation resulted in decreased basal expression of all studied genes. Under stimulation with LPS, an increased CASP1 expression was observed in TC-HSMAM1 cells compared to unstimulated cells, while the expression of IL1B and NLRP3 remained unaltered. Nevertheless, IL1B expression was increased, and CASP1 expression was reduced in LPS-stimulated TC-HSMAM1 cells when compared to LPS-stimulated control THP-1 cells. Stimulation of Cas9-TC-HSMAM1 cells with LPS resulted in increased expression of all studied genes compared to unstimulated cells. In addition, NLRP3 expression was found to be elevated in Cas9-TC-HSMAM1 cybrids compared to the TC-HSMAM1 cells. Nevertheless, CASP1 and IL1B expression was shown to have no significant differences from the TC-HSMAM1 cells, as confirmed by IL-1β cell secretion.
CONCLUSIONS Our results suggest that the presence of the m.15059G>A mutation in the MT-CYB gene may increase the formation of NLRP3 inflammasomes in macrophages due to an increased expression of NLRP3 and IL1B genes in normal physiological conditions.
This work was supported by Russian Science Foundation Grant #22-25-00274.
GW34-e0400
Victoria Khotina, Vasily Sinyov, Vasily Sukhorukov
Institute of General Pathology and Pathophysiology
OBJECTIVES Mitochondrial dysfunction can arise from a range of factors, including mutations in mtDNA. Among these mutations, the m.15059G>A mutation in the MT-CYB gene is of particular interest due to its impact on the cytochrome b protein, which plays a critical role in the electron transport chain of mitochondria. The m.15059G>A mutation leads to the formation of a truncated variant of cytochrome b, which can impair its functional activity and consequently perturb the mitochondrial bioenergetic profile of monocytes and macrophages. Monocytes and macrophages have high bioenergetic demands and rely on the proper functioning of mitochondria to carry out their diverse functions including phagocytosis and cytokine production. The bioenergetic profile of these cells is regulated by the balance between mitochondrial respiration and glycolysis. Any perturbations to this balance, such as those resulting from the m.15059G>A mutation, can have profound impacts on the function and behavior of these cells. While the exact mechanisms underlying this association remain unclear, we aimed to investigate the association between the m.15059G>A mutation in the MT-CYB with impaired mitochondrial respiration and bioenergetic profile in monocytes.
METHODS Human monocyte-like cell line THP-1, TC-HSMAM1 cybrids with m.15059G>A mutation (MT-CYB), and TC-HSMAM1 cells with eliminated m.15059G>A mutation (Cas9-TC-HSMAM1) by CRISPR/Cas9 editing were used. The Oxygraph+ System with S1 Clark-type polarographic oxygen electrode was used to measure oxygen consumption rate (OCR) of cells. Parameters of the cellular mitochondrial function was assessed using respiratory chain inhibitors (oligomycin A, FCCP, antimycin A).
RESULTS It was shown that the OCR traces of all studied lines displayed a similar profile. However, basal OCR was found to be significantly reduced in Cas9-TC-HSMAM1 cells compared to the control THP-1 cells and TC-HSMAM1 cybrids (P<0.05). Conversely, under treatment with oligomycin A and antimycin A, TC-HSMAM1 cybrids demonstrated an increase in OCR compared to control THP-1 cells and Cas9-TC-HSMAM1 cybrids. Analysis of mitochondrial bioenergetic profiles revealed that TC-HSMAM1 cybrids exhibited significantly increased parameters such as non-mitochondrial OCR and proton leak OCR in comparison to control THP-1 cells (P<0.001). However, these parameters did not differ significantly between Cas9-TC-HSMAM1 and THP-1 cells. Furthermore, parameters including ATP-linked OCR, maximal OCR, and reserve capacity OCR did not demonstrate statistically significant differences between THP-1 and TC-HSMAM1 cells. Nevertheless, there was a decrease in maximal OCR observed in Cas9-TC-HSMAM1 cells as compared to control THP-1 cells and TC-HSMAM1 cybrids (P<0.05). Evaluation of the cellular Bioenergetic Health Index (BHI), calculated using a logarithmic dependence of reserve capacity OCR, ATP-linked OCR, non-mitochondrial OCR, and proton leak OCR, revealed that BHI was reduced in TC-HSMAM1 cybrids compared to the control. Conversely, BHI was found to be significantly increased in Cas9-TC-HSMAM1 cybrids when compared to TC-HSMAM1.
CONCLUSIONS The m.15059G>A mutation in the MT-CYB gene has been shown to impact various parameters related to the bioenergetic functions of mitochondria, including non-mitochondrial OCR, proton leakage, and BHI.
This work was supported by Russian Science Foundation Grant #22-65-00005.
GW34-e0401
Victoria Khotina1, Mariam Bagheri-Ekta2, Arthur Lee1,2, Vasily Sukhorukov1,2
1Institute of General Pathology and Pathophysiology
2Petrovsky National Research Centre of Surgery
OBJECTIVES PERK is a transmembrane protein located in the endoplasmic reticulum (ER) of cells, and it plays an essential role in the regulation of ER-stress response. Recent studies have shown that ER stress and unfolded protein response induction alter the expression of genes related to lipid metabolism. Moreover, it has been shown that an increase of cellular cholesterol in vascular smooth muscle cells from atherosclerotic plaques triggers ER-stress and activates PERK signaling. However, it is not fully understood how PERK is involved in altering lipid metabolism in macrophages and in the formation of foam cells, a hallmark of atherosclerosis. The main aim of this study was to evaluate the impact of PERK on macrophage cholesterol accumulation and the regulation of expression of genes related to lipid metabolism.
METHODS Human monocyte-like cell line THP-1 was used as control. CRISPR/Cas9 editing was used to generate THP-1 cell line with PERK gene knockout (THP-1 PERK −/−). Macrophage-like phenotype differentiation was induced by phorbol 12-myristate 13-acetate (PMA). Foam cell formation was induced by cell incubation with low-density lipoproteins (LDL). LDL was isolated from plasma from atherosclerotic patients using preparative ultracentrifugation technique. Cholesterol accumulation was measured by spectrophotometry. The expression of ATP binding cassette subfamily A member 1 (ABCA1), acetyl-CoA acetyltransferase 1 (ACAT1), fatty acid synthase (FASN), neutral cholesterol ester hydrolase 1 (NCEH1) and cluster of differentiation 36 (CD36) genes was assessed by qPCR.
RESULTS To induce PERK knockout, THP-1 cells were transfected using two sgRNAs and a plasmid containing an expression cassette with the sequence of the puromycin resistance gene. A stable clone of cells with the most effective PERK knockout was obtained after 4 weeks of puromycin selection. PERK gene knockout in the obtained clone was confirmed by qPCR and Sanger sequencing. Incubation of THP-1 macrophages with atherogenic LDL induced an increase in intracellular cholesterol level compared to untreated control (P<0.001). In turn, incubation of THP-1 PERK −/− cells with atherogenic LDL had no impact on intracellular cholesterol accumulation. It was found that the basal expression of ABCA1, FASN, NCEH1 and CD36 was decreased in THP-1 PERK −/− macrophages compared to control cells (P<0.001). Incubation of THP-1 macrophages with atherogenic LDL led to an increased expression of the ACAT1 and NCEH1, as well as a decreased expression of ABCA1, FASN and CD36 compared to the untreated cells (P<0.001). At the same time, decreased expression of FASN, NCEH1 and CD36 in response to cholesterol accumulation was observed in THP-1 PERK −/− macrophages compared to the untreated cells (P<0.05), while the expression of ACAT1 and ABCA1 had no changes. Moreover, the expression level of ACAT1 had no difference between THP-1 and THP-1 PERK −/− cells under cell incubation with atherogenic LDL.
CONCLUSIONS Thus, it was found that PERK may be involved in the regulation of signaling pathways of ABCA1, FASN, NCEH1 and CD36. The obtained results may indicate a possible involvement of PERK in the intracellular lipid metabolism of macrophages, contributing in foam cell formation process.
This work was supported by Russian Science Foundation Grant #22-25-00393.
GW34-e0407
Jingyi Zhang1,2, Ziyang Xue1,2, Mingke Chang1,2, Xinxin Feng1,2, Yifan Cai1,2, Liang Bai1,3, Weirong Wang1,3, Sihai Zhao3, Enqi Liu1,3, Rong Wang1,2
1Institute of Cardiovascular Science, Translational Medicine Institute, Xi’an Jiaotong University Health Science Center, Xi’an 710061, China
2Department of Laboratory Animal Science, School of Basic Medical Sciences, Xi’an Jiaotong University, Xi’an, Shaanxi 710061, China
3Laboratory Animal Center, Xi’an Jiaotong University, Xi’an, Shaanxi 710061, China
OBJECTIVES Phenotypic switching of vascular smooth muscle cells (VSMCs) plays an essential role in the development of atherosclerosis. Protein inhibitor of activated STAT (Pias) regulates VSMCs phenotype via acting as sumo E3 ligase to promote protein sumoylation. Our previous study indicates that Pias3 expression decreased in atherosclerotic lesions. Therefore, this study was aimed to explore the role of Pias3 on VSMCs phenotype switching during atherosclerosis.
METHODS ApoE −/− and ApoE −/− Pias3 −/− double-deficient mice were fed with high-fat/high-cholesterol diet to induce atherosclerosis. Aorta tissues and primary VSMCs were collected for assessing plaque formation and VSMCs phenotype. In vitro, Pias3 was overexpressed in A7r5, a VSMCs cell line, by transfecting with Pias3 plasmid. Quantitative PCR, immunoblotting, immunoprecipitation, etc. were used to analyze the effect of Pias3 on VSMCs phenotypic switching.
RESULTS Pias3 deficiency significantly exacerbated atherosclerotic plaque formation and promoted VSMCs phenotypic switching to synthetic state. In vitro, overexpressing Pias3 in VSMCs increased the expression of contractile markers (myosin heavy chain 11, calponin 1), whereas decreased the level of synthetic marker (vimentin). Additionally, Pias3 overexpression impeded PDGF-BB-induced VSMCs proliferation and migration. Immunoprecipitation and mass spectrometry results showed that Pias3 enhanced the sumoylation and ubiquitination of vimentin, as well as shortened its half-life. Moreover, the ubiquitination level of vimentin could be impaired by 2-D08, a sumoylation inhibitor. It suggests that Pias3 might accelerate the ubiquitination-degradation of vimentin by promoting its sumoylation.
CONCLUSIONS These results indicated that Pias3 might ameliorate atherosclerosis progression by suppressing VSMCs phenotypic switching and reducing vimentin protein stability.
GW34-e0415
Linqiang Xi
Cardiac Pacing and Electrophysiology/Department of Cardiac Electrophysiology and Remodeling, The First Affiliated Hospital of Xinjiang Medical University, Urumqi 830054, Xinjiang, China
OBJECTIVES Energy metabolism in atrial remodeling has been implicated in the development of atrial fibrillation. Renault oxazine inhibits fatty acid oxidation and increases glucose oxidation, without causing an increase in glycolysis or lactic acid release. It improves the coupling between glycolysis and glucose oxidation, and can improve the REDOX balance and mitochondrial function, thereby suppressing the occurrence of atrial fibrillation. This study is aimed at understanding the specific mechanism by which Renault oxazine reshapes energy metabolism in atrial fibrillation.
METHODS Male SD rats were randomly divided into 3 groups (n=12): control group, ANgII group, and ANgII+Renault oxazine group. Blood samples were taken weekly and electrocardiogram, ultrasound, and electrophysiological examinations were carried out before sacrificing the rats for analysis. HL-1 cells were divided into five groups: control group, ANgII treatment group, deferoxamine treatment group, ANgII plus Renault oxazine group, and ANgII and Renault lamictal and deferoxamine combination treatment group. Western blotting was used to analyze the expression of key enzymes related to glucose and lipid metabolism.
RESULTS ANgII-induced atrial remodeling was associated with a significant increase in left atrial diameter and a high rate of atrial fibrillation. However, treatment with Renault oxazine significantly improved these parameters. Western blotting confirmed the existence of molecular expression disorders related to glucolipid metabolism in the ANgII treatment group HL-1 cells. HIF-1α and PPAR-gamma expression were significantly lower in this group, while Renault oxazine did not show any statistically significant differences from the control group. Treatment with Renault oxazine resulted in no significant differences in the expression of key enzymes related to glucose and fatty acid intake, when compared to the control group.
CONCLUSIONS Renault oxazine improves the energy metabolism of rat atrial fibrillation induced by ANgII remodeling by inhibiting the HIF-1α-PPAR-gamma pathway.
GW34-e0422
Jishou Zhang, Menglong Wang, Jun Wan
Renmin Hospital of Wuhan University
OBJECTIVES Inflammation plays a critical role in the development of hypertension and vascular remodeling. Resolvin E1 (RvE1), as one of the specialized proresolving lipid mediators, promotes inflammation resolution by binding with a G protein-coupled receptor, ChemR23. However, whether RvE1/ChemR23 regulates hypertension and vascular remodeling is unknown.
METHODS Hypertension in mice was induced by Angiotensin II (AngII) infusion (750 ng/kg/min) and RvE1 (2 μg/kg/day) was administrated through intraperitoneal injection. We knocked down ChemR23 expression in mice by intravenously injecting adeno-associated virus-9 (AAV9) encoding shRNA against ChemR23 (AAV9-ChemR23).
RESULTS We found that RvE1 lowered blood pressure, reduced aortic media thickness, attenuated aortic fibrosis, mitigated vascular smooth muscle cell (VSMC) phenotypic transformation and proliferation, which were all reversed by the knockdown of ChemR23 that was mainly expressed on VSMCs. Moreover, RvE1 reduced the aortic infiltration of macrophages and T cells, which was also reversed by ChemR23 knockdown. RvE1 inhibited Ccl5 expression in VSMCs via an AMPKα/Nrf2/canonical NF-κB pathway, thereby reducing the infiltration of macrophages and T cells. The AMPKα/Nrf2 pathway also mediated the effects of RvE1 on VSMC phenotypic transformation and proliferation. In hypertensive patients, the serum levels of RvE1 and other eicosapentaenoic acid (EPA)-derived metabolites were significantly decreased.
CONCLUSIONS RvE1/ChemR23 ameliorated hypertension and vascular remodeling by activating AMPKα/Nrf2 signaling, which mediated immune cell infiltration through inhibiting canonical NF-κB/Ccl5 pathway and regulated VSMC proliferation and phenotypic transformation in VSMCs. RvE1/ChemR23 may be a potential therapeutic target for hypertension.
GW34-e0423
Yijia Shao1, Xiang Liu2, Fang Wu1, Long Chen3
1Department of Geriatrics, The First Affiliated Hospital, Sun Yat-sen University, Guangzhou
2Department of Cardiac Surgery, Guangdong Cardiovascular Institute, Guangdong Provincial People’s Hospital, Guangdong Academy of Medical Sciences, Guangzhou
3The International Medical Department of Shenzhen Hospital, Southern Medical University, Shenzhen
OBJECTIVES Trimethylamine-N-Oxide (TMAO) is recognized as a novel marker and mediator of atherosclerotic cardiovascular disease (ASCVD). Endothelial progenitor cells (EPCs) are crucial for maintaining vascular homeostasis. Impaired EPC numbers and function correlate with increased adverse cardiovascular events. The aim of this study was to determine the effect of TMAO on late EPCs (LEPCs) and its underlying molecular mechanism.
METHODS In vitro migration and tubulogenic capacities of LEPCs were attenuated by TMAO in a dose-dependent manner, accompanied with inhibition of manganese superoxide dismutase (MnSOD) and mitochondrial damage. TMAO induced mitochondrial damage provoked the proinflammatory responses (increased levels of IL-6, IL-1b, ICAM-1, E-sel and TNF-α) and autophagic cell death (confirmed by Western blot immunofluorescent staining and transmission electron microscopy) in LEPCs.
RESULTS Overexpression of MnSOD through adenovirus transfection reversed TMAO-related LEPCs dysfunction. To study the effect of TMAO on LEPCs-mediated vascular repair in vivo, hind limb ischemia model was established in nude mice and LEPCs were injected in the ischemic hind limb. Laser Doppler imaging of mouse ischemic hindlimbs at 21 days indicated that TMAO treatment inhibited LEPCs-mediated blood flow recovery, which was restored by MnSOD overexpression. Tissue analyses further revealed consistent alterations in capillary density determined by CD31 staining.
CONCLUSIONS TMAO induces mitochondrial damage in LEPCs via MnSOD suppression, which leads to cell dysfunction, proinflammatory activation and autophagic cell death in vitro and impaired LEPCs-mediated revascularization in vivo. Overexpression of MnSOD restores TMAO-induced LEPCs dysfunction and further enhances LEPCs-mediated revascularization in the ischemic hind limbs in nude mice.
GW34-e0432
Zikun Duan1, Jing Gan1, Yulin Li2, Fan Wu1, Zhuofeng Lin1
1The First Affiliated Hospital, School of Pharmaceutical Sciences, Wenzhou Medical University
2Beijing Institute of Heart, Lung, and Blood Vessel Diseases, Anzhen Hospital of Capital Medical University
OBJECTIVES Bone morphogenetic protein 9 (BMP9) is a member of the TGF-β family of cytokines with pleiotropic effects on glucose metabolism, pro-fibrosis, and lymphatic development. However, the role of BMP9 in myocardial infarction (MI) remains elusive.
METHODS The expressional profiles of BMP9 in cardiac tissues and plasma samples of subjects with MI were determined by immunoassay or immunoblot. The roles of BMP9 in MI were determined by evaluating the impact of BMP9 deficiency and replenishment with adeno-associated virus-mediated BMP9 (AAV-BMP9) expression in mice with MI.
RESULTS Circulating BMP9 levels and its cardiac contents are markedly increased in humans and mice with MI and are negatively associated with cardiac functions. BMP9 deficiency causes a marked exacerbation of left ventricle dysfunction, increases infarct size, and cardiac fibrosis in mice with MI. In contrast, replenishment of BMP9 strikingly attenuates these adverse effects via two independent mechanisms, improving lymphatic drainage function, thereby leading to decrease cardiac edema, and triggering the mitochondrial production of 2,4-Dienoyl-CoA reductase (DECR1), a rate-limiting enzyme involved in β-oxidation of unsaturated fatty acids in mitochondria, which in turn promotes cardiac mitochondrial bioenergetics, then mitigating MI-induced cardiomyocyte injuries. Furthermore, DECR1 deficiency exacerbated MI-induced cardiac injury in mice, whereas this adverse effect was restored when treated with AAV-DECR1. In addition, DECR1 deletion strikingly abrogates the beneficial impact of BMP9 against MI-induced cardiac dysfunction and damage in mice.
CONCLUSIONS These results indicated that BMP9 protects against MI via fine-tuning the multiorgan crosstalk among the liver, lymph, and heart.
GW34-e0434
Zhongming Li1,2, Kai Wang1, Yinzhang Ding3, Yansong Li1, Di Xu1
1Department of Geriatrics, The First Affiliated Hospital of Nanjing Medical University, No. 300 Guangzhou Road, Nanjing, Jiangsu 210029, China
2Department of Echocardiography, Jiangsu Province Hospital of Chinese Medicine, Affiliated Hospital of Nanjing University of Chinese Medicine, No. 155 Hanzhong Road, Nanjing, Jiangsu 210004, China
3Department of Cardiology, The First Affiliated Hospital of Nanjing Medical University, No. 300 Guangzhou Road, Nanjing, Jiangsu 210029, China
OBJECTIVES Myocardial fibrosis is an essential pathological feature of ventricular remodeling in post-myocardial infarction (MI). Dapagliflozin (DAPA), the sodium-glucose cotransporter 2 inhibitor (SGLT2i), is a new type of anti-glycemic agent, whose cardiovascular protective mechanisms have been the focus of recent research. However, the effects of dapagliflozin on myocardial fibrosis post-myocardial infarction are still unclear. It is objected to investigating the effects of dapagliflozin on myocardial fibrosis in post-myocardial infarction mice and the regulation of the Nrf2/HO-1 signaling pathway.
METHODS MI or sham-operated models were established in C57BL/6 mice, individually treated with saline (10 mL/kg) or 1 mg/kg DAPA by gavage once a day for 28 days since the operation day. In vitro, cardiac fibroblasts were isolated and cultured from 1 to 3 days C57BL/6J mice, and the cell model was constructed by oxygen-glucose deprivation (OGD). The in vitro models were treated with or without 1 μM DAPA. The Nrf2 inhibitor ML385 was applied to investigate whether it reverses the effect of dapagliflozin on the OGD cell model and to explore the potential mechanism of dapagliflozin to improve myocardial fibrosis. Cardiac structure and function, cardiac fibrosis, and oxidative stress were measured.
RESULTS Our results revealed that DAPA significantly improved cardiac function and remodeling, reduced cardiac fibrosis, and ameliorated oxidative stress in the post-MI model. The Nrf2/HO-1 signaling pathway was suppressed in the MI group compared to the Sham and Sham+DAPA groups and activated in the post-infarction mice treated with DAPA. Similar observations were made in the OGD model after treatment with DAPA. Moreover, the Nrf2 inhibitor ML385 reverses the effect of dapagliflozin on the OGD-induced cardiac fibroblasts.
CONCLUSIONS Dapagliflozin ameliorated cardiac function, myocardial fibrosis, and oxidative stress in mice post-myocardial infarction by a mechanism that may be related to the activation of the Nrf2/HO-1 signaling pathway. In an OGD-induced cardiac fibroblast model, dapagliflozin ameliorated myocardial fibrosis by activating the Nrf2/HO-1 signaling pathway.
GW34-e0440
Dmitry Kashirskikh1,2,3, Nelya Chicherina1,4, Vadim Cherednichenko3, Igor Sobenin2,5, Alexander Orekhov1,2,3
1Institute for Atherosclerosis Research
2Laboratory of Angiopathology, Institute of General Pathology and Pathophysiology
3Laboratory of Cellular and Molecular Pathology of Cardiovascular System, Avtsyn Research Institute of Human Morphology of FSBI “Petrovsky National Research Centre of Surgery”
4Sirius University of Science and Technology
5Laboratory of Medical Genetics, National Medical Research Center of Cardiology
OBJECTIVES Pathological processes in the human body lead to various modifications of native low-density lipoproteins (LDL), and desialylation is a well-known atherogenic modification. Desialylated LDL (desLDL) are highly susceptible to oxidation, aggregation, and self-association, which can contribute to the development of atherosclerosis. Moreover, desLDL particles are actively phagocytosed by macrophages and smooth muscle cells in the aorta. Although atherosclerosis has been extensively studied in mouse models, there is currently no appropriate model to study LDL desialylation in mice. Present study aims to establish a mouse model that accurately simulates the desialylation of LDL.
METHODS Control group of C57Bl/6J wild-type mice (n=70 in each group) was injected with saline, while the experimental group was subjected to a single injection of immobilized Vibrio cholerae neuraminidase with an enzymatic activity of 0.2 U/mL. At each time point (1–7 days after a single injection), 10 mice from both groups were randomly selected for sacrifice. The control group (n=30 in each group) received saline injections for 6 weeks, while the experimental group underwent a series of injections of immobilized Vibrio cholerae neuraminidase for 6 weeks. At 2, 4, and 6 weeks, 10 mice were randomly selected from both the control and experimental groups. The sialic acid content in LDL samples was determined using the Warren method. Blood lipids were measured using commercial kits. Atherosclerotic burden in the mouse aorta was quantified by staining with Oil Red O and hematoxylin-eosin.
RESULTS A single injection of 100 μL of immobilized Vibrio cholerae neuraminidase solution at a concentration of 0.2 U/mL reduced LDL sialic acid levels by an average of 25±7.2% for 5 days. On days 1 to 5 after a single injection of immobilized neuraminidase, there was a significant decrease in LDL sialic acid content compared to the control group (P<0.05). The reductions were 34.3±4.2%, 27.4±4.3%, 18.2±2.8%, 28.0±1.8%, and 17.0±6.2% on days 1 to 5, respectively. Additionally, the sialic acid content in LDL was restored to initial values in the experimental group on days 6–7. Decreased sialic acid LDL levels were observed at 2, 4, and 6 weeks after the start of the series of injections of immobilized neuraminidase (14.3±1.4%, 38.26±5.8%, and 29±3.7% reductions, respectively) compared to the control group. No atherosclerotic lesions were observed in the aortic cross-sections stained with hematoxylin-eosin or in aortas stained with Oil Red O in C57Bl/6J wild-type mice after subchronic administration of immobilized Vibrio cholerae neuraminidase at a concentration of 0.2 U/mL for 6 weeks. Furthermore, LDL desialylation did not lead to significant changes in the levels of blood lipids in the experimental group compared to the control group of C57Bl/6J wild-type mice.
CONCLUSIONS Our findings demonstrate that LDL desialylation, a characteristic previously found in patients with atherosclerosis, can be successfully modeled in wild-type mice Notably, the administration of immobilized neuraminidase did not result in the development of atherosclerotic lesions in the aortic intima or alterations in blood lipid levels in wild-type mice. This work was supported by the Russian Science Foundation (Grant No. 22-25-00391).
GW34-e0441
Dmitry Kashirskikh1,2,3, Nelya Chicherina1,4, Vadim Cherednichenko3, Igor Sobenin2,5, Alexander Orekhov1,2,3
1Institute for Atherosclerosis research
2Laboratory of Angiopathology, Institute of General Pathology and Pathophysiology
3Laboratory of Cellular and Molecular Pathology of Cardiovascular System, Avtsyn Research Institute of Human Morphology of FSBI “Petrovsky National Research Centre of Surgery”
4Sirius University of Science and Technology
5Laboratory of Medical Genetics, National Medical Research Center of Cardiology
OBJECTIVES Atherosclerosis is a multifactorial disease characterized by the formation of atherosclerotic plaques in arterial walls. Desialylation of low-density lipoproteins (LDL), a modification of LDL particles, is known to be a key contributor to the initiation and progression of atherosclerosis. However, the lack of suitable animal models has impeded the study of this phenomenon. The present study sought to investigate the impact of LDL desialylation on the development of atherosclerotic lesions in ApoE−/− mice, with the aim of establishing a reliable animal model for exploring this modification.
METHODS The control group of ApoE−/− mice received a series of saline injections, while the experimental group of ApoE−/− mice (n=10 in each group) received a series of injections of 20 mU of neuraminidase for 6 weeks. Another experimental group of ApoE−/− mice received a series of saline injections and a high-fat diet (HFD) for 6 weeks. Animals were sacrificed 6 weeks after the start of the experiment. Atherosclerotic lesions were identified by Oil Red O staining.
RESULTS It was shown that in the experimental group of ApoE−/− mice, the area of atherosclerotic lesions was increased by 29.6±8.9% after a series of injections of neuraminidase compared to the control group of mice on a chow diet (P<0.01). Furthermore, LDL desialylation in ApoE−/− mice resulted in a 48.5±5.95% increase in the area of atherosclerotic lesions compared to the control group of mice on a chow diet (P<0.001). Atherosclerotic lesions were predominantly observed in the aortic arch and surrounding areas of large artery branching points, including the brachiocephalic, left carotid, and left subclavian arteries. In addition, atherosclerotic lesions were also present spontaneously throughout the aorta.
CONCLUSIONS The findings of this study suggest that LDL desialylation in ApoE−/− mice significantly increases the development of atherosclerosis. Our results demonstrate that LDL desialylation in vivo in ApoE−/− mice results in an increase in the area of atherosclerotic lesions, similar to that observed in ApoE−/− mice fed an atherogenic diet. This work was supported by the Russian Science Foundation (Grant No. 22-25-00391).
GW34-e0442
Dmitry Kashirskikh1,2,3, Igor Sobenin2,4, Alexander Orekhov1,2,3
1Institute for Atherosclerosis Research
2Laboratory of Angiopathology, Institute of General Pathology and Pathophysiology
3Laboratory of Cellular and Molecular Pathology of Cardiovascular System, Avtsyn Research Institute of Human Morphology of FSBI “Petrovsky National Research Centre of Surgery”
4Laboratory of Medical Genetics, National Medical Research Center of Cardiology
OBJECTIVES Numerous studies have established a clear association between desialylation of low-density lipoproteins (LDL) and atherosclerosis. The subchronic administration of immobilized neuraminidase may mimic the pathological reduction in LDL sialic acid levels previously observed in individuals with atherosclerosis, and enable the examination of its deleterious effects. The combination of ApoE−/− mice and neuraminidase injections presents an opportunity to investigate whether sustained LDL desialylation promotes the development of atherosclerosis. The aim of this study was to investigate the effect of LDL desialylation on blood lipid levels in ApoE−/− mice.
METHODS The experimental group of ApoE−/− mice (n=10) received a series of injections of 20 mU of neuraminidase for 6 weeks, while the control group of ApoE−/− mice (n=10) received a series of saline injections. In addition, there was another experimental group of ApoE−/− mice fed a high-fat diet (HFD) and received saline injections for a duration of 6 weeks. At the end of the 6-week experiment, the animals were sacrificed. The levels of blood lipids, including total cholesterol, LDL cholesterol (LDL-C), high-density lipoprotein (HDL) cholesterol (HDL-C) and triglycerides (TG), were assessed at the endpoint of a 6-week experiment in mice.
RESULTS It was revealed that plasma levels of total cholesterol and LDL-C in ApoE−/− mice significantly increased by 22.4±4.6% (P<0.001) and 28±1.1% (P<0.01), respectively, after the 6-week series of neuraminidase injections compared to the lipid levels in the control ApoE−/− mice. Furthermore, there were no significant differences in the levels of TG and HDL-C between the experimental group of ApoE−/− mice receiving neuraminidase injections and the control group of ApoE−/− mice on a chow diet. The ApoE−/− mice on the HFD, which consisted of mice on an atherogenic diet, exhibited elevated plasma levels of total cholesterol and LDL-C by 40.8±3.6% and 46±10.5% (P<0.001), respectively, compared to the control group of ApoE−/− mice on a chow diet. Moreover, TG levels did not change significantly, while HDL-C was significantly reduced by 54.3±8.6% (P<0.001) in the ApoE−/− mice on the HFD group compared to the values of the control ApoE−/− mice on a chow diet.
CONCLUSIONS The development of atherosclerosis in ApoE−/− mice subjected to a series of neuraminidase injections was characterized by notable elevations in the levels of total cholesterol and LDL-C in the plasma. These alterations closely resembled the changes observed in the group of mice maintained on an atherogenic diet. Our findings highlight the potential influence of neuraminidase on cholesterol metabolism and suggest a potential role in the pathogenesis of atherosclerosis. This work was supported by the Russian Science Foundation (Grant No. 23-45-00031).
GW34-e0443
Dmitry Kashirskikh1,2,3, Igor Sobenin2,4, Alexander Orekhov1,2,3
1Institute for Atherosclerosis Research
2Laboratory of Angiopathology, Institute of General Pathology and Pathophysiology
3Laboratory of Cellular and Molecular Pathology of Cardiovascular System, Avtsyn Research Institute of Human Morphology of FSBI “Petrovsky National Research Centre of Surgery”
4Laboratory of Medical Genetics, National Medical Research Center of Cardiology
OBJECTIVES Modifications of native low-density lipoproteins (LDL) play a critical role in the development of atherogenesis. Desialylated LDL is one of the modifications found in individuals with atherosclerosis. Notably, desialylated LDL has been identified as being more prone to self-association and accumulation within the intima. The enzymatic process responsible for LDL desialylation is primarily mediated by neuraminidases. Therefore, studying the alterations in the expression patterns of endogenous neuraminidases within different types of atherosclerotic lesions and healthy areas can significantly contribute to our understanding of the underlying mechanisms involved in atherosclerosis. The objective of this study was to evaluate the expression of endogenous neuraminidases in atherosclerotic lesions of patients with atherosclerosis.
METHODS The expression levels of lysosomal (NEU1), cytosolic (NEU2), plasma (NEU3), and mitochondrial (NEU4) neuraminidase genes were assessed using Real-time qPCR. Autopsy aorta samples were collected, including fatty streaks, lipofibrous plaques, and fibrous plaques. Unaffected human aortic intima samples served as the control. RNA isolation from the samples was performed using TRIzol, followed by DNase I treatment for DNA elimination. RNA was converted into cDNA using a two-step RT-PCR procedure.
RESULTS It was found that NEU1 gene transcripts were elevated in fatty streaks by 2-fold (P=0.05) when compared to healthy tissues. However, NEU1 mRNA levels were observed to be lower in fibrous and lipofibrous plaques. Lipofibrous plaques exhibited a 5-fold decrease in NEU2 and NEU3 gene expression (P=0.01) compared to healthy tissue. Additionally, lipofibrous plaques displayed increased NEU4 mRNA expression by 2-fold (P=0.005) compared to the control.
CONCLUSIONS Our study revealed distinct alterations in the expression levels of neuraminidase genes (NEU1–NEU4) in different stages of atherosclerotic plaque development. Fatty streaks exhibited an upregulation of NEU1 gene transcripts, suggesting increased neuraminidase activity in these early lesions. However, NEU1 mRNA was found to be less expressed in fibrous and lipofibrous plaques, suggesting a downregulation of NEU1 gene expression during plaque progression. Notably, lipofibrous plaques showed a significant decrease in NEU2 and NEU3 gene expression, indicating a potential role for these neuraminidases in the advanced stages of plaque formation. Interestingly, NEU4 mRNA levels were elevated in lipofibrous plaques, indicating a potential involvement of mitochondrial neuraminidase in the pathogenesis of these plaques. These findings provide valuable insights into the dynamic regulation of neuraminidase genes during atherosclerotic plaque development and highlight their potential significance in plaque pathophysiology. Further investigations are warranted to elucidate the specific functional implications of these neuraminidase alterations in the context of atherosclerosis. This work was supported by the Russian Science Foundation (Grant No. 23-45-00031).
GW34-e0446
Jinyao Liu1, Yumiko Oba1, Yaowei Chang1, Seiko Yamano2
1Graduate School of Medicine, Yamaguchi University
2Science Research Center, Institute of Life Science and Medicine, Yamaguchi University
OBJECTIVES Fatty liver disease has increased rapidly, and the risk of developing cardiovascular events in metabolic disorder-associated fatty liver disease (MAFLD) is higher than in non-alcoholic fatty liver disease. On the other hand, about 70% of adults are experienced drinker (nonheavy alcohol consumption), and the factors related to metabolic disorders, such as nutritionally unbalanced diet and non-excessive alcohol consumption, account for a considerable proportion of the population which may have a synergistic effect on the progression of liver disease. In the present study, the risk of sudden cardiac death in advanced MAFLD caused by an atherosclerosis-inducing diet (low-carbohydrate, high-protein, high-fat diet: AD) in parallel with non-excessive drinking are confirmed. Sympathetic nerve activation in the heart, myocardial fibril formation, and gap junction protein-related gene variations are assessed at the same time.
METHODS Thirteen-week-old hyperlipidemic spontaneous transgenic mice (apolipoprotein E/low-density lipoprotein receptor double-knockout mice: AL) were divided into four groups and maintained for 16 weeks i.e., AL mice on a standard chow diet (SCD) with or without ethanol (Et) treatment and AL mice on an AD with or without Et treatment. Age-matched male C57BL/6J mice on SCD without Et treatment served as controls. Blood is used to evaluated liver function and hyperlipidemia. Arrhythmia provocation test (acute restraint and intraperitoneal epinephrine injection) is used to confirm the risk of sudden cardiac death in advanced MAFLD. Liver and left ventricular (LV) tissues are used to evaluated liver and LV sympathetic nerve activation, fatty liver, hepatic inflammatory response and fibrogenesis (formation of fibrous bridges indicating progressive MAFLD), myocardial fibrogenesis and gap junction protein synthesis-related mRNA expression with pathology tests and RT-PCR.
RESULTS A combination of nonheavy alcohol consumption and AD for 16 weeks caused obesity, hyperlipidemia, hepatomegaly and liver dysfunction, and cardiac hypertrophy. It caused increased intrahepatic fat deposition, ballooning of hepatocytes, hepatic lobular inflammation, increased Kupffer cells and formation of hepatic fibrous bridges accompanying with high arrhythmia induction rate with two of 8 mice died. Regarding the onset mechanism, we focused on sympathetic nerve activation. In liver, hepatic Kupffer cells and stellate cells exhibited sympathetic activation with increased inflammatory response and fibrogenesis, which may be one of the pathogenesis mechanisms of progressive MAFLD. In LV, myocardium showed cardiac sympathetic nerve activity with myocardial interstitial fibril formation, up-regulated gap junction protein synthesis-related genes, which may be one of the mechanisms for the high risk of fatal arrhythmias.
CONCLUSIONS MAFLD caused by a combination of nonheavy alcohol consumption with a nutritionally imbalanced diet may be associated with the high risk of sudden cardiac death.
GW34-e0450
Zheng Jia, Zheng-jiang Xing, Hong-lin Zou, Jie Wei, Fan-di Meng, Ying Xie
Kunming Medical University Affiliated to Yan’an Hospital
OBJECTIVES Coronary atherosclerotic heart disease (CAD), as a major cardiovascular disease, is the leading cause of death worldwide. The Klotho/FGF23 axis is involved in the occurrence and development of cardiovascular diseases, but the role and underlying mechanism of the Klotho/FGF23 axis in CAD are still unclear.
METHODS Blood samples from 67 CAD patients undergoing coronary artery bypass grafting were collected, and the levels of Klotho and FGF23 were detected by ELISA. Cardiomyocytes from SD rats aged 0–3 days were isolated and cultured. Klotho, FGF23 and cardiomyocyte markers α-actin (α-SA), myosin heavy chain (MHC) and cardiac troponin I (cTnI) were detected by immunofluorescent staining. The expression of Klotho and FGF23 mRNA was detected by qRT-PCR. Cell apoptosis and cell cycle were detected by flow cytometry. Cell viability was detected by CCK-8. Western blotting was used to detect the expression of ERK/MAPK pathway-related proteins and the production of cytokines.
RESULTS The level of Klotho increased after CABG in patients with coronary heart disease, while the level of FGF23 decreased. The cardiomyocytes were integral in shape and structure, stable in beating after 15 days of culture, and positive for α-SA, MHC and cTnI. After lti-Klotho and lti-FGF23 were transfected into cardiomyocytes, fluorescent staining showed that the transfection was successful. The mRNA expression levels of Klotho and FGF23 were significantly higher than those in the NEG (empty vector) group showed to qRT-PCR results. Compared with the NEG group, the Klotho overexpression (Klotho) group had a higher Klotho positive rate and a lower FGF23 positive rate, while the FGF23 overexpression (FGF23) group had a higher Klotho positive rate and a lower Klotho positive rate displayed in the results of immunofluorescence staining. However, the expressions of p-ERK1/2 and p-P38 decreased in the Klotho group, but increased in the FGF23 group. In addition, overexpression of Klotho can inhibit cardiomyocyte apoptosis, increase the proportion of S phase, promote cell proliferation, increase transforming growth factor β1 (TGF-β1), nuclear factor κB (NF-κB), angiotensin-II (AT-II) and activator protein-1 (AP-1), overexpression of FGF23 had opposite effects, while ERK agonist (TPA) and inhibitor (U0126) reversed the effects of Klotho and FGF23.
CONCLUSIONS Klotho was negatively correlated with the expression of FGF23. The Klotho/FGF23 axis can regulate cardiomyocytes through the ERK/MAPK pathway and play an important role in the progression of CAD.
GW34-e0458
Yuto Kawase1, Kana Shimizu1,2, Masafumi Funamoto1,2, Yoichi Sunagawa1,2,3, Yasufumi Katanasaka1,2,3, Toshihide Hamabe-Horiike1,2,3, Satoshi Shimizu1,2,3, Philip Hawke4, Maki Komiyama2, Koji Hasegawa1,2, Tatsuya Morimoto1,2,3
1Division of Molecular Medicine, School of Pharmaceutical Sciences, University of Shizuoka, Shizuoka 422-8526, Japan
2Clinical Research Institute, Kyoto Medical Center, National Hospital Organization, Kyoto, Japan
3Shizuoka General Hospital, Shizuoka, Japan
4Laboratory of Scientific English, School of Pharmaceutical Sciences, University of Shizuoka, Shizuoka 422-8526, Japan
OBJECTIVES Cardiac remodeling induced by hypertrophic stresses such as hypertension or myocardial infarction is a compensatory mechanism associated with cardiomyocyte hypertrophy and cardiac fibrosis. Continuation of these responses eventually result in heart failure (HF). The histone acetyltransferase p300 plays an important role in the development of HF and may be a target for HF therapy. The phenolic phytochemical 6-shogaol, a pungent component of raw ginger, has various bioactive effects. However, its effect on HF has not been investigated. This study examined the effect of 6-shogaol on cardiomyocyte hypertrophy and fibrosis phenotype in vitro and on the development of heart failure in vivo.
METHODS To investigate the inhibitory effect of 6-shogaol against p300-HAT activity, an in vitro HAT assay was performed using a recombinant p300-HAT domain and core histones. Next, primary cultured cardiomyocytes and cardiac fibroblasts were treated with 1 μM 6-shogaol and then stimulated with phenylephrine (PE) or transforming growth factor-beta (TGF-β), respectively. Immunofluores staining, quantitative PCR (qPCR) analysis, and western blotting (WB) were performed on the cardiomyocytes. Measurement of L-proline incorporation, qPCR analysis, and WB were carried out on the cardiac fibroblasts. C57BL/6J mice were subjected to transverse aortic constriction (TAC) surgery, and then given a daily oral administration of 1 mg/kg 6-shogaol for 8 weeks. Echocardiographic analysis, histological staining, and WB were performed.
RESULTS An in vitro p300-HAT assay revealed that 6-shogaol significantly suppressed histone acetylation. In cardiomyocytes, immunofluores staining revealed that 6-shogaol significantly suppressed PE-induced increase in cell surface area. qPCR analysis demonstrated that 6-shogaol significantly suppressed PE-induced increases in the mRNA levels of hypertrophic response genes such as ANF and BNP. 6-shogaol also suppressed PE-induced increase in histone acetylation. Next, in primary cultured cardiac fibroblasts, 6-shogaol suppressed TGF-β-induced increases in L-proline incorporation, the mRNA and protein expression levels of α-smooth muscle actin, and histone H3K9 acetylation. Finally, echocardiography showed that 6-shogaol prevented the TAC-induced increase in the posterior wall thickness and decrease in the systolic function. Histological staining revealed that 6-shogaol suppressed the increases in the cardiomyocyte cross-sectional area and the area of perivascular fibrosis induced by TAC surgery. 6-shogaol also suppressed TAC-induced increase in histone H3K9 acetylation.
CONCLUSIONS These results indicate that 6-shogaol, a ginger extract, suppressed both PE-induced cardiomyocyte hypertrophy in cardiomyocytes and TGF-β-induced differentiation into myofibroblasts in cardiac fibroblasts. 6-shogaol also suppressed pressure overload-induced HF by inhibiting p300-HAT activity. Clinical studies are required to assess the potential of 6-shogaol as a heart failure treatment.
GW34-e0460
Yaowei Chang1, Jinyao Liu1, Yumiko Oba1, Seiko Yamano2
1Yamaguchi University Graduate School of Medicine, Ube, Yamaguchi, Japan
2Science Research Center, Institute of Life Science and Medicine, Yamaguchi University, Ube, Japan
OBJECTIVES Atherosclerosis is a chronic arterial disease and a major cause of vascular death. Dysfunction of endothelial progenitor cells (EPCs), with their ability to replace old and injured cells and differentiate into healthy and functional mature endothelial cells, contributes to the development of atherosclerosis. Using apolipoprotein E/low-density lipoprotein receptor double-knockout (AL) mice, the influences of a combination of alcohol consumption with an atherosclerosis-inducing diet (AD, a low-carbohydrate–high-protein-high-fat atherogenic diet) on EPCs and their regulatory factors were assessed.
METHODS We maintained 13-week-old male AL mice on an AD with or without ethanol (Et) treatment for 16 weeks. Age-matched male C57BL/6J (WT) mice on a standard chow diet (SCD) without ethanol treatment served as controls. The aorta (from the aortic root to the iliac bifurcation) were harvested after systemic perfusion with Hanks’ balanced salt solution via the left ventricle. Flow cytometry was performed to analyze aortic EPCs accumulation identified as CD34+/CD45- cells in the whole of the aortic cell suspension. The EPCs regulatory factors (CD133, FLK-1, and NRP-1) were evaluated with RT-PCR.
RESULTS AD without Et treatment induced the increased EPCs in the whole of the aortic cell suspension, however, the combination of AD and Et suppressed this increase accompanying with CD133, FLK-1 and NRP-1 mRNA expression down-regulations which up-regulated by AD.
CONCLUSIONS This study indicated that the synergistic effect of alcohol consumption and an atherosclerosis-inducing diet inhibited the increases in EPCs and its regulatory factors mRNA expressions, resulted in the dysfunction of EPCs recruitment and differentiation, which may be responsible for the progression of atherosclerosis.
GW34-e0474
Yanru Duan1, Yunhui Du2
1Department of Pathophysiology, School of Basic Medical Sciences, Capital Medical University, Beijing 100069, P. R. China
2Beijing Anzhen Hospital, Capital Medical University, Beijing Institute of Heart, Lung and Blood Vessel Diseases, Beijing 100029, P. R. China
OBJECTIVES APN, adiponectin, is a vascular protective molecule secreted by adipocytes. APPL1, adiponectin receptor-binding protein-like 1, functions as an adaptor and is also involved in vascular protection. APN exerts its vascular protective effects by interacting with APPL1. However, the downstream target genes that mediate the protective effects of this signaling axis on vascular endothelium in the context of diabetes remain unclear.
METHODS High-fat-diet (HFD)-induced diabetic vascular injury and hindlimb ischemia models were used for in vivo function demonstration. Diabetic medium-cultured human umbilical vein endothelial cells (HUVECs) and 293 T cells were utilized for in vitro molecular/cellular investigation. Transcription factor expression detection kits, transcription factor activity assay kits, and mass spectrometry techniques were employed to mechanism exploration.
RESULTS APPL1 knock-out mice were fed normal or HFD for 10 weeks. Eight weeks after HFD, mice were randomized to receive vehicle or APN treatment for another two weeks. Aortic vessels were prepared for transcriptome analysis. Bioinformatics analysis indicates that the Tight Junction (TJ) pathway exhibited the most prominent changes. Using a global signaling transduction network showed that TJ-related genes had the highest degree values. Among these genes, PCR results indicated that Occludin was the sole gene inhibited in endothelial cells treated by HG/HL and promoted by APN treatment in an APPL1-dependent manner. The protein level of Occludin was reduced in diabetic condition, and APN treatment upregulated the expression of Occludin in APPL1-mediated manner in vivo and vitro. Endothelial specific OCLN knockdown blunted APN-mediated blood flow recovery after femoral artery ligation in vivo, and abolished APN’s effects upon HG/HL-induced permeability, and apoptosis in vitro. Mechanically, we discovered MYB, HDAC1, and FOXO1 play a role in the regulation of Occludin. This finding was validated by CHIP-QPCR. Western blot analysis revealed that the APN-induced increase in Occludin protein level was abolished after knockdown of MYB and FOXO1. However, knockdown of HDAC1 in endothelial cells did not alter the expression of Occludin mediated by APN. Finally, CHIP-qPCR showed that both MYB and FOXO1 were involved in the enrichment of Occludin promoters mediated by APN.
CONCLUSIONS Our findings provide the first evidence that the APN/APPL1 signaling axis promotes the expression of Occludin through the transcription factors MYB and FOXO1, ultimately exerting a protective effect on vascular endothelium. This suggests that Occludin may serve as a novel therapeutic target for treating diabetic vascular endothelial injury.
GW34-e0485
Mengqiu Dang, Junyan Jin
Department of Cardiology, Cardiovascular Key Laboratory of Zhejiang Province, The Second Affiliated Hospital, College of Medicine, Zhejiang University, Hangzhou, People’s Republic of China
OBJECTIVES Cardiac hypertrophy is a key remodeling during diabetic cardiomyopathy, leading to heart dysfunction. Recent studies have demonstrated that mDia1 regulated the filamentous/globular (F/G)-actin ratio in response to pressure overload in mice. However, the mechanisms of mDia1 remain not fully understood in the progression of diabetic cardiomyopathy.
METHODS High-fat diet (HFD) for 16 weeks was used to induced diabetic cardiomyopathy. Male wild-type (WT) and mDia1-knockout (mDia1KO) mice were subjected to HFD and normal diet (ND). Echocardiography was used to accessed cardiac function. Pathological staining and mRNA expression of ANP, BNP as well as β-MHC were performed to access cardiac remodeling. Proteomic profiling and Glycometabolomics used to evaluate molecular changes.
RESULTS Blood glucose increased after HFD for 8 weeks. The expression of mDia1 was significant increased in HFD mice. mDia1 knockout decreased cell area, IVRT and E/A ratio meanwhile increased left ventricular diastolic volum. Whereas, LVEF was not changed after disruption of mDia1 as well as FS. A quantitative proteomics analysis revealed significant increases of mitochondrial pyruvate carrier 2 (MPC2) after mDia1 knockout in mice which was the unique entry point for the glycolytic end-product pyruvate to the mitochondria. Furthermore, results of glycometabolomics indicated that mDia1 knockout increased the products of glucose metabolism and tricarboxylic acid cycle including D-glucose-6-phosphate, alpha-ketoglutaric acid, fumaric acid malic acid and Adenosine monophosphate as well as adenosine diphosphate.
CONCLUSIONS Our results suggested that mDia1 deteriorates diabetic cardiomyopathy via decreasing of MPC2 expression, which is resulted to reducing glucose metabolism.
GW34-e0489
Changjin Li
Department of Cardiovascular Medicine, Changhai Hospital, Naval Medical University, Shanghai 200433, China
OBJECTIVES To explore the identification of clotting and fibrinolysis-related genes (CFRG) in atherosclerosis (AS) and analyze immune infiltration using bioinformatics techniques, aiming to determine the impact of CFRG in AS.
METHODS We performed differential expression analysis (AS samples vs normal samples) to obtain differentially expressed genes (DEGs) in GSE100927 dataset. Coagulation and fibrinolysis related differentially expressed genes (CFR-DEGs) were obtained by overlapping DEGs and CFRGs. Further, generalized linear model (GLM), grandient boosting machine (GBM) and randomForest (RF) algorithm were implemented to build a diagnostic model. We further performed immune infiltration and gene set enrichment analysis (GSEA) based on diagnostic genes.
RESULTS We identified 2211 DEGs associated with AS. Then, 13 CFR-DEGs were obtained via venn diagram. Subsequently, 5 feature genes (TNF, SERPINA3, F12, PLAU and CSF3) were identified via machine learning. Moreover, 4 diagnostic biomarkers associated with coagulation and fibrinolysis, including F12, PLAU, SERPINA3 and TNF, were screened via receiver operating characteristic (ROC) analysis. The immune infiltration and Gene Set Enrichment Analysis (GSEA) analysis suggested that these diagnostic biomarkers were related to the function of cytokine interaction pathway and some differential immune cells. Finally, we found significant lower expression of CSF3 and SERPINA3 in AS group compared to the normal group by RT-qPCR.
CONCLUSIONS This study identified four CFRG in AS: IF12, PLAU, SERPINA3, and TNF. Analyses of drug networks and immune cell infiltration shed light on the molecular mechanisms of CFRG in AS, providing potential directions for AS treatment.
GW34-e0495
Mizuho Yamamoto1, Masafumi Funamoto1,2, Yoichi Sunagawa1,2,3, Sho Uehara1, Yasufumi Katanasaka1,2,3, Toshihide Hamabe-Horiike1,2,3, Kana Shimizu1,2, Satoshi Shimizu1, Koji Hasegawa1,2, Tatsuya Morimoto1,2,3
1Division of Molecular Medicine, School of Pharmaceutical Sciences, University of Shizuoka
2Division of Translational Research, National Hospital Organization Kyoto Medical Center
3Shizuoka General Hospital
OBJECTIVES Epigenetic regulatory mechanisms such as histone post-translational modifications are involved in the development of heart failure (HF). The histone acetyltransferase p300 accelerates gene transcription via epigenetic mechanisms, including chromatin remodeling, in which chromatin changes structurally from heterochromatin to euchromatin. Acetylation of the histone tail domain has been intensively studied, including the domains H3 lysine 9 (H3K9) and 14 (H3K14). Recently, H3 lysine 122 (H3K122) has been reported to be a novel p300 histone acetylation site. H3K122 is located at a globular domain on the surface of the histone. H3K122 acetylation attenuates histone-DNA binding and dramatically activates gene transcription. However, nothing is known about the acetylation of the histone tail and globular domains in left ventricular hypertrophy (LVH) or HF. Therefore, the present study aims to examine how the acetylation of the histone tail and globular domains changes during the transition from the LVH to HF stages in Dahl rats, a model of hypertension-induced HF.
METHODS To analyze the acetylation levels of H3K9 and H3K122 during the transition from LVH to HF in salt-sensitive Dahl rats, Western blotting (WB) was performed. Next, a chromatin-immunoprecipitation (ChIP) assay was performed to examine the acetylation levels of H3K9 and H3K122 on the promoters of cardiac hypertrophic response genes, ANF and BNP. In addition, we analyzed the recruitment of p300 onto the promoters of the cardiac hypertrophy response genes using a ChIP assay. Finally, we investigated the expression levels of BRG1, a chromatin remodeling factor, and the binding of BRG1 to p300 during the transition from LVH to HF. To examine the recruitment of BRG1 on the promoter of hypertrophic response genes, a ChIP assay was performed.
RESULTS WB revealed that acetylation of H3K9 was increased in LVH, and that of H3K122 was increased in HF. ChIP assay showed that acetylation of H3K9 was increased in LVH and acetylation of H3K122 was increased in HF on the cardiac hypertrophic response gene promoter and that recruitment of p300 to the hypertrophic response factor promoters was comparable in LVH and HF. IP-WB results showed that the binding of BRG1 to p300 was significantly increased in HF. Furthermore, in vivo ChIP assay showed that the recruitment of BRG1 to the hypertrophic response factor promoters was increased considerably in HF compared to LVH.
CONCLUSIONS These data suggest that the acetylation of the histone globular domain H3K122 is enhanced during the transition from LVH to HF and that the formation of the p300/BRG1 complex is involved in the acetylation of H3K122. These findings may contribute to the discovery of a novel epigenetic regulation mechanism for HF, which may, in turn, lead to a much-needed new treatment for the disease.
GW34-e0535
Yan-Yun Pan
The First Affiliated Hospital of Zhejiang Chinese Medical University
OBJECTIVES During the pathogenesis of early pulmonary arterial hypertension (PAH), pulmonary arterial adventitial fibroblast act as an initiator and mediator of inflammatory processes that predispose vessel walls to excessive vasoconstriction and pathogenic vascular remodeling. Emerging studies report that Yin Yang-1 (YY-1) plays important roles in inflammatory response and vascular injury. Our recent study finds that activation of CD40L-CD40 signaling promotes pro-inflammatory phenotype of pulmonary adventitial fibroblasts. However, whether YY-1 is involved in CD40L-CD40 signaling-triggered inflammatory response in pulmonary adventitial fibroblasts and its underlying mechanism is still unclear.
METHODS Here, we show that soluble CD40L (sCD40L) stimulation promotes YY-1 protein expression and suppresses anti-inflammatory cytokine, IL-10 expression in pulmonary adventitial fibroblasts, while YY-1 knockdown prevents sCD40L-mediated reduction of IL-10 expression via enhancing IL-10 gene transactivation. Further, we find that sCD40L stimulation significantly increases H3K27me3 modification on IL-10 promoter in pulmonary adventitial fibroblasts, and YY-1 knockdown prevents the effect of sCD40L on IL-10 promoter through reducing recruitment of EZH2, a histone methyltransferase, binding to IL-10 promoter.
RESULTS Moreover, we find that sCD40L stimulation promotes YY-1 protein, but not mRNA expression, via decreasing m6A methylation on YY-1 mRNA to suppress YTHDF2-medicated mRNA decay. Overall, this in-depth study shows that activation of CD40L-CD40 signaling upregulates YY-1 protein expression in pulmonary adventitial fibroblasts, which results in increasing YY-1 and EZH2 binding to the IL-10 promoter region to enhance H3K27me3 modification, eventually leading to suppress IL-10 transactivation.
CONCLUSIONS This study firstly uncovers the roles of YY-1 on CD40L-CD40 signaling-triggered inflammatory response in pulmonary adventitial fibroblasts.
GW34-e0536
Xianjun Wu
First Affiliated Hospital of Zhejiang Chinese Medical University
OBJECTIVES Cardiovascular disease is a serious threat to human health because of its high mortality and morbidity rates. At present, there is no effective treatment. In Southeast Asia, traditional Chinese medicine is widely used in the treatment of cardiovascular diseases. Quercetin is a flavonoid extract of Ginkgo biloba leaves. Basic experiments and clinical studies have shown that quercetin has a significant effect in the treatment of cardiovascular diseases. However, its precise mechanism is still unclear. Therefore, it is necessary to exploit the network pharmacological potential effects of quercetin on Cardiovascular disease.
METHODS In present study, a novel network pharmacology strategy based on pharmacokinetic filtering, target fishing, Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis, compound-target-pathway network structured was performed to explore the anti-cardiovascular disease mechanism of quercetin.
RESULTS The outcomes showed that quercetin possess favorable pharmacokinetic profiles, which have interactions with 47 cardiovascular disease-related targets and 12 KEGG signaling pathways to provide potential synergistic therapeutic effects. Following the construction of Compound-Target-Pathway (C-T-P) network, and the network topological feature calculation, we obtained top 10 core genes in this network were AKT1, IL1B, TNF, IL6, JUN, CCL2, FOS, VEGFA, CXCL8, and ICAM1. KEGG pathway enrichment analysis indicated that quercetin produced the therapeutic effects against cardiovascular disease by systemically and holistically regulating many signaling pathways, including Fluid shear stress and atherosclerosis, AGE-RAGE signaling pathway in diabetic complications, TNF signaling pathway, MAPK signaling pathway, IL-17 signaling pathway, PI3K-Akt signaling pathway.
CONCLUSIONS The outcomes showed that quercetin possess favorable pharmacokinetic profiles, which have interactions with 47 cardiovascular disease-related targets and 12 KEGG signaling pathways to provide potential synergistic therapeutic effects. Following the construction of Compound-Target-Pathway (C-T-P) network, and the network topological feature calculation, we obtained top 10 core genes in this network were AKT1, IL1B, TNF, IL6, JUN, CCL2, FOS, VEGFA, CXCL8, and ICAM1. KEGG pathway enrichment analysis indicated that quercetin produced the therapeutic effects against cardiovascular disease by systemically and holistically regulating many signaling pathways, including Fluid shear stress and atherosclerosis, AGE-RAGE signaling pathway in diabetic complications, TNF signaling pathway, MAPK signaling pathway, IL-17 signaling pathway, PI3K-Akt signaling pathwa
GW34-e0540
Tan Nannan, Li Weili, Zhang Yawen, Ling Guanjing, Wei Yan, Wang Xiaoping
Beijing University of Chinese Medicine
OBJECTIVES Doxorubicin (DOX) is the cornerstone of clinical cancer therapy, but the clinical application is restricted due to the doxorubicin-induced cardiotoxicity (DIC). Restoring or enhancing autophagy flux is one of the most promising therapeutic strategies for DIC patients. Recent studies have found that abnormal mitochondrial-lysosomal contact could affect lysosomal function, and then lead to autophagy disorder. TBC1D15 plays a key role in this process. The role of TBC1D15 in myocardial infarction and cardiac ischemia-reperfusion models has been confirmed. Qishen granule (QSG) has a strong cardioprotective effect, but the pharmacological mechanism in DIC has not been elucidated. This study aimed to investigate whether QSG could protect against DIC from heart injury through activating autophagy via TBC1D15-mediated mitochondria-lysosome contact.
METHODS A DIC model was established via tail vein injection with DOX (5 mg·kg−1) weekly for 4 weeks and a DOX-induced H9C2 cell injury model was induced by 1 μmol/L DOX. In vivo, the mice were divided into five groups, control group, model group, QSG-low group, QSG-high group, and Pravastatin (positive drug) group. Echocardiographic assessment, histological examination, mRFP-GFP-LC3 double-labeled adenovirus and western blot analysis of TBC1D15 were collectively implemented to evaluate the cardioprotective effects of QSG. In vitro, the effects of over expressed TBC1D15 and QSG were evaluated by analyzing cell viability with a microplate reader using cell counting kit-8 (CCK-8). siRNA TBC1D15 was transfected to H9C2 with QSG, and then western blot was used for detecting the level of LC3 and P62, lysosome-sensor probe was used to detect the acidification of lysosome, which further verified whether QSG exerted its effect through TBC1D15.
RESULTS Overexpression of TBC1D15 could reverse cell viability, mitochondrial membrane potential and lysosomal acidification, and restore autophagy. QSG improved cardiac function and decreased serum levels of myocardial injury markers in vivo. Moreover, QSG could improve myocardial pathological damage, reduce the level of reactive oxygen species in myocardial tissue, reduce myocardial cell apoptosis, and repair mitochondrial cristae rupture. The TBC1D15 level in the DOX group was decreased and was increased after administration of QSG. The results of mRFP-GFP-LC3 double labeled adenovirus showed that the accumulation of autolysosomes in the QSG group was reduced. When TBC1D15 was knocked down by siRNA, the beneficial effect of QSG on mitochondria, autophagy and lysosomal acidification was disappeared.
CONCLUSIONS Qishen granule can improve Doxorubicin-induced cardiac dysfunction in mice by improving lysosomal acidification and restoring autophagy, and the mechanism is related to increasing the level of TBC1D15.
GW34-e0545
Bing Yan, Zhu Mei, Haixu Song, Dan Liu, Xiaolin Zhang, Chenghui Yan, Yaling Han
National Key Laboratory of Frigid Zone Cardiovascular Disease, Cardiovascular Research Institute and Department of Cardiology, General Hospital of Northern Theater Command, Shenyang 110016, China
OBJECTIVES CREG (Cellular repressor of E1A-stimulated genes) is a glycoprotein, which first described as a transcription repressor that represses the transcription and cellular transformation induced by the E1A oncoprotein. Recently, CREG is identified as a vascular protective molecule, and it is significantly downregulated in atherosclerotic vessels. Macrophages transform into foam cells upon taking-in lipids is crucial for atherogenesis, but no role for CREG in foam cell formation has previously been reported.
METHODS CREG expression was examined in western diet-fed atherosclerosis-prone ApoE−/− mice. CREG expression was examined after exposure of macrophages (Raw264.7 cells and bone marrow–derived macrophages) to OxLDL (oxidized low-density lipoprotein) in vitro. Foam cell formation, lysosomal maturation, and expression of scavenger receptor A (SRA) was examined in CREG deficiency (using CREG siRNA) macrophages (Raw264.7 cells and bone marrow–derived macrophages) in vitro. Further we generated macrophage-specific CREG-deficient mice in ApoE−/− background. Atherosclerotic lesion in whole aorta and aortic root were examined after ApoE−/− CREGfl/fl and ApoE−/− CREGfl/flLyz2cre mice fed with western diet for 16 weeks. SRA expression in aortic root lesions was also examined by immunostaining.
RESULTS CREG expression was markedly reduced in mice atherosclerotic plaques when compared with nonatherosclerotic vessels (Target protein levels fold, normal vs atherosclerotic vessel 1.000.06 vs 0.420.06, n=3, P<0.05; Relative transcript levels fold, normal vs atherosclerotic vessel 1.000.08 vs 0.670.09, n=3, P<0.05). Exposure of macrophages to OxLDL downregulated CREG in vitro (Target protein levels fold, control vs OxLDL 1.000.07 vs 0.530.05, n=3, P<0.05; Relative transcript levels fold, control vs OxLDL 1.000.09 vs 0.700.07, n=3, P<0.05). Furthermore, CREG deficiency in macrophages increased the expression of SRA (Target protein levels fold, siNC vs siCREG 1.000.21 vs 1.450.07, n=3, P<0.05). But CREG deficiency did not markedly change SRA mRNA levels (Relative transcript levels fold, siNC vs siCREG 1.000.01 vs 1.370.18, n=3, P=0.10). CREG deficiency also increased modified low-density lipoprotein uptake (Dii-Ac-LDL staining fold, siNC vs siCREG 0.990.23 vs 2.420.46, n=3, P<0.05) and foam cell formation (Oil red O staining fold, siNC vs siCREG 1.000.39 vs 2.810.39, n=3, P<0.05). Further we found CREG deficiency decrease lysosome staining and lysosomal marker protein LAMP-2 (Target protein levels fold, siNC vs siCREG 1.000.14 vs 0.540.08, P<0.05). Compared with ApoE−/− CREGfl/fl wild type control ApoE−/− CREGfl/fl Lyz2cre mice developed more atherosclerotic lesions in whole aorta (Lesion area % of total area, ApoE−/− CREGfl/fl vs ApoE−/− CREGfl/flLyz2cre 37.769.05 vs 49.786.235, n=7, P<0.05) and aortic root area (Lesion area % of total area, ApoE−/− CREGfl/fl vs ApoE−/− CREGfl/flLyz2cre 28.576.58 vs 48.7710.89, n=7, P<0.05), with increased SRA expression in aortic root lesions.
CONCLUSIONS Our study demonstrated that macrophage CREG reduction caused by OxLDL decreases SRA degradation and lysosomal maturation, which promotes foam cell formation and the progression of atherosclerosis. These results suggest CREG might be a novel therapeutic target for inhibition of atherosclerosis.
GW34-e0547
Yaohan Tang, Chenhui Yan, Yaling Han
National Key Laboratory of Frigid Zone Cardiovascular Disease, Cardiovascular Research Institute and Department of Cardiology, General Hospital of Northern Theater Command, Shenyang 110016, China
OBJECTIVES Acute myocardial infarction (AMI) refers to myocardial ischemia and hypoxia caused by coronary artery obstruction. Myocardial mitochondrial function is impaired during ischemia-hypoxia, which leads to insufficient mitochondrial function and ATP production, and then affects the function of myocardial cells. Voltage-dependent anion-selective channel 3 (VDAC3), a major mitochondrial outer membrane protein, is known to be essential for mitochondrial function. However, the role of VDAC3 in acute myocardial infarction has not been reported. The aim of this study is to clarify the role and mechanism of VDAC3 in AMI.
METHODS AMI was induced by permanent ligation of the left anterior descending coronary artery. The autophagy of myocardium was observed by transmission electron microscope. Overexpression or knockdown of VDAC3 was performed in vivo using adeno-associated viruses. Plasmid and siRNA were used to over-express and knock down VDC3 in HL-1 cells, respectively. Western Blot and RT-PCR were used to detect the expression of VDAC3, LC3, P62, BNP, IL-6 and IL-1β. CCK-8 kit was used to detect cell viability.
RESULTS The protein and mRNA levels of VDAC3 were significantly decreased at 3 d, 7 d, 14 d and 28 d after AMI (P<0.05), and the decrease was most obvious at 28 d. AAV-VDAC3 overexpression significantly reduced mortality (16.7 vs 46.1%, P<0.05), improved cardiac function, and reduced infarct size and fibrosis area in AMI mice. After VDAC3 knockdown by adeno-associated virus, sh-VDAC3 significantly increased the mortality (27.3 vs 77.3%, P<0.05) and cardiac function, as well as the area of myocardial infarction and fibrosis in AMI mice. Transmission electron microscopy results showed that autophagy was significantly reduced in AAV-VDAC3 group after MI. Western Blot results also showed that LC3 expression was significantly decreased, while autophagy was significantly increased in sh-VDAC3 group after MI. Consistent with the animal results, LC3 expression in HL-1 cells was significantly decreased in VDAC3 overexpression group after CoCl2 stimulation, while VDAC3 knockdown had opposite results. When HL-1 cells were exposed to CoCl2-induced hypoxia, 3-MA, an autophagy inhibitor, significantly rescued siVDAC3-induced cell death, indicating that VDAC3 reduced cardiomyocyte death by inhibiting autophagic cell death.
CONCLUSIONS VDAC3 can improve cardiac function and reduce mortality in mice with AMI by inhibiting autophagic cell death. VDAC3 has the potential to be a therapeutic target for AMI.
GW34-e0549
Qingbin Hou, Chenhui Yan, Yaling Han
National Key Laboratory of Frigid Zone Cardiovascular Disease, Cardiovascular Research Institute and Department of Cardiology, General Hospital of Northern Theater Command, Shenyang 110016, China
OBJECTIVES White adipose tissue is an important endocrine organ of human body, but how its modulates vascular function and atherosclerosis through endocrine mechanisms remains poorly understood. In previous studies, it was found that CREG1, as a glycoprotein molecule, regulates homeostasis of vascular wall cells and inhibits inflammation of vascular histiocytes and macrophages. Whether it can also play an anti-atherosclerotic role in adipose tissue is unknown.
METHODS We generated APOE−/− CREGfl/fl AdipQ cre and APOE−/− CREGfl/fl, APOE−/− and APOE−/− CREGtg mice, Susceptibility to atherosclerosis was evaluated in whole aortae and cross sections of the aortic sinus in male and female mice fed a high-fat Western diet for 14 weeks, starting at 8 weeks.
RESULTS CREG1 was downregulated in white adipose cell of APOE−/− mice fed a Western diet compared with chow diet. On Western diet, APOE−/− CREGfl/fl AdipQ cre mice exhibited marked increase in atherosclerotic lesion formation, expanded immune and inflammatory cell content in aortas, and increased necrotic core content compared with APOE−/− CREGfl/fl. APOE−/− CREGtg mice compared with APOE−/− mice, Contrary to the above results. Mechanistically, we show that CREG1 loss specifically decreased the expression of AMPKγ2, Its leads to increased expression of the transcription factor SREBP1, which promotes cholesterol metabolism, and triggers an inflammatory response. We further show that ox-LDL (oxidized LDL) treatment decreased adipose cell CREG1, its promoted the degradation of AMPKγ2.
CONCLUSIONS Adipose cell CREG1 delays atherogenesis by inhibiting proinflammatory recruitment and transcriptional activation of SREBP1 in part by upregulating AMPKγ2, implicating that targeting CREG1 in white adipose cell may serve as a novel therapeutic strategy to treat atherosclerosis.
GW34-e0550
Qingbin Hou, Yisi Liu, Chenhui Yan, Yaling Han
National Key Laboratory of Frigid Zone Cardiovascular Disease, Cardiovascular Research Institute and Department of Cardiology, General Hospital of Northern Theater Command, Shenyang 110016, China
OBJECTIVES Abdominal aortic aneurysm (AAA) is a serious and fatal disease, but until now there has not been an effective drug to treat AAA. MicroRNAs are small noncoding RNAs that modulate gene expression by negatively regulating translation of target genes. Although the role of several miRNAs in vascular smooth muscle cells (VSMCs) has been extensively characterized, the function of miRNA-3154 is still unknown.
METHODS Ang II-induced murine AAA model was used to study VSMC phenotypic transition, proliferation and migration in vivo. RNA-seq analysis, cell migration assay, cell invasion assay, collagen gel contraction assay, immunoblotting, protein interactome analysis, immunofluorescence, immunohistochemistry, Flourescence in situ hybridization, luciferase Assay were performed to clarify the phenotype and elucidate the molecular mechanisms.
RESULTS We screened for miRNAs whose expression is modulated in VSMCs by AngII-induced murine AAA model, and among the hits, we selected miR-3154. We found that miR-3154 was expressed in various tissues, primary murine cells, and pathological murine and human vascular specimens. Through gain- and loss-of-function approaches, we determined that miR-3154 affects VSMC proliferation, migration, differentiation, and contractility. The alterations of those properties were dependent upon epigenetic regulation of key VSMC differentiation genes. Notably, Pax7 was found to be a direct target of miR-3154 and able to modulate the Myocd, a potent cardiac and smooth muscle tissue-specific transcriptional coactivator of serum response factor (SRF), plays a vital role in inducing smooth muscle differentiation. Finally, in vivo tail intravenous injection antagomir miR-3154 and AAV-cmv-pax7 prevented the formation of abdominal aortic aneurysms in an AngII-induced model.
CONCLUSIONS miR3154 maintains the contractile phenotype of VSMCs through Pax7-Myocd signaling in vitro and in vivo. Its modulation in the context of disease could be exploited for therapeutic purposes.
GW34-e0552
Qingbin Hou, Jinping Jiang, Kun Na, Xiaolin Zhang, Dan Liu, Quanmin Jing, Chenghui Yan, Yaling Han
National Key Laboratory of Frigid Zone Cardiovascular Disease, Cardiovascular Research Institute and Department of Cardiology, General Hospital of Northern Theater Command, Shenyang 110016, China
OBJECTIVES Both primary Sjogren’s syndrome (pSS) and acute myocardial infection (AMI) are intricately linked to one another. However, their common mechanism is not fully understood. Herein, we examined the underlying network of molecular action associated with the development of this complication.
METHODS pSS (GSE40611) and AMI (GSE66360 discover cohort, DC) datasets were retrieved from the Gene Expression Omnibus (GEO) database, and differentially expressed genes (DEGs) were assessed via the R-based ‘limma’ packages. Subsequently, we explored the physiological roles of DEGs using Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG). Protein–protein interaction (PPI) axis was constructed using the Search Tool for the Retrieval of Interacting Genes (STRING) database, followed by visualization via Cytoscape. Hub gene selection employed the cytoHubba plugin, and validation was performed using GSE7451 for pSS and GSE66360 for AMI. Hub gene set enrichment (GSEA) and immune invasion analyses were carried out, and transcription factor (TF)- and TF-miRNA modulator axes for hub genes were constructed accordingly.
RESULTS Overall, we observed 453 highly- and 45 scarcely-expressed genes between the AMI patients and controls in the GSE66360 DC. In addition, we noted 796 highly- and 876 scarcely-expressed genes between pSS patients and controls in the GSE40611 dataset. The intersection of DEGs from both datasets identified 49 common highly expressed and 2 identical scarcely expressed genes. Moreover, the following pathways were markedly enriched among pSS and AMI patients: inflammatory response axis (P=1.3E-06), direct modulation of interleukin-1 beta synthesis (P=1.9E-06), fungus detection (P=9.8E-06), direct modulation of NF-kappaB transcription factor activity (P=1.2E-05), direct modulation of interleukin-6 synthesis (P=1.9E-05), MyD88-based toll-like receptor axis (P=4.4E-05), direct modulation of T cell proliferation (P=5.3E-05), direct modulation of apoptosis (P=8.1E-05). Following verification, S100A8, IGSF6, MNDA, NCF2, and MMP9 were recognized as hub genes, and the area under the curve (AUCs) of all genes were >0.7 for both pSS and AMI. Based on our GSEA analysis, the adaptive immune response network was strongly associated with enhanced IGSF6, MMP9, S100A8, MNDA, and NCF2 contents in the GSE66360 DC and GSE40611 datasets. Functional enrichment of these five hub genes is adaptive immune response. The Type 1 T helper cell showed the most association among all cell types related to AMI and pSS. Lastly, using TF-gene interaction and TF-miRNA co-modulatory network analyses, we identified 27 common TFs and 20 identical TF-miRNAs.
CONCLUSIONS Herein, we revealed common networks involving pSS and AMI etiologies. Knowledge of these networks and hub genes can enhance research into their associated mechanism and development of future robust therapy.
GW34-e0553
Zheming Yang1,2, Chenghui Yan1, Yaling Han1,2
1College of Medicine and Biological Information Engineering, Northeastern University, Shenyang, Liaoning 110167, China
2National Key Laboratory of Frigid Zone Cardiovascular Disease, Cardiovascular Research Institute and Department of Cardiology, General Hospital of Northern Theater Command, Shenyang 110016, China
OBJECTIVES Sorafenib (sor) is a widely used chemotherapeutic drug to treat hepatocellular carcinoma. However, it often results in significant cardiac dysfunction, which limits its use. Recent research has suggested that this side effect may be a result of necroptosis, a unique type of programmed cell death. Previous studies have shown that the cardiac homeostasis regulatory protein cellular repressor of E1A stimulated genes (CREG) is crucial in maintaining the heart’s physiological and pathological processes, this study explores CREG’s potential use in reducing sor-induced cardiac dysfunction by investigating its effect on necroptosis.
METHODS Mice with CREG knockout, CREG overexpression and mice pretreated with infusion of recombinant 0.3 mg/kg·d CREG protein were treated with sor, cardiac function was detected by animal ultrasound technology. Sor-induced myocardial injury were investigated by histological analyses, quantitative RT-PCR, western blotting. HepG2 cell lines and prepared primary mouse cardiomyocytes were treated with sor and various other treatments. Cardiomyocyte necroptosis was detected by flow cytometry, western blotting, and transmission electron microscopy (TEM). Immunofluorescence staining, JC-1 staining and western blotting were used to detect mitophagy. Mitochondrial Ca2+ uptake was detected by Rhod-2 probes using confocal imaging. Morphological changes in lysosome, mitochondria and mitochondria-associated endoplasmic reticulum membranes were imaged using TEM and confocal microscopy.
RESULTS The study found that the expression of CREG in the heart of mice treated with sor was reduced, further in vitro studies indicated that sor caused the degradation of CREG via the lysosomal pathway. Loss of function research showed that CREG deficiency aggravated the sor-induced cardiac dysfuntion, while function acquisition research suggested that CREG overexpression had the opposite effect. In vitro experiments have also found that overexpression of CREG reduced sor-induced necroptosis, while silencing CREG aggravated sor-induced necroptosis. More importantly, our results showed that treatment of mice with CREG recombinant protein effectively reduced the sor-induced cardiac dysfuntion. Mechanically, CREG increased the expression of mitofusin-2 (MFN2) by reduced excessive mitophagy induced by sor, which in turn inhibits sor-induced myocardial necroptosis. Moreover, we found that CREG did not impact sor-induced death of liver cancer cells.
CONCLUSIONS The overexpression of CREG could rescue sor-induced cardiomyocyte necroptosis without disturbing the anti-tumor effects. CREG protein may have clinical applications in the prevention of sor-induced cardiac dysfuntion.
GW34-e0559
Jinping Jiang, Xiaolin Zhang, Chenhui Yan, Yaling Han
National Key Laboratory of Frigid Zone Cardiovascular Disease, Cardiovascular Research Institute and Department of Cardiology, General Hospital of Northern Theater Command, Shenyang 110016, China
OBJECTIVES Abdominal aortic aneurysm (AAA) is a chronic vascular inflammatory disorder for which there is currently no effective drug therapy. S100A12 plays a significant role in regulating inflammatory processes and immune responses. However, the role of S100A12 in the pathogenesis of AAA remains unclear. In this study, we identified how S100A12 affects AAA and the underlying mechanisms.
METHODS For in vivo experiments, lysozyme M promoter S100A12 transgenic mice (S100A12Tg) were generated and infused with angiotensin II (Ang II) for four weeks. Reverse Transcription-Polymerase Chain Reaction (RT-PCR), western blotting, immunohistochemistry, and gelatin zymography were performed to examine the aortas of each group. RAW264.7 cells were stimulated in vitro by human recombinant S100A12 protein, plasmid, and relative RNA interference. STAT3 inhibitor S31-201 was intraperitoneally injected in S100A12Tg mice and infused with angiotensin II for four weeks to induce AAA formation. Finally, an enzyme-linked immunosorbent assay was used to assess S100A12 levels in patients with AAA and normal controls.
RESULTS S100A12 greatly increased the incidence of Ang II-induced AAA in vivo, leading to AAA characterized by inflammation, oxidative stress, elastin fragmentation, degradation of the extracellular matrix, and increased expression of matrix metalloproteinase 9 (MMP9). Expression of the inflammatory factor MCP-1, oxidative stress marker COX2, and MMP9 in RAW264.7 cells increased with S100A12 stimulation. We demonstrated that S100A12 stimulated macrophages to promote inflammation, oxidative stress, and extracellular matrix degradation by activating the TLR4/JAK2/STAT3 signaling pathway. The incidence of Ang II-induced AAA was significantly lower in the S3I-201 group than that in the control group. Plasma S100A12 levels were significantly higher in patients with AAA than those in healthy subjects.
CONCLUSIONS S100A12 aggravated AAA by promoting inflammation, oxidative stress, and extracellular matrix degradation in macrophages via activation of the TLR4/JAK2/STAT3 signaling pathway.
GW34-e0560
Hanlin Wu, Jing Wang, Yuxin Bu, Jia Li, Yiming Li, Quanmin Jing, Chenghui Yan, Dan Liu, Yaling Han
National Key Laboratory of Frigid Zone Cardiovascular Diseases, Department of Cardiology and Cardiovascular Research Institute of PLA, General Hospital of Northern Theater Command, Shenyang, Liaoning Province 110016, China
OBJECTIVES Pentamethylquercetin (PMQ) is a natural polymethyl flavonoid that possesses anti-apoptotic and other biological properties. Abdominal aortic aneurysm (AAA), a fatal vascular disease with a high risk of rupture, is associated with phenotypic switching and apoptosis of medial vascular smooth muscle cells (VSMCs). This study aimed to investigate the protective effects of PMQ on the development of AAA and the underlying mechanism.
METHODS ApoE−/− mice were continuously infused with angiotensin II (Ang II) for 4 weeks to develop the AAA model. Intragastric administration of PMQ was initiated 5 days before Ang II infusion and continued for 4 weeks. VSMCs of mice from each group were analyzed to understand the effects of PMQ on apoptosis.
RESULTS PMQ dose-dependently reduced the incidence of Ang II-induced AAA, aneurysm diameter enlargement, elastin degradation, VSMC phenotypic switching and apoptosis. Furthermore, PMQ also inhibited phenotypic switching and apoptosis in Ang II-stimulated VSMCs. PMQ exerted protective effects by regulating GSK-3β phosphorylation via the C/EBPβ/PTEN/AKT axis. AAV-mediated overexpression of PTEN reduced the therapeutic effects of PMQ in the AAA model mice, suggesting that the effects of PMQ on Ang II-mediated AAA formation are related to the C/EBPβ/PTEN/AKT/GSK-3β axis.
CONCLUSIONS PMQ inhibits VSMCs phenotypic switching and apoptosis by bounding to C/EBPβ at Lys253 with hydrogen bond to regulate C/EBPβ nuclear translocation and PTEN/AKT/GSK-3β axis, thereby inhibiting Ang II-induced AAA formation. Thus, PMQ might be a potential therapeutic agent for the treatment of AAA.
GW34-e0561
Hanlin Wu, Ting Zhou, Jing Wang, Yuxin Bu, Zheming Yang, Chenghui Yan, Dan Liu, Yaling Han
National Key Laboratory of Frigid Zone Cardiovascular Diseases, Department of Cardiology and Cardiovascular Research Institute of PLA, General Hospital of Northern Theater Command, Shenyang, Liaoning Province 110016, China
OBJECTIVES Type 2 diabetes is prevalent worldwide. Cardiovascular complications in patients with diabetes include diabetic cardiomyopathy (DCM). Ubiquitin-conjugating enzyme E2 I (UBC9) is the only SUMO-E2 enzyme that plays a key role in cardiomyocytes. The present study aimed to elucidate the role and mechanism of action of UBC9 in the development of DCM.
METHODS We established myocardial-specific UBC9 knockout (UBC9-CKO) and adeno-associated virus-overexpressing mouse models. Additionally, a mouse model of DCM was established using high-fat diet feeding and low-dose streptozotocin injection. This mechanism was explained by the stimulation of cultured cardiomyocytes with palmitic acid.
RESULTS The transcript and protein levels of UBC9 were significantly decreased in the myocardium of patients with DCM. Myocardial-specific UBC9 knockout aggravated cardiac dysfunction, alleviated myocardial fibrosis and hypertrophy, and promoted mitophagy and mitochondrial damage. Meanwhile, UBC9 overexpression exhibited the opposite effects. UBC9 exerted mitochondrial protective effects independent of SUMOylation. UBC9 regulated RUNX2 degradation by directly binding to NEDD4, thus regulating PSEN2 to exert protective effects against mitophagy. Moreover, the effect of UBC9 on mitochondrial function was reversed upon PSEN2 knockdown.
CONCLUSIONS UBC9 regulates mitophagy through the NEDD4/RUNX2/PSEN2 pathway and alleviates DCM development by improving myocardial mitochondrial dysfunction. These findings offer novel perspectives on the application of UBC9 as a therapeutic target for DCM
GW34-e0562
Yuxin Bu, Chenhui Yan, Dan Liu, Yaling Han
National Key Laboratory of Frigid Zone Cardiovascular Disease, Cardiovascular Research Institute and Department of Cardiology, General Hospital of Northern Theater Command
OBJECTIVES Trim55 (Tripartitemotif-containing55), is a member of the Trims family, is mainly expressed in myocardium and skeletal muscle. Trim55 plays an important role in promoting the development of mouse heart embryo. However, the role of Trim55 in myocardial infarction (MI) and its related molecular mechanism has not been reported.
METHODS The MI model was established in C57BL/6J mice, Trim55 gene knockout and Trim55 gene overexpression mice using by permanently ligating the left anterior descending coronary artery. At 3 and 28 days after MI, cardiac function, fibrosis and cardiomyocyte apoptosis were evaluated through echocardiography, HE staining, Masson staining, TUNEL staining and Western blot. In vitro, primary rat cardiomyocytes (NRCMs) were isolated and cultured. NRCMs were treated with Trim55 overexpressed or knockdown adenovirus, and then cells were stimulated with hypoxia. The effects of Trim55 on cardiomyocyte apoptosis were determined by quantitative PCR, Western blot, flow cytometry and TUNEL staining.
RESULTS The transcription and protein levels of Trim55 in the border zone of myocardium post-MI were significantly increased. Trim55 gene knockout could significantly improve the cardiac systolic function, decrease cardiomyocytes apoptosis and infarct size after MI. However, Trim55 overexpression could led to the opposite effect. In vitro, overexpression of Trim55 promoted cardiomyocytes apoptosis by inhibiting HO-1 protein expression. On the contrary, Trim55 knockdown caused the opposite effect. In addition, Trim55 was proved to be the direct target of forkhead box transcription factor 3 (Foxo3) in cardiomyocytes.
CONCLUSIONS Trim55 promoted cardiomyocytes apoptosis by regulating the expression of HO-1, thereby aggravating myocardial injury after MI.
GW34-e0563
Jing Wang, Dan Liu, Xiaoxiang Tian, Chenhui Yan, Yaling Han
National Key Laboratory of Frigid Zone Cardiovascular Disease, Cardiovascular Research Institute and Department of Cardiology, General Hospital of Northern Theater Command, Shenyang 110016, China
OBJECTIVES Vascular calcification (VC) is a common pathological feature of atherosclerosis, chronic kidney disease, vascular injury and aging. VC is characterized by different mineral deposits accumulating in blood vessels and valves. Cellular repressor of E1A-stimulated genes (CREG) is a small molecule secreted glycoprotein and is consisted by 220 amino acids. Studies have shown that CREG is involved in regulating cell growth and differentiation. The aim of this study is to clarify the role of CREG participates in the development of VC.
METHODS Male C57BL/6J, CREG transgenic (CREGTG), and CREG smooth muscle cell-specific knockout (CREGSMKO) mice were used to establish a VC model induced by VitD3 injection. Alizarin red staining, Von Kossa staining, and western blotting were used to assess the severity of VC. In vitro, mouse primary vascular smooth muscle cells (VSMCs) were isolated and cultured, VSMCs were infected with CREG overexpressing adenovirus or CREG small interfering RNA, followed by phosphate (Pi) stimulation, real-time PCR, western blotting, Alizarin red staining were used to assess the severity of VSMCs calcification.
RESULTS In vivo, the mRNA and protein levels of CREG were significantly decreased in VitD3-induced VC. CREGSMKO mice aggravated VC by increasing the expressions of the osteogenic markers runt-related transcription factor 2 (Runx2) and osteopontin (OPN), and inhibiting SMA expression. In contrast, CREGTG mice inhibit VC by inhibiting the expressions of Runx2 and OPN, and increasing SMA expression. In vitro, CREG expression was significantly decreased in Pi-induced VSMCs. CREG knockdown exacerbated Pi-induced VSMCs differentiation and calcification, CREG overexpression inhibited VSMCs differentiation and calcification induced by Pi stimulation. Mass spectrum and western blotting revealed that FHL2 was a downstream molecular of CREG, and was involved in the regulation of CREG in VC. Moreover, CREG could regulate FHL2 protein expression in lysosome-dependent pathway.
CONCLUSIONS Our study identified that CREG was a novel endogenous inhibitor of VC by regulating FHL2 protein expression. This study could provide a promising target for VC therapy.
GW34-e0564
Ting Zhou1,2, Hanlin Wu2, Chunying Liu2, Xiaodong Jia2, Dan Liu2, Chenghui Yan2, Haixu Song2, Chenhui Yan1, Yaling Han2
1National Key Laboratory of Frigid Zone Cardiovascular Disease, Cardiovascular Research Institute and Department of Cardiology, General Hospital of Northern Theater Command, Shenyang, Liaoning Province 110016, China
2Department of Cardiology, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei Province 430022, China
OBJECTIVES Pathological cardiac hypertrophy is a major risk factor for heart failure (HF) and a leading cause of morbidity and mortality worldwide. However, the molecular mechanisms underlying pathological cardiac hypertrophy remain largely unclear. In the present study, we investigated the possible underlying effects of UBC9 on cardiac hypertrophy and heart failure, and whether the effects depend on SUMOylation.
METHODS UBC9 expression was analyzed in samples of 30 human hypertrophic hearts compared to 166 normal controls using RNA sequencing analysis of GSE141910 from the Gene Expression Omnibus (GEO, https://www.ncbi.nlm.nih.gov/geo). Real-time polymerase chain reaction (RT-PCR), western blotting, and immunohistochemistry (IHC) were utilized to analyze the expression of UBC9 in samples of 4 recruited human hypertrophic hearts and 4 normal controls. Mice with cardiac-specific over-expression and low-expression of UBC9 by intravenous injection of adeno-associated virus 9 (AAV9)–encoding UBC9 or shUBC9 under the control of cardiac troponin T (cTnT) promoter were subjected to sham or transverse aortic construction (TAC) surgery for 8 weeks to induce heart failure in vivo. Isolated Neonatal Mouse Cardiomyocytes (NMCMs), rat cardiomyocyte line (H9C2) and human cardiomyocyte line (AC16), transfected with UBC9 adenovirus or UBC9 siRNA followed by phenylephrine (PE 500 μM) treatment for 48 h, were utilized to identify the effects of UBC9 on cardiac hypertrophy in vitro. Unbiased screening was performed on myocardial samples from UBC9flox/floxαMyHCcre and UBC9flox/flox embryonic mice, or Myh7-cre UBC9flox/flox and UBC9flox/flox adult mice, using protein mass spectrometry (MS) analysis and further analyzed by Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis to identify the potential downstream targets of UBC9. A dual-luciferase reporter gene experiment was conducted to confirm the transcription factor of LIM and cysteine-rich domain 1 (LMCD1).
RESULTS UBC9 mRNA and protein expression increased in the hearts of patients with hypertrophic cardiomyopathy (HCM) and pressure overload-induced mice, as well as in the NMCMs, H9C2, and AC16 treated with PE. Besides, UBC9 translocated from cytoplasm to nuclear in the cardiomyocytes treated with PE. Cardiac cell-specific UBC9 over-expression alleviates cardiac hypertrophy, cardiac fibrosis, and dysfunction in vivo and in vitro. Cardiac cell-specific UBC9 knockout aggravates cardiac hypertrophy and heart failure in vivo and in vitro. Mechanistically, UBC9 inhibits the expression of downstream target-LMCD1, and the activation of calcineurin-Nuclear Factor of the Activated T Cell (NFAT)-Myocyte-enriched calcineurin interacting protein 1.4 (MCIP1.4) signaling cascade by directly binding to forkhead box O3 (FOXO3), a transcription factor of LMCD1, and promoting FOXO3 dephosphorylation and nuclear translocation, independent of SUMOylation.
CONCLUSIONS This study provides a new insight that UBC9, as a novel negative regulator of pathological cardiac hypertrophy and a potential intervention target in cardiac hypertrophy and HF, may contribute to exploring effective therapeutic strategies for pathological cardiac hypertrophy treatment.
GW34-e0568
Zhu Mei1,2, Ziping Song1, Chenghui Yan1, Yaling Han1,2
1National Key Laboratory of Frigid Zone Cardiovascular Disease, Cardiovascular Research Institute and Department of Cardiology, General Hospital of Northern Theater Command, Shenyang, Liaoning 110016, China
2College of Medicine and Biological Information Engineering, Northeastern University, Shenyang, Liaoning 110167, China
OBJECTIVES The cardioprotective effect of AMPK in the progression of MI are attributed to phosphorylation of the α subunit at Thr172. However, whether the AMPKγ2 subunit plays an important role in MI remains unclear.
METHODS AMPKγ2 plasma levels in patients with MI were tested using ELISA. LAD surgery was performed to establish a mouse MI model. Transwell and scratch assays were performed to evaluate the macrophage migration ability. Lyz2-cre AMPKγ2 and adeno-associated virus carrying the F4/80 promoter for macrophage-specific overexpression AMPKγ2 mice were employed to investigate the role of AMPKγ2 in macrophages derived from MI in vivo.
RESULTS AMPKγ2 is highly expressed in macrophages. AMPKγ2 levels were decreased in monocytes of myocardial infarction patients and downregulated by inflammatory stimuli of IFN-γ in RAW264.7 cells and in bone marrow-derived macrophages (BMDM) extracted from mice subjected to MI. Lyz2-cre AMPKγ2 mice developed worse cardiac dysfunction post MI by boosting macrophage infiltration. Overexpression of AMPKγ2 repressed macrophage migration and inflammation in vivo and in vitro. The CXCL16 was a key contributor to macrophage migration and inflammation, which was alleviated by AMPKγ2 administration via transcriptional regulation. Moreover, the CXCL16/CXCR6 and NF-κB-MCP1/IL-6 pathways jointly formed a positive feedback loop. AMPKγ2 repressed the CXCL16/CXCR6 axis by restraining YY1 expression to rescue cardiac dysfunction after MI and restrain macrophage infiltration dependent on AMPKα1 subunit-mediated AMPK activity. A769662, but not the classic AMPK agonist metformin can rescue deficiency of AMPKγ2 induced-the myocardial dysfunction. AMPKγ2 phosphorylated smurf2S384 to promote YY1 degradation via the ubiquitin proteasome pathway. Finally, IFN-γ downregulated HOXA5 to repress AMPKγ2 transcriptional activity and protein expression in vitro and in vivo.
CONCLUSIONS AMPKγ2 inhibits the YY1-CXCL16-CXCR6 pathway, which plays an important role in cardiac dysfunction following MI, and may be a potential therapeutic target for myocardial remodeling after MI.
GW34-e0571
Zhen Zhang1,2, Yanqiu Xing1,2
1Department of Geriatric Medicine, Qilu Hospital of Shandong University
2Key Laboratory of Cardiovascular Proteomics of Shandong Province
OBJECTIVES Aging is widely accepted as an independent risk factor for cardiovascular disease, which contribute to the increasing morbidity and mortality in the elderly population. Lysine β-hydroxybutyrylation (Kbhb) was discovered as a novel post-translational protein modifications (PTM) in which β-hydroxybutyrate (BHB) is covalently attached to lysine ε-amino groups. Recent studies have revealed that histone Kbhb may contribute to tumor progression, diabetic cardiomyopathy progression, and postnatal heart development. However, to date, there have been no studies reporting a global analysis of Kbhb proteins in aging hearts or elucidating the underlying mechanisms of this modification in the aging process. Herein, we conducted quantitative proteomics and Kbhb PTM omics to comprehensively elucidate the alterations of global proteome and Kbhb modification in the hearts of aged mice.
METHODS In this study, twenty C57BL/6J mice at 3- and 22-months of age were adopted as young controls (WTY, n=10) and aged models (WT-A, n=10), respectively. Various measurements and analyses were conducted during the experiment, including body weight, heart weight, grip strength, echocardiography, serum parameters, and myocardial morphology. In the end, three hearts from each group were used as biological replicates for high-throughput and quantitative data-dependent acquisition (DDA) proteomics analysis. Kbhb modification omics analyse was conducted using affinity enrichment followed by high-resolution high-throughput mass spectrometry (LC-MS/MS) analysis.
RESULTS Our results revealed a decline in grip strength and cardiac diastolic function in 24-month-old aged mice compared to 3-month-old young mice. Through LC-MS/MS we identified a total of 1710 β-hydroxybutyrylated lysine sites in 641 proteins in the cardiac tissue of young and aged mice. Additionally, we observed 183 Kbhb sites were identified in 134 proteins, which exhibited significant differential modification in aged hearts (fold change >1.5 or <1/1.5, P<0.05). Notably, the Kbhb-modified proteins were primarily found in energy metabolism pathways, such as fatty acid elongation, glyoxylate and dicarboxylate metabolism, citrate cycle (TCA cycle), and oxidative phosphorylation. Furthermore, these Kbhb-modified proteins were predominantly localized in mitochondria.
CONCLUSIONS For the first time, our study provides a global proteomic profile and Kbhb modification landscape of cardiomyocytes in aged hearts. These findings open up new possibilities for treating cardiac aging and aging-related cardiovascular diseases by reversing abnormal Kbhb modifications.
GW34-e0589
Jiaxin Xu, Haixu Song, Chenhui Yan, Yaling Ha
National Key Laboratory of Frigid Zone Cardiovascular Disease, Cardiovascular Research Institute and Department of Cardiology, General Hospital of Northern Theater Command, Shenyang, China
OBJECTIVES S100A12 is a calcium-binding protein belonging to the S100 subfamily of myeloid-related proteins that acts as an alarmin to induce a pro-inflammatory innate immune response. Cardiac hypertrophy is a key biological response to injurious stresses such as pressure overload and when excessive can lead to heart failure. Previous studies revealed that S100A12 markedly expressed in the serum from HF patients, which suggested that S100A12 is a prognostic marker for several adverse events in HF patients. Although it has been linked to several chronic inflammatory diseases, however, its role in myocardial interstitial fibrosis remains unknown. In order to study this issue, we generated the transgenic (TG) mice which specific expressed the human S100A12 in myeloid cells and further explored that the level of S100A12 was markedly upregulated in the blood and heart from HF patients. Furthermore, we explored that S100A12 highly expression in macrophages is involved in Ang II-induced adverse cardiac fibrosis and dysfunction in vivo and in vitro.
METHODS Angiotensin II (1000 ng kg−1 min−1, Ang II) was administered to wild-type (WT) and transgenic (Tg) mice expressing human S100A12 in myeloid cells for 28 d. Humanized S100A12 markedly aggravated Ang II-induced myocardial fibrosis compared with that in WT mice. Furthermore, thereceptor for advanced glycation end products (RAGE) blocker (Tranilast, 5 mg kg−1 d−1) administration significantly attenuated Ang II-induced myocardial fibrosis and dysfunction in the Tg mice. Previous studies indicated that S100A12 is associated with cystic fibrosis, idiopathic pulmonary fibrosis, and renal fibrosis. In addition, the findings in our study showed that S100A12 also promoted cardiac fibrosis and dysfunction. In vivo and in vitro study, S100A12 increased expression of GSK-3β and Smad3 phosphorylation which associated with cardiac fibrosis. Using immunoprecipitation experiments, we show that GSK-3β is able to bind to RAGE. Primary Mouse Embryonic Fibroblasts (MEFs) isolation and culture. Mechanistically, S100A12 (200 pg/mL) significantly promoted proliferation, migration, collagen production, and phenotypic transdifferentiation of primary mouse embryonic fibroblasts (MEFs).
RESULTS S100A12 expressed in macrophages exaggerated Ang II-induced cardiac fibrosis in mice. Silencing RAGE (RAGE-siRNA, 20 μmol/L) ameliorated the potent fibrotic role of S100A12 by phosphorylation of glycogen synthase kinase-3β (GSK-3β), indicating the involvement of the RAGE/GSK-3β/Smad3 cascade in S100A12-mediated cardiac fibrosis. Notably, circulating and cardiac S100A12 levels were higher in patients with heart failure than in normotensive individuals.
CONCLUSIONS S100A12, in the presence of Ang II, promotes cardiac fibrosis and dysfunction; Using RAGE blocker develop less cardiac fibrosis and dysfunction; In CHF patients, high levels of S100A12 were detected in serum and heart. These findings indicate that S100A12 could be a key regulator of cardiac fibrosis and dysfunction in CHF patients. Ang II-induced myocardial fibrosis and dysfunction are largely mediated by the S100A12-RAGE-GSK-3β axis. The inhibition of S100A12 may represent a new therapeutic target for the treatment of myocardial fibrosis.
GW34-e0591
Yiming Li, Yuying Li, Chenhui Yan, Yaling Han
National Key Laboratory of Frigid Zone Cardiovascular Disease, Cardiovascular Research Institute and Department of Cardiology, General Hospital of Northern Theater Command
OBJECTIVES The inflammatory response and repair of myocardial tissue after myocardial infarction (MI) play an important role in the prognosis of MI; in which macrophages play an important role. In this study, we have attempted to elucidate whether CREG1, a widely expressed differentiation homeostasis regulatory protein, is involved in the repair of myocardial injury after MI by regulating macrophage function.
METHODS Western blotting, immunofluorescence, flow cytometry, and qPCR were used to study the expression profile of CREG1. The miRNA chip and transcription factor chip were used to screen CREG1 target genes. Myeloid-specific knockout mice and transgenic mice were used to verify the role of CREG1 in MI in mice.
RESULTS Deletion of CREG1 in macrophages led to further reduction of cardiac function after MI and increased inflammatory cell infiltration in the cardiac tissue in the early stage of MI. The cardiac function of Creg1 tg mice was improved compared to that of Creg1 wt/wt mice after MI. Reduced CREG1 expression led to increased expression of pro-inflammation-related genes in macrophages; conversely, increased CREG1 expression led to inactivation of macrophage-related genes. CREG1 can regulate the inflammation pathway via the miR-125a/99b/let-7e-5p cluster. Overexpression of miR-125a-5p in CREG1-deficient bone marrow derived macrophages (BMDMs) alleviated the excessive activation of macrophage inflammation. Conversely, inhibition of miR-125a/99b/let7e-5p activated the inflammatory pathway and increased macrophage migration in BMDMs overexpressing CREG1. We found that HOXA5 might bind to the promoter region of miR-125a/99b/let-7e-5p after CREG1 expression was increased. ETS1, as a transcription repressor, inhibits the expression of CREG1 in macrophages after MI. Meanwhile, serological studies on patients with MI suggest that the expression of miR125a-5p, HOXA5, and CREG1 are significantly correlated.
CONCLUSIONS The loss of CREG1 in macrophages causes an increase in cardiac inflammation in the acute phase of MI through downregulation of HOXA5-miR-125a-5p/99b/let7e-5p, which subsequently aggravates cardiac tissue repair and pathological ventricular remodeling.
GW34-e0592
Yiming Li, Chenghui Yan, Yaling Han
National Key Laboratory of Frigid Zone Cardiovascular Disease, Cardiovascular Research Institute and Department of Cardiology, General Hospital of Northern Theater Command
OBJECTIVES Aortic dissection (AD) is a serious and life-threatening condition which allows blood to enter the vessel wall and causes aorta rapture. NETosis is a type of programmed cell death induced by neutrophils, which is characterized by the release of neutrophil extracellular traps (NETs). In this study, we attempted to elucidate whether NETs components are involved in the occurrence and development of AD and found out treated target.
METHODS The protein, RNA and transcription factor chip were used between humanic dissected and normal aorta to screen target differentially expressed genes. Pathological and serological verification of the local and systemic expression of target genes in AD patients. Construction of S100A12tg/tg AD mouse model using BAPN drinking+AngII micropump method. Ultrasound, micro-CT, and PET-CT were used to evaluate aortic remodeling status in vivo. Western blotting, immunofluorescence, flow cytometry, and qPCR were used to study the expression level of neutrophil and vascular smooth muscle cell (VSMC) in vitro.
RESULTS Neutrophil Elastase (NE), one of the key components of NETs, was significant overexpressed in AD than that in non-AD human aorta. Human AD aorta underwent structural changes and a large amount of collagen deposition, indicating VSMC phenotypic modulation happened. There was a significant overexpression of S100A12 and NE in the mass spectrometry detection of human AD and adjacent aortic proteins. Serological studies had found that the expression level of S100A12 in the serum of AD patients reaches its peak around 24 hours after onset, and the content was about three times that of the normal population. Using BAPN+AngII to establish AD mice model, myeloid overexpression S100A12 mice showed higher negative vascular remodeling rate and rupture rate compared to C57 mice. WB and PCR showed high expression of TLR4 in the aorta of S100A12 transgenic AD mice, which is consistent with the results of human AD aorta. S100A12 can promote the expression and degranulation of NE through TLR4-PAD4 axis, while siTLR4 and siPAD4 can partially reduce the expression level of NE. However, when co-culturing neutrophils and VSMCs, only specific inhibition of NE activity can better save the migration, apoptosis and Phenotypic Modulation of VSMCs induced by NETosis, rather than other links of NETosis. After intraperitoneal injection of Sivelestat, a specific inhibitor of NE activity, at different stages of the AD mice model, PET-CT showed that Sivelestat could reduce the level of local inflammation in the aorta of mice, reduce the incidence of AD, improve vascular remodeling and prolong the survival time of mice.
CONCLUSIONS S100A12 overexpression in neutrophils promotes NE expression and degranulation through TLR4-PAD4 axis. Released activated NE can promote VSMC migration, apoptosis and phenotype transformation, while specific inhibition of NE activity can save this effect.
GW34-e0593
HaiXu Song, Jiayin Li, Zhu Mei, Xiaoxiang Tian, Dan Liu, Chenhui Yan, Yaling Han
National Key Laboratory of Frigid Zone Cardiovascular Disease, Cardiovascular Research Institute and Department of Cardiology, General Hospital of Northern Theater Command, Shenyang, China
OBJECTIVES Megakaryocytes are the precursors of platelets, abnormal megakaryocyte maturation and platelet production will lead to platelet-related diseases, and destroy the dynamic balance of hemostasis and bleeding. Cellular repressor of E1A-stimulated genes 1 (CREG1), is a small molecular weight secreted glycoprotein widely expressed in mature tissue cells, and has ability to maintain tissue and cell maturity, and promotes tissue differentiation, however, the role of CREG1 in megakaryocytes differentiation and platelet production is not clear.
METHODS In this study, we generated a megakaryocyte/platelet-specific creg1 knockout mouse (Creg1pf4-cre ). Creg1-floxed mice (Creg1fl/fl ) were crossed with a transgenic line expressing Cre recombinase under the megakaryocyte/platelet-specific pf4 promoter. To investigated the role of CREG1 in the regulation of megakaryocyte maturation and platelet production by using the megakaryocyte/platelet-specific knockout mice. The results showed that Creg1fl/fl mice had normal platelet lifespans by detecting the proportions of biotin-labeled platelets at different time points. We also found that CREG1 deficiency significantly impaired thrombopoiesis by used anti-CD42b monoclonal antibodies to deplete platelets.
RESULTS The platelets from peripheral blood were detected by annexin V staining and western blot, the results indicated that there was no difference in apoptosis between Creg1fl/fl mice and Creg1pf4-cre mice. Fetal livers were isolated from Creg1fl/fl and Creg1pf4-cre embryos and incubated with IL-3 and TPO for 5 days. The results showed that Creg1pf4-cre megakaryocytes produced fewer proplatelets and morphologically less differentiated than Creg1fl/fl megakaryocytes by using immunofluorescence confocal microscopy. Mechanistic studies showed that the MEK-ERK1/2 pathway plays a critical role in the megakaryocyte differentiation and proplatelets production, Creg1-deficient megakaryocytes resulted in inactivation of MEK-ERK1/2 pathways. We further demonstrated that CREG1 interacted with MEK-ERK1/2 pathway directly. Immunoprecipitation analysis and immunofluorescent staining revealed that CREG1 directly interacted with MEK1. PAK/LIMK1/Cofilin pathway is required for the regulation of megakaryocyte cytoskeleton protein recombination, polyploidy, and DMS polarization. Western blot analysis revealed that phosphorylation of PAK1 and LIMK1 was impaired in Creg1pf4-cre megakaryocytes cultured from bone marrow, further leading to a significantly decreased in Cofilin phosphorylated protein. Further, we determined the core promoter of CREG1 gene in 293T cells, and used the dual-luciferase report assay to discover that C/EBPβ (CCAAT enhancer binding protein beta) combined with the core promoter of CREG1. C/EBPβ is an important transcriptional regulator involved in the proliferation, differentiation and maturation of blood cells, adipocytes and mammary cells, especially in hematopoietic differentiation. Luciferase assay demonstrated that C/EBPβ significantly up-regulated the activity of the core promoter of CREG1, and promoted the differentiation of megakaryocyte.
CONCLUSIONS This study uncovers the role of CREG1 in regulation of megakaryocyte mature and thrombopoiesis, and provides a possible theoretical basis for the prevention and treatment of thrombocytopenia.
GW34-e0594
Dan Liu, Xiaoli Cheng, Haixu Song, Chenhui Yan, Yaling Han
National Key Laboratory of Frigid Zone Cardiovascular Disease, Cardiovascular Research Institute and Department of Cardiology, General Hospital of Northern Theater Command, Shenyang 110016, China
OBJECTIVES Cardiovascular diseases and cancers are two important causes of human death. Doxorubicin (DOX) is currently one of the most used broad-spectrum anthracycline anticancer drugs. However, some cancer patients may experience significant myocardial damage after the use of doxorubicin, which limits its application in cancer patients. At present, there is no effective prevention and treatment for DOX-induced cardiotoxicity. Therefore, clarifying the mechanism of DOX-induced cardiotoxicity and finding intervention targets will provide the basis for clinical prevention and treatment of DOX-induced cardiotoxicity. Cellular repressor of E1A-stimulated genes (CREG1) is an important cardioprotective factor, which plays an important role in maintaining cardiomyocyte differentiation and homeostasis regulation. However, the roles and mechanisms of CREG1 in DOX-induced cardiotoxicity have not been reported.
METHODS In vivo, the intraperitoneal injection of DOX was used to establish a mouse DOX-induced cardiotoxicity model, the mRNA and protein expression of CREG1 in the myocardium was examined using real-time PCR and western blot. To clarify the role of CREG1 in the development of DOX-induced cardiotoxicity, CREG1 transgenic mice, cardiac-specific CREG1 knockout mice and its littermate controls were used to establish DOX-induced cardiotoxicity model. HE staining, Masson staining, WGA staining and western blot were applied to examine fibrosis, myocardial hypertrophy and ferroptosis of myocardium. In vitro, neonatal mouse cardiomyocytes (NMCMs) were cultured and stimulated with DOX, CREG1 overexpression adenovirus and small interfering RNA was used to examine the role of CREG1 on the ferroptosis of NMCMs.
RESULTS In vivo, the mRNA and protein expression of CREG1 were significantly reduced in DOX-treated myocardium. CREG1 transgenic mice significantly alleviated the myocardial damage induced by DOX, and CREG1 deficiency in heart aggravated the DOX-induced cardiotoxicity. The abnormal increase in ferroptosis and impaired mitochondrial function were shown in the DOX-treated heart tissues. CREG1 transgenic mice inhibited the ferroptosis and improved mitochondrial function, and CREG1 deficiency aggravated the ferroptosis and mitochondrial dysfunction induced by DOX. In vitro, the mRNA and protein of CREG1 was reduced in DOX-treated NMCMs. CREG1 overexpression reduced the ferroptosis of cardiomyocytes induced by DOX, CREG1 knockdown aggravated the ferroptosis of cardiomyocytes induced by DOX. Mechanically, CREG1 inhibited the mRNA and protein expression of pyruvate dehydrogenase kinase 4 (PDK4). The effect of CREG1 overexpression on ferroptosis of cardiomyocytes was reversed by PDK4 overexpression.
CONCLUSIONS CREG1 alleviated DOX-induced cardiotoxicity by inhibiting ferroptosis of cardiomyocytes. Our findings might help clarify new roles of CREG1 in the development of DOX-induced cardiotoxicity.
GW34-e0595
Xiaolin Zhang, Minghui Cheng, Xiaoxiang Tian, Dan Liu, Chenhui Yan, Yaling Han
National Key Laboratory of Frigid Zone Cardiovascular Disease, Cardiovascular Research Institute and Department of Cardiology, General Hospital of Northern Theater Command, Shenyang, China
OBJECTIVES Atherosclerosis is one of the most common forms of cardiovascular disease. Recent studies have shown that the inflammatory process in the vessel wall accounts for a key role in the development of atherosclerotic plaques. S100A12 has been shown to activate inflammation in different experimental autoimmune disease models. Due partly to its cellular origins, the exact role of S100A12 in the atherosclerotic plaque remains elusive. The aim of the present study was to evaluate the impact of S100A12 on atherosclerosis and to elucidate the underlying molecular mechanisms.
METHODS S100A12 gene expression and plasma levels and their correlation with disease severity were evaluated in coronary artery disease (CAD) patients. Cellular sources of S100A12 and its biological effects were evaluated in vitro and in vivo in Lyz-S100A12/ApoE−/− and littermate mice fed western diet for 16 weeks. RNA interference technology was used to knockdown RAGE, PDCD4 and NF-κB in vivo.
RESULTS S100A12 in coronary artery and plasma levels was increased in CAD patient which associated with the degree of atherosclerosis, disease severity scores and adverse outcomes of cardiovascular event. Moreover, macrophages were identified as the main S100A12 producing cell in the human coronary atherosclerotic plaque. Macrophage specific S100A12 expression aggravated the atherosclerotic lesion size in the aorta, modulated local and systemic inflammatory responses and significantly upregulated the PDCD4-NF-κB expression in the atherosclerotic plaques compared to those found in littermate mice in vivo. In vitro PDCD4-specific siRNA abolished NF-κB-induced macrophage modulated local and systemic inflammatory responses. Finally, we found S100A12 markedly promoted the binding of CEBP-β to PDCD4 promoter and modulated signaling pathway. Tranilast, blocking S100A12 interaction RAGE, ameliorated vascular inflammation and alleviated plaque regression or prevented progression of established atherosclerosis compared with the ApoE−/− mice.
CONCLUSIONS Taken together, S100A12 released by macrophages promoted the inflammation responses and increased atherosclerosis through activating the PDCD4-NF-κB signal pathway. This suggested that macrophage specific S100A12 expression within the atherosclerotic plaque may be promising early clinical diagnosis or therapy target for the occurrence and development of CAD.
GW34-e0597
Xiaodong Jia, Kun Na, Zhu Mei, Chenhui Yan, Yalin Han
National Key Laboratory of Frigid Zone Cardiovascular Disease, Cardiovascular Research Institute and Department of Cardiology, General Hospital of Northern Theater Command, Shenyang 110016, China
OBJECTIVES In recent years a large number of clinical studies have found that serum APN levels in patients with HF increased, positively correlated with severity of the disease, then put forward “adiponectin paradox” point of view. So, as a cardioprotective factor, APN significantly increases the pathological mechanism and What is the biological function? Is it to combat or aggravate heart function impairment? The correlation between the increase of APN serological level and the severity of HF can make it a biomarker for predicting poor prognosis of HF? These issues have not been clarified so far.
METHODS APN expression in plasma and adipose tissue of mice at different time points was detected by ELISA and Western blot to evaluate the relationship between APN expression and cardiac function. Western blot was used to detect the expression of AMPKα2 and pAMPK protein in the adipose tissue of HF mice, and to evaluate the correlation between the expression and activity of AMPKα2 in the adipose tissue of HF mice before and after HFD feeding and the expression of APN. The 3T3L1 fat progenitor cell lines were used to Compound C and Si-AMPK Compound 2, respectively, for low expression of AMPK and AMPK Compound 2. Established of C57BL6J male mouse and adipoq-ko mouse model of HF by isoprenaline and fed normal diet for 8 weeks.
RESULTS After 20 weeks of feeding with HFD, the body weight of mice showed an obvious decreasing trend. Cardiac ultrasonography showed that LVEF decreased from 53.03±2.80% before feeding to 38.08±1.92% at the end of 28 weeks (P<0.01). FS decreased from 26.98±1.81% to 18.44±1.07% (P<0.01). At 14 weeks of modeling, the plasma APN concentration of the model mice was 2928±129 ng/mL, lower than that of the control mice (P<0.05). The plasma APN concentration of the control mice was 4134±455 ng/mL. The expression of APN in adipose tissue was significantly increased 7 days after Compound C injection compared with the control group. The plasma APN concentration of mice in the Compound C injection group (9963±736 pg/mL) was significantly higher than that in the control group (4377±319 pg/mL), with statistical significance (P<0.05). At the same time, mRNA level detection showed that Compound C intervention did not increase the expression of APN mRNA in the adipose tissue of mice.
CONCLUSIONS Animal and cytological studies confirmed that there was a two-way regulatory relationship between APN and AMPK, and inhibition of AMPK, especially AMPKα2, could up-regulate the expression of APN protein. In mice with HF, adipoq-ko group heart function is worse than C57+ISO group. Experiments have shown that APN has a protective effect on the heart. But, when the heart failure is severe, the APN index is also significantly higher. Then the “adiponectin paradox” point of view needs further study.
GW34-e0600
Cheng Yu, Weihong Lin, Lianglong Chen
Department of Cardiology, Fujian Heart Center, Provincial Institute of Coronary Disease, Affiliated Union Hospital of Fujian Medical University
OBJECTIVES Cell proliferation of vascular smooth muscle cells (VSMCs) accounts for the pathogenesis of vascular neointima formation. Recent studies have reported that RNA modification plays a crucial role in the progression of neointima formation. Acetylated nucleobase ac4C, catalyzed by N-acetyltransferase 10 (NAT10), is well conserved in the enzymatic modification of RNA and is widely distributed in mRNA. However, the biological functions and underlying regulatory mechanisms of mRNA ac4C modifications in CVDs, including vascular remodeling, remain unknown.
METHODS The atherosclerotic segments of the left anterior descending coronary arteries (LAD) and control internal mammary coronary arteries (IMA) were collected. Immunohistochemical staining (IHC), western blotting (WB), HPLC-MS/MS, and Dot blot were performed to detect the level of NAT10 and ac4C modification in LAD and IMA. The rat carotid artery balloon injury model and mouse carotid wire injury were used to study postinjury neointima formation in vivo. Cell proliferation assay, cell migration assay, cell apoptosis assay, HPLC-MS/MS, dot blot, and immunofluorescent staining were conducted to probe the biological functions of NAT10. The potential mechanisms of NAT10 in vascular neointima formation were clarified by RNA-seq, RNA binding protein immunoprecipitation (RIP), acetylated mRNA immunoprecipitation (acRIP), proteomics, etc.
RESULTS IHC and WB showed that NAT10 was highly expressed in LAD compared with IMA. NAT10 protein expression, assessed at days 7, 14, and 28 post balloon injury, was gradually increased in injured carotid arteries compared with sham arteries. Knockdown of rat carotid vascular NAT10 expression suppressed the vascular neointima formation, reduced cell proliferation, and promoted cell apoptosis. Consistently, local overexpression of NAT10 by adenovirus infection significantly exacerbated the balloon injury-induced neointima formation in rats. VSMC-specific deletion of NAT10 dramatically inhibited neointima formation in the vascular wire injury model. Loss- and gain-of-function NAT10 assays also showed that NAT10 promotes the proliferation, migration, and cell apoptosis of VSMCs. HPLC-MS/MS and Dot blot assay results showed that the abundance of ac4C modifications was significantly increased in the sample from human atherosclerotic arteries, rat-injured carotid arteries. RNA-seq, NAT10 RIP, acRIP, and proteomics analysis revealed that NAT10, through an acetylate-modified domain, mediated the stability of Itgb1 mRNA by binding to its mRNA 3′ UTR region and up-regulating its mRNA ac4C modification. CCK-8 and Ki67 staining showed that Itgb1 knockdown significantly abolished the promotion effect of NAT10 ectopic expression. The wound healing and transwell assay demonstrated that the enhancing effect of NAT10 on the invasive potential of VSMC was abrogated by Itgb1 silencing. Besides, the inhibitor of NAT10, remodelin, was applied in vivo which showed an inhibitory effect on vascular neointima formation.
CONCLUSIONS Our results suggested that NAT10 maintained Itgb1 mRNA stability through mRNA ac4C modification resulting in increased IGTB1 expression, and aggravated postinjury neointima formation in vivo.
GW34-e0606
Guanxing Pan1,2, Zhiyong Qi3, Xiaowen Wu4, Lin Chang2, Wei Zhang2, Jianjun Zhang2,5, Haoxuan Zhong3, Si Zhang2, She Chen2, Zhongren Ding2,4
1Department of Pharmacy, The First Affiliated Hospital of Fujian Medical University, Fuzhou, China
2Department of Biochemistry and Molecular Biology, School of Basic Medical Sciences, Fudan University, Shanghai, China
3Department of Cardiology, Zhongshan Hospital, Fudan University, Shanghai, China
4School of Pharmacy, Tianjin Medical University, Tianjin, China
5Department of Clinical Laboratory, Shanghai Pudong Hospital, Fudan University Pudong Medical Center, Shanghai, China
OBJECTIVES Atherosclerosis, the main pathological basis of cardiovascular diseases including coronary artery disease (CAD) and stroke, has been proved to be a chronic inflammatory disease. The role of nucleotide-binding oligomerization domain 2 (NOD2) in atherosclerosis remains undetermined despite its pivotal role in immunity and inflammation.
METHODS NOD2 in platelets, peripheral blood mononuclear cells (PBMCs), and exosomes isolated from blood samples of patients with CAD and atherosclerotic mice was analyzed by qPCR and Western blot (WB). Washed platelets were activated by agonists and NOD2 in supernatant exosomes was analyzed by WB. The uptake of platelet-derived exosomes by human monocytic THP-1 cells was visualized by immunofluorescent staining. The effects of NOD2 on THP-1 cell migration, adhesion, differentiation, and inflammatory response were investigated using a NOD2-overexpressing lentiviral vector or NOD2 antagonist GSK669, followed by WB analysis of the underlying mechanism. GSK669 was intravenously administrated to ApoE−/− mice fed on a high-fat diet (HFD) to seek the therapeutic effects of GSK669 on atherosclerosis.
RESULTS Unexpectedly, we found that platelets from patients with CAD and atherosclerotic mice express less NOD2 at the protein level; interestingly, the PBMCs from the same patients and mice show higher NOD2 expression. The NOD2 change is not caused by transcriptional regulation. We also observed the higher NOD2 protein contents in the plasma exosomes of patients. In line, activated platelets release NOD2-containing exosomes, which increase NOD2 expression in THP-1 monocytes. NOD2 overexpression in THP-1 monocytes enhances cell migration and inflammatory response without changing cell adhesion and differentiation. Consistently, NOD2 antagonist GSK669 attenuates the migration and inflammatory response of THP-1 monocytes. Mechanically, NOD2 triggers the phosphorylation of receptor-interacting protein 2 (RIP2), further enhancing the JNK/p38 axis during THP-1 differentiation. Intravenous administration of GSK669 significantly reduces atherosclerotic lesions and macrophage infiltration in hyperlipidemic ApoE−/− mice.
CONCLUSIONS Platelet NOD2 exacerbates atherosclerosis. Mechanistically, activated platelets secret exosomes that transfer NOD2 protein to monocytes and enhance monocyte migration and inflammatory response through the RIP2/JNK/p38 signaling cascade. GSK669, the NOD2 receptor antagonist, shows the potential as a therapeutic agent for atherosclerosis.
GW34-e0630
Kaifan Niu, Zhixiang Wang, Tianbao Ye, Liang Liu, Xia Lu, Xian Jin
Shanghai Sixth People’s Hospital Affiliated to Shanghai Jiao Tong University School of Medicine
OBJECTIVES Accumulation of senescent cells and production of senescence-associated secretory phenotype (SASP) are associated with many cardiac pathologies. The precise function and mediation of senescent cells, especially senescent cardiomyocytes, under myocardial ischemia remains poorly understood. Nuclear receptor subfamily 4 group A member 1 (NR4A1), an important regulator of biological processes such as oxidative stress and inflammatory response, may serve as a potential target in mediating cellular senescence.
METHODS Myocardial ischemia/reperfusion (I/R) injury was constructed in mice by left anterior descending coronary artery ligation for 45 minutes, followed by reperfusion for 24 hours. Hypoxia/reoxygenation injury were conducted with neonatal rat cardiomyocytes (NRCMs) with hypoxia for 6 hours, followed by reoxygenation for 18 hours. NR4A1 knockout mice were utilized to determine the function of NR4A1, while adeno-associated virus 9 (AAV9) and adenovirus (Adv) were employed to manipulate the expression of NR4A1 in mice hearts and cultured NRCMs, respectively. Senescence-associated ß-galactosidase (SA-ß-gal) staining was used to detect senescent cells. Senescent markers p16 and p21 were examined through immunofluorescence (IF) and western blot (WB), and SASP was measured by real-time quantitative polymerase chain reaction (RT-qPCR) and enzyme-linked immunosorbent assay (ELISA). Morphological alterations of cardiomyocytes were observed by wheat germ agglutinin (WGA) staining in vivo and IF staining of cardiac troponin T (cTnT) in vitro. Live cell imaging of NRCMs were taken to evaluate cardiomyocyte contractility function, while cardiac systolic and diastolic function in mice were assessed by transthoracic echocardiography. Coimmunoprecipitation (Co-ip) and nucleoplasmic separation were performed to explore the underlying mechanism of NR4A1 on cardiomyocyte senescence.
RESULTS Cellular senescence was evident in injured myocardial tissues after I/R, as shown by SA-ß-gal staining. In vitro experiments proved cardiomyocyte senescence was markedly increased in NRCMs after H/R, as significant elevation of senescent markers p16 and p21 were observed. Transcriptome analyses discovered cellular senescence in strong correlation with inflammatory response, and NR4A1, reduced after I/R, may be a potential regulator of these biological processes. Remarkably elevated senescent markers were observed in NR4A1 knockout I/R mice, while NR4A1 overexpression attenuated I/R induced myocardial senescence. With addition of D-galactose as positive control, it is affirmed NR4A1 overexpression protected against cardiomyocyte senescence. Live cell imaging and cytoskeletal staining demonstrated that senescent cardiomyocytes exhibited impaired contractility with hypertrophic morphology, while cardiomyocytes overexpressing NR4A1 showed a less severe decline in contractile function, along with milder morphological alterations. Overall, NR4A1 knockout led to exacerbated cardiac function, and myocardial specific NR4A1 overexpression exerted protective effects after I/R. To explore the precise function and possible mechanism of NR4A1 in regulating cardiomyocyte senescence, markers of SASP were evaluated to reveal significantly increased levels of pro-inflammatory SASP factors in NR4A1 knockout I/R mice, including IL-1β, CXCL1, CCL2, while noticeably lower levels in NR4A1 overexpressing I/R mice, suggesting NR4A1 may contribute to weakened inflammatory response. As a prominent reduction in the elevation of NF-κB p65 phosphorylation was observed in NR4A1 overexpressing I/R mice compared to wildtype I/R mice, IF and nucleoplasmic separation were further employed to reveal NR4A1 overexpression inhibited p65 nuclear translocation. Furthermore, co-ip confirmed interaction between NR4A1 and NF-κB as well as significantly stronger interaction after NR4A1 overexpression after both I/R and H/R.
CONCLUSIONS Cardiomyocyte senescence exacerbated in response to I/R injury can be attenuated by NR4A1, at least partly through inhibition of NF-κB signaling. Anti-senescence treatment of cardiomyocytes can be a novel therapeutic strategy for myocardial ischemia and infarction.
GW34-e0636
Xi Zhang, Dan Liu, Ziqi Liu, Haixu Song, Xiaolin Zhang, Quanmin Jing, Chenhui Yan, Yaling Han
National Key Laboratory of Frigid Zone Cardiovascular Disease, Cardiovascular Research Institute and Department of Cardiology, General Hospital of Northern Theater Command, Shenyang 110016, China
OBJECTIVES Members of the S100 protein family have been reported to function as endogenous danger signals (alarmins) playing an active role in tissue inflammation and repair when released from myeloid cells. S100A12 represents a novel biomarker and therapeutic target that regulates cardiovascular disease after its release. However, the role of S100A12 in the etiology of acute myocardial infarction (AMI) is not well understood. Neutrophil extracellular traps (NETs) are implicated in the pathogenesis of AMI and in the externalization of some S100 family members. Here, we investigated the effect of S100A12 on neutrophils’ function and myocardial injury after AMI.
METHODS Since S100A12 is a human specific molecule and not expressed in mice, we constructed transgenic (TG) mice expressing S100A12 in myeloid cells. Myocardial infarction was induced by ligating the left anterior descending coronary artery (LAD). Cardiac function was assessed by echocardiography. Neutrophils from TG and WT mice bone marrow were isolated and cultured. In vitro experiments we used the overexpressing plasmid to increase the expression of S100A12 and small interfering RNA (siRNA) to inhibit the expression of S100A12. We used Western blotting, quantitative RT-PCR, immunofluorescence, immune-histochemistry and Masson’s trichrome staining to explore the function of S100A12. The levels of S100A12 and NETs related markers (Such as MPO) were detected in the blood by ELISA. The levels of NETosis- and apoptosis-associated proteins were analysed by western blot. The formation of NETs was visualized using differential interference contrast (DIC) method.
RESULTS We found significant increase of S100A12 level after AMI in vitro and in vivo. Moreover, S100A12 markedly increased neutrophil infiltration and exacerbated the damage of myocardial injury in TG mice compared to WT mice. Within the infarct zone, more markers of NETs were observed in TG mice, whereas intraperitoneal injection of DNase I decreased the damage effect of NETs. By ChIP and dual-luciferase reporter assay we found that Hypoxia-inducible factor-1α (HIF1α) directly bound to the promoter of the S100A12 gene and regulated S100A12 expression. We used the overexpressing plasmid of S100A12 and observed that S100A12 enhanced the level of NETosis. Whereas, siRNA-mediated S100A12 silencing reduced the production of NETs. Mechanistically, S100A12 binds with the receptor for advanced glycation end products (RAGE) and increases NETs formation and siRNA-mediated gene of RAGE ameliorated the effect of S100A12. We next used tandem mass tag (TMT) labeling quantitative proteomic technology and discovered that annexin a5 (ANXA5) was a receptor with significantly different expression level in TG compared with WT mice. In vitro experiments demonstrated that there existed a direct interaction between ANXA5 and RAGE. Importantly, S100A12-ANXA5-RAGE pathway was confirmed to be involved in the formation of NETs in neutrophils. We found and verified that GATA1 and STAT3 are transcription factors that play an important role among them.
CONCLUSIONS Our findings reveal a critical role of S100A12 in regulating neutrophil activation and NETs formation, resulting in aggravated myocardial injury after AMI. In such a process, S100A12 regulates NETosis via the Hif1α-S100A12-ANXA5/RAGE pathway. S100A12 is a useful molecular marker and possible target for treatment for AMI.
GW34-e0651
Yujie Zhang
National Key Laboratory of Frigid Zone Cardiovascular Disease and Department of Cardiology, The General Hospital of Northern Theatre Command
OBJECTIVES Qiliqiangxin Capsules (QLQX), a traditional Chinese patent medicine, has been approved for clinical treatment of heart failure. However, protective effects and molecular mechanism of QLQX on Ang II-induced chronic heart failure remains little known.
METHODS Chemical constituents and potential targets of QLQX were obtained via the Traditional Chinese Medicine Systems Pharmacology (TCMSP). The DisGeNET, and GeneCards databases were used to collect chronic heart failure-related target genes. Based on common targets related to both QLQX and chronic heart failure, a protein interaction network was generated using the STRING database. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses were performed. Network diagrams of the active component–target and protein–protein interactions (PPI) were constructed using Cytoscape. Finally, the most important signaling pathway was validated experimentally in C57 mice administrated by Angiotensin II (1000 ng kg−1 min−1) for 28 days.
RESULTS The target and mechanism of QLQX in the treatment of CHF were analyzed based on network pharmacology: The results of the study showed that pharmacological studies yielded 700 targets for CHF and 239 targets for QLQX. The target of QLQX was combined with the target of chronic heart failure to obtain 99 overlapping targets, which have different high expression in different cells in the heart: Immunocytes (52), endothelial cells (51), cardiomyocytes (34), smooth muscle cells (33) and fibroblasts (31). A protein-protein interaction network with 99 nodes and 1747 edges was obtained by topological screening. The average node degree value was 38, in which the node represented the target, the edge represented the association between two targets, and the degree value represented the intensity of the association. The top ten targets of correlation strength were Interleukin-6 (IL-6), Albumin (ALB), Threonine kinase 1 (AKT1), Vascular endothelial growth factor (VEGFA), Tumor necrosis factor (TNF), Nitric oxide synthase 3 (NOS3), Epidermal growth factor (EGF), Mitogen activated protein kinase 8 (MAPK8), Chemokine-8 (CXCL8) and Endothelin1 (EDN1). It is suggested that they play an important role in QLQX anti CHF. By analyzing KEGG signaling pathway of nodes in the network, we found that the top 10 signaling pathways related to CHF were AGE-RAGE, IL-17, blood flow shear stress and atherosclerosis, HIF-1, TNF, NF-kappa B, cGMP-PKG, JAK-STAT, MAPK, and AMPK signaling pathways. It is suggested that QLQX may play a role in the treatment of CHF through the above pathways. Among these signaling pathways, AGE-RAGE was selected for validation in C57 mice. The results indicated that QLQX may inhibit inflammatory processes in chronic heart failure via the AGE-RAGE signaling pathway.
CONCLUSIONS This study provides a strategy for understanding the mechanism of QLQX against chronic heart failure by combining network pharmacology and experimental validation.
GW34-e0652
Yujie Zhang
National Key Laboratory of Frigid Zone Cardiovascular Disease and Department of Cardiology, The General Hospital of Northern Theatre Command
OBJECTIVES Cardioembolic stroke (CS) contributes significantly to a growing proportion of ischemic strokes (IS). Furthermore, the incidence of CS is anticipated to increase manifold in the upcoming decade. Recent studies suggest that inflammatory immune response and fibrosis in the atrial substrate lead to atrial fibrillation (AF)/CSs. Nonetheless, the regulatory mechanisms related to immune infiltration and fibrosis in CS have received insufficient attention in academic research.
METHODS Initially, the GEO database was accessed to obtain gene expression profiles of control and CS samples. The identification of immune- and fibrosis-associated genes was conducted separately through the ImmPort and DisGeNet databases, respectively. Twenty-seven hub genes associated with CS were acquired through differential analysis as well as WGCNA; the analyses were conducted on whole blood samples obtained from GSE58294. Moreover, in order to obtain the key gene MMP9, a core subnetwork comprising twenty-seven hub genes was identified and subsequently intersected with immune- and fibrosis-related differential expression genes (DEGs) (ImmDEGs and FibDEGs).
RESULTS Twenty-two herbal ingredients were predicted based on MMP9. Eventually, the results revealed that MMP9 docks well with isoliquiritigenin (ISL), shikonin, and 1,4-naphthoquinone. Furthermore, herbs rich in ISL, shikonin, and 1,4-naphthoquinone, suppress immune infiltration and alleviate myocardial fibrosis, ultimately leading to an improvement in CS by regulating MMP9.
CONCLUSIONS The proposed strategy offers a systematic approach to elucidate phenotype-target-herb compound associations based on CS, thereby providing novel perspectives for identifying lead compounds from Chinese herbs.
GW34-e0677
Zihui Zhang1, Heng Ma1,2
1Xi’an Key Laboratory of Stem Cell and Regenerative Medicine, Institute of Medical Research, Northwestern Polytechnical University, Xi’an, Shaanxi 710072, China
2Department of Physiology and Pathophysiology, School of Basic Medicine, Fourth Military Medical University, Xi’an, Shaanxi 710032, P.R. China
OBJECTIVES Myocardial ischemia/reperfusion (I/R) injury is characterized by cell death via various cellular mechanisms upon reperfusion. As a new type of cell death, ferroptosis provides new opportunities to reduce myocardial cell death. Ferroptosis is known to be more active during reperfusion than ischemia. However, the mechanisms regulating ferroptosis during ischemia and reperfusion remain largely unknown.
METHODS The contribution of ferroptosis in ischemic and reperfused myocardium were detected by administered of Fer-1, a ferroptosis inhibitor to C57BL/6 mice, followed by left anterior descending (LAD) ligation surgery. Ferroptosis was evaluated by measurement of cell viability, ptgs2 mRNA level, iron production, malondialdehyde (MDA) and 4-hydroxynonenal (4-HNE) levels. H9C2 cells were exposed to hypoxia/reoxygenation to mimic in vivo I/R. We used LC-MS/MS to identify potential E3 ligases that interacted with frataxin in heart tissue. Cardiac-specific overexpression of frataxin in whole heart was achieved by intracardiac injection of frataxin, carried by adeno-associated virus serotype 9 (AAV9) containing cardiac troponin T (cTnT) promoter, then cardiac function as well as ferroptosis were measured.
RESULTS We showed that regulators of iron metabolism, especially iron regulatory protein, were increased in the ischemic myocardium or hypoxia cardiomyocytes. In addition, we found that frataxin, which is involved in iron metabolism, is differentially expressed in the ischemic and reperfused myocardium and involved in the regulation of cardiomyocytes ferroptosis. Furthermore, we identified an E3 ligase, NHL repeat-containing 1 (NHLRC1), that mediates frataxin ubiquitination degradation. Cardiac-specific overexpression of frataxin ameliorated myocardial I/R injury through ferroptosis inhibition.
CONCLUSIONS Through a multi-level study from molecule to animal model, these findings uncover the key role of frataxin in inhibiting cardiomyocyte ferroptosis and provide new strategies and perspectives for the treatment of myocardial I/R injury.
GW34-e0678
Wang Xiaoqi1,2, Yan Xiangyu1,2, Chen Yujia1,2, Mang Ge1,2, Zhang Maomao1,2
1Department of Cardiology, The Second Affiliated Hospital of Harbin Medical University
2The Key Laboratory of Myocardial Ischemia
OBJECTIVES Neutrophils have been implicated as intense responders of Myocardial ischemia reperfusion (MI/R) injury for immunometabolic rewiring locally. Metabolic stress derived protein lactylation plays an important role in immune response. However, the functions of lactylation in neutrophils post MI/R remain unknown. We aimed to investigate the mechanism of lactylation involving in remote immunometabolic reprogramming in neutrophils post-MI/R.
METHODS We utilized single-cell transcriptome of bone marrow and circulating neutrophils exposed to various MI/R periods to explore immunometabolic pattern. We combined with proteomic and lactyl proteomics analysis to identify the S100a9K26 as a major lactylated target. Mouse models including Padi4-knockout, Tlr4-knockout, and S100a9K26R mutant mice, followed by multiple molecular biological methodologies, were used to demonstrate the role of S100a9K26 lactylation (S100a9K26la) in MI/R. Serum S100a9K26la levels were measured in acute myocardial infarction (AMI) patients after percutaneous coronary intervention (PCI) to evaluate the clinical relevance.
RESULTS We demonstrated that inflammatory and glycolytic process in neutrophils were activated in the distant bone marrow, far from the injury heart. We provided the first evidence that S100a9k26la driven by glycolysis elevated in neutrophiles along their trajectory post MI/R. Mechanistically, lactylation of S100a9 amplified the inflammation through the inhibition of S100a9 degradation, promotion of neutrophil extracellular traps (NETs) dependent S100a9 release and boost of neutrophils hyperactivition via toll like receptor 4 (TLR4). Blockade of S100a9K26la inhibits inflammation and improves cardiac function post MI/R. Finally, we demonstrated that K26-Lactylated s100a9 is associated with cardiac death in AMI patients.
CONCLUSIONS Our results show that innate immune signaling of neutrophils begins in the bone marrow, implicate glycolysis-derived S100a9K26 lactylation in governing S100a9 degradation and release. We provide evidence that S100a9K26la may serve as a novel therapeutic target and clinical prognostic biomarker for MI/R.
GW34-e0682
Yihui Wang1,2,3, Tang Gao1,2,3, Wenyuan Wang1,2,3, Lingling Xu1,2,3, Yishu Song1,2,3, Lufang Wang1,2,3, Li Zhang1,2,3, Mingxing Xie1,2,3
1Department of Ultrasound Medicine, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology
2Clinical Research Center for Medical Imaging in Hubei Province
3Hubei Province Key Laboratory of Molecular Imaging
OBJECTIVES Heart transplant rejection (HTR) significantly affects the survival time and living quality of postoperative patients. How to effectively control it has always been an important scientific problem. Inducing immune tolerance and noninvasive monitoring it through ultrasound imaging is an extremely promising method for HTR treatment. However, there is still no mature strategy.
METHODS This study proposed the induction of immune tolerance by transfusion of engineered macrophages and the implementation of ultrasound molecular imaging to monitor graft rejection. Secondly, a mouse heart transplantation model was established, and the engineered macrophages were transfused back to induce tolerance and explore the mechanism of induction of immune tolerance. Finally, in vivo bio-orthogonal reactions were used to achieve ultrasound molecular imaging of the engineered macrophages and monitor the rejection reaction.
RESULTS In vitro experiments, RAW264.7 cells were induced to express PD-L1 at 48 h using interferon IFN-γ, and the immunofluorescence results showed that the PD-L1 level was significantly higher in the treated group compared with the control group. In addition, macrophages were incubated with rapamycin nanoparticles for 8 h. Under the stimulation of an inflammatory environment simulated by lipopolysaccharide, macrophages showed a significant increase in drug release at 36 h, while the non-inflammatory group showed a stable drug release level. In vivo experiments, azide-labelled macrophages were infused back into the tail vein of transplanted mice and DBCO-CY5 was injected 24 h later. The fluorescence intensity of the transplanted target site was observed to increase in a time-dependent manner, with a dense fluorescence signal at 5 h. The statistical results were highly significant when compared to the other time points.
CONCLUSIONS Bio-orthogonal glycolytic markers successfully enable non-invasive monitoring of graft rejection sites, and engineered macrophages are biologically active and can respond to the inflammatory microenvironment at the rejection site to target the release of rapamycin in order to induce immune tolerance with PD-L1.
GW34-e0687
Yuan Song1,2,3, Yihui Wang1,2,3, Lingling Xu1,2,3, Wuqi Zhou1,2,3, Wenyuan Wang1,2,3, Tang Gao1,2,3, Li Zhang1,2,3, Mingxing Xie1,2,3
1Department of Ultrasound Medicine, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology
2Clinical Research Center for Medical Imaging in Hubei Province
3Hubei Province Key Laboratory of Molecular Imaging
OBJECTIVES Myocardial ischemia-reperfusion injury (MIRI) has been shown to be closely associated with ferroptosis. Ferroptosis, as a way of cell death caused by iron ion overload, often leads to lipid peroxidation, and then causes rapid stress of lipid droplets (LDs). AIE-based multimodal probes have not been reported to monitor the dynamic changes of LDs in ferroptosis. Therefore, this study intends to use AIE probe to guide lipid drop multimodal imaging in ferroptosis to warn the risk of MIRI, in order to provide a new strategy for the early diagnosis of MIRI.
METHODS In vitro, three different cell lines, Hela, RAW264.7 and H9c2, were cultured with an ferroptosis induction agent (Erastin). The lipid imaging performance of AIE was evaluated by comparing the AIE probe with the commercial lipid trap BODIPY. Secondly, the dynamic relationship between LDs content and lipid peroxidation level in continuous time points after ferroptosis was analyzed and established. In addition, hypoxia and reoxygenation of cardiomyocytes were simulated to further evaluate the correlation between LDs and ferroptosis. Finally, a mouse model of myocardial ischemia-reperfusion injury was constructed. Combined with multimodal imaging such as fluorescence and ultrasound, AIE probe monitoring LDs metabolism was explored at the organismal level to verify the early diagnosis ability of AIE probe on MIRI.
RESULTS The probe is highly sensitive to LDs and can reflect lipid distribution more accurately than BODIPY. The probe study showed that Erastin induced LDs content increased gradually within 9 h and the intracellular lipid peroxidation level was lower. However, the LDs content tended to decrease after 12 h and reached the peak of ferroptosis within 24 h. In addition, the continuous frozen sections and staining of the ischemic myocardium in mice showed that the distribution of LDs could not only accurately indicate the target site of myocardial injury, but also reflect the degree of myocardial injury in mice. In particular, the echocardiography was used to evaluate the myocardial function of mice. it could accurately reflect the pathological status of the myocardium in early MIRI by ultrasonic measurement and quantification.
CONCLUSIONS AIE probe successfully realized early multimodal noninvasive monitoring of MIRI, and for the first time reflected its correlation with ferroptosis through guided LDs imaging, which could further clarify the pathological course of MIRI, especially the metabolic level of ferroptosis, and provide a promising new idea for the early diagnosis of MIRI.
GW34-e0688
Xiaoqiang Tang1,2,3
1Key Laboratory of Birth Defects and Related Diseases of Women and Children of MOE, State Key Laboratory of Biotherapy, West China Second University Hospital, Sichuan University, No.17 People’s South Road, Chengdu, Sichuan 610041, China
2Health Commission Key Laboratory of Chronobiology, Sichuan University, No.17 People’s South Road, Chengdu, Sichuan 610041, China
3Development and Related Diseases of Women and Children, Key Laboratory of Sichuan Province, West China Second University Hospital, Sichuan University, No.17 People’s South Road, Chengdu, Sichuan 610041, China
OBJECTIVES Cardiovascular aging and remodelling have become the key risk factors for cardiovascular diseases and contribute to increased mortality globally. Metabolic dysregulation is the fundamental mechanism underlying cardiovascular aging and diseases. Metabolic dysregulation reprogrammes epigenetic modifications by regulating the metabolite pool to remodel the cardiovascular tissues. However, the epigenetic mechanisms underlying cardiovascular aging and diseases remain largely unknown.
METHODS Our previous studies used clinical samples, animal models, and multiple-omics strategies to elucidate the epigenetic mechanisms underlying cardiovascular aging and diseases. We mainly focused on crotonylation and acetylation to understand the epigenetic mechanisms underlying cardiovascular aging.
RESULTS Our findings have elucidated that: 1) The mitochondrial metabolic enzyme ECHS1 regulated histone crotonylation to drive the reprogramming of cardiomyocyte transcriptome and aging-related cardiac hypertrophy, 2) The deacetylases SIRT2-4 regulated the balance between the anti-oxidative and prooxidative system within the mitochondria to regulate cardiovascular aging and diseases, 3) The deacetylase SIRT2 and metabolic sensor AMPK formed a positive feedback loop to maintain metabolic homeostasis and repressed cardiovascular aging and remodelling.
CONCLUSIONS Our studies have decoded some key epigenetic mechanisms underlying cardiovascular aging and diseases. These findings were published in journals such as Circulation (2017, 2021), Eur Heart J (2017, 2023), J Exp Med (2022), and Signal Transduct Target Ther (2022, 2023). Two of these papers were selected as the Top 10 Basic Research on Cardiovascular Diseases in China, three were highlighted in EHJ/F1000/JACC, and these papers were cited over 2600 times.
GW34-e0708
Jiawen Chen, Tiankai Shan, Tianwen Wei, Liansheng Wang
The First Affiliated Hospital of Nanjing Medical University
OBJECTIVES Cancer therapies inhibiting CHK1-dependent pathways have been shown to be effective in wide variety of tumors, especially combined with gemcitabine. However, the incidence of related cardiac complications limits the further clinical application. This study aimed to elucidate the mechanism of CHK1 inhibition with gemcitabine clinical treatment induced cardiotoxicity.
METHODS Pharmacological CHK1 inhibition and cardiomyocyte-conditional knock out of CHK1 was characterized in conjunction with gemcitabine treatment in murine models. Proteomics analysis was performed to clarify the mechanisms of the combination therapy-induced cardiotoxicity. Changes in cardiac morphology, function and molecule were examined. CHK1 subcellular location was observed on mitochondria by tissue mitochondria isolation assay and immunofluorescent. Overexpression of CHK1 in mitochondria was achieved by mitochondria-targeting sequence directed AAV9 viruses in vivo. Further protein docking analysis and CO-IP was conducted to verify the interaction of SIRT3 and CHK1 in mitochondria. NMCM and adult mice were used for confirming the role of SIRT3-dependent michondrial redox homeostatsis in the combination therapy-induced cardiotoxicity.
RESULTS Both CHK1 pharmacological inhibition and knock out with gemcitabine induced weight loss, heart atrophy and cardiac dysfunction in mice, with the enrichment of dysregulated inflammatory genes identified by proteomics. Further experiments revealed that the united therapy resulted in respiratory chain dysfunction, mitochondrial redox imbalance and cardiomyocyte pyroptosis. We primarily found that CHK1 was localized in mitochondria. Restoration of mitochondrial CHK1 in cKO mice regained mitochondrial redox homeostatsis, inhibited cardiomyocyte pyroptosis and mitigated united therapy induced cardiotoxicity. Mechanically, mitochondrial CHK1 directly binded SIRT3 while CHK1 inhibition or knock out and gemcitabine treatment decreased SIRT3 expression. Overexpression SIRT3 in mice with united therapy maintained mitochondrial function, alleviated pyroptosis and improved cardiac function.
CONCLUSIONS We concluded that mitochondrial CHK1 plays a critical role for mitochondrial redox homeostatsis through SIRT3. Targeting SIRT3 may be a new therapeutic approach for the prevention and clinical treatment of CHK1 pathway inhibition with gemcitabine induced cardiotoxicity.
GW34-e0728
Jianghui Chen, Rongfeng Huang, Haoran Xin, Meiyu Zhou, Lihua Li, Zhihui Zhang, Min-Dian Li
Department of Cardiovascular Medicine, Southwest Hospital, Army Medical University
OBJECTIVES Irregular eating patterns are recently discovered new risk factors for dyslipidemia and hepatic lipid metabolism disorders, which are positively correlated with the risk of cardiovascular diseases. Time-restricted feeding (TRF) that controls the eating time window can improve blood pressure and blood lipids in patients with metabolic syndrome. Meal timing resets circadian clocks in peripheral tissues, particularly in the liver. However, it remains largely uncharacterized whether and how meal timing organizes circadian rhythms beyond the transcriptome. Therefore, in-depth exploration of the mechanism of the peripheral circadian clock system adapting to feeding rhythms will provide new strategies and targets for the prevention and treatment of the risk of cardiovascular diseases related to circadian clock disorders.
METHODS We systematically profiled the proteomes of non modified proteins and those containing the four major PTMs in livers from TRF female mice. Samples were derived from the same cohort of mice and collected every 4 hours for two complete diurnal cycles. We obtained profiles of the rhythmic proteins or modified proteins from each time-restricted feeding regimen and compared the phase regulatory pattern between day/sleep time-restricted feeding (DRF) and night/wake time-restricted feeding (NRF) mice. Trans-omics integrative analyses were performed to reveal essential circadian protein features related to meal timing.
RESULTS We detected robust daily rhythms in 43% of phospho-proteins, 31.9% of lipids, 26.1% of non-modified proteins, 24.6% of ubiquitylated proteins, 17.1% of glycosylated proteins and 2.2% of succinylated proteins. Integrative analyses revealed that clock regulation of fatty acid metabolism represents a key circadian feature that is reset by meal timing.
CONCLUSIONS Together, this dataset represents a comprehensive resource detailing the proteomic and lipidomic responses by the liver to alterations in meal timing, which can provide some references for the prevention and treatment of the risk of cardiovascular diseases related to circadian clock disorders.
GW34-e0734
Jianxin Zhang, Rongfeng Huang, Haoran Xin, Lihua Li, Zhihui Zhang, Min-Dian Li
Department of Cardiovascular Medicine, Southwest Hospital, Army Medical University
OBJECTIVES Circadian desynchrony results in adipose dysfunctions, which further leads to metabolic syndrome such as obesity, hypertension and dyslipidemia. High-fat diet (HFD) feeding rewires circadian rhythms of feeding behavior and peripheral organs. While the liver has been extensively studied, it remains largely unknown how adipose tissue adapt their circadian rhythms upon diet-induced obesity and the flexibility towards weight loss after dietary intervention.
METHODS Eight-week old of wild type male mice on HFD for 13 weeks and achieved an average body weight of 50 grams. Then these diet-induced obese (DIO) mice were randomly assigned to two groups, one group was fed HFD for 7 days and the other was fed matched low-fat diet (LFD). The heart tissue was dissected from mice at Zeitgeber time 0 h (9:00 am, ZT0) and ZT12 (21:00 pm).
RESULTS We detected about 200 and 2500 diurnal genes in HFD and LFD, respectively. Pathway analysis revealed that rhythmic pathways in HFD are represented by circadian rhythm, ribosome biogenesis, nucleosome organization, whereas those in LFD are represented by myeloid cell function. Remarkably, the majority of the circadian clock genes except Clock exhibit robust diurnal rhythm in the adipose tissue of HFD-fed mice. Analysis of mRNAs and proteins in another cohort of HFD-fed mice confirmed that Clock lost rhythmicity at the transcript level but not in proteins. Diet reversal to LFD specifically restores diurnal difference in Clock transcripts in adipose tissue. We matched transcriptomics data with global profiling of neutral lipids, and found that lipid metabolism catalyzed by triglycerol hydrolase Ces1d is a key circadian feature that is activated by diet reversal.
CONCLUSIONS Our work defines the circadian signatures in the adipose tissue of diet-induced obese mice, and their flexibility upon dietary intervention, thereby shedding light on potential clock-modulated tissue-specific pathways during obesity.
GW34-e0735
Cheng-Lin Zhang1, Lei Zhao2, Lei He3, Yu Huang3
1Department of Pathophysiology, Shenzhen University
2School of Biomedical Sciences, Chinese University of Hong Kong
3Department of Biomedical Sciences, City University of Hong Kong
OBJECTIVES Endothelial nitric oxide synthase (eNOS) monomerization/dimerization plays crucial role in regulating endothelium-dependent relaxations (EDR). Growing evidence has revealed that autophagic dysregulation is involved in the pathogenesis of diabetic endothelial dysfunction. However, whether autophagy regulates eNOS activity through controlling eNOS monomerization/dimerization remains elusive.
METHODS Autophagy and autophagic flux was evaluated through WB or through the mCherry-GFP-LC3 system. eNOS dimer and monomer were evaluated through low temperature WB. Autophagy and autophagic flux were induced by torin, rapamycin, as well as calorie restriction. Vascular tone was measured by the wired myograph system.
RESULTS The present study shows that autophagic flux was impaired in the endothelium of diabetic db/db mice as well as in human endothelial cells exposed to advanced glycation end products or oxidized low-density lipoprotein. Inhibition of autophagic flux by chloroquine or bafilomycin A1 reduces eNOS dimerization and lowers nitric oxide bioavailability through raising mitochondrial reactive oxygen species (mtROS). Overexpressing transcription factor EB (TFEB), a master regulator of autophagy and lysosomal biogenesis, restored autophagic flux, lowered endothelial cell ROS level, increased eNOS dimerization, and improved vascular function in db/db mouse aortas. Inhibition of mammalian target of rapamycin kinase (mTOR) increased TFEB nuclear localization, reduced mtROS accumulation, facilitated eNOS dimerization, and improved EDR in db/db mice. Moreover, calorie restriction also increased aortic TFEB expression, improved autophagic flux, and restored EDR in db/db mice.
CONCLUSIONS Taken together, the present study reveals that autophagy deficiency-mtROS-eNOS monomerization axis is involved in the endothelial dysfunction of db/db mice. Restoration of autophagy by targeting TFEB represents a novel approach for the treatment of diabetic endothelial dysfunction.
GW34-e0752
Wenwu Zhu, Wei Du, Bing Han
Division of Cardiology, Xuzhou Central Hospital, Xuzhou Clinical School of Xuzhou Medical University, Xuzhou Institute of Cardiovascular Disease
OBJECTIVES The ideal treatment for myocardial infarction should be able to effectively improve the oxygen supply in the infarct area and reduce the death of myocardial cells. In this study, we designed a nano-delivery system of MnO2@INN@Alb (IHM) used as an effective hypoxia mitigation and pyroptosis control strategy for improving cardiac activity in patients with myocardial infarction, and ultimately reducing mortality in patients with myocardial infarction.
METHODS INN@Alb nanoparticles were first prepared to improve the solubility of INN order to reach the effective concentration for pyroptosis inhibition. Then we verified the encapsulation of each element after modifying INN@Alb with KMnO4 and PEG.
RESULTS We have demonstrated that the synthesized IHM can efficiently produce O2 in the presence of acid (pH<6.5) and H2O2. IHM actively reverses hypoxia, scavenges reactive oxygen species (ROS) and stimulate angiogenesis in the ischemic myocardium of AMI rat. Also, IHM can significantly reverse the occurrence of cardiomyocyte pyroptosis and repress pyroptotic signaling pathway. Moreover, IHM significantly attenuated apoptosis of cardiomyocytes and reduced the infarct size and fibrosis area in ischemic myocardium. Subsequently, the remodeling of the left ventricle was significant alleviated, and heart function of AMI rat was markedly improved and reduced the infarct size and fibrosis area in ischemic myocardium.
CONCLUSIONS This IHM nanoparticle provides a new way to improve the existing treatment and is a potential new venue to treat patients with acute myocardial infarction in the near future.
GW34-e0762
Chaoqun Ma1, Qiang Xu2
1National Key Laboratory of Frigid Zone Cardiovascular Diseases (NKLFZCD), Cardiovascular Research Institute and Department of Cardiology, General Hospital of Northern Theater Command
2Department of Cardiology, Navy 905 Hospital, Naval Medical University
OBJECTIVES Pulmonary arterial hypertension (PAH) is a progressive and life-threatening disease characterized by pulmonary vascular remodeling, which may cause right heart failure and even death. Accumulated evidence has confirmed that microRNA-26 family members play critical roles in cardiovascular disease; however, their function in PAH remains largely unknown. Our study aims to elucidate the role and underlying mechanism of the miR-26a-5p in the development of PAH.
METHODS The expression levels of miR-26 family members were investigated through quantitative reverse transcription polymerase chain reaction (qRT-PCR). Chromatin immunoprecipitation (ChIP) analysis and luciferase reporter assays were used to explore the upstream regulator of miR-26a-5p expression in pulmonary arterial smooth muscle cells (PASMCs). Furthermore, the regulatory role of miR-26a-5p in PASMC autophagy was detected through tandem mRFP-GFP-LC3B fluorescence microscopy and western blots. Gain and loss of function assays were performed to explore the effect of miR-26a-5p on PASMC proliferation and migration, as determined by cell counting kit-8, 5-Ethynyl-2′-deoxyuridine (EdU) staining, wound-healing and transwell assays. Mechanistically, dual-luciferase reporter gene assays and western blots were used to identify the direct downstream targets of miR-26a-5p. Moreover, hypoxia-induced PAH rat models were established. Right ventricular hypertrophy and pulmonary vascular remodeling were evaluated using hemodynamic measurements and histological analysis followed by intraperitoneal injection administration of adeno-miR-26a-5p in vivo.
RESULTS Among the miR-26 family, miR-26a-5p was identified as the most downregulated member in plasma from PAH patients. Meanwhile, the expression of miR-26a-5p was decreased in the hypoxia-induced PASMC autophagy models, as well as lung tissues of PAH patients. Furthermore, ChIP analysis and luciferase reporter assays revealed that hypoxia inducible factor 1α (HIF-1α) specifically interacted with the promoter of miR-26a-5p and inhibited its expression in PASMCs. Tandem mRFP-GFP-LC3B fluorescence microscopy demonstrated that miR-26a-5p inhibited hypoxia-induced PAMSC autophagy, characterized by reduced formation of autophagosomes and autolysosomes. Consistently, miR-26a-5p obviously inhibited the conversion of LC3B-I to LC3B-II and p62 degradation in hypoxia-induced PASMCs. In addition, miR-26a-5p overexpression potently inhibited PASMC proliferation. Mechanistically, 6-phosphofructo-2-kinase/fructose-2,6-biphosphatase 3 (PFKFB3), unc-51 like autophagy activating kinase 1 (ULK1) and ULK2 were identified as direct targets of miR-26a-5p, involving in hypoxia-induced PASMC dysfunction. Meanwhile, PFKFB3 could further enhance the phosphorylation level of ULK1 and promote autophagy in PASMCs. Moreover, intratracheal administration of adeno-miR-26a-5p markedly alleviated right ventricular hypertrophy and pulmonary vascular remodeling in hypoxia-induced PAH rat models in vivo.
CONCLUSIONS These results indicate that the HIF-1α/miR-26a-5p/PFKFB3/ULK1/2 axis plays critical roles in the regulation of hypoxia-induced PASMC autophagy and proliferation. MiR-26a-5p may represent as an attractive biomarker for the diagnosis and treatment of PAH.
GW34-e0763
Chaoqun Ma1, Dingyuan Tu2, Qiang Xu3
1National Key Laboratory of Frigid Zone Cardiovascular Diseases (NKLFZCD), Cardiovascular Research Institute and Department of Cardiology, General Hospital of Northern Theater Command, Shenyang, Liaoning 110000, P.R. China
2Department of Cardiology, The 961st Hospital of Joint Logistic Support Force of PLA, Qiqihaer, Heilongjiang 161000, China
3Department of Cardiology, Navy 905 Hospital, Naval Medical University, Shanghai 200052, P.R. China
OBJECTIVES Heart failure (HF) is the end stage of various cardiovascular diseases with high mortality rate. Novel diagnostic and therapeutic biomarkers for HF are urgently warranted. This study aims to identify potential hub genes and immunological characteristics of HF by the combination of bioinformatics analysis and machine learning.
METHODS The gene expression profiles of 124 HF patients and 135 nonfailing donors (NFDs) were obtained from six datasets in the Gene Expression Omnibus (GEO) public database. We applied robust rank aggregation (RRA) with weighted gene co-expression network analysis (WGCNA) method to identify critical hub genes in HF development. Furthermore, three machine learning methods were employed to discover novel diagnostic markers in HF, including best subset regression, regularization technique, and support vector machine-recursive feature elimination (SVM-RFE). Besides, immune infiltration was investigated in HF by single-sample gene set enrichment analysis (ssGSEA).
RESULTS Through the combination of RRA with WGCNA method, 39 shared genes were recognized to be associated with HF. Through integrating the three machine learning methods, FCN3 and SMOC2 were identified as novel diagnostic markers for HF. In addition, differences in immune infiltration signature were also found between HF patients and NFDs. Immune cell infiltration analysis found that aDCs, B cells, Macrophages, Mast cells, Th2 cells, and Treg may be involved in the HF development. Similarly, there was a different infiltration of immune-related functions in APC co inhibition, cytolytic activity, HLA, inflammation promoting, T cell co inhibition, T cell co stimulation, Type I IFN response between HF and NFDs. Moreover, we explored the potential associations between the expression of two diagnostic markers and immune response in the pathogenesis of HF.
CONCLUSIONS In summary, FCN3 and SMOC2 can be used as diagnostic markers of HF, and immune infiltration plays an important role in the initiation and progression of HF.
GW34-e0764
Chaoqun Ma1, Dingyuan Tu2, Qiang Xu2
1National Key Laboratory of Frigid Zone Cardiovascular Diseases (NKLFZCD), Cardiovascular Research Institute and Department of Cardiology, General Hospital of Northern Theater Command, Shenyang, Liaoning 110000, P.R. China
2Department of Cardiology, Changhai Hospital, Naval Medical University, Shanghai 200433, P.R. China
OBJECTIVES Cuprotosis is a newly discovered programmed cell death which acts by modulating tricarboxylic acid cycle. Emerging evidence has showed that cuprotosis-related genes (CRGs) play critical roles in the development of multiple diseases. However, the mechanism of cuprotosis involved in the occurrence and progression of heart failure (HF) has not been investigated yet.
METHODS Six HF microarray datasets (GSE16499, GSE26887, GSE42955, GSE57338, GSE76701, and GSE79962) were downloaded from the Gene Expression Omnibus (GEO) database to screen differentially expressed CRGs between HF patients and nonfailing donors (NFDs) using R package “limma”. Furthermore, three regularised linear methods-least absolute shrinkage and selection operator (LASSO) regression, RIDGE regression, and elastic net (EN) regression were applied to identify key diagnostic CRGs features by the “glmnet” package, and model performance was assessed by root mean squared error (RMSE). In addition, the potential biological functions of diagnostic CRGs were investigated by utilizing Gene Ontology (GO) function enrichment, Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment, together with immune infiltration analysis.
RESULTS Nine CRGs were differentially expressed in heart tissues from HF patients and NFDs. In addition, a three-CRG diagnostic signature, DLAT, SLC31A1, and DLST, which can easily distinguish HF patients and NFDs, was established by cross-combination of three machine learning algorithms, including LASSO, RIDGE and EN regression. Moreover, function enrichment and immune infiltration analysis demonstrated that the three diagnostic CRGs expression was strongly correlated to alterations of energy metabolism and immune microenvironment in HF.
CONCLUSIONS Our study discovered for the first time that cuprotosis was strongly related to the pathogenesis of HF. A three-CRG diagnostic signature for HF (DLAT, SLC31A1, and DLST) associated with energy metabolism and immune signaling pathways was identified, providing new insights into the underlying mechanisms and effective treatments of HF.
GW34-e0769
Bofang Zhang1,2,3, Hong Jiang1,2,3, Jing Chen1,2,3
1Department of Cardiology, Renmin Hospital of Wuhan University, Wuhan, China
2Cardiovascular Research Institute of Wuhan University, Wuhan, China
3Hubei Key Laboratory of Cardiology, Wuhan, China
OBJECTIVES Maladaptive ventricular remodelling greatly impacts the life quality and long-term prognosis of acute myocardial infarction (AMI) patients. The activation and differentiation of cardiac fibroblast is the core of ventricular remodelling after infarction and initiate the heart’s pathological remodelling process. LncRNAs have been confirmed to be critical in regulating AMI, besides, recently published studies disclosed that several LncRNAs play key roles in regulating the activation of fibroblast. Our present study is to investigate LncRNA that regulates fibroblast activation in AMI and uncover the potential mechanisms.
METHODS RNA-sequencing and bioinformatical analyses were conducted to identify the candidate LncRNAs for AMI. Adenovirus for LncRNA knockout and overexpression were constructed to alter the PVT1 expression of cardiac fibroblast in vitro. Fibroblast-specific knockout and overexpression mice were also constructed to alter PVT1 expression in vivo. Cardiac fibroblast characteristics measurement, including proliferation, migration, secretion, and differentiation, were performed to verify the ability of PVT1 to regulate the activation of fibroblasts. Cardiac morphological, structural, functional, and hemodynamic changes as well as the degree of fibrosis were also measured to assess the effect of PVT1 on AMI. RNA-pulldown+Mass spectrometry and RNA immunoprecipitation were performed to identify the specific binding factors of PVT1.
RESULTS RNA-sequencing results revealed the expression of PVT1 was significantly and persistently elevated from 1 to 7 days after AMI. Besides, RNA-FISH and q-PCR analysis uncovered PVT1 was mainly enriched in the nucleus of the cardiac fibroblast. Thereafter, gain- and loss-of-function experiments were performed both in vitro and in vivo. Functionally, PVT1 knockout dramatically inhibited the proliferation, migration, secretion, and differentiation capability of fibroblast, as well as reduced infarct size, improved cardiac performance and alleviated cardiac fibrosis by mitigating fibroblast activation. However, enforced PVT1 expression resulted in the opposite effects. Mechanistically, RNA-pulldown+Mass spectrometry indicated histone methylase Suv39h1 was a direct target of PVT1, which was also confirmed by RIP assays. In addition, Chip-PCR analysis also demonstrated that Suv39h1 could directly bind to the promoter region of the NF-κB/p65, thus altering the level of H3K9me3 and inhibiting its transcription. Moreover, subsequent rescue experiments further verified that PVT1 regulated the expression of NF-κB/p65 to exaggerate AMI-induced fibrosis in a Suv39h1-dependent manner.
CONCLUSIONS This study demonstrates that PVT1-Suv39h1-NF-κB/p65 pathway is a promising therapeutic target for the treatment of AMI and the subsequent maladaptive ventricular remodelling by ameliorating the activation of cardiac fibroblast.
GW34-e0770
Chenhuiyu Qian1, Zijian Huang1, Yuhan Cao2, Cong Fu1
1Department of Cardiology, The First Affiliated Hospital of Wannan Medical College
2Department of Nephrology, The First Affiliated Hospital of Wannan Medical College
OBJECTIVES The mechanism of myocardium regeneration after myocardial infarction (MI) is still unknown. Exosome-based therapy is a promising cell-free approach for repairing the ischemic myocardium. This study investigated the protective effect of cardiomyocyte derived exosomes carrying myeloid triggering receptor 2 (TREM-2) on myocardium regeneration after MI.
METHODS First, Plasma samples from healthy controls and patients with myocardial infarction were collected to measure plasma exosomes TREM-2 levels. TREM2 expression in hypoxic myocardium derived exosomes was detected in vitro. Myocardial infarction models were established in wild type (WT) mouse and TREM2 gene knockout (TREM-2−/−) mouse by ligating the left anterior descending branch (LAD). Plasma exosomes from WT MI mouse (WT-P-exo) and TREM-2−/− MI mouse (KO-P-exo) were collected. Before ligation of anterior descending branch, exosomes were injected into the myocardium (TREM2−/− mice injected with WT-P-exo, WT mice injected with KO-P-exo). Exosomes inhibitor GW4869 was injected to inhibiting exosomes secretion. The AKT signal pathway inhibitor LY294002 intraperitoneal injected in myocardium 2 hours before surgery. The expression levels of TREM2, Cyclin A2, AKT, and p-AKT were detected in the infarcted and peri-infarcted areas. In addition, the therapeutic efficacy of intramyocardially injected exosomes for MI treatment was evaluated through functional and histological analyses. Macrophage scavengers was used to eliminate the impact of macrophage TREM-2.
RESULTS First, Expression of TREM-2 and Cyclin A2 mRNA in the peri-infarct area of WT mice increased after MI. The expression of TREM-2 derived from hypoxic cardiomyocytes exosomes was elevated compared to normal cardiomyocytes. The TREM-2 expression in plasma exosomes was also increased in MI patients. Injection of GW4869 resulted in a significant decrease in the expression of TREM-2, Cyclin A2 and p-AKT proteins in the peri-infarct area in WT MI mice. Immunofluorescence, HE and Masson staining demonstrated that injection of GW4869 reduced the average fluorescence intensity of CyclinA2 in the infarct margin area, promoted myocardial damage and increased fibrosis area of WT MI mice. The ultrasound results showed that injection of GW4869 aggravated heart damage in WT MI mice. In TREM-2 knock out (TREM-2−/−) mice, the expression of cyclin A2 and p-AKT was no elevated. Injection of WT-P-exo resulted in a significant increase in the expression of TREM-2, Cyclin A2 and p-AKT proteins in the peri-infarct area in TREM-2−/− MI mice, while injection of KO-P-exo did not affected the expression of TREM-2, Cyclin A2 and p-AKT proteins in the peri-infarct area in WT MI mouse. Injection of WT-P-exo into the TREM2−/− MI mice added the average fluorescence intensity of CyclinA2 in the infarct margin area and alleviated myocardial damage and decreased fibrosis area and improved cardiac function. In addition, intraperitoneal injection of LY294002 decreased the levels of TREM-2, Cyclin A2 and p-AKT proteins in the peri-infarct area in WT MI mice and WT-P-exo injected TREM2−/− mice. Further, LY294002 reduced the average fluorescence intensity of CyclinA2 in the infarct margin area, promoted myocardial damage, increased fibrosis area and decreased cardiac function in WT MI mice and WT-P-exo injected TREM2−/− mice.
CONCLUSIONS Exosomes carrying TREM-2 were released from cardiomyocytes to promote the regeneration of cardiomyocytes in infarction margin. The expression of CyclinA2 was increased in infarction margin which makes cardiomyocytes re-enter the cell cycle. It suggested a potential target to intervene myocardial regeneration after MI.
GW34-e0774
Yi Zhong1,2,3, Xueke Li1,2,3, Luyang Yi1,2,3, Yishu Song1,2,3, Wuqi Zhou1,2,3, Wenqu Li1,2,3, Qiaofeng Jin1,2,3, Tang Gao1,2,3, Li Zhang1,2,3, Mingxing Xie1,2,3
1Department of Ultrasound Medicine, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430022, China
2Clinical Research Center for Medical Imaging in Hubei Province, Wuhan 430022, China
3Hubei Province Key Laboratory of Molecular Imaging, Wuhan 430022, China
OBJECTIVES Ischemia-reperfusion injury (IRI) after heart transplantation is a clinically significant sterile inflammation. It can induce primary graft dysfunction within 24 hours after heart transplantation and is the main pathogenic mechanism of post-transplant cardiac dysfunction and failure, leading to death and disability. Alleviating IRI is of great significance for improving graft survival. Recent studies have shown that ferroptosis plays a key role in the initiation of IRI after heart transplantation. Ferroptosis-induced release of damage-associated molecular patterns (DAMPs) promotes the adhesion of neutrophils to coronary endothelial cells through toll-like receptor 4-dependent signaling pathways, thereby triggering harmful inflammatory reactions in the transplanted heart. Therefore, inhibiting ferroptosis can reduce the release of DAMPs and alleviate IRI. As a promising cell-free therapy, extracellular vesicles derived from bone marrow mesenchymal stem cells (BMSC-EVs) can effectively attenuate IRI. However, whether BMSC-EVs can alleviate transplant heart IRI by inhibiting ferroptosis remains unknown, and the underlying mechanisms need to be elucidated. The aim of this study is to reveal the molecular mechanisms by which BMSC-EVs alleviate cardiac allograft IRI by inhibiting ferroptosis.
METHODS In this study, a heterotopic heart transplantation model was established using BALB/c mice as donors and C57BL/6 mice as recipients. Allografts were collected at 2 h, 5 h, 8 h, 24 h, 48 h, and 72 h after reperfusion to investigate the timing of ferroptosis occurrence. After heart transplantation, mice were intravenously injected with BMSC-EVs, and the control group received an equal volume of saline to evaluate the therapeutic effect of EVs. MicroRNA databases, EV databases, and dual luciferase reporter gene experiments were used to identify miRNAs targeting ferroptosis-related genes in BMSC-EVs. The molecular mechanisms by which BMSC-EVs inhibit ferroptosis in transplanted hearts and cardiac cells through their miRNAs were further explored using gene overexpression/knockdown, qRT-PCR, western blotting, flow cytometry, and biochemical analysis. Additionally, histological analysis was performed to assess the efficacy of BMSC-EVs in attenuating cardiac allograft IRI.
RESULTS Biochemical analysis and qRT-PCR showed that compared to the control group, the levels of ACSL4, BECN1, Hmox1, iron, and malondialdehyde were significantly elevated in the cardiac allografts at 24 h after reperfusion, indicating that 24 h after reperfusion is the optimal timepoint to observe ferroptosis in cardiac transplant mice. BMSC-EVs significantly inhibited ferroptosis in cardiac allografts, as demonstrated by the significant reduction in ACSL4, BECN1, Hmox1, iron, and malondialdehyde. Histological analysis also indicated that BMSC-EVs significantly alleviated cardiomyocyte apoptosis and tissue damage, and reduced the levels of inflammatory factors in myocardial tissue. Based on TargetScan and ENCORI database analysis, miR-144-3p, which is rich in BMSC-EVs, was found to inhibit the expression of ACSL4. The direct binding of miR-144-3p to ACSL4 was validated through dual luciferase reporter gene experiments. Gene overexpression and knockdown experiments confirmed that ACSL4 promotes ferroptosis in cardiomyocytes during the process of hypoxia-reoxygenation. Moreover, cell co-incubation experiments with BMSC-EVs demonstrated that BMSC-EVs inhibited ACSL4 and upregulated GPX4. This effect was weakened when BMSC-EVs containing miR-144-3p inhibitor were used, indicating that BMSC-EVs can inhibit ferroptosis in cardiomyocytes during the process of hypoxia-reoxygenation by suppressing ACSL4 through miR-144-3p.
CONCLUSIONS This study revealed that BMSC-EVs can inhibit ferroptosis and alleviate cardiac allografts IRI by delivering miR-144-3p to suppress ACSL4. This novel mechanism by which BMSC-EVs exert their therapeutic effects will provide new strategies for developing more potent engineered BMSC-EVs.
GW34-e0779
Hanwen Zhang, Qi Chen
Department of Pathophysiology, Nanjing Medical University, Nanjing 211166, China
OBJECTIVES Macrophage-elicited inflammation is an essential player in obesity-associated heart disease. However, the underlying mechanism remains obscure. Our study aimed to identify the key macrophage population involved in obesity-induced cardiac dysfunction and investigate the molecular mechanism that contributes to the inflammatory response.
METHODS We conducted an epidemiological study of 8567 patients with cardiovascular diseases (CVDs) to identify etiological factors involved in obesity-induced cardiac dysfunction. We used lineage tracing to evaluate the contribution of resident and monocyte-derived macrophages in diet-induced obesity mice. Single-cell RNA-sequencing analysis of CD45+CD11b+F4/80+ cardiac macrophages from normal and obese mice were performed to explore the heterogeneity of macrophages. We then used a dual recombinase system (Cre and Dre) to specifically ablate CCR2+ macrophages. We further investigated the role of CCR2 in modulating obesity-induced cardiac inflammation and dysfunction by macrophage specific CCR2 deletion or CCR2 antagonist treatment. The molecular mechanism underlying the pro-inflammatory effect of CCR2+ macrophages was finally assessed at in vivo and in vitro by CUT&TAG, ChIP-PCR, luciferase assay, and macrophage-specific lentivirus transfection.
RESULTS Our population study and mouse model experiments reveal that inflammation, particularly monocyte-derived macrophage infiltration, exacerbates obesity-associated cardiac dysfunction in humans and mice. We identify 17 macrophage clusters in normal and obese hearts. We further show that the Ccr2 cluster undergoes functional transition from homeostatic maintenance to pro-inflammation. Consistently, inducible ablation of CCR2+ macrophages or selective deletion or inhibition of macrophage CCR2 prevents cardiac dysfunction. At the mechanistic level, we demonstrate that obesity-induced functional shift of CCR2-expressing macrophages is mediated by the CCR2/ATF3/Lysozyme C/NF-κB signaling. We uncover a noncanonical role for lysozyme C as a transcription activator, binding to the RelA promoter and driving NF-κB signaling, and strongly promotes inflammation and cardiac dysfunction in obesity.
CONCLUSIONS We present here a revelation of macrophages in cardiac dysfunction under metabolic challenge. In addition, we elucidate the contribution of CCR2+ macrophages to cardiac inflammation and dysfunction through the regulation of ATF3/Lysozyme C/NF-κB signaling. Our findings suggest that lysozyme C may represent a potential target for diagnosis of obesity-induced inflammation and treatment of obesity-induced heart disease.
GW34-e0827
Victoria Khotina1, Mariam Bagheri-Ekta2, Arthur Lee1,2, Vasily Sukhorukov1,2
1Institute of General Pathology and Pathophysiology
2Petrovsky National Research Centre of Surgery
OBJECTIVES PERK is an endoplasmic reticulum-resident transmembrane protein that plays a crucial role in the unfolded protein response (UPR), which is activated in response to ER stress. While PERK’s primary function is to regulate protein folding and maintain ER homeostasis, emerging evidence suggests that PERK may be also involved in the regulation of inflammatory pathways in macrophages. However, there is a lack of data on PERK-mediated regulation of the inflammatory response in macrophages during atherosclerosis. The main aim of this study was to evaluate the impact of PERK on the regulation and activation of the inflammatory pathways in macrophages.
METHODS Human monocyte-like cell line THP-1 was used as control. CRISPR/Cas9 editing was used to generate THP-1 cell line with PERK gene knockout (THP-1 PERK −/−). Macrophage-like phenotype differentiation was induced by phorbol 12-myristate 13-acetate (PMA). Pro-inflammatory response was induced by cell incubation with lipopolysaccharides (LPS). The expression of C-Jun N-Terminal Kinase 1 (JNK1A2), nuclear factor kappa B subunit 1 (NFKB1), peroxisome proliferator activated receptor gamma (PPARG), tumor necrosis factor alpha (TNFA), interleukin 1 beta (IL1B) and interleukin 6 (IL6) genes were assessed by qPCR.
RESULTS To induce PERK knockout, THP-1 cells were transfected using two sgRNAs and a plasmid containing an expression cassette with the sequence of the puromycin resistance gene. A stable clone of cells with the most effective PERK knockout was obtained after 4 weeks of puromycin selection. PERK gene knockout in the obtained clone was confirmed by qPCR and Sanger sequencing. It was observed that there was no difference in the basal expression of JNK1A2, NFKB1, TNFA, IL1B and IL6 between THP-1 and THP-1 PERK −/− macrophages. However, the basal expression of PPARG in THP-1 PERK −/− was significantly increased compared to the control cells (P<0.001). Upon pro-inflammatory stimulation of THP-1 macrophages with LPS, there was an upregulation in the expression of JNK1A2, NFKB1, TNFA, IL1B and IL6, accompanied by a downregulation of PPARG compared to the untreated cells (P<0.01). On the other hand, the expression levels of JNK1A2, NFKB1, PPARG showed no significant differences in THP-1 PERK −/− macrophages compared to the untreated cells following LPS stimulation. However, LPS stimulation of THP-1 PERK −/− macrophages resulted in a decreased expression of TNFA and an increased expression of IL1B and IL6 compared to the untreated cells (P<0.01). Interestingly, the expression level of IL6 was decreased in THP-1 PERK −/− cells compared to the control cells upon LPS stimulation (P<0.01), while the expression of IL1B showed no significant differences.
CONCLUSIONS Our findings suggest that PERK may play a role in regulating the expression of pro-inflammatory genes, particularly JNK1A2, NFKB1, PPARG, TNFA and IL6, in macrophages during inflammatory responses. This indicates that PERK-mediated signaling in macrophages may have a significant impact on modulating inflammation, which is a crucial factor in the development and progression of atherosclerotic plaques.
This work was supported by Russian Science Foundation Grant #22-25-00393.
GW34-e0845
Yilin Zhou, Xiaomin Wei, Yuan Han, Enge Shao, Zhiwen Xiao, Yanbing Wang, Xiaoran Shi, Xinzhong Li
Department of Cardiology, Nanfang Hospital, Southern Medical University
OBJECTIVES Myocardial infarction (MI) leads to reactive oxygen species (ROS) production, which triggers DNA damage response (DDR) and cell cycle arrest in the cardiomyocytes (CMs) of infarcted area, thereby limiting the efficiency of endogenous myocardial regeneration and interfering with cardiac repair. Telomeric DNA, located at the ends of chromosomes, is particularly vulnerable to ROS-induced damage, while effective repair methods for the damage are currently lacking. This study aims to investigate whether telomerase reverse transcriptase (Tert), which has a protective effect on telomeric DNA, can effectively repair telomeric DNA damage caused by ROS, inhibit DDR-induced cell cycle arrest, and promote myocardial regeneration after MI.
METHODS The telomere length and telomerase activity were assessed using TEL-CY3 immunofluorescence staining and ELISA. Primary CMs were isolated and various interventions such as siRNA knockdown, adenoviral overexpression, and specific inhibitors were used to modulate the expression and function of Tert. Additionally, CM-specific Tert knockout mice generated using the Cre-loxP system or mice with adenovirus-mediated Tert overexpression were used for in vivo experiments. WB, qPCR, immunofluorescence and other techniques were employed to validate the impact of Tert on telomeric DNA damage repair and CM cell cycle re-entry. Echocardiography and histological staining were performed to evaluate heart function and cardiac remodeling. Subsequently, DNA pulldown and immunoblotting experiments were conducted to explore the upstream and downstream mechanisms of Tert regulation.
RESULTS The telomere length and telomerase activity in mouse CMs were found to be negatively correlated with age. Overexpression of Tert enhances telomerase activity and lengthens telomeres, thereby repairing DNA damage induced by ROS and promoting CM proliferation in vitro. In vivo experimental results demonstrate that enhancing Tert significantly improves cardiac function and prognosis by alleviating CM DNA damage post-MI. In terms of mechanism, DNA pulldown experiments identified hnRNPA2B1 as a potential upstream regulator of Tert in CMs. Further immunoblotting analysis revealed that Tert protects DNA from ROS-induced damage by inhibiting ATM phosphorylation and blocking activation of the Chk1/p53/p21 pathway.
CONCLUSIONS HnRNPA2B1-activated Tert could repair telomeric DNA via extending the length of telomeres and enhancing telomerase activity, thus blocking the ATM/Chk1/p53/p21signaling pathway, inducing the cell cycle re-entry to achieve the cardiac regenerative repair after MI.
GW34-e0852
Hao Zhang1,2,3, Zhuqing Li2, Wei Cai4, Yanxin Wang2, Jiaxin Feng2, Ning Yang1, Chengzhi Lu2, Yuming Li1
1Clinical School of Cardiovascular Disease, Tianjin Medical University, Tianjin, China
2The First Central Clinical School, Tianjin Medical University, Tianjin, China
3Center for Disease Control in Hanzhong, Hanzhong, Shaanxi, China
4Characteristic Medical Center of People’s Armed Police Force, Tianjin, China
OBJECTIVES Researchers have suggested the hypotensive effect of Sodium-dependent glucose transporters-2 inhibitor (SI), which may be a new target to improve hypotensive effect of renal denervation (RDN). We used spontaneously hypertensive rats (SHRs) to confirm this conjecture and explore relevant possible pathways.
METHODS 7 Wistar-Kyoto (WKY) rats and 35 SHRs were randomly divided into 6 groups. The blank control group (WKY Control, W group) was 7 WKY rats, while hypertension group (SHR control group, S group), the hypertensive sham operation group (Sham group), the RDN group (RDN group, R group), the SGLT-2 inhibitor control group (SI group) and RDN combined with sodium-dependent glucose transporters-2 inhibitor group (SIR group) was 7 SHR rats. Before RDN of R and SIR group, the SIR group was gavaged with dapagliflozin (DAPA), and the SI group was gavaged at the same time. Sham group was treated with sham operation. One week after the operation, the rat tail manometry and echocardiography were performed, and the peripheral blood inflammatory cells were detected by flow cytometry before sample collection. After sample collection, the serum was tested by enzyme linked immunosorbent assay (ELISA), Including interleukin-6(IL-6), Angiotensin-II (Ang-II), renin and norepinephrine (NE). Pathological testing included HE staining, Masson staining of myocardial tissue, tyrosine hydroxylase (TH) immunohistochemistry and C-Fos immunofluorescence staining of hypothalamus tissue.
RESULTS We found that RDN operation after intragastric administration of DAPA could significantly reduce systolic blood pressure in SHRs, better than RDN alone (mean systolic blood pressure: 133.5±9.093 mmHg versus 162.8±5.355 mmHg), which was independent of its hypoglycemic effect (Pearson correlation coefficient<0.2). At the same time, in pathological results of SIR group, it was found that compared with R group, there was a more significant improvement in the intensity of TH staining in hypothalamus tissue, closer to the normal level of cross-sectional area in myocardial cells (SIR versus W group: 262.2±21.58 versus 182.7±21.67 μm2), a stronger sympathetic inhibition in the brain (C-Fos mean fluorescence intensity SIR versus R group: 0.4880±0.03613 versus 0.3153±0.06217) and a lower fibrotic area in myocardial cells (SIR versus R group: 6.186±0.7125% versus 12.67±1.394%), and the proportion of inflammatory mononuclear cell subsets and the level of inflammatory factors were improved as well. While DAPA reduced inflammation and sympathetic nerve overexcitation alone, it could not completely reverse blood pressure and cardiac function. Similarly, the amelioration of inflammatory status and blood pressure reduction of rats treated with RDN alone was not better than that of rats treated with RDN combined with DAPA.
CONCLUSIONS The addition of DAPA before RDN operation can reduce the blood pressure of SHRs, and the effect was far better than RDN alone, and the level of related inflammatory factors also decreased correspondingly. At the same time, it was considered that the enhancement of hypotensive effect of DAPA in RDN was caused by inhibiting sympathetic activity and reducing inflammatory reactions. The possible common pathway of RDN and DAPA involves multiple inflammatory and blood pressure related signaling pathways, we still need further experimental verification, our study may have important clinical implications for RDN treatment in the future.
GW34-e0875
Yang Li, Yu Xue, Haixu Song, Chenghui Yan, Yaling Han
Department of Cardiology, General Hospital of Northern Theater Command
OBJECTIVES Guanxin Shutong (GXST) capsule is a Chinese medicinal formula that has been used clinically for the treatment of coronary heart disease in China. However, the molecular mechanisms of the cardioprotective effect are still unclear.
METHODS This study explores the cardioprotective effect and mechanism of GXST using the hypoxia reoxygenation (H/R)-induced myocardial injury model. In the in vivo study, an animal model of myocardial ischemia-reperfusion (MI/R) was induced by coronary occlusion. Fourteen male C57BL/6 mice were randomized into two groups: MI/R group (n=7), MI/R group treated with GXST (n=7). MI/R was induced by ligation of the left anterior descending coronary artery for 30 minutes, followed by ligature removed to restore coronary blood flow. GXST capsule (1.5 g/(kg·d)) or saline were administered via direct gastric gavage for 56 day after surgery. Mice were sacrificed and the hearts were harvested for histopathology and western blot analysis. In the in vitro experiments, H9c2 cells were incubated with GXST (200 μg/mL) capsule to explore the direct effects of CXST following exposure to CoCl2. Apoptosis and autophagy were measured to assess the protective effect of GXST. Proteins related to autophagy, apoptosis and AMPK/mTOR signalling pathways were detected using Western blotting.
RESULTS In vivo, GXST capsule significantly decreased levels of autophogy, apoptosis index, infarct size and myocardial fibrosis. In vitro, apoptosis and autophagy induced by H/R were significantly reduced after treating the H9C2 cells with GXST capsule. Moreover, GXST capsule also suppressed the AMPK/mTOR signalling pathway. Furthermore, the AMPK inhibitor Compound C (at 0.5 mmol/L), and GXST significantly inhibited CoCl2-induced autophagy and apoptosis, while AICAR (an AMPK activator, at 0.5 mmol/L) increased cardiomyocyte apoptosis and autophagy and abolished the anti-apoptotic effect of GXST capsule. Similar phenomena were also observed on the expressions of apoptotic and autophagic proteins, demonstrating that GXST reduced the apoptosis and autophagy in the CoCl2-induced injury model via inhibiting the AMPK/mTOR pathway.
CONCLUSIONS GXST capsule plays a cardioprotective role by alleviating apoptotic and autophagic cell death through inhibition of the AMPK/mTOR signaling pathway.
GW34-e0903
Wenjiao Gu
Second Hospital of Lanzhou University
OBJECTIVES The purpose of the present study was to study the effects of adenosine disodium triphosphate (ADTP) and alendronate sodium (AL), two extragenous PPi sources, on Atherosclerotic calcification (AC) in ApoE KO mice and to explore the potential its mechanism by examination of PPi content, lipid profiles and inflammation status.
METHODS We established AC models on ApoE KO mice, ADTP and AL were intraperitoneally injected for 2 months respectively. After the experiment, the cross-sectional samples of the aortic root were stained with von kossa staining kit to investigate the effects of ADTP and AL on AC in ApoE KO mice. Based on the work of previous section, serum lipid profiles, contents of pyrophosphate and inflammatory factors were examined in the blank, model and experimental groups of mice to explore the possible mechanisms of ADTP and AL on the inhibition of AC.
RESULTS We found that ApoE KO mice developed severe AC accompanied with hyperlipidemia, inflammation and with low level of PPi in serum. Compared with that in ApoE KO group, ADTP and AL dose-dependently reduced the aortic calcification lesions, and ADTP and AL had no effect on the contents of the serum TC, TG, LDL-C and HDL suggesting that the inhibitory effects of ADTP and AL on AC may not be related to the contents of lipid profiles. In addition, mice in ApoE KO group had low PPi content, meanwhile ADTP and AL increased the PPi content, the results suggested that deficiency of PPi in serum of ApoE KO mice might contribute to the AC and that complementation of PPi with ADTP and AL might be the important mechanism underlying their inhibition of AC. Finally, we found that ADTP and AL decreased the content of TNF-α and IL-6 in the serum of ApoE KO mice in dose-dependently manner suggesting that inhibition of inflammation by ADTP and AL might be another critical mechanism underlying their attenuation of AC.
CONCLUSIONS ADTP and AL, two exogenous PPi sources, reduced AC in ApoE KO mice by increasing the PPi level and inhibiting the inflammation.
GW34-e0915
Yi Yang1,2, Xinquan Wang1,2, Dan Wang1,2, Tao Luo1,2, Peijian Wang1,2
1Department of Cardiology, The First Affiliated Hospital, Chengdu Medical College
2Key Laboratory of Aging and Vascular Homeostasis of Sichuan Higher Education Institutes, The First Affiliated Hospital, Chengdu Medical College
OBJECTIVES Diabetic cardiomyopathy is a major cause of mortality in patients with diabetes mellitus, but specific strategies for the prevention and therapy of cardiomyopathy in diabetic population still remains elusive. G protein-coupled receptor 35 (GPR35), a novel member of GPRs, plays important roles in cardiac physiology and pathology. To date, however, the role of GPR35 in diabetic hearts has not been identified yet.
METHODS A leptin receptor-deficient (db/db) C57BLKS mouse, a mouse model of type 2 diabetes, was used in our study. Neonatal murine ventricular myocytes (NMVMs) were isolated from wild-type (WT) and GPR35 knockout (GPR35−/−) mice and cultured separately in normal-glucose medium, high-glucose medium (HG), high fatty acid (HF), and HG plus HF medium (HG-HF).
RESULTS Here, we found that GPR35 was increased in diabetic NMVMs and hearts in a time-dependent manner, respectively. The impaired cardiac function, aggravated cardiac hypertrophy, and deteriorated myocardial fibrosis in diabetic hearts were significantly attenuated by GPR35−/−. Moreover, in vitro results showed that GPR35 silencing inhibited the apoptosis of cardiomyocytes and the generation of ROS induced by HG-HF, through increasing mitochondrial morphology and function and subsequently activating the antioxidant system. Finally, our results indicated that HG-HF induced the upregulation of GPR35 by inhibiting optic atrophy 1 (OPA1) expression in diabetic cardiomyocytes.
CONCLUSIONS Collectively, our findings provide the evidence that GPR35−/− improves cardiac function and fibrosis in diabetic mice, suggesting that ablation of GPR35 may be pharmacological approach to alleviate diabetic cardiomyopathy.
GW34-e0941
Zijian Huang1, Chenhuiyu Qian1, Yuhan Cao2, Cong Fu1
1Department of Cardiology, Yi Ji Shan Hospital Affiliated to Wan Nan Medical College
2Department of Nephrology, Yi Ji Shan Hospital Affiliated to Wan Nan Medical College
OBJECTIVES Recent studies have identified MDSCs as an important player after acute myocardial infarction. However, the mechanism remains unclear. Our study indicated that ticagrelor can exert anti-inflammatory effects after acute myocardial infarction by altering the phenotype of MDSCs.
METHODS Phenotypic changes of MDSCs were analyzed by Flow Cytometry with or without ticagrelor treatment in vitro. The expression of P2y12 on MDSCs was detected by qRT-PCR and Western Blot. A mouse AMI model was established by left anterior descending coronary artery (LADCA) ligation. Phenotype of MDSCs were analysed by Flow Cytometry with or without ticagrelor gavage in vivo. The expression of TNF-α and IL-10 were detected by qRT-PCR and Western Blot. Myocardial injury degree was detected by HE staining. Cardiac systolic function was evaluated by transthoracic echocardiography.
RESULTS No statistical difference in mRNA and protein expression of P2y12 receptor in MDSCs in vitro compared to macrophages. In AMI model mice, the percentage of MDSCs was increased in the circulation and infarcted heart and decreased in the spleen. The ratio of PMN-MDSC in the circulation and infarcted heart was significantly increased while M-MDSC was not changed compared with the control and sham groups. Meanwhile, PMN-MDSC but not M-MDSC decreased significantly in the spleen of AMI mice. Ticagrelor significantly reduced the proportion of PMN-MDSC in the infarcted heart and circulation. The proportion of M-MDSC increased in a dose-dependent manner. The proportion of PMN-MDSC and M-MDSC in the spleen increased. In addition, ticagrelor treatment increased IL-10 expression and decreased TNF-α expression in infarcted hearts. Ticagrelor treatment also improved cardiac function after MI. Splenectomy in AMI mice reduced the proportion of PMN-MDSC and increased the proportion of M-MDSC in circulation and infarcted hearts, reduced cardiac inflammation level and improved cardiac function. AMI mice circulating-derived MDSC injection increased the PMN-MDSC ratio and decreased the M-MDSC ratio in infarcted hearts and circulation in Splenectomy AMI mice and increased TNF-α expression and cardiac injury and decreased IL-10 expression in infarcted hearts. In contrast, ticagrelor-treated AMI mice circulating-derived MDSCs injection significantly increased the M-MDSC ratio and decreased the PMN-MDSC ratio in the infarcted heart and circulation in Splenectomy AMI mice compared with non-ticagrelor-treatment-MDSC injection. The cardiac inflammation level was also decreased and cardiac function improved.
CONCLUSIONS Ticagrelor treatment promotes the spleen-derived MDSC differentiate to M-MDSC after AMI and the mobilization of M-MDSC to heart, which reduces cardiac inflammation and improves cardiac function.
GW34-e0948
Yunpeng Zhang1, Yang Zhao2, Tong Liu1
1Tianjin Key Laboratory of Ionic-Molecular Function of Cardiovascular Disease, Department of Cardiology, Tianjin Institute of Cardiology, The Second Hospital of Tianjin Medical University, Tianjin
2Department of Radiology, Tianjin Institute of Urology, The Second Hospital of Tianjin Medical University, Tianjin
OBJECTIVES Doxorubicin (DOX) is the first-line standard treatment of numerous cancers. However, DOX may lead to irreversible degenerative cardiomyopathy, congestive heart failure, and various arrhythmias, known as doxorubicin-induced cardiotoxicity (DIC), which limits the clinical applications. Studies reported that ferroptosis plays an important role on the development of DIC. Targeted intervention of ferroptosis with the specific inhibitors, such as ferrostatin-1 (Fer-1), could effectively prevent DIC. In recent years, catalytic nanozymes, such as cerium oxide (CeO2), have received much attention due to their strong antioxidant properties. Our previous research found that the transferrin receptor (TFR1) protein expression was significantly upregulated in DIC mice. In this study, we synthesized a biomimetic CeO2-based nanozyme by biomineralization with transferrin (Tf) proteins as the template, and then we assessed its targeted therapy for DIC via Tf-TFR1 mediated endocytosis.
METHODS The physicochemical and biological properties of NPs were characterized. Cellular uptake, cytotoxicity test, intracellular mitochondrial membrane potential (MMP), reactive oxygen species (ROS), mitochondrial ferrous ions and lipid peroxidation toxicity were verified with rat cardiomyocytes line H9c2 cells. Afterward, the male C57BL/6N mice were randomly divided into control, DOX, DOX+Fer-1, and DOX+NPs groups. Heart structure and function were measured by echocardiographic, electrocardiography and epicardial electrical labelling. Hematoxylin and eosin to observe the morphological changes. Western blot determines mitochondrial homeostasis regulation-related proteins and ferroptosis signalling pathway. Finally, the biodistribution and biocompatibility of CeO2@Tf was assessed.
RESULTS Transmission electron microscopy images revealed the encapsulated CeO2@Tf was ≈4 nm in diameter and remain stable in deionized water. The X-ray photoelectron spectroscopy revealed the ratio of Ce (III) to Ce (IV) ions was roughly 1:2.5. Meanwhile CeO2@Tf NPs showed good catalase (CAT) mimetics and superoxide dismutase (SOD) mimetics enzyme activity. The viability of H9c2 cells were not influenced by NPs even at high doses of Ce up to 200 μM. The nanozyme significantly reversed cardiac structural and electrical remodelling and reduced myocardial necrosis. Meanwhile, RNA sequencing of ventricular tissue revealed increased cardiac Hmox1 mRNA levels in DIC mice. Further, it demonstrated that nanozyme intervention not only significantly suppressed the oxidative stress, mitochondrial lipid peroxidation and mitochondrial membrane potential damage, but also inhibited activation of Nrf2/Hmox1 signalling pathway to restore mitochondria-dependent ferroptosis. Furthermore, the nanozyme could be cleared by the hepatobiliary systems, and showed a good biocompatibility.
CONCLUSIONS The current study showed that biomimetic mineralization of hybrid CeO2-based nanozyme against DIC were mediated by effect of antioxidant stress and the inhibition of Nrf2/Hmox1 signalling pathway, thereby maintaining mitochondrial homeostasis and function, and re-storing mitochondria-dependent ferroptosis. This provides some insights into the role of ferroptosis in DIC. Furthermore, the nanozyme could be a promising prevention and treatment candidate for clinical translation as a novel cardiomyocyte ferroptosis protector to mitigate DIC and improve the prognosis and quality of life of cancer patients.
GW34-e0982
Dong Guo, Mingming Zhang, Lang Hu, Yan Li
Department of Cardiology, Tangdu Hospital, Airforce Medical University
OBJECTIVES Lipid droplet (LD) accumulation is a notable feature of obesity-induced cardiomyopathy. Multiple studies have revealed the presence of excess LDs in the hearts of obese patients, and the extent of LD accumulation is directly related to cardiac function impairment. However, molecular details of LD biogenesis and pathological processes governing cardiac LD content remains poorly understood. Reticulon 3 (RTN3), an endoplasmic reticulum (ER)-localized protein, was recently identified as a key modulator of lipid metabolism in adipocytes. Therefore, we investigated the role of RTN3 in cardiac lipid metabolism and its regulatory mechanism in the development of obesity-induced cardiomyopathy.
METHODS Mice were fed high-fat diet (HFD; 60 kcal% fat) to induce obesity. Cardiac-specific RTN3 knockout/overexpression mice were developed using Cre/loxp system or adeno-associated virus 9 intramyocardial injection. Multiple analyses, including serial echocardiography, lipidomics, transmission electron microscopy, and oil red O staining were performed in vivo. Primary neonatal rat ventricular cardiomyocytes (NRVCs) were isolated and treated with palmitate in vitro. Adenovirus was used to upregulate or downregulate RTN3 in NRVCs. Long-term live-cell monitoring, fluorescent imaging, co-immunoprecipitation (Co-IP), chromatin immunoprecipitation, and liquid chromatography-tandem mass spectrometry (LC-MS/MS) were employed to elucidate the mechanism by which RTN3 affected cardiac lipid metabolism.
RESULTS HFD-fed mice exhibited significantly elevated cardiac LD amount and RTN3 expression. Cardiac-specific RTN3 overexpression induced obvious cardiac LD accumulation and functional impairment in normal diet-fed mice, phenotyping the detrimental effects of HFD. In contrast, RTN3 ablation ameliorated HFD-induced cardiac dysfunction and intramyocardial lipid accumulation, suggesting the important role of RTN3 in cardiac lipid metabolism. Then we investigated the linkage between RTN3 and myocardial lipid accumulation in vitro in NRVCs. Live cell imaging revealed that RTN3 overexpression significantly accelerated LD biogenesis in cardiomyocytes. Co-IP coupled with LC-MS/MS analysis identified fatty acid binding protein 5 (FABP5), a famous intracellular carrier for long-chain fatty acid (FA), as an interaction partner of RTN3. Silencing FABP5 blunted LD boosting induced by RTN3 overexpression. Moreover, the channeling of FA into LDs induced by RTN3 overexpression were also attenuated by FABP5 knockdown. Further investigations discovered that the 1st–65th amino acids of RTN3 were indispensable for the interaction between RTN3 and FABP5 and could induce a parallel effect on LD content and FAs channeling compared with full length RTN3. Finally, the mechanism underlying the upregulation of RTN3 was explored. We found that lipid overload-induced RTN3 upregulation was due to the increased expression of CCAAT/enhancer binding protein α (C/EBPα), which positively regulates RTN3 transcription by binding to its promoter region.
CONCLUSIONS Our study reveals a novel mechanism contributing to LD biogenesis in cardiomyocytes. ER-localized RTN3 directly interacts with FABP5 and facilitates FABP5-mediated FA directional transport to the ER, thereby promoting LD biogenesis. Under lipid overload conditions, C/EBPα was upregulated and then positively regulates RTN3 transcription, which ultimately led to the overactivated LD biogenesis and cardiac lipid accumulation. These findings advance our current understanding of the pathophysiology of obesity-induced myocardial lipid accumulation and cardiac dysfunction and lay the groundwork for novel strategies to treat cardiac dysfunction in obese patients.
GW34-e0984
Ya Qi Huang, Kuang Peng, Jie Wu
The First Affiliated Hospital of University of South China
OBJECTIVES This study aimed to observe whether ferroptosis is involved in cardiac and renal fibrosis in Dahl salt-sensitive hypertensive rats under long-term high-salt load.
METHODS Ten 7-week-old male Dahl salt-sensitive rats were adaptively fed for 1 week and then randomly divided into two groups: the normal-salt group fed with regular chow containing 0.3% NaCl and the high-salt group fed with high-salt chow containing 8% NaCl. After 8 weeks of feeding, non-invasive blood pressure measurements were performed using a rat tail artery blood pressure measurement device, and data were observed and recorded weekly to assess the success of the model. After the modeling period, histological changes in the heart and kidney tissues were observed using HE staining, and collagen fiber changes in the heart and kidney tissues were evaluated using Masson staining. The morphological features of mitochondrial Ferroptosis in cardiomyocytes and podocytes were observed under electron microscopy. Prussian blue staining was used to observe iron deposition in the tissues. Immunofluorescence analysis was conducted to observe the expression of GPX4 and xCT in the tissues. Iron content and MDA levels in the tissues were measured using colorimetric assays, and the expression levels of GPX4 and xCT in the tissues were detected using Western blot analysis.
RESULTS The systolic and diastolic blood pressures of the high-salt group rats were significantly higher than those of the normal-salt group (P<0.01). Histological staining with HE and Masson’s trichrome revealed that the myocardial tissues of the high-salt group rats exhibited cellular hypertrophy, disorganized arrangement, inflammatory cell infiltration in the interstitium, and increased collagen fiber proliferation around the interstitium and blood vessels (P<0.05). In the kidney tissues, partial glomeruli showed increased volume, sclerosis, disorganized arrangement, and atrophy of some renal tubules. Inflammatory cell infiltration and abundant collagen fiber proliferation were observed in the glomeruli, interstitium, and around blood vessels of the high-salt group rats (P<0.05), while the heart and kidney tissue structures of the normal-salt group rats appeared normal. Electron microscopy examination revealed that the heart and kidney tissues of the high-salt group rats exhibited mitochondrial shrinkage, decreased or absent cristae, and increased density of the mitochondrial double membrane, whereas no such changes were observed in the normal-salt group rats. Prussian blue staining of the heart and kidney tissues of the high-salt group rats showed deposition of blue particles, indicating iron deposition in the tissues, whereas little blue staining was observed in the heart and kidney tissues of the normal-salt group rats. The high-salt group exhibited elevated tissue iron content and MDA levels compared to the normal-salt group (P<0.05). Immunofluorescence analysis revealed reduced expression of xCT and GPX4 in the myocardium and kidneys of the high-salt group compared to the normal-salt group (P<0.05), and Western blot analysis similarly showed decreased expression of xCT and GPX4 in the myocardium and kidneys of the high-salt group compared to the normal-salt group (P<0.05).
CONCLUSIONS Ferroptosis, characterized by mitochondrial changes, is observed in the damaged heart and kidney tissues of Dahl salt-sensitive hypertensive rats. The downregulation of xCT/GPX4, proteins associated with Ferroptosis, suggests the involvement of Ferroptosis in the organ damage of salt-sensitive hypertension.
GW34-e1043
Feng Wang
Department of Cardiology, First Affiliated Hospital of Bengbu Medical College
OBJECTIVES The clinical application of doxorubicin (Dox) in tumor chemotherapy is limited by time-dependent and dose-dependent cardiotoxicity. Hence, there is an urgent need to elucidate doxorubicin cardiotoxicity and to solve the difficult problem in clinical application. It has been verified that serum and glucocorticoid-regulated kinase 1 (SGK1) possess cardioprotective effects.
METHODS H9c2 cells were treated with 1 μM doxorubicin for 24 h to establish doxorubicin cardiotoxicity, so as to determine the biological role of SGK1 in doxorubicin cardiomyopathy and to elucidate the underlying molecular mechanism. SGK1 level in doxorubicin-treated H9c2 cells was assessed by performing Western blot assay and RT-qPCR. CCK-8 assay and TUNEL staining were employed to evaluate the cell viability and cell apoptosis. Besides, apoptosis-related proteins were measured by Western blot assay to analyze cell apoptosis. Additionally, the release of TNF-α, IL-1β, IL-6, and IL-10 and the levels of ROS, MDA, and SOD were detected to reflect inflammation and oxidative stress. Moreover, Western blot assay was adopted for determination of ERS-associated proteins.
RESULTS Upregulation of SGK1 alleviated doxorubicin-induced cardiotoxic injury, cell apoptosis, inflammation and oxidative stress in H9c2 cells. Moreover, SGK1 overexpression mitigated doxorubicin-induced ERS in H9c2 cells. The suppressing effects of SGK1 on doxorubicin-induced cardiotoxic injury, apoptosis, inflammation, oxidative stress and ERS in H9c2 cells were partially abolished upon GRP78 overexpression.
CONCLUSIONS To conclude, upregulation of SGK1 may alleviate doxorubicin cardiotoxicity by repressing GRP78-mediated ERS.
GW34-e1058
Nikita Nikiforov1,2,3, Tatiana Kirichenko1,2, Marina Kubekina3, Yegor Chegodaev1,2, Alexander Zhuravlev1,2, Leonid Ilchuk3, Alexander Orekhov1,2,4
1Laboratory of Angiopathology, The Institute of General Pathology and Pathophysiology, 8 Baltiyskaya Street, Moscow 125315, Russia
2P. Avtsyn Research Institute of Human Morphology, Federal State Budgetary Scientific Institution “Petrovsky National Research Centre of Surgery”, 3 Tsyurupa Street, Moscow 117418, Russia
3Center for Precision Genome Editing and Genetic Technologies for Biomedicine, Institute of Gene Biology, Russian Academy of Sciences, 34/5 Vavilova str., Moscow 119334, Russia
4Institute for Atherosclerosis Research, Osennyaya Street 4-1-207, Moscow 121609, Russia
OBJECTIVES The pro-inflammatory reaction of human monocytes and macrophages in response to a pathogen is characterized by a subsequent attenuation and the tolerance formation. We decided to study the pro-inflammatory response of monocytes from patients with preclinical atherosclerosis and the subsequent decrease in the pro-inflammatory activity of cells during differentiation into macrophages.
METHODS The study included 46 healthy patients with normal intima-media thickness (IMT) of the carotid arteries and 26 patients with atherosclerotic plaque and thickened IMT. CD14+ monocytes were isolated from blood, stimulated with 1 μg/mL LPS for 1 day and cultured for 7 more days without LPS. Secretion of IL-1β, IFN-α2, IFN-γ, TNF-α, CCL2, IL-6, IL-8, IL-10, IL-12p70, IL-17A, IL-18, IL-23 and IL-33 were measured in supernatants at each stage by ELISA.
RESULTS We found no differences in the generation of cytokines by monocytes in response to LPS. However, further in the course of cell differentiation into macrophages, we found that the cells of patients with atherosclerosis are characterized by increased secretion of the cytokines CCL2 and IL-6 on days 1–6 in culture. Moreover, after changing the medium, 7-day-old macrophages from atherosclerotic patients retained increased secretory activity compared to macrophages from healthy donors. This was manifested in increased basal secretion of the cytokines CCL2, IL-6, and IL-8. The degree of secretion of these cytokines significantly (P<0.01) correlated directly with IMT.
CONCLUSIONS Macrophages derived from LPS-stimulated monocytes from patients with asymptomatic atherosclerosis are characterized by prolonged and increased pro-inflammatory activity. The causes of chronification of inflammation in the vascular wall may lie at the level of circulating monocytes.
Supported by RSF (Grant No. 22-15-00273).
GW34-e1073
Zheng Yin1,2,3, Jun Wan1,2,3
1Department of Cardiology, Renmin Hospital of Wuhan University, Wuhan, China
2Cardiovascular Research Institute, Wuhan University, Wuhan, China
3Hubei Key Laboratory of Cardiology, Wuhan, China
OBJECTIVES Hypertension is one of the most notorious chronic diseases, which leads to approximately 30% of all deaths worldwide. Hypertensive vascular remodeling is defined as the changes in vascular function and structure induced by persistent hypertension. Maresin-1 (MaR1), one of metabolites from Omega-3 fatty acids, has been reported to promote inflammation resolution in several inflammatory diseases. This study aims to investigate the effects of MaR1 on hypertensive vascular remodeling.
METHODS We constructed an Angiotensin II (Ang II)-infused mouse model of hypertension, and evaluated the effects of MaR1 on blood pressure and hypertensive vascular remodeling. In addition, rat-derived vascular smooth cells (VSMCs) were used to construct vascular inflammation model in vitro. The biobehavior of VSMCs in response to Ang II was evaluated through CCK-8, EdU and wound healing assay.
RESULTS In vivo, MaR1 administration significantly decreased the systolic and diastolic blood pressure, decreased the medial thickness of aorta, attenuated aorta fibrosis, suppressed phenotype transition of VSMCs. In vitro, MaR1 treatment significantly modulated VSMC phenotype, and inhibited VSMC proliferation and migration. Moreover, MaR1 could attenuate NLRP3 inflammasome activation and GSDMD-related pyroptosis both in vivo and in vitro. Mechanically, the blocking of Nrf2/HO-1 pathway could reverse the protective effects of MaR1 on VSMC phenotype switching, proliferation, migration through the suppression of pyroptotic cell death.
CONCLUSIONS MaR1 may attenuate hypertensive vascular remodeling by inhibiting VSMC pyroptosis through the Nrf2/HO-1 signaling pathway. Overall, MaR1 may be a novel therapeutic strategy for the treatment of hypertension in the future.
GW34-e1075
Ya Liu, Xiaokang Chen, Dongdong Sun, Chao Zhang, Wujian Liu, Wei Eric Wang
Department of Geriatrics, Southwest Hospital, Third Military Medical University (Army Medical University), Chongqing, China
OBJECTIVES The metabolic phenotype of proliferative cardiomyocytes differs significantly from non-proliferative cardiomyocytes. Several studies indicated that the metabolic features include the augment of intermediate metabolite NAD+ in proliferative cardiomyocytes. However, whether and how NAD+ regulates adult cardiomyocyte proliferation is unknown.
METHODS The pro-proliferative effect of NAD+ was detected with Ki67 and PH3 immunostaining in both cultured cardiomyocytes and heart tissues. NAD+ was manipulated by supplementation of NAD+ precursor NMN or inhibition of NAD+ synthesis by FK866. The therapeutic effect of NAD+ against myocardial infarction (MI) was examined by echocardiograph.
RESULTS The NAD+ level in cardiomyocytes reduced during the first postnatal week, coinciding with the timeframe when cardiomyocytes transit from a proliferative state to a quiescent state. Supplementation of NAD+ promoted the proliferation of cultured postnatal 1 (P1) and P7 cardiomyocytes in-vitro, while inhibition of NAD+ synthesis inhibited the proliferation. Supplementation of NAD+ prolonged the proliferative window period of cardiomyocytes in neonatal mouse. It also promoted cardiomyocyte proliferation in adult post-MI heart, and enhanced cardiac function and improved long-term survival of the post-MI mice. Mechanistically, NAD+ regulated cardiomyocyte proliferation through a SIRT1-dependent manner. Inhibition of SIRT1 blocked the pro-proliferative effect of NAD+. Among the histone proteins related to SIRT1, the expression of H3K9ac profoundly decreased under the supplementation of NAD+, which could be partially rescued by SIRT1 inhibitor. Decreased H3K9ac lead to a downregulation of the cell cycle inhibitor p27.
CONCLUSIONS The ratio of cytoplasmic NAD+/NADH influences cardiomyocyte proliferation both in-vitro and in-vivo, through regulating SIRT1 deacetylase which suppresses cell cycle inhibitory gene p27 by modifying H3K9ac. Supplementation of NAD+ might be a potential therapeutic strategy for treating MI.
GW34-e1081
Qian Chen1, Xixiang Tang2, Suhua Li1
1Department of Cardiovascular Medicine, The Third Affiliated Hospital, Sun Yat-sen University, Guangzhou 510630, China
2VIP Medical Service Center, The Third Affiliated Hospital, Sun Yat-sen University, Guangzhou 510630, China
OBJECTIVES Lipotoxicity is a well-established contributor to cardiomyocyte death and heart damage, with ferroptosis being identified as a crucial death mode in cardiomyocyte disease. However, the role and underlying mechanisms of lipotoxicity in cardiomyocyte ferroptosis remain poorly understood. Therefore, this study aims to explore the potential role and mechanism of lipotoxicity in cardiomyocyte ferroptosis.
METHODS To establish an in vitro lipotoxic cell model, Rat H9c2 cardiomyocytes were treated with palmitic acid (PA). Ferrostatin-1 (Fer-1) and Liproxstatin-1 (Lip-1) were added to the cell model to inhibit ferroptosis. To explore the role of STING in H9c2 cardiomyocytes, siRNA transfection was employed to knockdown STING expression. In addition, an in vivo lipotoxic SD rat model was established using a high-fat diet (HFD), and ferroptosis was intervened by intraperitoneal injection of ferroptosis inhibitor Fer-1. STING expression in rats was knocked down by tail vein injection of myocardial-specific interference adeno-associated virus (AAV), or systemically inhibited by intraperitoneal injection of STING inhibitor H151. Doppler ultrasound technology was used to monitor changes in rat cardiac structure and function. HE and MASSON staining were employed to observe myocardial pathology and fibrosis changes in rats. Cell viability and death were measured using the CCK-8 assay kit and the Calcein-AM/PI assay kit, respectively. BODIPY™ 581/591 C11 was used to observe the level of lipid peroxidation in cells. The end-products of lipid peroxidation, malondialdehyde (MDA), and glutathione (GSH) levels in H9c2 cells/myocardium were detected by the thiobarbituric acid reactive substances (TBARS) kit and GSH assay, respectively. The levels of ferroptosis related proteins (ACSL4, SCL7A11, TFRC and GPX4) and STING pathway related proteins (STING, TBK/p-TBK, p65/p-65 and IRF3/p-IRF3) in cell and heat tissue were detected by Western blot analysis.
RESULTS In vivo experiments revealed that ferroptosis inhibitors significantly reduced the level of death and lipid peroxidation of PA-induced lipotoxic cardiomyocytes. Additionally, they increased the level of GSH and GPX4 in cardiomyocytes. The time-dependent activation of the STING pathway following PA stimulation was also observed. Knockdown the expression of STING could reduce PA-induced cell death and lipid peroxidation levels while restoring the expression of GPX4. Furthermore, the hearts of HFD rats showed worse structure and function compared to normal control rats. Specifically, HFD rats exhibited more disordered myocardial structure and arrangement, significantly increased fibrosis level, and reduced GSH levels. In addition, the protein levels of SCL7A11 and GPX4 were decreased, while STING protein level was increased in the hearts of HFD rats. In HFD rats with myocardial specific knockdown of STING, GPX4-GSH antioxidant capacity was restored, and the level of lipid peroxidation was reduced, improving the structure and function of the heart, disorder of myocardial structure arrangement and significantly reducing the level of myocardial fibrosis.
CONCLUSIONS The findings suggests that lipotoxicity can induce ferroptosis in cardiomyocytes through the activation of the STING pathway, providing new targets, and strategies for the treatment of lipotoxicity cardiomyopathy.
GW34-e1091
Ziyu Guo1, Ao Wang2, Yanxiang Gao3, Jingang Zheng3
1Department of Cardiology, Peking University China-Japan Friendship School of Clinical Medicine, Beijing, China
2Department of Physiology and Pathophysiology, School of Basic Medical Sciences, State Key Laboratory of Vascular Homeostasis and Remodeling, Peking University
3Department of Cardiology, China-Japan Friendship Hospital
OBJECTIVES After cardiac ischaemia, an insufficient myocardial perfusion often occurs despite flow is successfully and completely restored in an upstream artery. This “no-reflow phenomenon” is caused by coronary microvascular dysfunction and is linked to a poor clinical outcome. This study aimed to assess the dynamic change of the CFR before and after ischemia reperfusion by pulsed-wave Doppler measurements.
METHODS Use a modified parasternal long-axis (PLAX) view to examine the left anterior descending coronary artery. CFR is defined as the ratio of the maximal flow velocity induced by a metabolic or pharmacologic stimulus to the resting left coronary artery (LCA) flow velocity. Measure the coronary flow velocity and CFR again in 1 hour, 3 hours, 5 hours, 8 hours and 24 hours, 48 hours after reperfusion, respectively. Compare the measurements to before IR measurements.
RESULTS Before the IR surgery, the baseline CFR and the mice had a normal CFR value near 2.14±0.43. After the IR surgery, CFR was significantly decreased in reperfusion 1 hour compared to the before IR surgery (1.18±0.14 vs 2.14±0.43), indicating that microcirculation was still not immediately restored even after opening the criminal vessels. As the reperfusion time was prolonged, the CFR values improved, but remained lower than before the procedure. What’s more, there was no significant difference between the CFR of reperfusion for 24 h and the CFR of reperfusion for 1 h. And there were no significant changes in cardiac function of the left ventricle when the CFR was significantly reduced in the mice.
CONCLUSIONS After ischemia reperfusion, the 1-hour CFR was significantly decreased than pre-operation. The CFR has gradually recovered over time, but remains below normal. The systolic function was preserved. Therefore, it is important to establish a practical guide to help doctors detect early microvascular dysfunction and study the progression of cardiovascular disease over time.
GW34-e1097
Xiaoming Xu
Department of Cardiology, Xijing Hospital, Fourth Military Medical University
OBJECTIVES Fibronectin type III domain containing 4 (FNDC4) displays a high homology with FNDC5 which has a wide range of cardiac metabolic protection. Recent findings indicated that FNDC4 was highly expressed in liver, and was involved in fatty metabolism and inflammation. In this study, we investigated the effects of FNDC4 on a mouse model of myocardial infarction and reperfusion (MI/R) injury.
METHODS MI/R models were established by ligation of the anterior descending branch followed by removal of the knot at 30 min, to investigate the change of FNDC4 expression. FNDC4-knockout (FNDC4−/−) and overexpression of myocardial FNDC4 were induced to clarify the effects of FNDC4 by determining myocardial infarct size, cardiomyocyte apoptosis and cardiac function. Cardiomyocytes apoptosis induced by hypoxia/reoxygenation treatment in vitro was further detected to access the effect of FNDC4. A nonbiased approach via the RNA sequencing (RNA-seq) analysis was performed to identify the underlying mechanism.
RESULTS Downregulation of myocardial FNDC4 expression was observed in MI/R mice in contrast to sham mice. FNDC4 deficiency further exacerbated cardiac function, augmented cardiac infarct size and increased cardiomyocyte apoptosis, compared with wide type (WT) mouse. While FNDC4 overexpression by intramyocardially injected with recombinant adeno-associated virus 9 encoding FNDC4 (rAAV9-Fndc4-3Flag) significantly improved cardiac function, reduced infarct size and cardiomyocyte apoptosis compared to mice injected with control virus. Results from in vitro studies demonstrated FNDC4 overexpression in NRVMs markedly decreased cardiomyocyte apoptosis induced by hypoxia/reoxygenation injury evidenced by expression of cleaved caspase 3, proportion of TUNEL-positive nuclei and annexin V+/PI+/- cells, as well as cell viability. By performing RNA-sequence analysis, a ERK1/2/Nrf2/HO-1&NQO1 axis was identified and was responsible for the beneficial effects of FNDC4 on cardiomyocytes.
CONCLUSIONS We discover and confirm for the first time that FNDC4 mediates cardioprotection in a MI/R injury model through ERK1/2/Nrf2/HO-1&NQO1 pathway. These findings show that FNDC4 is a potential therapeutic target for MI/R injury.
GW34-e1107
Fan Pan
West China Hospital of Sichuan University
OBJECTIVES Commercial biological Artificial heart valves (BHVs), which are made by cross-linking Glutaraldehyde with pig pericardium (PP), bovine pericardium (BP) and other animal tissues, have been widely used in interventional valve replacement. However, Glutaraldehyde cross-linked biological valve has many problems that limit its service life, the most important of which is thrombosis and calcification. We use sulfonated Betaine Zwitterion hydrogel to modify the Glutaraldehyde crosslinked biological valve, and evaluated the antithrombotic and anti-calcification properties of the valve leaf modified by this method.
METHODS
1. In vitro antithrombogenicity assay: To evaluate the antithrombogenic properties of the valve leaflets with the copolymer, an arteriovenous shunt model in rabbits was used to conduct an ex vivo perfusion experiment. After 2 hours of circulation, the samples were gently removed from the catheter and photographed. The samples with thrombus were also collected for quantitative analysis.
2. Subcutaneous implantation assay: The male Sprague Dawley rats (approximately 50 g) were anesthetized with pentobarbital sodium solution. Surgical incisions (10 mm) were made at the rat’s back, and then the samples were implanted subcutaneously. After 90 days, the samples were harvested and used to evaluate the degree of calcification.
RESULTS
1. The weight of the thrombus in the control group was 3.52±0.64 mg, while the weight of the thrombus in the experimental group was 0.95±0.12 mg. The experimental group was superior to the control group in terms of antithrombotic activity.
2. The calcium content in the control group is 202.75±3.16 μg/mg, while the calcium content in the experimental group is 65.78±2.12 μg/mg, indicating that the experimental group has better anti calcification capacity.
CONCLUSIONS Sulfonated betaine zwitterion hydrogel can improve the antithrombotic and anti-calcification ability of artificial biological valve.
GW34-e1112
Peinan Ju, Qing Yu, E Guangxi, Yawei Xu, Jianhui Zhuang, Wenhui Peng
Department of Cardiology, Shanghai Tenth People’s Hospital, School of Medicine, Tongji University, Shanghai, China
OBJECTIVES Sleep is vital to health. Sleep arousal is associated with the increased mortality of cardiovascular disease in the elderly. Our study is to determine the role of sleep in myocardial infarction (MI) and to dissect the underlying mechanisms.
METHODS Sleep fragmentation (SF) model were used to simulate long-term sleep disorders. Left anterior descending coronary artery ligation mouse model were used to cause myocardial infarction in mice. Echocardiography was used to evaluate cardiac function after MI. Several morphological staining experiments were conducted to observe infarct area and fibrosis in heart. Flow cytometry was used to evaluate the infiltration of leukocytes after MI in the heart, as well as emergency hematopoiesis in bone marrow. Dihydrorhodamine 123 was used to detect oxidative stress levels in cardiac neutrophils. Bone marrow transplantation (BMT), Ly6G neutralizing antibodies, and SiglecF neutralizing antibodies were used to explore whether SF exacerbate MI through hematopoiesis and neutrophils. Adoptive cell transfer and competitive BMT were conducted to clarify the effect of SF on proliferation and differentiation of Granulocyte-Macrophage Progenitor (GMP) into neutrophils. High-throughput sequencing was used to explore the impact of sleep disorders on GMP during acute MI, search for related signaling pathways and differential expressed genes. Myeloid-specific Cebpe deletion mice and GMP adoptive transfer were conducted to clarify the specific mechanism by which Cebpe in GMP regulates sleep disorders and exacerbates MI. Chromatin immunoprecipitation-Sequencing (Chip-Seq) of transcription factor Cebpe was conducted to find related genes regulated by GMP through Cebpe.
RESULTS Mice with SF had higher mortality after MI. The cardiac function was decreased, infarct area was increased in sleep fragmented mice. SF also significantly aggravated MI-induced pathological cardiac fibrosis and collagen I synthesis. In addition, SF resulted in a significant increase in the number of neutrophils in infarcted heart, mainly due to an increase in SiglecFhigh neutrophils. Neutrophils in the infarcted heart of SF mice produce more reactive oxygen species (ROS). Neutralization of neutrophils or SiglecFhigh subsets of neutrophil can significantly alleviate the MI progression in SF mice. Infiltrated neutrophils in the heart of SF mice are mainly derived from GMP in bone marrow. The amount of GMP increases in the bone marrow of SF mice, which is more significant after MI. Transcription factor Cebpe was increased in GMP after long term SF. GMP-specific Cebpe deletion alleviated the progression of MI in SF mice.
CONCLUSIONS Our findings suggest that long term sleep disorder promotes the increase of bone marrow GMP Cebpe expression in the acute phase of MI, accelerates the proliferation and differentiation of GMP into neutrophils, leads to more neutrophil released to the infarcted area and transformed into SiglecFhigh subtype, and finally aggravates MI through the synthesis of ROS and collagen I.
GW34-e1113
Bolin Li1,2, Yue Wu1
1Department of Cardiovascular Medicine, The First Affiliated Hospital, Xi’an Jiaotong University, Xi’an, Shaanxi, China
2Department of Cardiovascular Medicine, The Affiliated Hospital of Southwest Jiaotong University, Chengdu, Sichuan, China
OBJECTIVES Deep vein thrombosis (DVT) is a common vascular disease of which pulmonary embolism is the most serious complication. In obese humans, impaired fibrinolysis plays an important role during thrombus formation. Plasminogen activator inhibitor 1 (PAI1) functioning primarily as a negative regulator of fibrinolysis has increased significantly in obesity. And liver that sense obesity-induced metabolic stress was identified as a major determinant of circulating PAI1 in obese individuals. But the mechanism of increased PAI1 during obesity is not clear. Here we explored the upstream regulator of the PAI1 and how it functions in fibrinolysis and DVT.
METHODS Mice were fed with normal diet or high fat diet for 4 months. Then, each group of mice were randomly selected for inferior vena cava stenosis model for evaluating DVT burden, and the remaining mice for blood sampling for evaluating plasm fibrinolytic capacities by fibrin formation and lysis test. To explore the upstream regulator of PAI1, Bioinformatic analysis on transcriptomics and targeted bile acid metabolomics in human, baboon and mice liver samples were performed. Then, the expression of PAI1 was assessed in mice and MPH treated by FXR agonist as well as in MPH derived from FXR knock out (KO) mice. FXR KO mice were randomly selected for DVT modeling and blood sampling to measure plasm fibrinolytic capacities and DVT burden. Ultimately, Obese mice treated with Tropifexor were used to assess plasm fibrinolytic capacities and DVT burden.
RESULTS Obese mice had significantly reduced fibrinolysis and increased DVT burden compared to lean mice. And the levels of plasma PAI1 were also higher in obese participants and mice, due largely to their increased hepatocyte expression. Transcriptomics and targeted bile acid metabolomics analysis of human, baboon and mice liver samples suggest that increased PAI1 expression may be related to decreased FXR transcription activity in obesity; and similar results were obtained in MPH treated by palmitic acid. We focused on the FXR-PAI1 pathway, which may mitigate the reduction in fibrinolysis in obesity. FXR activation could inhibit mRNA and protein expression of PAI1 in both mice and MPH, and FXR KO could block the decline of PAI1 in MPH induced by FXR agonist. The fluorescent enzyme activity was significantly reduced after FXR activation in both AML12 and HEK293 cells transfected with plasmids containing PAI1-1 promoter area. FXR KO mice present significantly increased plasma PAI1 as well as further impaired fibrinolysis and increased DVT burden. Tropifexor activating FXR in obese mice lowered plasma PAI1 as well as further alleviated fibrinolysis and DVT burden.
CONCLUSIONS We identify FXR acting as a novel regulator of PAI1 and playing a new role in fibrinolysis and DVT. Clinical trial drugs targeting FXR in hepatocytes may suggest therapeutic strategies for improving fibrinolysis and lowering the risk of DVT in obesity.
GW34-e1120
Jianshu Chen
Lanzhou University Second Hospital
OBJECTIVES Endoplasmic reticulum stress (ERS) and mitochondrial autophagy are closely associated with myocardial damage in heart failure with preserved ejection fraction (HFpEF). The endoplasmic reticulum is involved in regulating mitochondrial function through the mitochondria-associated membrane (MAM). Here, we identified the role of leucine-rich repeat kinase 2 (LRRK2) in ER–mitochondrial tethering, which is essential for the treatment of myocardial damage caused by HFpEF.
METHODS With WKY as the control group, female SHR constructed HFpEF model, and rat tail blood pressure measurement, hemodynamics, echocardiography and exercise fatigue test to explore the successful establishment of HFpEF model. The protein expression of ATF6, LRRK2 and Parkin was changed by using different doses of LRRK2 inhibitor GNE-7915 in HFpEF rat model, and the changes of ERS and mitochondria were observed by electron microscopy.
RESULTS ERS transcription factor 6 (ATF6) had specific sequence pairing with the LRRK2 promoter region. Compared with WKY and SHR rats, HFpEF rats showed decreased lung dry weight/wet weight ratio and a large number of red blood cells clustered in the HE-stained alveolar cavity of the lung tissue. The ratio of type I/III collagen in myocardial tissue of HFpEF rats increased, and the ratio of type I/III collagen in myocardial tissue decreased after the intervention of GNE7915. The cross-sectional area of cardiomyocytes in HFpEF rats was larger than that in the other three groups. Compared with WKY and SHR rats, transmission electron microscopy showed that the mitochondrial arrangement of myocardial tissue in HFpEF rats was disordered, the shape was irregular, the original structure was lost, the mitochondrial ridge was blurred, some mitochondria vacuolated, and the endoplasmic reticulum was swollen. Western blot results showed that the expression of LRRK2 after the intervention of GNE-7915 was lower than that in the non-intervention group of HFpEF.
CONCLUSIONS MAM and the expression of LRRK2 were increased in HFpEF rats. ERS ATF6 may play a regulatory role in mitochondrial autophagy through the MAM complex LRRK2 pathway and participate in myocardial damage by HFpEF.
GW34-e1135
Zhuqing Li, Chengzhi Lu
Tianjin First Center Hospital
OBJECTIVES Heart Failure with Preserved Ejection Fraction (HFpEF) is a multifaceted disease, intricately linked with sympathetic nervous system activation. Despite advancements, therapeutic options for HFpEF remain constrained, largely due to a dearth of fitting preclinical models. Renal denervation (RDN), an emerging intervention, holds potential in attenuating sympathetic activity and ameliorating HFpEF high-risk factors such as hypertension and insulin resistance. Our study is designed to establish a multifactorial HFpEF mouse model and assess the therapeutic implications of RDN.
METHODS Aged 12-month-old male C57BL/6N mice were exposed to a high-fat diet (60% calories from lard) and L-NAME (0.5 g/L in drinking water) over a 5-week period to generate a multi-hit HFpEF model. The efficacy of the model was assessed through a range of methods, encompassing echocardiography, histology, exercise exhaustion tests, serum NT-proBNP measurements, and intraperitoneal glucose tolerance tests. Upon successful model establishment, mice were randomized to receive either bilateral renal denervation or a sham operation. The subsequent impact of RDN on the aforementioned parameters was then evaluated.
RESULTS In C57BL/6N mice, a cardiometabolic HFpEF phenotype was effectively induced through a multi-hit approach encompassing a high-fat diet, L-NAME administration, and aging. This potent combination triggered obesity and glucose intolerance, culminating in the emergence of characteristic HFpEF features such as cardiac hypertrophy, fibrosis, preserved fractional shortening with impaired deformation, atrial enlargement, lung congestion, hypertension and decreased exercise endurance. Intervention with RDN mitigated this cardiometabolic dysregulation and enhanced cardiac function, as demonstrated by reductions in cardiac hypertrophy, myocardial fibrosis, atrial weight, NT-proBNP levels (sham-RDN vs. RDN: 687±18 vs. 473±27, P<0.05), lung congestion, and improved exercise endurance (sham-RDN vs. RDN: 320±23% vs. 411±19%, P<0.05).
CONCLUSIONS Our study suggests that RDN effectively mitigates cardiac function and structure in a multi-hit HFpEF mouse model, indicating its potential as a promising therapeutic strategy for HFpEF.
GW34-e1136
Qingbo Lv1,2, Min Shang1, Wenbin Zhang1,2, Guosheng Fu1,2
1Key Laboratory of Cardiovascular Intervention and Regenerative Medicine of Zhejiang Province, Sir Run Run Shaw Hospital, Zhejiang University School of Medicine, Hangzhou, People’s Republic of China
2Department of Cardiology, Sir Run Run Shaw Hospital, School of Medicine, Zhejiang University, Hangzhou, People’s Republic of China
OBJECTIVES Metabolic modulation is a promising target in preventing adverse remodeling of pressure overload induced cardiac hypertrophy. Previous studies have well documented the metabolic remodeling during the cardiac hypertrophy, mainly focusing on the glycolysis, FA oxidation, and BBCA metabolism. Little is known about the role of proline in regulating cardiac metabolism. In this study, we mainly demonstrated the importance of proline metabolism in pressure-overloaded induced cardiac remodeling.
METHODS In this study, we initially established the TAC model and the PE-induced neonatal rat ventricular myocytes (NRVM) hypertrophy model as in vivo and in vitro experimental models, respectively. We examined proteins, mRNA, and other factors in cardiac tissues to ascertain the changes in relevant metabolic enzymes. Subsequently, the construction of small interfering knockdown system and over-expression viruses allowed us to regulate the expression of the target gene, PRODH, in in vitro cell experiments and to explore its function in the development of cardiomyocytes hypertrophy. Following this, we employed the CRISPR-CAS9 technology to construct cardiac conditional PRODH knockout mice (PRODH-cKO) and cardiac-specific overexpression viruses mediated by AAV9, thus achieving in vivo regulation of PRODH expression. Mechanistically, we utilized unbiased transcriptome profiling, metabolic flux analysis, and non-targeted LC-MS, among other techniques, to systemically analyze the transcriptomics and metabolomics of cardiac tissue or cardiomyocytes. This thorough investigation revealed changes in metabolic enzymes and substrates in cardiac remodeling.
RESULTS In this study, we initially established the TAC model and the PE-induced neonatal rat ventricular myocytes (NRVM) hypertrophy model as in vivo and in vitro experimental models, respectively. We examined proteins, mRNA, and other factors in cardiac tissues to ascertain the changes in relevant metabolic enzymes. Subsequently, the construction of small interfering knockdown system and over-expression viruses allowed us to regulate the expression of the target gene, PRODH, in in vitro cell experiments and to explore its function in the development of cardiomyocytes hypertrophy. Following this, we employed the CRISPR-CAS9 technology to construct cardiac conditional PRODH knockout mice (PRODH-cKO) and cardiac-specific overexpression viruses mediated by AAV9, thus achieving in vivo regulation of PRODH expression. Mechanistically, we utilized unbiased transcriptome profiling, metabolic flux analysis, and non-targeted LC-MS, among other techniques, to systemically analyze the transcriptomics and metabolomics of cardiac tissue or cardiomyocytes. This thorough investigation revealed changes in metabolic enzymes and substrates in cardiac remodeling.
CONCLUSIONS Our research rigorously investigated the role of PRODH expression in pathological cardiac remodeling through in vivo and in vitro experiments. The absence of PRODH led to a deterioration of cardiac function, whereas the overexpression of PRODH could improve the TCA cycle’s energy supply by promoting the utilization of proline by cardiomyocytes, and stimulate glutathione synthesis by synthesizing glutamate. In summary, our study is the first to systematically demonstrate the role of proline metabolic cycling in pathological cardiac remodeling. This not only reveals the metabolic remodeling changes in pathological cardiac remodeling but also provides new therapeutic targets for the treatment of cardiac remodeling and heart failure.
GW34-e1139
Bingjun Lu
Army Medical University Southwest Hospital
OBJECTIVES The aim of this study was to compare the molecular characteristics of myocardial cell populations with different proliferative abilities in order to reveal the molecular mechanisms underlying changes in myocardial cell proliferation, and to identify potential targets for promoting cardiomyocyte cytokinesis.
METHODS In this study, weighted gene co-expression network analysis was employed to classify genes into 9 expression modules based on their expression patterns during cardiac development. Gene functional enrichment analysis and Pearson correlation analysis were then combined to screen and identify genes associated with cell cycle regulation during heart development.
RESULTS Gene functional enrichment analysis revealed that genes in the blue and turquoise modules may be involved in cell cycle regulation during heart development. Further analysis using Pearson correlation identified 3807 genes significantly correlated with the expression patterns of Ki67 and Ccnb1 genes. By analyzing myocardial cell populations with different regenerative capacities after cardiac tissue injury, proliferative cardiomyocyte populations were identified, and 825 genes were found to be significantly upregulated. In addition, a comparison between nuclei division and cytokinesis myocardial cell subpopulations led to the identification of 207 genes that may promote cardiomyocyte cytokinesis.
CONCLUSIONS In conclusion, this study compared and analyzed the molecular characteristics of myocardial cell populations with differing proliferative abilities from multiple perspectives. Thirty novel genes that potentially promote cardiomyocyte cytokinesis were identified. These findings provide new insights and therapeutic targets for the treatment and prevention of future heart diseases. By analyzing gene expression modules and cell cycle regulation during cardiac development, we can better understand the mechanisms of cardiac cell proliferation and division, which can contribute to the development of innovative treatment methods and drugs.
GW34-e1140
Xiaoyan Zhu, Lin Yuan, Hu Liang
Academy of Integrative Medicine, Shanghai University of Traditional Chinese Medicine, Shanghai 201203, China
OBJECTIVES Oroxylin A (OrA) is a flavonoid obtained from the traditional Chinese medicine Scutellaria baicalensis. Overwhelming pieces of evidence expound on the anti-inflammatory, anti-bacterial, anti-viral, and anti-cancer potentials of this flavonoid, which makes it an engrossing compound for research. However, its effect on platelet activation remains unclear. The present study investigated the effects of OrA on platelet activation and thrombus formation in vitro and in vivo.
METHODS We detected the effects of OrA on platelet aggregation and ATP release induced by various agonists using aggregometer in vitro. Clot retraction and spreading were imaged by digital camera and fluorescence microscope. PAC-1 and CD62P was measured by Flow cytometry. Platelet adhesion on collagen-coated surface under flow conditions was performed using the BioFlux 200 setup. We also evaluated the antithrombotic effect of OrA using mouse mesenterial arteriole thrombosis model induced by FeCl3.
RESULTS OrA concentration-dependently suppressed platelet aggregation at the concentration range of 25–200 μM induced by ADP, thrombin and collagen. OrA presented the superior antiplatelet activity on ADP-induced platelet aggregation, and the IC50 value of OrA for ADP-induced platelet aggregation was 25±5.0 μM. OrA also inhibited ATP release, platelet spreading and clot retraction. We examined PAC-1 and CD62P, two platelet activation markers, via flow cytometry and found that OrA decreased expression of PAC-1 and CD62P in platelets. Finally, administration of OrA also suppressed platelet adhesion on collagen in flow condition and decreased thrombus formation in mouse mesenteric arterioles.
CONCLUSIONS In conclusion, we for the first time demonstrated that OrA suppressed platelet activation and thrombus formation, highlighting its therapeutic potential in cardiovascular diseases.
GW34-e1145
Ryosuke Tsukabe1, Satoshi Shimizu1,2, Yoichi Sunagawa1,2,3, Yasufumi Katanasaka1,2,3, Toshihide Hamabe1,2,3, Masafumi Funamoto1,2, Kana Shimizu1,2, Koji Hasegawa1,2, Tatsuya Morimoto1,2,3
1Division of Molecular Medicine, School of Pharmaceutical Sciences, University of Shizuoka, Shizuoka 422-8526, Japan
2Division of Translational Research, Clinical Research Institute, Kyoto Medical Center, National Hospital Organization, Kyoto 612-8555, Japan
3Shizuoka General Hospital, Shizuoka 420-8527, Japan
OBJECTIVES Hypertrophic stimuli activate cardiac transcription factors such as the zinc finger protein GATA4 and modulate gene expression in cardiomyocytes. Acetylation is a critical GATA4 activation mechanism mediated partly by intrinsic histone acetyltransferases (HAT) such as transcriptional coactivator p300. However, the details of the regulatory mechanism of GATA4 transcriptional activity remain unclear. In this study, we investigated whether GATA4 self-association is involved in transcriptional activation during the development of cardiomyocyte hypertrophy.
METHODS To investigate whether GATA4 undergoes homodimerization in vitro, a GST pull-down assay was performed with a 35S-labelled GATA4 protein using GST-GATA4. To identify the dimerization region necessary for the homodimerization of GATA4, we performed a series of GST pull-down assays. Moreover, we investigate whether GATA4 forms a multimer, full-length GATA4 produced by E. coli was subjected to size exclusion column chromatography, followed by Western blotting using an anti-GATA4 antibody. To determine how many GATA4 units formed a multimer, HEK293T cells were co-transfected with FLAG-GATA4, HA-GATA4, and 3xMyc-GATA4. Nuclear extracts prepared from these cells were immunoprecipitated with an anti-FLAG antibody followed by an anti-HA antibody, and Western blotting was performed using an anti-Myc antibody. HEK293 cells were transfected with ANF-luc or ET-1-luc, GATA4 or acetyl-deficient GATA4, and p300 or HAT-deficient p300, and then reporter gene assay was performed. Finally, HEK293 cells were transfected with FLAG-GATA4 and HA-GATA4, and the deletion mutant containing GATA4 multimerization region peptide (GMP). Nuclear extracts prepared from these cells were immunoprecipitated with anti-FLAG antibody followed by Western blotting. Cultured cardiomyocytes were transfected with GMP and then treated with PE. These cells were immunostaining with anti-MHC.
RESULTS A GST pull-down assay showed that GATA4 is bound to itself in vitro and that amino acids 308–326 are required for this binding. Size exclusion column chromatography showed that GATA4 formed multimers. Immunoprecipitation-Western blotting revealed that GATA4 formed, at least, a trimer. A reporter assay showed that p300 enhanced the promoter activities of ANF and ET-1. The self-association of GATA4 was disrupted by both acetyl-deficient GATA4 and HAT-deficient p300. Overexpression of GMP prevented p300-induced GATA4 self-association but not binding with p300 nor acetylation of GATA4. Chromatin immunoprecipitation and DNA pull-down assays showed that GMP did not inhibit p300-induced DNA binding of GATA4. The inhibition of GATA4 self-association using GMP suppressed phenylephrine-induced hypertrophic transcriptional activity and cardiomyocyte hypertrophy in cultured cardiomyocytes.
CONCLUSIONS These results suggest that GATA4 multimerization plays an essential role in hypertrophic gene transcription and that inhibiting this multimerization suppresses hypertrophic responses in cardiomyocytes. Future investigation of compounds that inhibit GATA4 multimerization may lead to the development of novel heart failure therapies.
GW34-e1181
Chaowei Hu1, Zhiyong Du1, Lijie Han1, Jing Li2, Huahui Yu1, Yu Wang1, Yanwen Qin1,3
1Key Laboratory of Upper Airway Dysfunction-Related Cardiovascular Diseases, Beijing Anzhen Hospital, Capital Medical University, Beijing Institute of Heart, Lung and Blood Vessel Diseases, Beijing 100029, China
2Heart Center & Beijing Key Laboratory of Hypertension, Beijing Chaoyang Hospital, Capital Medical University, Beijing 100020, China
3Key Laboratory of Remodeling-Related Cardiovascular Diseases, Beijing Anzhen Hospital, Capital Medical University, Beijing Institute of Heart, Lung and Blood Vessel Diseases, Beijing 100029, China
OBJECTIVES Homozygote familial hypercholesterolemia (HoFH) is a rare metabolic disorder attributed to the expression and functional defects of low-density lipoprotein receptor (LDLR), which result in extremely high level of LDL-c and ultimately metabolic dysfunction in patients. Intestinal flora have been uncovered to be implicated in various metabolic disorders. However, whether gut microbiota participates in the development of HoFH remains largely unknown.
METHODS To investigate this issue, we performed comprehensive metagenomic and metabolomic analyses of LDLR knockout hamsters, and fecal microbiota transplantation from LDLR−/− to germ-restriction hamsters.
RESULTS The global gut microbial characteristics including richness and diversity in LDLR−/− group were similar to those in controls. It was identified that Staphylococcus hominis, Kushneriaavicenniae and Bacillus dielmoensis were significantly enriched in LDLR knockout hamsters, whereas Bacillus halotolerans and Bacillus drentensis were reduced. In addition, LDLR−/− hamsters showed profound alterations in metabolomic phenotypes as compared to the control group. LDLR−/− hamsters exhibited enhanced abundance of Glycerolipids, Hormones and Organic Acid, as well as its derivatives such as Mag (18:1), 7-Ketocholesterol and Palmitoylethanolamide (PEA). Moreover, by fecal transplantation from LDLR−/− hamsters to germ-restriction recipients, elevated cholesterol level was observed to be transferrable through gut microbiota, and the direct causal impact of gut microbiota on serum metabolomics shifts of the host was also demonstrated. Here it was further confirmed that, the circulating level of circulating PEA, which was enriched in LDLR−/− donors, was significantly augmented in serum of recipients following fecal microbiota transplantation. In vitro, PEA administration elicited markedly elevated expression of 3-hydroxy-3-methyl glutaryl coenzyme A reductase (HMGCR) in Human Hepatocellular Carcinoma (Huh7) cell line.
CONCLUSIONS The present results demonstrate that gut microbiota in LDLR−/− hamsters lead to enhanced cholesterol levels through modulating metabolites. Meanwhile, it provides insights into a potential role of LDLR−/− specific intestinal microbiota-associated PEA as a key factor in regulating cholesterol synthesis. These findings describe a novel causal role of aberrant gut microbiota and metabolomics in contributing to the pathogenesis of HoFH.
GW34-e1200
Runmin Sun, Qiongying Wang, Jing Yu
Lanzhou University Second Hospital
OBJECTIVES The present study was designed to investigate whether mitochondrial dysfunction and oxidative stress in HTN-injured cardiomyocytes are associated with SIRT3 SUMOylation and whether SIRT3 deSUMOylation can ameliorate the related cardiomyocyte injury.
METHODS The human cardiomyocyte AC16 cell line was selected as the study target. The SIRT3-K288R cell line construction was verified by immunoprecipitation and the effect of Ang-II on SIRT3 SUMOylation was explored; the expression of SOD2, Acetyl-SOD2 (MnSOD), protein acetylation level, inflammatory vesicles NLRP3, etc. in different subgroups were determined by Western Blot to explore the SUMOylation modifications and SIRT3 deacetylation in HTN-related cell injury.
RESULTS
(1) Immunoprecipitation results showed that the level of SIRT3 SUMOylation was reduced after SIRT3-K288R transfection; compared to the control group, the level of SIRT3 SUMOylation was increased in the presence of AngII;
(2) Western Blot results showed that compared to the HTN cell injury model, the cardiomyocytes in the SIRT3-K288R intervention group had increased Acetyl-SOD2/SOD2 levels were increased (P<0.05) and protein acetylation levels were decreased in the SIRT3-K288R intervention group compared with the HTN cell injury model, indicating that SIRT3 increased the deacetylation effect on mitochondria and increased cellular antioxidant stress enzyme activity.
CONCLUSIONS In the HTN-associated cell injury model, the level of SIRT3 class ubiquitination was significantly increased, and the removal of SUMOylation modification by viral transfection could improve oxidative stress, protein acetylation levels and associated inflammatory responses in HTN-injured cardiomyocytes, indicating that SIRT3 SUMOylation may play an important mechanism in HTN-associated cell injury, and reducing SIRT3 SUMOylation modification may be a possible direction for future research.
GW34-e1217
Liangpeng Li, Qiao Liao, Chunyu Zeng
Department of Cardiology, Amy Medical Center (Daping Hospital), Army Medical University
OBJECTIVES Stimulating adult cardiomyocyte proliferation is a promising strategy for treating heart failure post myocardial infarction (MI). The cardiac metabolic pattern switching from glucose utilization to fatty acids oxidation are key factors contributing to the decreased proliferation potential of cardiomyocyte after birth. However, the underlying mechanisms are unclear, the potential targets for therapeutic purpose are still lacking.
METHODS We used Trimetazidine (TMZ), a metabolic modulator which inhibits fatty acids oxidation, to treat neonatal C57/BL6J mice, adult mouse and Bama minipig MI. The glucose utilization and fatty acids oxidation of cardiomyocytes were measured by XF24e Seahorse analyzer. Cardiomyocyte proliferation was evaluated by Ki67/Ph3, time lapse live cell imaging, and Myh6-MerCreMer/Mosaic analysis with double markers (MADM) transgenic mice. The cardiac function, scar size were also assessed. RNA-sequencing are performed on neonatal mouse heart to elucidate the transcriptional profile.
RESULTS
1. TMZ repressed fatty acid oxidation. As revealed by Seahorse analyzer, TMZ treatment inhibited fatty acids oxidation and enhanced glucose utilization in both neonatal and adult rodent cardiomyocyte.
2. TMZ promotes cardiomyocyte proliferation in postnatal mice. In neonatal mice, TMZ increased the heart weight and gross volume while decreased the cross-section area of cardiomyocyte, Ki67 and Ph3 positive cardiomyocytes were significantly elevated by TMZ treatment.
3. TMZ stimulate cardiomyocyte proliferation in adult mice and pigs. In adult mice and Bama minipig MI, TMZ improved the cardiac function, limited the scar size, and increased the number of Ki67 and Ph3 positive cardiomyocytes 4 week post MI. In Myh6-MerCreMer/MADM mice MI, the cardiomyocytes with single-color fluorescence, which represent authentic proliferated cardiomyocyte, were increased by TMZ. Time-lapse live cell video also showed that the cytokinesis was elevated in TMZ treated group compared with control.
4. ECM/SMAD4/BMP10 are potential mechanism underlying TMZ’s effect. RNA-seq revealed that in postnatal mouse heart treated with or without TMZ, “ECM” and “integrin binding” were most enriched GO terms. Immunostaining also confirmed that Collagen I is decreased by TMZ treatment. RNA-seq revealed that Bone Morphogenetic Protein 10 (BMP-10) is the top one in foldchange among up-regulated genes, administration of recombinant BMP10 mimic the stimulation effect of TMZ on cardiac proliferation. Transactor predicting tool ChEA3 suggested that SMAD4 is potential regulators of BMP10. The nuclear localization of SMAD4 is indeed increased by TMZ compared with control in postnatal mouse cardiomyocyte.
The nuclear translocation of SMAD4 is regulated by its phosphorylation. As integrin signaling are known molecular sensing ECM mechanical stress and transduced into intra-cellular signaling, we speculate that the effector down-stream of integrin signaling may regulate SMAD nuclear localization. We also found that the inhibition of ROCK1, the kinase which is activated while ECM component bind to integrin, reduced SMAD4 nuclear localization and the subsequent BMP10 production.
CONCLUSIONS We firstly showed that in both mouse and swine model, metabolic alteration by TMZ ameliorated cardiac remodeling post MI by promoting cardiomyocyte proliferation. This effect is potentially associated with ECM remodeling/SMAD4/BMP10 signaling. These findings revealed the mechanism by which metabolism regulate cardiac regeneration and provide potential targets for therapeutic purpose.
GW34-e1218
Jia Li
Changhai Hospital
OBJECTIVES Heart failure is the ultimate stage in the development of various cardiovascular diseases, and morbidity and mortality have increased rapidly in recent years. Splicing factors control alternative splicing events, forming a complex regulatory network to regulate biological process which is associated with heart failure. Our research aimed to comprehensively analyze and identify variable splicing factors and alternative splicing regulatory networks in heart failure utilizing RNA-seq data and machine learning algorithms.
METHODS Three RNA-seq raw sequencing datasets contain heart failure and non-heart failure donors were retrieved and downloaded from the GEO database, and the gene expression matrices of the three datasets were obtained by bioinformatics analysis. The batch effects were removed by the SVA method and combined into one expression matrix. The SpliceAid 2 database was used to obtain the expression of splicing factors from the expression matrix. The differentially expression alternative splicing events in heart failure patients compared with non-heart failure donors was identified by calculating the percent spliced-in (PSI) value to quantify, and the regulatory network of splicing factors and variable splicing events was constructed. Functional enrichment analysis was performed for differential alternative splicing genes. Furthermore, heart failure samples were clustered unsupervised according to differential variable splicing events. Finally, characteristic splicing factor genes were selected through integrating LASSO and SVM-RFE algorithms, and validated in a microarray dataset.
RESULTS A total of 4055 differential alternative splicing events and 3142 DEAS related genes were identified. A total of 30 alternative splicing factors were identified from expression matrix according to the SpliceAid 2 database, of which 12 splicing factors showing higher expression levels in heart failure samples compared to non-heart failure samples. In addition, we identified the splicing regulation network of which contain 203 significant SF-ASE pairs in heart failure. The functional enrichment analysis result revealed that the differentially alternative splicing genes showed high enrichment in PPAR signaling pathway, regulation of actin cytoskeleton, muscle contraction and Z-disc. Furthermore, heart failure patients were divided to two distinct DASE-based clusters by unsupervised clustering method. Finally, four splicing factor genes were selected as characteristic genes using two machine learning algorithms in heart failure.
CONCLUSIONS Our study provided a systematic portrait analysis of alternative splicing regulatory network and splicing factors in heart failure, and uncovered splicing networks that are valuable in deciphering the underlying mechanisms of AS in heart failure. Heart failure characteristic SF genes were determined, which were remarkably linked to RNA metabolism process.
GW34-e1223
Huahui Yu1,2, Yunhui Du1,2, Xiaolu Jiao1,2, Qianwen Lv1,2, Fan Li1,2, Yu Wang1,2, Yanwen Qin1,2
1Beijing Anzhen Hospital, Capital Medical University
2Beijing Institute of Heart Lung and Blood Vessel Disease
OBJECTIVES Myocardial infarction (MI) contributes substantially to CVD-associated mortality. Despite the success of current therapies for MI, many patients still develop adverse cardiac remodeling and heart failure. ANGPTL8 is a secretory protein, which plays a critical role in cardiovascular diseases, including atherosclerosis. However, the direct role of ANGPTL8 in the progression of myocardial remodeling after MI is still unknown. Thus, we explored the contribution of ANGPTL8 to myocardial remodeling after MI and its underlying mechanisms.
METHODS The MI model was induced by the ligation of LAD coronary artery, and the cardiac function of mice was determined 28 days later by echocardiography. By using ANGPTL8 knockout mice, we determined the role of ANGPTL8 on post-MI in vivo. Cardiac fibrosis was measured by Masson staining. and the molecular markers of remodeling were evaluated by Western blot and immunohistochemistry. The mice primary cardiac fibroblasts with ANGPTL8 overexpression were used to detect the mechanism of ANGPTL8 promoted the progression of myocardial remodeling after MI.
RESULTS ANGPTL8 mRNA and protein levels were increased in mice hearts after 28-day MI. Genetic deletion of ANGPTL8 reduced pathological cardiac remodeling and increased survival rate. ANGPTL8 knockout significantly attenuated fibroblast activation, collagen production and fibrosis, cardiomyocyte hypertrophy and improved impaired heart function, at infarcted border zone after MI. Mechanistically, ANGPTL8 was shown to induce cardiac fibroblast-myofibroblast transformation via the transforming growth factor (TGF)-β level.
CONCLUSIONS In conclusion, MI upregulates cardiac ANGPTL8 expression. ANGPTL8 knockout could attenuate cardiac fibrosis at infarcted border zone and improve impaired heart function, suggesting that ANGPTL8 may be a novel therapeutic target against pathologic remodeling in MI patients.
GW34-e1243
Jie Wang, Yanmin Zhang
National Regional Children’s Medical Centre (Northwest); Key Laboratory of Precision Medicine to Pediatric Diseases of Shaanxi Province; Xi’an Key Laboratory of Children’s Health and Diseases, Shaanxi Institute for Pediatric Diseases; Xi’an Children’s Hospital, Affiliated Children’s Hospital of Xi’an Jiaotong University, Xi’an, Shaanxi 710003, China
OBJECTIVES Complement C1q binding protein (C1QBP) is primarily localized in mitochondrial matrix and associated with mitochondrial oxidative phosphorylative function. C1QBP deficiency presents as a mitochondrial disorder involving multiple organ systems. Recently, disease associated C1QBP mutations have been identified in patients with a combined oxidative phosphorylation deficiency (COXPD) taking an autosomal recessive inherited pattern. The clinical spectrum ranges from intrauterine growth restriction to childhood (cardio) myopathy and late-onset progressive external ophthalmoplegia. We previously identified a homozygous C1QBP-L275F mutation in two COXPD brothers with cardiac hypertrophy.
METHODS The patient and his mother specific induced pluripotent stem cells (iPSC) carrying a homozygosis and heterozygous C1QBP-L275F mutation respectively were generated from PBMCs. Thus PBMCs were reprogrammed using non-integrative sendai viral vectors containing reprogramming factors OCT4, SOX2, KLF4 and C-MYC. Cardiomyocytes were derived from iPSCs named as Proband-Homo-iPSC-CMs and Mother-Hetero-iPSC-CMs respectively. Then the hypertrophic phenotypes of cardiomyocytes were investigates using confocal microscope, Image J and Real-time-PCR. And mitochondrial function studies using transmission electron microscopy and XFe 24 Extracellular Flux Analyzer. Finally, C1qbp-L272F gene knockin mice are generated to investigate the potential mechanism at the whole, organ and molecular levels at 4 months old.
RESULTS Generated iPSCs expressed pluripotent markers, trilineage differentiation potential, carrying C1QBP-L275F mutation, and have a normal karyotype. iPSC-CMs derived from iPSC expressed cardiomyocyte marker CTNT and α-actine. Successfully differentiated iPSC-CMs were using for further investigations. Thus firstly Proband-Homo-iPSC-CMs showed cardiomyocyte area larger and the cardiac hypertrophic marker genes ANP, BNP, and MYH6 was significantly up-regulated compared with Mother-Hetero-iPSC-CMs. Secondly mitochondrial structures were vacuole-impaired, the number and area of mitochondria decreased significantly in Proband-Homo-iPSC-CMs. Furthermore, mitochondrial fusion/fission dynamics disturbance and a decrease in complex I and complex IV subunits was present in Proband-Homo-iPSC-CMs. High-resolution respirometry respiration results showed reduced basal and maximal respiration, proton leakage, and ATP production were found in Proband-Homo-iPSC-CMs. Finally, echocardiographic measurements was performed, and thickness of interventricular septum and posterior left ventricular wall were present in homozygosis C1qbp-L272F mice. Similarly, cardiac hypertrophy markers of Anp and Bnp present an up-regulation trend in homozygosis C1qbp-L272F mice. The results of cardiac electron microscopy showed that the cardiac fiber arrangement of homozygous C1qbp mutant mice was disordered, mitochondrial swelling and partial mitochondrial ridge disappeared, and the expression of mitochondrial dynamic was significantly abnormal compared with that of wild type group.
CONCLUSIONS We successfully established the C1QBP-L275F iPSC-CMs and C1qbp-L272F gene knockin mice. The cardiac hypertrophy was recurrented in this two models. Moreover mitochondrial structure, function, energy metabolism changes was exhibited and potential mechanism was explored. C1QBP/C1qbp mutation-specific iPSC-CMs and mouse models can provide a tool for future research on the mechanism of mitochondrial cardiomyopathy.
GW34-e1246
Yingchao Gong1, Chenyang Jiang1, Guosheng Fu1, Peng Zhang2, Dongwu Lai1
1Department of Cardiology, Key Laboratory of Cardiovascular Intervention and Regenerative Medicine of Zhejiang Province, Sir Run Run Shaw Hospital, Zhejiang University School of Medicine, Hangzhou, China
2Central Laboratory, Putuo Hospital, Shanghai University of Traditional Chinese Medicine, Shanghai, China
OBJECTIVES Cardiac pathological remodeling following myocardial infarction (MI) can result in heart failure and sudden cardiac death. Mtus1A (Mitochondrial tumor suppressor 1A) is located in mitochondrion and its deficiency is associated with mitochondrial disorder. However, the precise involvement of Mtus1A in MI remains unclear. In this study, we aimed to reveal the critical role of Mtus1A in post-MI cardiac remodeling.
METHODS We performed single-cell sequencing analysis to clarify the expression of Mtus1 in cardiomyocytes subjected to ischemic stress. The impact of Mtus1 on cardiac functions and the integrity of MAMs (mitochondria-associated endoplasmic reticulum membranes) was investigated in cardiomyocyte-specific Mtus1 gene knockout mice and in neonatal rat ventricular myocytes through siRNA transfection. Additionally, immunoprecipitation-mass spectrometry was employed to identify the interactive proteins of Mtus1A. In order to investigate their roles in MI, adeno-associated virus serotype 9 (AAV9) carrying Mtus1A or shRNA targeting Fbxo7 (F-box protein 7) was employed.
RESULTS Our study revealed a significant downregulation of Mtus1 expression in the hearts of both patients and mice with MI. Additionally, we identified the localization of Mtus1A to the MAMs both in vitro and in vivo settings. Furthermore, loss of Mtus1A disrupted the association between endoplasmic reticulum and mitochondria, resulted in impairment of heart function and mitochondrial structure under basal conditions. At the mechanistic level, our study demonstrated that Mtus1A acted as a scaffold protein to maintain the stability of the IP3R1-Grp75-VDAC1 complex in MAMs. Additionally, we observed that Mtus1A can be stabilized through K6-linked ubiquitination by Fbxo7. At the functional level, our finding indicated that overexpression of Mtus1A mitigates cardiac function induced by left anterior descending ligation by preserving the connection between the ER and mitochondria.
CONCLUSIONS Taken together, our findings suggest that Mtus1A is essential for the maintaining the structural soundness of the IP3R1-Grp75-VDAC1 complex and facilitating the interaction between the ER and mitochondria. Disruption of this pathway leads to mitochondrial dysfunction and exacerbates MI-induced cardiac dysfunction.
GW34-e1261
Yao Zhang, Wei Zhao, Rongjie Tang, Yingxin Huo, Feng Sun, Qiufang Lian
Xianyang Hospital of Yan’an University
OBJECTIVES To observe the effect of allicin (ALL) on renal fibrosis in salt-sensitive hypertensive rats and explore the mechanism of ALL based on Klotho/TGF-β1 signaling pathway.
METHODS Twenty-four male Dahl salt-sensitive rats, 8 weeks old, were randomly divided into normal-salt (NS), high-salt (HS) and high-salt plus ALL (HS+ALL) groups after 1 week of acclimatization feeding. They were given 0.3% NaCl, 8% NaCl and 8% NaCl with ALL (16 mg/kg, p.o), respectively, once a day for 4 weeks. Systolic blood pressure (SBP) was measured using a non-invasive blood pressure monitor, 24-hour urine was collected, the kidney weight coefficient (kidney weight/rat weight) was calculated, renal tissue was collected. The collagen volume of kidney (CVF) was calculated. The contents of interleukin-6 (IL-6) and tumor necrosis factor-α (TNF-α) in kidney tissue were determined by ELISA, the mRNA and protein levels of Collagen I, Fibronectin, Klotho and TGF-β1 in kidney tissue were detected by qRT-PCR and Western Blot.
RESULTS
1. Compared with the NS group, SBP was increased (P<0.05), urine creatinine and protein levels were increased (P<0.05), and renal weight coefficient was increased in the HS group rats (P<0.05). These were reversed in part in HS+ALL group (P<0.05).
2. Compared with the NS group, H&E staining showed severe renal injury, compensatory hypertrophy of some glomeruli, widening of peritubular space, and significant inflammatory cell infiltration, Masson staining showed significant renal fibrosis and increased CVF (P<0.05), levels of IL-6 and TNF-α in renal tissue were increased (P<0.05), Collagen I and Fibronectin mRNA and protein content in kidney tissue were increased (P<0.05) in the HS group. The above indicators were reversed in part in HS+ALL group (P<0.05).
3. Compared with the NS group, the mRNA and protein expression levels of Klotho in kidney tissue decreased in the HS group (P<0.05), the mRNA and protein expression levels of TGF-β1 in kidney tissue increased (P<0.05). The above indicators were reversed in part in HS+ALL group (P<0.05).
CONCLUSIONS ALL can improve renal dysfunction and renal fibrosis caused by salt-sensitive hypertension, which is related to its regulation of Klotho/TGF-β1 signaling pathway.
GW34-e1265
Yingxin Huo, Wei Zhao, Rongjie Tang, Yao Zhang, Feng Sun, Qiufang Lian
Xianyang Hospital of Yan’an University
OBJECTIVES To observe the effect of allicin (ALL) on cardiac fibrosis in salt-sensitive hypertension and explore the mechanism of ALL based on the TGF-β1/Smad2/3 signaling pathway.
METHODS Twenty-four male Dahl salt-sensitive rats, 8 weeks old, were randomly divided into normal-salt group (NS), high-salt group (HS) and high-salt group plus ALL group (HS+ALL) after 1 week of adaptive feeding, and were given sodium chloride containing 0.3% NaCl, 8% NaCl and 8% NaCl and ALL (16 mg/kg, p.o) once a day for 4 weeks, respectively. Systolic blood pressure (SBP) was measured by non-invasive blood pressure monitor, left ventricular ejection fraction (LVEF), left ventricular end-systolic volume (LVESV) and left ventricular end-diastolic volume (LVEDV) were measured by small animal ultrasound, rat hearts were taken, heart mass index (HMI) and left ventricular mass index (LVMI) were calculated, ELISA was performed to determine the contents of interleukin-6 (IL-6) and tumor necrosis factor-α (TNF-α) in kidney tissue, qRT-PCR and Western Blot were performed to detect the mRNA and protein levels of Collagen I, TGF-β1, p-Smad2 and p-Smad3 in kidney tissue.
RESULTS
1. Compared with the NS group, SBP was increased (P<0.05), LVEF was decreased, LVESV and LVEDV were increased, HMI and LVMI were increased in the HS group rats. These were reversed in part in HS+ALL group (P<0.05).
2. Compared with the NS group, HE staining in the HS group showed disturbed arrangement of LV myocytes and significant inflammatory cell infiltration, increased interstitial collagen deposition in LV cells and increased CVF (P<0.05), increased IL-6 and TNF-α content in kidney tissue, increased mRNA and protein content of Collagen I in kidney tissue (P<0.05). The above indicators were reversed in part in HS+ALL group (P<0.05).
3. TGF-β1 mRNA and protein expression levels were increased in renal tissue of the HS group compared with the NS group (P<0.05), in addition, the protein levels of p-Smad2 and p-Smad3 were also increased in renal tissue (P<0.05). The above indicators were reversed in part in HS+ALL group (P<0.05).
CONCLUSIONS ALL can improve cardiac hypofunction and cardiac fibrosis caused by salt-sensitive hypertension, which is related to its inhibition of TGF-β1/Smad2/3 signaling pathway.
GW34-e1268
Ming Bai1,2,3,4, Jingjing Zhang5,6, Shuwen Hu4, WenJun Zhang1,2,3,4, Fei Zhao7, Xiaowei Niu1,2,3,4
1Department of Cardiology, The First Hospital of Lanzhou University, Lanzhou 730000, China
2Gansu Key Laboratory of Cardiovascular Diseases, Lanzhou 730000, China
3Gansu Clinical Medical Research Center for Cardiovascular Diseases, Lanzhou 730000, China
4The First School of Clinical Medicine of, Lanzhou University, Lanzhou 730000, China
5Gansu Provincial Maternity and Child-care Hospital/Gansu Provincial Central Hospital, Medical Genetics Center, Lanzhou 730000, China
6Gansu Provincial Clinical Research Center for Birth Defects and Rare Diseases, Lanzhou 730000, China
7TEDA International Cardiovascular Hospital, Department of Electrophysiology, Tianjin 300000, China
OBJECTIVES Myocardial ischemia reperfusion (MI/R) injury often leads to adverse cardiovascular outcomes in patients with acute myocardial infarction. Effective strategies for preventing and treating myocardial ischemia-reperfusion (MI/R) injury are still lacking in clinical practice. Our previous research has conducted the network pharmacology analysis, which identified Angelica sinensis polysaccharide (ASP), a major water-soluble component of Angelica sinensis, with potential cardioprotective effects. In this study, we aim to elucidate the mechanism of action of ASP by integrating bioinformatics analysis with both in vivo and in vitro experiments.
METHODS In the in vitro experiments, we established a hypoxia/reoxygenation (H/R) model using H9c2 rat cardiomyocyte cell line. In the in vivo experiments, we induced myocardial ischemia-reperfusion (MI/R) injury by ligating the left anterior coronary artery in rats. Myocardial samples were collected for transcriptome sequencing and subsequent bioinformatics analysis.
RESULTS
ASP decreases H/R-induced cell pyroptosis: Compared to the H/R model group, pre-treatment with ASP significantly mitigated the inhibitory effects of H/R on H9c2 cell viability. ASP treatment markedly reduced the release of lactate dehydrogenase (LDH) and the percentage of TUNEL-positive cells induced by H/R. ELISA analyses demonstrated that ASP intervention decreased the levels of inflammatory cytokines (IL-1β, IL-18, and TNF-α) induced by H/R. Western blot analysis revealed that ASP treatment significantly downregulated the expression levels of NLRP3 and N-GSDMD compared to the H/R group. Additionally, ASP intervention inhibited the activity of Caspase-1.
ASP alleviates inflammatory injury following MI/R: In the rat MI/R model, pre-treatment with ASP significantly reduced the infarct size, levels of myocardial enzymes (LDH and cTNT), and ST-segment elevation on the electrocardiogram. HE staining showed that ASP treatment significantly reduced inflammatory cell infiltration and improved the morphology of myocardial tissue. Echocardiographic analysis demonstrated that ASP therapy significantly ameliorated heart dysfunction with increased ejection fraction and fractional shortening. Consistent with the in vitro results, ASP treatment markedly decreased the percentage of TUNEL-positive cells induced by MI/R, as well as the release of inflammatory cytokines (IL-1, IL-18, and TNF-α). ASP intervention also reduced the activity of Caspase-1 and downregulated the expression levels of NLRP3 and N-GSDMD.
Bioinformatics analysis: Differential expression analysis, gene set enrichment analysis (GSEA), Gene Ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis, and protein-protein interaction (PPI) network analysis revealed that the FN1/NF-κB/NLRP3 signaling pathway could be a potential molecular mechanism.
Investigation of the molecular mechanism: The Western blot analysis and rescue experiments showed that ASP significantly downregulated the FN1/NF-κB/NLRP3 signaling pathway. Additionally, the administration of the NLRP3 agonist nigericin significantly reversed the protective effects of ASP against MI/R injury.
CONCLUSIONS Our findings suggest that ASP protects the myocardium against ischemia/reperfusion injury by inhibiting cardiomyocyte pyroptosis. The FN1/NF-κB/NLRP3 inflammasome pathway mediates the cardioprotective effects of ASP. These results offer new insights into the development of ASP-related drugs, and provide a theoretical basis for exploring novel strategies for treating MI/R injury.
GW34-e1273
Dong Han1, Tingwen Zhou2, Yongjun Wang2, Feng Cao1
1National Clinical Research Center for Geriatric Diseases, 2nd Medical Center, Chinese PLA General Hospital, Beijing 100853, China
2Department of Cardiovascular Surgery, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430022, Hubei Province, China
OBJECTIVES Calcification of the aortic valve leads to increased leaflet stiffness and consequently results in the development of calcific aortic valve disease (CAVD). However, the underlying molecular and cellular mechanisms of calcification remain unclear. Here, we identified a novel aortic valve calcification-associated PIWI-interacting RNA (piRNA) (AVCAPIR) that increases valvular calcification and promotes CAVD progression.
METHODS Using piRNA sequencing, we identified piRNAs contributing to the pathogenesis of CAVD that we termed AVCAPIRs. High-cholesterol diet (HCD)-fed ApoE−/− mice with AVCAPIR knockout were used to examine the role of AVCAPIR in aortic valve calcification (AVC). Gain- and loss-of-function assays were conducted to determine the role of AVCAPIR in the induced osteogenic differentiation of human primary valvular interstitial cells (hVICs). To dissect the mechanisms underlying the AVCAPIR-elicited procalcific effects, we performed various analyses, including an RNA pulldown assay followed by liquid chromatography-tandem mass spectrometry (LC-MS/MS), methylated RNA immunoprecipitation sequencing (MeRIP-seq), quantitative reverse transcription polymerase chain reaction, Western blotting, and RNA sequencing (RNA-seq). RNA pulldown and RNA immunoprecipitation (RIP) assays were used to study piRNA interactions with proteins.
RESULTS We found that AVCAPIR was significantly upregulated during AVC and exhibited potential diagnostic value for CAVD. AVCAPIR deletion markedly ameliorated AVC in HCD-fed ApoE−/− mice, as shown by reduced thickness and calcium deposition in the aortic valve leaflets, improved echocardiographic parameters (decreased peak transvalvular jet velocity and mean transvalvular pressure gradient, as well as increased aortic valve area), and diminished levels of osteogenic markers (Runx2 and Osterix) in the aortic valves. These results were confirmed in osteogenic medium-induced hVICs. Using unbiased protein-RNA screening and molecular validation, we found that AVCAPIR directly interacts with FTO, subsequently blocking its N6-methyladenosine (m6A) demethylase activity. Further transcriptomic and m6A modification epitranscriptomic screening and molecular validation confirmed that AVCAPIR hindered FTO-mediated demethylation of CD36 mRNA transcripts, thus enhancing CD36 mRNA stability through the m6A reader IGF2BP1. The AVCAPIR-dependent increase in CD36 in turn stabilizes CD36’s binding partner PCSK9, a procalcific gene, at the protein level, which accelerates the progression of AVC.
CONCLUSIONS We identified a potentially novel piRNA that induced AVC via an RNA epigenetic mechanism and provide novel insights into piRNA-directed theranostics in CAVD.
GW34-e1281
Lei Wang
Capital Medial University
OBJECTIVES Pathological cardiac hypertrophy leads to heart failure and is the leading causes of death from cardiovascular disease. Understanding the molecular mechanism of pathological cardiac hypertrophy will contribute to the prevention and treatment of heart failure. β-catenin is an important regulator of cardiac development and remodeling. The high β-catenin activity leads to cardiac hypertrophy and heart failure. Ubiquitination modification is essential to maintain the normal activity, localization, and degradation of β-catenin. However, which deubiquitinating enzymes participated in the regulating of β-catenin activity remain largely unknown.
METHODS Immuno-coprecipitation-mass spectrometry (IP-MS) was used to identify the BRISC deubiquitinating enzyme scaffolding protein ABRO1 was the interacting protein of β-catenin in cardiomyocytes. To confirm the direct interaction of ABRO1 and β-catenin, we performed co-immunoprecipitation and GST-pull down. The truncated text of ABRO1 and β-catenin were designed to recognize the interaction location of ABRO1 and β-catenin. To identify the β-catenin-ubiquitination modification site by BRISC complex, we performed ubiquitinating proteome. Then ubiquitination-modification-sites mutation of β-catenin were generated to confirm the ubiquitination modification sites induced by ABRO1. The role of ABRO1 in cardiac hypertrophy and heart failure was further evaluated in MYH6creABRO1flox/flox mice and cardiomyocyte-specific overexpression of ABRO1 mice in response to angiotensin II (Ang II). Immunofluorescent staining and qPCR were used to verify β-catenin nuclear trans-location and transcriptional activity in ABRO1 knockout and overexpression cardiomyocytes and heart. Moreover, we performed the rescue study in MYH6creABRO1flox/flox mice either by intravenous injection of AAV9-encoding ABRO1 under the cTnT promoter or using the β-catenin inhibitor ICG-001 to validate the biological function of ABRO1 and β-catenin interaction.
RESULTS We revealed that the BRISC deubiquitinating enzyme scaffolding protein ABRO1 was the interacting protein of β-catenin in cardiomyocytes. ABRO1 bound to β-catenin directly via the lysine at position 508 and exerted de-ubiquitination to decrease the activity of β-catenin by removing the K63 ubiquitin chain, therefore inhibiting β-catenin nuclear translocation and transcriptional activity. The protein level of ABRO1 in murine cardiomyocytes subject to Ang II and transverse aortic constriction stimulation and in hypertrophic myocardium tissues of patients with heart failure was significantly increased. Cardiac ABRO1 deficiency aggravated cardiac hypertrophy and heart failure under Ang II treatment by increasing β-catenin activity. Inversely, cardiac specific ABRO1 overexpressing attenuated Ang II induced cardiac hypertrophy and dysfunction. Restoration of ABRO1 expression via AAV9 vectors or inhibiting β-catenin activity in MYH6creABRO1flox/flox mice could attenuate Ang II-induced heart failure.
CONCLUSIONS We confirmed that ABRO1 inhibited cardiac hypertrophy by deubiquitinating and inhibiting β-catenin activation.
GW34-e1295
Tianxin Ye1, Jinxiu Yang1, Bo Yang2, Xingxiang Wang1
1Department of Cardiology, The First Affiliated Hospital, School of Medicine, Zhejiang University
2Department of Cardiology, Renmin Hospital of Wuhan University
OBJECTIVES Post-operative atrial fibrillation (POAF) is a common complication in patients undergoing cardiac surgery. The purinergic receptor P2X7 (P2X7R) is involved in ventricular arrhythmia after myocardial infarction (MI), whereas its effects on atrial fibrillation (AF) are unclear. This study was to assess the effect of P2X7R on atrial arrhythmogenic remodeling in the experimental sterile pericarditis (SP)-induced POAF model.
METHODS MI and POAF rat models were induced. Electrocardiogram, atrial electrophysiological protocol, histology, mRNA sequencing, real-time quantitative PCR, western blot, and Elisa assay were performed.
RESULTS The P2X7R was significantly up-regulated in the atrium both in the MI and SP rats. SP significantly increased AF susceptibility; reduced the protein expression of ion channels including Nav1.5, Cav1.2, Kv4.2, Kv4.3, and Kv1.5; caused atrial fibrosis; increased norepinephrine level in plasma; promoted the production of inflammatory cytokines and accumulation of immune cells; and activated NLRP3 inflammasome signaling pathway. P2X7R antagonist Brilliant Blue G (BBG) suppressed SP-induced alterations. The mRNA sequencing demonstrated that BBG prevented POAF mainly by regulation of the immune system. In addition, IL-1R antagonist anakinra also reduced AF incidence in SP-induced POAF.
CONCLUSIONS P2X7R inhibition prevents SP-induced atrial proarrhythmic remodeling, which is closely associated with the improvement of inflammatory changes, ion channel expression, atrial fibrosis, and sympathetic activation. The findings point to P2X7R inhibition as a promising target for POAF and perhaps other conditions.
GW34-e1298
Dong Han1, 2, Yongjun Wang2, Feng Cao1, 2
1National Clinical Research Center for Geriatric Diseases, 2nd Medical Center, Chinese PLA General Hospital, Beijing 100853, China
2Department of Cardiovascular Surgery, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430022, Hubei Province, China
OBJECTIVES Anthracycline cardiotoxicity remains a major challenge to the survival and life quality of cancer patients receiving anthracycline-based chemotherapy. Previous studies have indicated that heart and tumor could interact and influence the course of each other’s pathologies in patients with these two disease entities. Small extracellular vesicles (sEVs) critically orchestrate proximal and distant organ-organ communications and may play essential roles in heart-tumor interaction. However, whether cancer-secreted sEVs affect the progression of anthracycline cardiotoxicity via regulating the tumor cell-cardiomyocyte crosstalk has not yet been explored.
METHODS To decipher the potential interaction of breast cancer cells (BCCs) with cardiomyocytes in DOX-induced cardiotoxicity (DOXIC), a transwell based coculture system was employed. To determine whether BCC-sEVs were able to aggravate DOX-induced myocardial injury in vivo, sEVs derived from DOX-treated syngeneic E0771 breast cancer cells or normal control E0771 cells were intramyocardially injected into the myocardium of DOX-treated mice. To identify the specific functional miRNA in D-BCC-sEVs, differentially expressed miRNAs in D-BCC-sEVs and N-BCC-sEVS were revealed by miRNA sequencing. To investigate the mechanisms of exsomal miRNA’s function, we performed various analyses including quantitative reverse transcription polymerase chain reaction, Western blot, histology, cardiac function (by echocardiography), functional rescue experiments, RNA pulldown, RNA immunoprecipitation, luciferase reporter assay.
RESULTS Here, we demonstrate that doxorubicin (DOX)-treated breast cancer cells (D-BCCs) secreted sEVs that can exacerbate DOX-induced ferroptosis in cardiomyocytes and in an orthotopic mice model of breast cancer. The expression of miR-338-3p in D-BCCs and plasma-sEVs of DOX-treated tumor bearing mice was significantly upregulated. Incubation of cardiomyocytes with sEVs isolated from D-BCCs (D-BCC-sEVs) or overexpression of miR-338-3p alone intensified DOX-induced ferroptotic cell death in vitro. N6-methyladenosine was revealed to mediate the upregulation of miR-338-3p in D-BCCs through enhanced maturation. D-BCCs-enriched miR-338-3p was packaged in sEVs and transferred into cardiomyocytes in a RBMX-dependent manner. Anti-ferroptotic genes CP, SLC7A11, and GPX4 were experimentally established as targets of miR-338-3p in cardiomyocytes. Blockage of BCC-D-EXOs release with Rab27a silence or systematic anti-miR-338-3p antagomir treatment assuaged BCC-D-EXOs-induced myocardial ferroptosis and injury. Plasma sEVs isolated from DOX-induced cardiomyopathy patients potently enhanced DOX-induced ferroptosis in hiPSC-CMs, which was rescued by miR-338-3p inhibitor.
CONCLUSIONS Our findings uncovered for the first time that DOX-treated BCCs exacerbated DOXIC through releasing miR-338-3p-enriched sEVs. Therefore, targeting sEV-mediated tumor/cardiomyocyte pathological communication may offer a novel approach for the management of DOXIC.
GW34-e1305
Tianxin Ye1, Jinxiu Yang1, Bo Yang2, Xingxiang Wang1
1Department of Cardiology, The First Affiliated Hospital, School of Medicine, Zhejiang University
2Department of Cardiology, Renmin Hospital of Wuhan University
OBJECTIVES Depression, the most prevalent psychiatric disorder characterized by a depletion of pleasure, is closely associated with the occurrence and development of atrial fibrillation (AF), whereas the underlying mechanisms remain unclear. Activation of the P2X purinoceptor 7 (P2X7R) is involved in some cardiovascular diseases and participates in the development of depression, but little attention has been given to its role in AF. This study was to investigate the effects of P2X7R on depression-related AF.
METHODS Chronic unpredictable stress (CUS) or lipopolysaccharide (LPS) was carried out to induce rodent models of depression. Behavioral assessments, atrial electrophysiological parameters, Electrocardiogram (ECG) parameters, western blot, and histology were performed.
RESULTS Shortened effective response period (ERP), prolonged activation latency (AL), and increased inducibility of AF were observed in both LPS- and CUS-induced depression, along with the up-regulation of P2X7R in atria. CUS facilitated atrial fibrosis. CUS reduced HRV and increased the expression of TH and GAP43, representing autonomic dysfunction. Downregulation of Nav1.5, Cav1.2, Kv4.3, Kv1.5, Cx40, and Cx43 in CUS indicated the abnormalities in ion channels. In addition, the expression levels of TLR4, P65, P-P65, NLRP3, ASC, caspase-1, and IL-1β were remarkably elevated in depression. The pharmacological inhibitor (Brilliant Blue G, BBG) or genetic deficiency of P2X7R significantly mitigated depressive-like behaviors, and ameliorated electrophysiological deterioration and autonomic dysfunction in CUS-induced rats or mice. Moreover, BBG improved ion channel expression, atrial fibrosis, and attenuated atrial inflammation in the pathophysiological process of depression-related AF.
CONCLUSIONS Depression induces AF and promotes P2X7R-dependent activation of NLRP3 inflammasome, while pharmacological P2X7R inhibition or P2X7R genetic deficiency prevents atrial remodeling without interrupting normal atrial physiological functions. Our results point to P2X7R as an important factor in the pathology of depression-related AF.
GW34-e1349
Ruirui Chen, Yan Li
Department of Cardiology, Second Affiliated Hospital, Military Medical University of the Air Force
OBJECTIVES The amount of blood immune cells and the risk of heart failure (HF) are known to be positively associated. However, the causal impact of peripheral blood immune cells on HF has not been fully elucidated. We performed a Mendelian randomization (MR) study to examine the possible etiologic role of peripheral blood immune cells in HF.
METHODS A Mendelian randomization (MR) analysis was performed using genetic variants closely associated with circulating leukocyte, granulocyte, lymphocyte, and monocyte counts and monocyte subpopulations as instrumental variables (IV). A genome-wide association study (GWAS) dataset of 977,323 participants of European ancestry, including 47,309 HF cases and 930,014 controls, was collected to identify genetic variants in HF. A two-sample Mendelian randomization framework was implemented to examine the causal relationship between peripheral blood immune cells and HF. And then to find more effective ways to treat heart failure.
RESULTS Our findings indicated that higher monocyte count [odds ratio (OR), 0.914; 95% confidence interval (CI), 0.846–0.988; P=0.023] was causally associated with HF susceptibility. In addition, the absolute amount of CD14- CD16+ monocyte and CD14+ CD16- monocyte were negatively associated with the increased susceptibility to HF.
CONCLUSIONS Our findings show that the genetic predisposition to lower peripheral monocyte cell counts, especially CD14- CD16+ monocyte and CD14+ CD16- monocyte subtypes, can exert a causal effect on HF risk. This may play a guiding role in finding more effective drug treatment for heart failure.
GW34-e1357
Jian Zhang1, Ping Ren2, Poli Lin2, Boya Cao2, Jiali Chen3, Ping He1, Wei Wang4, Linghui Lu3, Junling Cao2,5
1School of Life Sciences, Beijing University of Chinese Medicine
2School of Chinese Materia Medica, Beijing University of Chinese Medicine
3School of Chinese Medicine, Beijing University of Chinee Medicine, Beijing
4School of Basic Medical Sciences, Guangzhou University of Chinese Medicine
5Dongfang Hospital, Beijing University of Chinese Medicine
OBJECTIVES Lonicerae Japonicae Flos (LJ), one of the genuine medicinal materials in China, is mainly used to treat inflammation related diseases clinically for hundreds of years. Inflammation plays an important role in the occurrence and development of myocardial infarction (MI) and clinical studies have shown that anti-inflammatory treatment can improve myocardial ischemia post MI. This study was undertaken to explore the anti-inflammation mechanism of Lonicerae Japonicae Flos extract (LJE) against myocardial injury post MI targeting lipoxygenases pathway.
METHODS LJE was extracted from the herb of LJ. Left anterior descending ligation was applied to induce an MI mice model. LJE was orally administered 24 h after MI for 7 days, and then cardiac function, myocardial pathology, serum cardiac markers CKMB and cTnI, and inflammatory factors TNF-α and IL-6 in myocardium were evaluated, respectively. In vitro, the injury model of H9C2 cardiomyocytes was induced by oxygen-glucose deprivation/reperfusion (OGD/R). Cell viability, LDH, TNF-α and IL-6 in cell supernatant were detected. The expression levels of lipoxygenases pathway related proteins ALOX5, ALOX12 and ALOX15 was detected by western blotting both in vivo and in vitro.
RESULTS The results of echocardiography and serum cardiac markers showed that LJE improved cardiac function and relieved the impairment of myocardial tissue after MI. Additionally, LJE treatment could reduce inflammation levels in myocardial tissue. Moreover, lipoxygenases pathway was activated after MI and it was inhibited by the treatment of LJE with the significant decrease of ALOX5. In vitro, LJE treatment improved the cell viability and reduced the release of LDH, TNF-α and IL-6. Moreover, the protein expression levels of ALOX5, ALOX12 and ALOX15 were significantly decreased with LJE treatment.
CONCLUSIONS LJE improves cardiac function and ameliorates myocardial injury post MI by inhibiting inflammation via regulating lipoxygenases pathway.
GW34-e1370
Hailin Zhang, Lishan Zeng, Dajun Chai
The First Affiliated Hospital, Fujian Medical University
OBJECTIVES Ischemic cardiomyopathy (ICM) is the leading cause of cardiovascular disease. Recent studies have shown that ketone bodies not only act as a “super fuel” for the heart, but also inhibit adverse cardiac remodeling by improving vascular endothelial function, reducing oxidative stress, and regulating inflammation. Autophagy is an important means of self-repair after injury, and cardiomyocyte autophagy levels are reduced after myocardial infarction. It is unclear whether ketone bodies can improve the structure and function of the ICM rat heart by increasing cardiomyocyte autophagy levels.
METHODS 1,3-Butanediol (1,3-Butanediol, 1,3-BD) is a precursor for ketone body metabolism and can be converted to β-hydroxybutyric acid (β-hydroxybutyric acid, β-HD) in the liver. β-HD is not only an energy substrate but also an important molecule for cell signaling. In this study, an ICM rat model was established by continuously ligating the left anterior descending coronary artery branch of standard SD rats for 5 weeks, and the non-infarcted rats were randomly divided into control, ICM, and 1,3-BD intervention groups, with 15 rats in each group. In the 1,3-BD group, 1,3-BD (1 mL/100 g) was administered by gavage daily for 4 weeks. The effects of 1,3-BD on cardiac structure and function and myocardial pathological damage in ICM rats were evaluated by cardiac ultrasound, hemodynamics, hematology, and histopathology at week 5 after left anterior descending ligation. Changes in mitochondrial ultrastructure and autophagy levels in myocardial tissue at the infarct border were observed by transmission electron microscopy and laser confocal microscopy, and possible regulatory mechanisms were investigated in combination with transcriptomic and metabolomic analyses and validated by immunoblotting.
RESULTS After 5 weeks of continuous ligation of the left anterior descending branch of the coronary artery, both cardiac ultrasound and invasive hemodynamic monitoring showed significantly enlarged left ventricular chambers and significantly reduced systolic and diastolic functions in the rats in the ICM group. 1,3-BD intervention significantly improved cardiac structure and function. Left ventricular ejection fraction: 50.39±5.94% vs 71.92±6.08% (both P<0.05). Compared with the rats in the ICM group, HE staining of the rats in the 1,3-BD group showed a reduction in the left ventricular chambers, wall thickening, and inflammatory cell aggregation, and Masson staining showed reduced collagen fiber deposition in myocardial tissue. Mitochondrial transmission electron microscopy confirmed that 1,3-BD significantly improved the structure and number of myocardial mitochondria in the marginal zone, and autophagic vesicles were significantly increased. Extracted proteins from myocardial cells in the marginal zone were analyzed by immunoblotting, and compared with the Ctr group, the levels of ATG7, ATG13, beclin1, SQSTM1/p62, ULK1 and LC3II/LC3I proteins were significantly decreased in the ICM group, and autophagy-related protein expression was significantly upregulated after 1,3-BD intervention; meanwhile, PI3K, AKT and FOXO3 proteins were significantly downregulated in the ICM group compared with the Ctr group, and 1,3-BD intervention significantly reversed the expression of PI3K, AKT and FOXO3 proteins, suggesting that 1,3-BD may enhance autophagy in injured cardiomyocytes through upregulation of PI3K/AKT/FOXO3 pathway.
CONCLUSIONS 1,3-BD significantly improved cardiac structural function, attenuated myocardial pathological injury and mitochondrial damage in ICM rats, and could upregulate myocardial autophagy levels in the limbic zone through activation of PI3K/AKT/FOXO3 signaling pathway. Exogenous supplementation of 1,3-BD not only provides richer energy support to the ischemic heart, but also improves the poor prognosis of ICM by enhancing autophagy of injured cardiomyocytes through ketone body β-Hb. Appropriate ketone body supplementation may be a potentially effective means of treating ICM.
GW34-e1372
Bingyu Zheng
The First Affiliated Hospital of Nanjing Medical University
OBJECTIVES Tyrosine kinase inhibitors (TKIs), a type of targeted therapy for non-small cell lung cancer, were reported to cause QT prolongation. In our clinical practice, we found that crizotinib administration increases the risk of QT prolongation. The aim of this study was to characterize the mechanism behind.
METHODS By using intracellular calcium indicator and voltage sensitive dye, we tested the effect of crizotinib treatment on electrophysiology of human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs). The effect of crizotinib administration In vivo was investigated in adult zebrafish. qRT-PCR and Western Blot were applied for detection of ion channels expression. With mass spectrometry-based proteome and phosphoproteome analysis, molecular mechanism of crizotinib induced QT prolongation was revealed.
RESULTS Up to 1 μM crizotinib treatment did not show toxicity to hiPSC-CMs viability 24 hours of crizotinib treatment led to prolonged action potential duration (APD90) compared with control in 1 Hz paced hiPSC-CMs. However, the calcium transients between two groups were of no significant difference. Adult zebrafish treated with 10 μM crizotinib presented prolongated QT intervals compared with control. qRT-PCR of ion channels involved in repolarization revealed that KCNH2 expression level was reduced in crizotinib exposure group. In addition, Cell membrane hERG expression were significantly decreased in crizotinib-treated group. Phosphorylated protein quantification combined with total protein quantification analysis suggested reduced histone acetyltransferases p300 phosphorylation. To study the relation between p300 and hERG expression, we used p300 overexpression plasmid, p300 siRNA and p300 inhibitor A-485 in human cardiomyocytes AC-16. Western blot showed that overexpression of p300 increased the cytomembrane hERG, while siRNA and A-485 reduced it. Further co-Immunoprecipitation demonstrated the interaction between p300 and hERG. Moreover, overexpression of p300 increased the hERG acetylation, decreased the hERG ubiquitination, thus reversed the crizotinib induced hERG degradation, while siRNA and inhibitor A-485 had the opposite effect.
CONCLUSIONS Previous studies have showed that crizotinib led to blockage of hERG. Our study provided a new prospective to explain crizotinib induced cytomembrane hERG down-regulation.
GW34-e1384
Xiuxiang Liu, Jie Liu
Key Laboratory of Arrhythmias of the Ministry of Education of China, Research Center for Translational Medicine, Shanghai Heart Failure Research Center, Department of Cardiovascular Surgery, Shanghai East Hospital, Tongji University School of Medicine, Shanghai 200120, China
OBJECTIVES Diabetes mellitus (DM) is currently a main public health problem, one of its complications is diabetic cardiomyopathy (DC), which is recognized as a microvascular disorder that leads to morbidity and mortality in diabetic patients. Endothelial cells play an important role in the maintenance of cardiac homeostasis and function, Transcription factor Klf11 highly expressed in endothelial cell and plays a protective role in many cardiovascular diseases, but the detailed effects of endothelial KLF11 in DC has not been reported. This study aimed to explore the role of endothelial cell KLF11 in the pathogenesis of DC and the underlying mechanisms.
METHODS Analysis of GEO profiles in hearts tissues from both Streptozotocin (STZ) induced diabetes and db/db background diabetes mouse models. Mouse model of DC was generated using a high-fat diet combined with a low-dose STZ injection (110 mg/kg) in vivo. mCMVECs cells with siRNA-KLF11 or siRNA-NOX2 were used to study endothelial cell KLF11 function in vitro. DHE staining was used to evaluate oxidative stress levels, NO Elisa kit was used to measure cultured medium Nitric Oxide concentration, qRT-PCR and western blot to further explore the mechanisms of KLF11 regulating oxidative stress through the NOX2/eNOS/NO pathway to protect diabetic cardiomyocyte injury. Coculture siRNA-KLF11 or siRNA-NOX2 cultured medium with MCM cell line were exposed in high glucose (HG) 25 mM to study cardiomyocyte hypertrophy in vitro. NOX2 knockout mice was generated by CDH5-specific nanoparticles loaded NOX2 gRNA Cas9 plasmids to rescue the phenotype of diabetic cardiomyopathy.
RESULTS Analysis of GEO profiles revealed that Klf11 is downregulated in hearts tissues from both Streptozotocin (STZ) induced diabetes and db/db background diabetes mouse models. Mouse model of DC was generated using a high-fat diet combined with a low-dose STZ injection (110 mg/kg) in vivo, we found significantly downregulated Klf11 expression in cardiac ECs accompanied by increased expression of oxidate stress genes (Cybb) in cardiac tissues and cardiac hypertrophy in T2DM mouse compared with sham operated mice. Moreover, we found significantly elevated Klf11 expression in mouse cardiac microvascular endothelial cells (mCMVECs) when exposed to high glucose (25 mM). Knockdown of Klf11 by siRNA in mCMVECs showed a significantly increased oxidative stress by DHE staining accompanied by decreased nitric oxide (NO) concentration in the medium upon high glucose exposure compared with scramble-siRNA transfected mCMVECs. Mechanistic study revealed a significantly upregulated NADPH oxidases 2 (Nox2) expression in mCMVECs upon Klf11 siRNA-knockdown. NOX2 converts oxygen into the superoxide anion which further reacts with NO to form perxynitrite (ONOO-). Perxynitrite is the main cause of eNOS uncoupling in ECs which leads to produces superoxide instead of NO and causing oxidate stress in ECs. Further study showed that Nox2 knockdown by siRNA significantly reversed increased oxidative stress and downregulated NO secretion upon Klf11 knockdown in mCMVECs with high glucose exposure as well as reduced the conditioned medium stimulated mouse cardiomyocytes (MCMs) hypertrophy. Finally, we generated a nanoparticle combined with cdh5 promoter driven Nox2 gRNA Cas9 plasmid to achieve ECs targeted Nox2 deletion in vivo. The results showed that EC-specific Nox2 knockdown protected against STZ induced ECs oxidative stress and CMs hypertrophy.
CONCLUSIONS These data demonstrate that KLF11 is a novel transcriptional suppressor of Nox2 in endothelial cells, constituting a potential molecular target for treatment of DC.
GW34-e1388
Lishan Zeng
The First Hospital, Fujian Medical University
OBJECTIVES This study investigates the mechanism by which the nuclear receptor LXR agonist T0901317 improves the development of calcific aortic valve disease (CAVD) through in vitro and in vivo experiments in mice, and proposes to demonstrate that T0901317 inhibits the osteogenic phenotype of aortic valve mesenchymal cells by regulating endoplasmic reticulum stress (ERS), thereby delaying the development of CAVD, and provides a new theoretical basis for further understanding of CAVD, as well as a new auxiliary assessment index and intervention target for the prevention and treatment of the disease.
METHODS Primary human aortic valve interstitial cells (hAVICs) were used to induce osteogenesis by giving calcification medium (OM), with comparison of T0901317. Immunofluorescence staining was used to detect the expression of the osteogenic markers Runx2 and OPN as well as the ERS marker PERK. Immunoblotting was used to detect the expression of proteins of the PERK-eIF2α-ATF4-Runx2 pathway. Alizarin staining was used to observe calcium salt deposition and calcium nodule formation. ApoE−/− mice were used to establish a CAVD model by high-fat diet (HFD) feeding for 24 weeks, with comparison of T0901317 oral administration. Ultrasound imaging system was used to assess the functional changes of aortic valve and heart structure. HE staining, Masson staining and Von Kossa staining were used to observe the thickness of aortic valve leaflets, degree of valve fibrosis, calcium salt deposition and calcium nodule formation. Immunofluorescence staining was used to detect the expression of Runx2, OPN and PERK.
RESULTS Immunofluorescence staining results showed that the expression of the OM component bone markers Runx2 and OPN as well as the ERS marker PERK was upregulated compared to the CTL group, while T0901317 intervention induced downregulation of Runx2, OPN and PERK expression. Immunoblotting results showed that the expression of PERK-eIF2α-ATF4-Runx2 pathway protein was increased in the OM group compared with the CTL group, while T0901317 intervention led to a decrease in the expression of this pathway protein. Alizarin staining showed that a large amount of calcium salt deposition and calcium nodule formation were seen in the OM group compared to the CTL group, while T0901317 intervention significantly improved these conditions. Echocardiographic results showed a significant increase in aortic orifice flow velocity and transvalvular pressure difference in the HFD-induced CAVD mice compared with the normal, drug control and model control groups, which was largely reversed in the T0901317 intervention group. After T0901317 treatment, increased valve fibrosis, calcium salt deposition and calcium nodule formation were improved, meanwhile the expression of Runx2, OPN and PERK was significantly reduced by immunofluorescence staining.
CONCLUSIONS LXR agonist T0901317 inhibits the transformation of human aortic valve interstitial cells to an osteogenic phenotype. T0901317 improves the ERS response in human aortic valve interstitial cells and slows the osteogenic response in human aortic valve interstitial cells via the PERK-eIF2α-ATF4-Runx2 pathway. T0901317 protects aortic valve structure and function in CAVD mice by a mechanism that may be related to the inhibition of osteogenic transition and ERS.
GW34-e1389
Poli Lin1, Ping Ren1, Boya Cao1, Jiali Chen2, Ping He3, Junling Cao4, Jian Zhang3
1School of Chinese Materia Medica, Beijing University of Chinese Medicine
2School of Chinese Medicine, Beijing University of Chinese Medicine
3School of Life Sciences, Beijing University of Chinese Medicine
4Dongfang Hospital, Beijing University of Chinese Medicine
OBJECTIVES Scrophulariae Radix (known as xuanshen, XS), is a traditional Chinese medicine which has strong anti-inflammatory effects and is widely used in the formulation of cardiovascular diseases. Toll-like receptor 4 (TLR4) pathway is an important inflammatory response pathway and a key regulator of the initiation and spread of inflammation after acute myocardial ischemia (AMI). This study aims to explore and verify the pharmacological function and the mechanism of XS and its effect component in regulating TLR4 pathway on AMI.
METHODS First, the components that may act on TLR4 target in XS were screened by network pharmacology, and further identified by molecular docking technology. Then, the efficacy of XS and the screened component on AMI was evaluated based on AMI mouse model which was established by 7 days left coronary artery ligation and hypoxia-inflammation compound induced H9C2 cardiomyocyte model which was treated by 1 μg/mL lipopolysaccharide and incubated with oxygen deprivation for 8 h. XS and Asn were given orally by gavage. Carvedilol and TAK-242, the TLR4 inhibitor, were given as positive control. Then, the regulation of XS and the screened component on proteins of TLR4 pathway was determined by western blot assay, including TLR4, nuclear factor kappa B (NF-κB), NOD-like receptor thermal protein domain associated protein 3 (NLRP3), gasdermin D (GSDMD) and interleukin-1β (IL-1β).
RESULTS By network pharmacology, L-Asparagine (Asn) was predicted as the potential effect component of XS in regulating TLR4 pathway. The result of molecular docking showed that Asn could bind to TLR4 and its co-receptor protein respectively by hydrogen bonds. XS and Asn inhibited the dilatation of the left ventricle and alleviated acute ischemic injury, including a significant increase of ejection fraction and fractional shortening, a significant decrease of creatine kinase MB and cardiac Troponin I in serum, and collagen volume fraction. Meanwhile, XS and Asn protected cardiomyocytes against inflammation damage through reducing inflammatory cell infiltration and the expression levels of tumor necrosis factor-α and interleukin-6. Furthermore, XS could significantly decrease the protein levels of TLR4, NLRP3, GSDMD and IL-1β in AMI mice. Asn could significantly decrease the protein levels of TLR4, NF-κB, NLRP3 and IL-1β in AMI mice. In vitro, XS and Asn significantly protected H9C2 cardiomyocyte against hypoxia and inflammatory injury. Moreover, XS could significantly decrease the protein levels of TLR4 and NF-κB in hypoxia-inflammation induced cell model. Asn could significantly decrease the protein levels of TLR4, NF-κB and NLRP3 in hypoxia-inflammation induced cell model.
CONCLUSIONS XS is effective in alleviating myocardial ischemic injury, improving cardiac function and inhibiting the excessive inflammation, possibly by regulating TLR4 pathway. Asn is the potential effect compound of XS and also could alleviate myocardial ischemic injury and inhibit the activation of TLR4 and its downstream proteins to inhibit the inflammation.
GW34-e1393
Yuangengshuo Wang, Wenyao Wang, Chunli Shao, Chen Li, Yitian Zheng, Yida Tang
Department of Cardiology and Institute of Vascular Medicine, Peking University Third Hospital
OBJECTIVES Calcific aortic valve disease (CAVD) is a structural heart disease with a high incidence and mortality rate but limited pharmacological treatment. Clinical studies have found that patients with CAVD exhibit a significant reduction of local thyroid hormone concentration, suggesting that thyroid dysfunction may play an essential role as an inducement factor in the occurrence and progression of CAVD. The purpose of this study is to explore the relationship and underlying mechanisms between thyroid function and CAVD.
METHODS In this study, we use both in vivo and in vitro models to investigate the role of thyroid hormone in CAVD development. C57BL/6J mice aged 10–12 weeks were randomly divided into four groups (n=6). The aortic valves in the surgical group were directly injured by a spring wire under the guidance of echocardiography. The T3 group was treated with triiodothyronine (5 μg/g·d) after surgery for 8 weeks. The PTU group was treated with 0.025% propylthiouracil (PTU) after surgery for 8 weeks. Sham surgery was performed similarly but without wire insertion into the left ventricle. Additionally, different concentration gradients of thyroid hormone were employed as interventions to investigate their impact on the osteogenic differentiation process in valvular interstitial cells. The EF, FS, aortic valve peak velocity, and effective orifice area (EOA) were recorded before and every 4 weeks after the operation. Von Kossa staining and Alizarin red staining were used to estimate the degree of calcification. qRT-PCR, Western blot, and Opal multiplex immunofluorescence were used to measure the RNA and protein expression levels.
RESULTS Compared to the sham-operated group, mice with wire-induced injury to the aortic valve demonstrated a significant increase in aortic valve peak velocity and a decrease in EOA. Among them, the T3 group had a larger EOA than the sham-operated group, more significantly than the PTU group. Results from Von Kossa staining, Sirius Red staining, and alkaline phosphatase staining indicated that the extent of valve calcification was more severe in the PTU group than in the sham-operated group and greater in the sham-operated group than in the T3 group. These results suggested that decreased thyroid hormone levels accelerated valve calcification and remodeling. Mechanistically, low thyroid hormone level decreased the expression of Med1 in vitro. Thyroid hormone deficiency can impair BMP/TGF-β signaling pathway transduction by affecting Med1 expression, which is associated with calcific aortic valve disease (CAVD) progression.
CONCLUSIONS Low thyroid hormone concentration is a significant inducement factor for CAVD. It can impede valvular interstitial cells’ repair function and promotes valve calcification and structural remodeling. This process may be achieved by affecting Med1 expression levels and thus impairing BMP/TGF-β signaling pathway transduction. Supplementation of thyroid hormones helps maintain normal valvular interstitial cell function and inhibits osteogenic differentiation.
GW34-e1398
Ruimin Liu
Beijing Anzhen Hospital, Capital Medical University
OBJECTIVES Obesity related cardiomyopathy is a worldwide health problem caused by lipid accumulation in the heart. Chemerin, an adipocyte-secreted protein, has been recently suggested to be linked to metabolic syndrome and cardiac function in obese and diabetes mellitus. This study aimed to investigate the potential roles of adipokine chemerin on high fat-induced cardiac dysfunction.
METHODS Six to eight-week-old male Rarres2 −/− mice and WT littermates were studied. Mice were fed with normal diet or high fat diet for 20 weeks in a 12-hour cycle environment. For calorimetric analyses, mice were acclimated to metabolic cages 1 week after fed with 16 weeks of HFD. Heart function was measured by echocardiography, HE staining and Masson trichrome staining were used for observing cardiac morphology and collagen deposition.
RESULTS Firstly we found that an increase of serum chemerin in HFD mice, which is mainly released in adipose tissues, induced CMKLR1 to express its primary receptor. This early occurrence may protect against cardiomyopathy brought on by obesity. Next, chemerin knockout mice were created to confirm this notion. When they were fed a high-fat diet, it was shown that the Rarres2 knock further generated lipotoxicity, which caused inflammation, glucose/insulin resistance, and cardiac dysfunction. Nevertheless, these abnormalities above were not observed in Rarres2 −/− mice under normal-diet feeding. Thirdly, using lipid-overloaded cardiomyocytes, we observed that chemerin supplementation lowered lipid accumulation, insulin resistance, and inflammation. Furthermore, knocking down the CMKLR1 gene could abolish these protective effects of chemerin in lipid-overloaded cardiomyocytes. Herein, our data demonstrate that adipocyte-derived chemerin functions as a critical protective regulator of cardiac function in the obese/or pre-diabetic heart.
CONCLUSIONS Together, these data obtained demonstrate that adipocyte-derived chemerin functions as an endogenous cardioprotective factor in the presence of obesity, alleviates lipid overload-induced lipotoxicity, insulin resistance, inflammation and improves cardiac dysfunction, it therefore indicates that chemerin and as a novel potential treatment approach for the obese related cardiomyopathy.
GW34-e1404
Chao Yang1,2, Yuxing Chen1,2, Xinyang Hu1,2, Wei Zhu1,2, Jian’an Wang1,2,3
1Department of Cardiology of the Second Affiliated Hospital, School of Medicine, Zhejiang University, Hangzhou, China
2Cardiovascular Key Laboratory of Zhejiang Province, Hangzhou, China
3Binjiang Institute of Zhejiang University (Research Center for Life Science and Human Health), Hangzhou 310053, China
OBJECTIVES The therapy of left ventricular mechanical unloading, such as transcatheter aortic valve replacement (TAVR) and left ventricular assist devices (LVAD) has significantly improved the prognosis for patients with heart failure. However, the detailed cellular and/or molecular mechanisms of recovery of cardiac function remain largely unknown. We aimed to build a cellular atlas of the heart that experienced stress and de-stress, focusing on myofibroblasts, precisely its cell fate of differentiation and function.
METHODS We conducted single-cell sequencing, ATAC sequencing on non-myocardial cells from mouse heart samples of pressure overload-induced heart failure and recovery, using TAC (transverse aortic arch constriction)/deTAC (with suture for the constriction removed 2 weeks after TAC) mouse model to identify the dynamic changes. The lineage tracing technique was also used to gain further insight into cell fate and reciprocal interactions. During this process, the macrophage subgroup was cleared using various approaches to investigate the cross-talk between myofibroblast and macrophage.
RESULTS We successfully generated a TAC/deTAC mouse model, which can faithfully simulate a reversal process of functioning for the heart that underwent a pressure overload stress (a hypertrophy process). Mouse multi-omics sequencing reveals the genetic changes, spatial distribution, and transcriptional regulation of cell populations at different stages. Among the different cell types, significant changes in the gene expression pattern were observed in fibroblasts. Aided by myofibroblast lineage tracing, flow cytometry examinations suggested that remaining in a quiescent state rather than showing a decrease in its number appeared as the central fate of myofibroblasts that accompanied the reversal of cardiac function when the over-loading was relieved. Surprisingly, however, simultaneous pressure unloading and macrophage clearance significantly reduced the number of myofibroblasts.
CONCLUSIONS With a reliable TAC/deTAC mouse model, our multi-omics data revealed dynamic changes in cell populations of the heart that underwent pressure overload stress followed by unloading; our data demonstrated a quiescent state rather than a reduction in number was the significant fate for the activated myofibroblasts for which macrophages were involved. These findings unveiled some novel mechanisms that had never been described before.
GW34-e1405
Xianbao Liu, Jinyong Chen, Jian’an Wang
The Second Affiliated Hospital, Zhejiang University School of Medicine
OBJECTIVES Calcific aortic valve disease (CAVD) poses a serious threat to the health of older adults, yet no available drugs have proven effective against its progression. A proper drug delivery system can be a potential solution to CAVD drug therapy. However, due to the rapid blood flow rate associated with aortic valve stenosis and the lack of specific markers, targeted drug delivery for CAVD are hard to achieve.
METHODS The expression of protease-activated-receptor-2 (PAR2) was analyzed, and a magnetic nanocarrier functionalized with PAR2-targeting hexapeptide and loaded with XCT790 for dual-active targeting therapy was developed. A parallel-plate flow chamber was designed and a mouse CAVD model was established to evaluated the targeting curative ability of the nanoplatform in vitro and in vivo.
RESULTS PAR2 expression in human CAVD samples and osteogenically differentiated valvular interstitial cells (VICs) was increased. Intravenous injection of our nanocarriers led to significantly higher enrichment within the calcified valves under magnetic-field guidance compared to those without PAR2-targeting peptides. Loaded with XCT790, an anti-calcification drug, our nanocarrier effectively inhibited osteogenic differentiation of VICs, and alleviated aortic valve calcification and stenosis in mouse model. Mechanical study revealed the pyruvate dehydrogenase kinase 4 (PDK4)-mediated metabolic reprograming is partly involved in the therapeutic effect of targeted nanoparticles on CAVD.
CONCLUSIONS This work presents the first effective targeted drug delivery system for the prevention and treatment of CAVD in induced animal model to the best of our knowledge. The composite drug delivery platform combining PAR2- and magnetic-targeting features may represent a promising therapeutic strategy for CAVD.
GW34-e1406
Ying Lin, Qifan Yang, Xiaoping Lin, Xianbao Liu, Wei Zhu, Jian’an Wang
The Second Affiliated Hospital, School of Medicine, Zhejiang University
OBJECTIVES Extracellular matrix (ECM) is closely involved in aortic valve development and interrupted ECM maturation contributes to aortic valve anomaly. Several ADAMTS (A Disintegrin and Metalloproteinase with Thrombospondin motifs) gene family members have been reported to be related to or cause aortic valve diseases. Therefore, we aimed to test whether the other ADAMTS gene family members also involved in the aortic valve diseases.
METHODS Morpholino based approach was applied for targeted gene phenotype-screening the outflow tract anomaly in zebrafish and the 19 ADAMTS family genes exon Sanger sequencing was carried out for a cohort BAV registry study (with 304 BAV patients and 301 controls with normal cardiac valve structure). Adamts gene specific genetic tracing mouse models were generated to evaluate the spatiotemporal expression pattern in the aortic valve, and different gene modified mouse models (including mutant gene knock-in and conditional gene knock-out) were generated using the CRISPR/Cas9 method to evaluate their roles in aortic valve development and valvular pathogenesis. Movat’s pentachrome staining was used to describe the altered ECM maturation process in cardiac valves. Bulk RNA-seq was also performed based on both genetic mouse heart tissue and iPSC induced endothelial cells generated from BAV patient peripheral blood cells to explore the potential molecular pathways, which was further confirmed in ex vivo embryonic outflow tract tissue culture study.
RESULTS We identified some novel ADAMTS genes that can be involved in cardiac outflow valve development, exon sequencing identify ADAMTS16 p.H357Q variant in an inherited BAV family. Therefore, the Adamts16-Cre; Rosa26-tdTomato mouse model was generated which showed Adamts16-expressing cells were primarily restricted to valvular endothelial cells (VECs) at E12.5 and then started to be expressed in valvular mesenchymal cells (VICs) at E13.5. The CRISPR/Cas9 based Adamts16 +/ - and Adamts16+/H355Q mice both exhibited an R-NC-type BAV phenotype, with progressive aortic valve thickening, whereas conditional Adamts16 knockout mouse models demonstrated that Adamts16 deficiency mainly in endothelial lineage cell recapitulated the BAV phenotype. ADAMTS16 deficiency demonstrated worse ECM deposition and loss of the ECM trilaminar structure. Bulk RNA sequencing using iPSC-induced endothelial cells (iPSC-ECs) and genetic mouse embryonic tissue unveiled enhanced focal adhesion signaling, which was accompanied by elevated fibronectin levels. Both in vitro iPSC-ECs culture and ex vivo embryonic tissue explant studies validated the altered focal adhesion signaling.
CONCLUSIONS Our present study identified a novel BAV-causing ADAMTS16 p. H357Q variant. ADAMTS16 deficiency led to BAV formation.
GW34-e1411
Pengqi Lin1, Quanwei Pei1, Bin Li1, Jiemei Yang2, Lina Zou1, Dezhan Su1, Junpei Zhang1, Hongpeng Yin1, Mbabazi Nadine1, Junjie Yang1, Zhenwei Pan3, Jingjie Li1, Dechun Yin1
1Department of Cardiology, The First Affiliated Hospital of Harbin Medical University, Harbin, China
2Department of Echocardiography, The First Affiliated Hospital of Harbin Medical University, Harbin, China
3Department of Pharmacology (State-Province Key Laboratories of Biomedicine-Pharmaceutics of China, Key Laboratory of Cardiovascular Medicine Research, Ministry of Education), College of Pharmacy, Harbin Medical University, Harbin, China
OBJECTIVES Current stress cardiomyopathy (SCM) theories revolve mostly around “catecholamine myocardial toxicity” and “sympathetic hyperactivation”. However, the role of the central nervous system (CNS) in the pathogenesis of stress cardiomyopathy remains unknown. Microglia can secrete glial transmitters in the central nervous system to support and trophic neurons and participate in the regulation of neural activities. The paraventricular nucleus of the hypothalamus is the central sympathetic outflow tract. In this study, we investigated the role of microglial activation in the paraventricular hypothalamic nucleus (PVN) in the development of SCM.
METHODS To create a SCM model, male Sprague-Dawley (SD) rats were immobilized under stress for 6 hours every day for 7 days. RNA sequencing, ventricular electrical stability, left ventricular morphology, and sympathetic nerve activity were measured. In addition, brain tissues were extracted to detect microglial activation and inflammatory cytokine expression.
RESULTS (1) RNA sequencing analysis showed that functions of differentially expressed genes were significantly enriched in neuroinflammatory pathway. (2) Microglial were significantly activated in PVN, as evidenced by an increase in their number as well as axon process length and branching. (3) In SCM, microglial in the PVN increased central sympathetic outflow by increasing the expression of inflammatory factors. (4) It is possible that inhibiting microglial activation could suppress central inflammation and increase the electrical stability of the heart in SD rats with SCM.
CONCLUSIONS Inhibiting activated microglial in the PVN possibly reduce ventricular fibrillation vulnerability in SCM rats by central neuroinflammatory pathway. These findings suggest that microglial are a potential target for sudden cardiac death prevention in SCM.
GW34-e1415
Lihong Yao, Mengkang Fan, Hongzhuan Sheng
Affiliated Hospital of Nantong University
OBJECTIVES Betaine, as a non-hazardous natural plant compound found in beetroot and seafood, acts as an osmolyte and a methyl donor. Betaine has been shown to have a variety of benefits in a variety of human diseases. However, its role in cardiac hypertrophy is unclear. Our aim is to investigate the role and regulatory mechanism of betaine in occurrence of pressure overload-induced cardiac hypertrophy.
METHODS C57 BL/6 mice were used for preparing pressure overloaded-induced cardiac hypertrophy model in vivo by TAC (transverse aortic constriction) surgery, and neonatal rat cardiomyocytes (NRCMs) were used for preparing cardiac hypertrophy model in vitro by phenylephrine (PE) stimulation. Adenovirus containing shRNA-HCAR1 (ADV-shHCAR1) were constructed to knockdown HCAR1 in NRCMs. The surface area of cardiomyocytes was detected by HE staining, WGA staining and actin-tracker staining, and the level of cardiac fibrosis was detected by masson staining. ELISA kit was used to determine the level of cAMP, real-time quantitative PCR was used to detect the level of cardiac hypertrophy related markers (ANP, BNP, MHC), and western blot was used to detect the changes of MHC and Hippo signaling pathway related molecules. Binding interaction between HCAR1 and betaine was predicted by AutoDock 4 and further validated with RNA-seq analysis.
RESULTS In vivo experiments revealed that betaine administration of male C57BL/6 mice for 4 weeks after transverse aortic constriction preserved ejection fraction and decreased the left ventricular posterior wall thickness, heart weight/tibia length, cross-sectional area and cardiac fibrosis area. Computer molecule docking screening analysis identified betaine as a potential ligand of HCAR1. Treatment with betaine significantly attenuated PE-induced hypertrophy in NRCMs, which was alleviated by transfection of ADV-shHCAR1. Transcriptome sequencing analysis and probative experiment proved that betaine took effects via regulating HCAR1/Hippo signaling pathway.
CONCLUSIONS 1. Betaine attenuated pressure overload-induced cardiac hypertrophy by inhibiting HCAR1 activation and activated Hippo signaling pathway. 2. Betaine was a candidate for the treatment of cardiac hypertrophy.
TRANSLATIONAL RESEARCH OF CARDIOVASCULAR DISEASE
GW34-e0093
Yusi Chen
The Second Xiangya Hospital of Central South University
OBJECTIVES Idiopathic pulmonary arterial hypertension (IPAH) is a potentially fatal pulmonary vascular disease with an extremely poor natural course of IPAH. The limitations of current treatment and the unclear etiology and pathogenesis of IPAH require new targets and avenues of exploration involved in the pathogenesis of IPAH. tRNA-derived small RNAs (tsRNAs), a new type of small noncoding RNAs, have a significant part in the progress of diverse diseases. However, the potential role of tsRNAs in IPAH remains unknown.
METHODS Small RNA microarray was implemented on three pairs of plasma of IPAH patients and healthy controls to investigate and compare tsRNAs expression profiles. Validation samples were used for real-time polymerase chain reaction (Real-time PCR) to verify several dysregulated tsRNAs. miRanda and TargetScan were adopted to determine the potential target genes and mechanisms of the validated tsRNAs in IPAH by bioinformatic analysis.
RESULTS Microarray detected 816 statistically differentially expressed tsRNAs, of which 243 tsRNAs were upregulated and 573 were downregulated in IPAH. Eight validated tsRNAs in the results of Real-time PCR were concordant with the small RNA microarray: four upregulated (tRF3a-AspGTC-9, 5′tiRNA-31-GluCTC-16, i-tRF-31:54-Val-CAC-1 and tRF3b-TyrGTA-4) and four downregulated (5′tiRNA-33-LysTTT-4, i-tRF-8:32-Val-AAC-2, i-tRF-2:30-His-GTG-1, and i-tRF-15:31-Lys-CTT-1). The Gene Ontology analysis has shown that the dysregulated tsRNAs are related to cellular macromolecule metabolic process, regulation of cellular process, regulation of cellular metabolic process. It is disclosed that potential target genes of dysregulated tsRNAs are involved in PPAR signaling, BMPR2 signaling, and HIF-1 signaling significant to the pathogenesis of IPAH by Kyoto Encyclopedia of Genes and Genomes.
CONCLUSIONS This study investigated tsRNA profiles in IPAH and found that the expression of eight dysregulated tsRNAs may become a novel type of bio-diagnostic indicators and possible targets for IPAH.
GW34-e0212
Ziwei Zhu, Xiaowei Zhang
Lanzhou University Second Hospital
OBJECTIVES Myocardial infarction (MI) causes a significant contributor to morbidity and mortality worldwide. However, the molecular mechanism remains unclear. Cold-inducible RNA-binding protein (CIRP) acts as a damage-associated molecular pattern, raising critical implications for the pathobiology of inflammatory diseases. We explored the underlying mechanism of CIRP as MI therapeutic target that may help patients reduce the mortality rate.
METHODS CIRP and cTnI were highly expressed in patients with acute MI, and this elevation was confirmed in rats with acute MI compared with the Sham group. Rats were randomly assigned into 4 groups: sham group, sham-Cirbp−/− group, WT group, and Cirbp−/− group, and the last two groups underwent left anterior descending (LAD) ligation.
RESULTS Cardiac function and myocardial injury were significantly better in the Cirbp−/− group than in the WT group after acute MI. The degree of neutrophil infiltration and the expression levels of C5a, ICAM-1, MMP-9, IL-1β, IL-6, and TNF-α was lower in the Cirp−/− group than the WT group. Consistent results were obtained in evaluating the expression of inflammatory-related proteins in tissue samples and cell cultures.30 minutes after LAD ligation, C23, a competitive inhibitor of CIRP, was administered intravenously to evaluate the effect of C23 on MI and found that C23 can improve cardiac function and have an anti-inflammatory effect after MI.
CONCLUSIONS CIRP is one of damage-associated molecular patterns that aggravates the myocardial injury of acute coronary syndrome, and that the use of CIRP specific antagonist C23 can alleviate the myocardial injury and ventricular remodeling.
GW34-e0239
Zhen Wang
Department of Ultrasound Medicine, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology
OBJECTIVES Cardiac dysfunction is a well-recognized complication of sepsis and is associated with the outcome and prognosis of septic patients. Evidence suggests that IL-12A participates in the regulation of various cardiovascular diseases, including heart failure, hypertension and acute myocardial infarction. However, the effects of IL-12A in sepsis-induced cardiac dysfunction remain unknown. Hence, this study aimed to determine the role of IL-12A in sepsis-induced cardiac dysfunction and explore its underlying mechanisms.
METHODS In our study, lipopolysaccharide (LPS) and cecal ligation and puncture (CLP) model were used to mimic sepsis, and cardiac IL-12A expression was assessed. In addition, IL-12A knockout mice were used to detect the role of IL-12A in sepsis-related cardiac dysfunction.
RESULTS We observed for the first time that IL-12A expression is upregulated in mice after LPS treatment and macrophages were the main sources of IL-12A. In addition, our findings demonstrated that IL-12A deletion aggravates LPS-induced cardiac dysfunction and injury, as evidenced by the increased serum and cardiac levels of lactate dehydrogenase (LDH) and cardiac creatine kinase-myocardial band (CK-MB). Moreover, IL-12A deletion enhances LPS-induced macrophage accumulation and drives macrophages toward the M1 phenotype in LPS-treated mice. IL-12A deletion also downregulated the activity of AMP-activated protein kinase (AMPK) but increased the phosphorylation levels of p65 (p-p65) and NF-κB inhibitor alpha (p-IκBα). In addition, IL-12A deletion aggravates CLP-induced cardiac dysfunction and injury. Treatment with the AMPK activator AICAR abolishes the deterioration effect of IL-12A deletion on LPS-induced cardiac dysfunction.
CONCLUSIONS IL-12A deletion aggravated LPS-induced cardiac dysfunction and injury by exacerbating the imbalance of M1 and M2 macrophages. Our data provide evidence that IL-12A may represent an attractive target for sepsis-induced cardiac dysfunction.
GW34-e0303
Yan Zhao, Hangtian Yu, Angwei Gong, Shuaidan Zhang, Bing Xiao
Department of Cardiology, The Second Hospital of Hebei Medical University, Shijiazhuang, Hebei, People’s Republic of China
OBJECTIVES The causal relationship between heart rate variability and cardiovascular diseases and the associated events is still unclear, and the conclusions of current studies are inconsistent. We aimed to explore the relationship between heart rate variability and cardiovascular diseases and the associated events with the Mendelian randomization study.
METHODS We selected normal-to-normal inter-beat intervals (SDNN), root mean square of the successive differences of inter-beat intervals (RMSSD), and peak-valley respiratory sinus arrhythmia or high-frequency power (pvRSA/HF) as the three sets of instrumental variables for heart rate variability. The outcome for cardiovascular diseases included essential hypertension, heart failure, angina pectoris, myocardial infarction, non-ischemic cardiomyopathy, and arrhythmia. Cardiac arrest, cardiac death and major coronary heart disease event were defined as the related events of cardiovascular diseases. The data for exposures and outcomes were derived from publicly available genome-wide association studies. Inverse variance weighted was used for the main causal estimation. Analyses of heterogeneity and pleiotropy were conducted using the Cochran Q test of Inverse variance weighted and MR-Egger, leave-one-out analysis, and MR-Pleiotropy Residual Sum and Outlier methods.
RESULTS The Inverse variance weighted method indicated that genetically predicted pvRSA/HF was associated with the increased risk of cardiac arrest (odds ratio 2.02, 95% confidence interval 1.25–3.28, P=0.004). The results were free of heterogeneity and pleiotropy. There were no outliers and the leave-one-out analysis proved that the results were reliable.
CONCLUSIONS This study provides genetic evidence that pvRSA/HF is causally related to cardiac arrest.
GW34-e0353
Yang Zhang1,2, Xiaoman Wang1, Xunkai Li1, Shuangjie Lv1, Bo Yan3, Houzao Chen1, Xiaoqiang Tang4
1State Key Laboratory of Medical Molecular Biology, Department of Biochemistry & Molecular Biology, Institute of Basic Medical Sciences, Chinese Academy of Medical Sciences & Peking Union Medical College, Beijing 100005, China
2Department of Cardiology, Zhongshan Hospital, Fudan University, Shanghai Institute of Cardiovascular Diseases, National Clinical Research Center for Interventional Medicine, Shanghai 200032, China
3Institute of Precision Medicine, Jining Medical University, Jining, Shandong 272067, China
4Key Laboratory of Birth Defects and Related Diseases of Women and Children of MOE, State Key Laboratory of Biotherapy, West China Second University Hospital, Sichuan University, Chengdu 610041, China
OBJECTIVES The mechanisms underlying ageing-induced vascular remodelling remain unclear. This study investigates the role and underlying mechanisms of the cytoplasmic deacetylase sirtuin 2 (SIRT2) in ageing-induced vascular remodelling.
METHODS The expression of Sirtuins was analyzed by bulk RNA-seq, quantitative real-time PCR, western blot, and immunofluorescence staining. Male C57BL/6 WT and Sirt2-KO mice were used to analyze the effects of SIRT2 on ageing- and stress-induced vascular dysfunction. Vascular function was evaluated by pulse-wave velocity and constriction-relaxation of aortas. Vascular remodeling was analyzed by hematoxylin-eosin staining and vascular transcriptome was analyzed by bulk RNA-seq. Mechanism study was performed with western blot, immunoprecipitation, dihydroethidium and MitoSOX stainings. Finally, the SIRT2 coexpression module in aortas was analyzed for predicting the age-related aortic diseases in humans.
RESULTS SIRT2 level was the highest among the Sirtuins in human and mouse aortas. In aged mouse aortas, SIRT2 enzymatic activity and protein level were decreased. Functional and histological experiments demonstrated that knockout of Sirt2 aggravated dysfunction (increased stiffness and reduced constriction-relaxation function) of the aorta in aged mice, which was coupled with the remodeling of the vascular medial layer. Further transcriptome and immunofluorescence staining analyses revealed the potential contribution of mitochondrial reactive oxygen species (mROS) to SIRT2 roles in ageing-related vascular dysfunction. Mechanism studies showed that Sirt2 knockout led to hyperacetylation and hyperphosphorylation (activation) of the ageing-controlling protein p66Shc, which is one of the ringleaders of mROS. Scavenging mROS with MnTBAP repressed Sirt2-knockout-induced aggravation of vascular remodeling and dysfunction in aged or angiotensin II (Ang II)-infused mice. Finally, we found that the SIRT2 coexpression module in aortas was reduced with ageing across species and was a significant predictor of age-related aortic diseases in humans.
CONCLUSIONS The deacetylase SIRT2 is a response to ageing that delays vascular ageing, and the cytoplasm–mitochondria axis (SIRT2–p66Shc–mROS) is important for vascular ageing. Therefore, SIRT2 may serve as a potential therapeutic target for vascular rejuvenation.
GW34-e0370
Ru Ying, Cheng-Long Wu, Ran Yin, Jia-Ling Yin
Department of Cardiology, The First Affiliated Hospital of Nanchang University
OBJECTIVES Atrial fibrosis is important as an atrial fibrillation (AF) substrate. The purpose of our study was to evaluate the effect of Chemokine C-X-C motif ligand-1 (CXCL1) on atrial fibrosis and atrial fibrillation and the role of endoplasmic reticulum stress (ERS) and TXNDC5 plays in the process.
METHODS CXCL1 levels in peripheral blood, left atrial blood and right atrial blood of 16 control patients and 29 AF patients were evaluated. We evaluated the effect of CXCL1 on rat atrial fibrosis by Masson staining and expressions of Collagen 1, collagen 3, TGF-β, GRP78, ATF6 and TXNDC5 using an organo-culture system. CXCL1 effects on rat cardiac fibroblasts were also investigated, and the role of ERS and TXNDC5 was investigated by ERS inhibitors and TXNDC5 shRNA.
RESULTS Serum CXCL1 levels were increased in peripheral blood, left atrial blood and right atrial blood in AF group compared with the control group. In Exvivo, CXCL1 promoted fibrosis in rat atrial muscle, as well as upregulated GRP78, ATF 6, TXNDC5 expression and increased collagen 1, collagen 3 and TGF-β1 secretion. Both ERS inhibitor 4-phenylbutyrate (4-PBA) and TXNDC5 shRNA could reverse the pro-fibrotic effect of CXCL1 in rat cardiac fibroblasts.
CONCLUSIONS CXCL1 promote atrial fibrosis by activing ERS and TXNDC5, which may further contribute to the progression of atrial fibrillation.
GW34-e0483
Kun Li, Ping Zhang
Beijing Tsinghua Changgung Hospital
OBJECTIVES Facioscapulohumeral muscular dystrophy (FSHD) is one of the most common muscular dystrophies. Over 95% of FSHD cases are caused by deletion of the subtelomeric macrosatellite repeats (D4Z4) on human chromosome 4q35. Directly sequencing these repetitive regions is difficult owing to the high similarity among repeat units and high GC content. The existing FSHD1 diagnostic approaches, including Southern blot and Molecular combing, are not widely used for the reason of time-consuming, labor-intensive, and high equipment requirements. We aim to develop an efficient and accurate procedure for the diagnosis of FSHD.
METHODS Whole genome sequencing was performed for ten individuals from a large FSHD family. Parametric linkage analysis was performed via Merlin (v1.1.2) with the following parameters: dominant model, an estimated population allele frequency of 1E-5, and penetrance of 90%. 4q and 10q haplotypes were characterized by alignment to the chm13 reference genome. A BLAT of the pLAM sequence were performed to identify the A/B haplotypes. We realigned the 250-bp paired end whole genome sequencing reads to the chm13 reference genome using BWA-MEM and then measured the read count for reads containing either the 4q-specific D4Z4 sequence of 4q-specific pLAM sequence and normalized counts to the read depth of the p-arm of chromosome 4. Ultra-long read Nanopore long read genome sequencing were applied to genotype the pathogenic allele.
RESULTS Rare variants-based linkage analysis identified one single 1.7 MB haplotype on chromosome 4q35.2, presenting in affected individuals and absent from unaffected family members. Parametric linkage analysis resulted in a LOD score of 3.228 for the region. All pedigree samples contained 4q-pLAM sequence suggesting at least one copy of an 4qA permissive haplotype. Normalized counts of reads containing 4q-specific pLAM sequence were comparable between the FSHD patients, while 4q-specific D4Z4 repeat sequence demonstrated fewer reads in pedigree samples. Ultra-long read Nanopore sequencing of one affected individual detected a pathogenic FSHD allele containing a 4qA permissive haplotype and 5 D4Z4 repeats.
CONCLUSIONS Genome-wide rare variants-based linkage analyses is a powerful tool for the detection of pathogenic regions in family study. Ultra-long read Nanopore sequencing is capable of genotyping pathogenic FSHD1 alleles.
GW34-e0546
Yaohan Tang, Chenghui Yan, Yaling Han
National Key Laboratory of Frigid Zone Cardiovascular Disease, Cardiovascular Research Institute and Department of Cardiology, General Hospital of Northern Theater Command, Shenyang 110016, China
OBJECTIVES The cellular repressor of E1A stimulated genes 1 (CREG1) has been previously found to have anti-inflammatory and cardiac effects in our laboratory, but the active region of CREG1 is still unclear. This has become an obstacle to the translation of CREG1 into small molecule peptide drugs. The aim of this study is to identify the active region of CREG1 that plays an anti-inflammatory and cardioprotective role in AMI, and to obtain an effective fragment with a smaller molecular weight.
METHODS In this study, four peptides containing different domains of CREG1 were synthesized. These peptides or human CREG1 recombinant protein were added to the macrophage cell line RAW264.7 and stimulated by lipopolysaccharide (LPS) to induce inflammation model. The effect of the active fragment on macrophage migration was examined by a scratch assay. AMI mice were treated with the four peptides or the full-length CREG1 protein by subcutaneous implantation of an implantable osmotic sustaine-release pump. Cardiac function was assessed by small animal ultrasound 28 days later.
RESULTS A peptide containing the N-terminal 30–80 amino acids (NC51) of CREG1 protein inhibited the expression of IL-1β and IL-6 in a concentration-dependent manner and had no effect on the viability of macrophages. Treatment with NC51 peptide significantly reduced LPS-induced macrophage migration; NC51 could down-regulate iNOS expression and up-regulate CD206 expression. Compared with the MI group, the EF and FS% of the hearts in NC51 group and CREG1 group were significantly increased, and the heart weight to body weight ratio and heart weight to tibia length ratio were significantly decreased, and the area of myocardial infarction and fibrosis were also significantly reduced. NC51 group and CREG1 group also reduced the content of CKMB in the serum of mice, but had no significant effect on AST and ALT. In addition, NC51 and CREG1 reduced the infiltration of CD68-positive macrophages and the expression of IL-1β and IL-6 in the myocardial tissue of the infarct border zone, and also inhibited the expression of iNOS, but increased the expression of CD206. Compared with the CREG1 group, the NC51 group and the CREG1 group had no significant differences in the above indicators. Mechanically, NC51 plays an inhibitory role in the inflammatory response of macrophages through ANXA1-mediated entry and depends on Sirt1/NF-κB pathway.
CONCLUSIONS NC51 (N30-80aa), a small active polypeptide fragment of CREG1 recombinant protein, can protect against macrophage inflammatory response and cardiac function injury in mice after AMI. Mechanistic studies suggested that NC51 entered macrophages through ANXA1 and exerted anti-inflammatory effects dependent on Sirt1/NF-κB. NC51 peptide may be a potential therapeutic peptide drug for inhibiting inflammation and fibrosis after myocardial infarction and alleviating heart failure.
GW34-e0709
Tianwen Wei, Tiankai Shan, Jiawen Chen, Liansheng Wang
The First Affiliated Hospital of Nanjing Medical University
OBJECTIVES The regenerative capacity of adult mammalian hearts is limited. While numerous studies have explored mechanisms of cardiomyocyte cell-cycle withdraw, the regulative network is still complex. This study aimed to explore whether CHK1 could promote cardiac regeneration and repair in pigs after myocardial infarction and the specific mechanisms.
METHODS I/R injury was induced in pigs, recombinant human checkpoint kinase 1 (rhCHK1) protein (1 mg/kg) encapsulated in hydrogel or equivalent hydrogel was injected into the infarct border zone. The therapeutic efficacy of rhCHK1 was evaluated by echocardiography, cardiac magnetic resonance imaging (MRI), hematological indices and immunofluorescence. Multi-omics analysis were performed to explore the potential mechanisms of rhCHK1. Further, protein interaction prediction and CO-IP experiments were conducted to verify the specific interaction pattern and domains between CHK1 and PKM2. Hipsc-CMs and porcine models were used for confirming whether rhCHK1 promotes cardiac regeneration through combining with the PKM2 C-domain.
RESULTS RhCHK1 protein stimulated cardiomyocytes (CMs) proliferation and reduced cardiac inflammation response at 3 days post operation, improved cardiac function and attenuated ventricular remodeling, and reduced the infarct area at 28 days after I/R injury. Mechanistically, multiomics sequencing analysis showed that glycolysis and mTOR pathways were significantly enriched in this process. Further CO-IP experiments and protein docking prediction showed that CHK1 directly bound to and activated the S37 and Y105 sites of PKM2 to promote metabolic reprogramming. We further constructed plasmids that knock out different CHK1 and PKM2 domains, and transfected them into H293T cells for CO-IP experiments. It was found that the 391–476 domain of CHK1 directly binds to the 157–400 amino acids of PKM2. Further hipsc-CMs in vitro and pig in vivo experiments both demonstrated that CHK1 stimulates CMs renewal and cardiac repair by activating PKM2 C-domain-mediated cardiac metabolic reprogramming and mTORC1 pathway.
CONCLUSIONS This study demonstrates that rhCHK1 could combine with PKM2 391–476 amino acid domain to mediate metabolic reprogramming and mTORC1 pathways to promote cardiomyocytes proliferation and myocardial repair. This study is the first to clarify the specific functional mechanism of CHK1 in adult Bama pigs. Our results underscore the potential of the local applied CHK1 protein kinase as a safe and simple feasible treatment option for repairing heart.
GW34-e0758
Zhongyin Zuo1,2, Sainan Li3, Fengqi Xuan3, Jie Zhang3, Zichen Liu3, Shibei Zhang4, Ming Liang4,5, Zulu Wang4,5
1Department of Cardiology, General Hospital of Northern Theater Command of Jinzhou Medical University, Shenyang 110016, China
2Department of Cardiology, Shanghai Fourth People’s Hospital Affiliated to Tongji University School of Medicine, Shanghai 200080, China
3Department of Cardiology, General Hospital of Northern Theater Command of China Medical University, Shenyang 110016, China
4Department of Cardiology, General Hospital of Northern Theater Command, Shenyang 110016, China
5National Key Laboratory of Frigid Zone Cardiovascular Diseases, General Hospital of Northern Theater Command, Shenyang 110016, China
OBJECTIVES Catheter ablation (CA) of idiopathic ventricular arrhythmias (IVAs) originating from the left ventricular summit (LVS) and the interventricular septum pose a serious challenge due to anatomical constraints. However, a novel ablation technique with guidewire has emerged as a promising approach for mapping and ablation of these arrhythmias.
METHODS Ablation models were established to simulate blood vessels of different diameters (1.17 mm, 2.24 mm) in isolated swine myocardial tissue, and guidewire ablation (GA) was performed with different power of 10 W, 15 W, 20 W, and 25 W during 60 sec to determine the maximum safe power without steam pop occurring for each diameter group. Then with the maximum safe power, GA was performed on each group for different duration of 10, 20, 30, 40, 50, and 60 sec, and generator impedance (GI) changes, steam pop events as well as lesion diameters were recorded to elucidate the correlation between ablation duration and lesion size. Finally, GA was performed on in vivo swine heart and the animals was dissected at 48 hours and 8 weeks later to verify the feasibility of the maximum safe power and the lesion formation.
RESULTS Occurrence of steam pop increased along with the raise of ablation power in both two groups, and the maximum safe power was 10 W for the 1.17 mm group and 15 W for the 2.24 mm group. Under the specific maximum safe power, as the ablation duration prolonged, the increase in lesion diameter and decrease in GI were seen in both groups. There was a stronger correlation between GI drop and lesion diameter in group with smaller vessel diameter (1.17 mm-10 W group: maximum lesion diameter & GI drop, R = 0.965, minimum lesion diameter & GI drop, R=0.951; 2.24 mm-15 W group: maximum lesion diameter & GI drop, R=0.872, minimum lesion diameter & GI drop, R=0.888). In vivo experiment, besides VF happened once, no other adverse events were seen, and lesions were found at both 48-hour and 8-week groups.
CONCLUSIONS The vessel diameter, power and duration of ablation have effects on lesion formation. As the vessel diameter decreases, the safe power decreases as well. Additionally, there is a stronger correlation between GI drop and lesion diameter in smaller vessels.
GW34-e0759
Zhongyin Zuo1,2, Sainan Li3, Fengqi Xuan3, Jie Zhang3, Zichen Liu3, Shibei Zhang4, Ming Liang4,5, Zulu Wang4,5
1Department of Cardiology, General Hospital of Northern Theater Command of Jinzhou Medical University, Shenyang 110016, China
2Department of Cardiology, Shanghai Fourth People’s Hospital Affiliated to Tongji University School of Medicine, Shanghai 200080, China
3Department of Cardiology, General Hospital of Northern Theater Command of China Medical University, Shenyang 110016, China
4Department of Cardiology, General Hospital of Northern Theater Command, Shenyang 110016, China
5National Key Laboratory of Frigid Zone Cardiovascular Diseases, General Hospital of Northern Theater Command, Shenyang 110016, China
OBJECTIVES Guidewire ablation (GA) is a novel alternative ablation method for the patients who had previous ablation failure. Howerer as a novel technology, the energy transmission process of GA is unknown.
METHODS (1) 2 groups of 1.17 mm (4F) and 2.24 mm (8F) tissues were divided, and then each group was further divided into four sub-groups of 10 W, 15 W, 20 W and 25 W on the basis of the ablation power. During the ablation process, the baseline impedance, generator impedance (GI) changes, steam pop events and their occurrence time were recorded, and the lesion diameters were measured after operation. (2) In accordance with the power, each group was divided into four sub-groups of 10 W, 15 W, 20 W and 25 W, and the same parameter changes were recorded, and the lesion diameters were measured as the study (1). (3) As the secure power obtained in the study (2), the myocardial tissues were segmented into 10 mm group and 20 mm group for the exposed length of the guidewire, and then segmented into 7 sub-groups base on ablation durations (10 s, 20 s, 30 s, 40 s, 50 s, 60 s and 90 s). (4) The influence of the infusion velocity of normal saline on the diameter of the lesion. Using the secure power and duration of GA obtained from the previous studies, GA was performed according to the normal saline infusion speed of 0, 1, 2, 3 and 4 mL/min, and the relationship between the lesion diameter and the saline infusion speed was analyzed.
RESULTS In the 1.17 mm (n=32) and 2.24 mm (n=32) simulating vessel diameter groups, we observed that steam pop rate raised with the increase of ablation power. The steam pop rates of 10 W (n=8), 15 W (n=8), 20 W (n=8) and 25 W (n=8) were 0% (0/8), 50% (4/8), 87.5% (7/8) and 100% (8/8) respectively, and the secure power of 1.17 mm group is 10 W possibly (1.17 mm group; 10 W, 0%; 15 W, 50%, P<0.01). In the 2.24 mm group, the steam pop rates were 0% (0/8), 0% (0/8), 0%, 50% (4/8) at 10 W, 15 W, 20 W and 25 W, therefore, we deem the safe power was 20 W (2.24 mm; 20 W, 0%; 25 W, 50%, P<0.01). (2) The steam pop rates raised with the increase of ablation power in the 10 mm (n=32) and 20 mm (n=32) groups. As the power of 10 W (n=8), 15 W (n=8), 20 W (n=8), and 25 W (n=8), the steam pop rates of 10 mm group were 0% (0/8), 12.5% (1/8) 62.5% (5/8) and 100% (8/8), the secure power was identified as 15 W (10 mm group; 15 W, 12.5%; 20 W, 62.5%, P<0.01). In the 20 mm group, the steam pop rates were 0% (0/8), 0% (0/8), 25% (2/8) and 75% (6/8) at 10 W, 15 W, 20 W and 25 W respectively, and the secure power might be 20 W (20 mm group: 20 W, 25%; 25 W, 75%, P<0.01). (3) The lesion diameter of 10 mm group (n=42) and 20 mm group (n=42) increased significantly within 10–40 seconds, (4) We recognized that 2 mL/min may be the best infusion speed.
CONCLUSIONS If the guidewire exposed was shorter and the target vessel diameter was smaller, high power and long duration ablation should be more cautious to decrease steam pop rate, and saline infusion should be provided to balance the effectiveness and safety of GA.
GW34-e0789
Tingting Lv, Ying Yang, Ping Zhang
Beijing Tsinghua Changgung Hospital, School of Clinical Medicine, Tsinghua University
OBJECTIVES It was reported that mexiletine benefited for type 2 long QT syndrome (LQT 2) patient carrying a G604C hERG (G604C) mutation while propranolol did not in our clinical practice. However, the possible mechanism of mexiletine on hERG—G604C mutant potassium channels current (IKr ) remains unclear.
METHODS Here hERG—WT (WT), G604 and heterozygous (WT/M) monoclonal stable cells were constructed in HEK293 cells. Western Blot assay and Real Time PCR assay were used to detect the levels of hERG protein and mRNA, western blot of membrane protein and immunofluorescence to evaluate protein trafficking, and whole-cell patch-clamp to detect IKr current.
RESULTS As a result, the expression of hERG mRNA and protein were both identical in WT and G604C group. HERG trafficking deficiency was presented in G604C group. IKr current decreased over 50% in WT/M group compared to WT group, and presented the loss of function in the G604C group. Incubation of increased concentration of propranolol didn’t change hERG mRNA and protein, but inhibited IKr current with a concentration-dependent manner. Mexiletine rescued hERG protein trafficking deficiency in WT/M group but not in G604C group, representing the transient effect of mexiletine on IKr current as a concentration-dependent inhibition while the chronic effect of mexiletine as promotion on G604C mutant hERG channel.
CONCLUSIONS Consequently, hERG—G604C mutation presented hERG trafficking deficiency and damaged IKr current, which can be improved by mexiletine in heterozygous (WT/M) HEK 293 cells.
GW34-e0812
Luyang Yi1,2,3, Xueke Li1,2,3, Wuqi Zhou1,2,3, Rui Wang1,2,3, Yishu Song1,2,3, Mengdan Ding1,2,3, Li Zhang1,2,3, Mingxing Xie1,2,3
1Department of Ultrasound Medicine, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology
2Hubei Province Clinical Research Center for Medical Imaging
3Hubei Province Key Laboratory of Molecular Imaging
OBJECTIVES Cardiac allograft vasculopathy (CAV) is a significant complication that arises from the phenotypic conversion of vascular smooth muscle cells (VSMCs) and has a severe impact on the prognosis of patients following heart transplantation. In this study, a novel ultrasound-triggered delivery system was developed, which involved a dextran nanoparticle microbubble complex encapsulating miR-145. The aim was to prevent CAV by modulating the expression of Krüppel-like factor 4 (KLF4) in VSMCs.
METHODS To prepare Spermine-dextran encapsulating miR-145 (SpeDex-miR-145), the reductive amine method was employed. Subsequently, SpeDex-miR-145 was conjugated to microbubbles (MB) through thiol bonding. A mouse model of cardiac allograft vasculopathy was established. On day 3 after successful surgery, SpeDex-miR-145@MB was administered via injection into the tail vein. Ultrasound irradiation was then conducted above the allograft through the abdominal wall. On postoperative day 28, graft vessels were subjected to quantitative RT-PCR analysis, HE staining, EVG staining, and immunofluorescence (IF). These techniques were used to assess graft vessel characteristics, miR-145 expression levels, and to identify the phenotype of vascular smooth muscle cells (VSMCs). This comprehensive analysis allowed for the evaluation of the therapeutic effects of SpeDex-miR-145@MB on graft vessels and the modulation of VSMC phenotype.
RESULTS The expression miR-145 was upregulated after ultrasound triggered SpeDex-miR-145@MB delivery. The expression levels of KLF-4 and KLF5, the upstream target genes of miR-145, were downregulated after ultrasound triggered SpeDex-miR-145@MB delivery. The VSMCs contractile marker SM22 was significantly upregulated after ultrasound triggered SpeDex-miR-145@MB delivery; while the expression of OPN, a synthetic marker of VSMCs, was downregulated significantly. The intima-media ratio was smaller after ultrasound triggered SpeDex-miR-145@MB delivery.
CONCLUSIONS The utilization of this nano-delivery system demonstrated effective delivery of miR-145 into VSMCs through ultrasound triggering. This targeted delivery successfully prevented the progression of CAV by reversing the phenotypic conversion of VSMCs. These findings indicate that the ultrasound nano-delivery system holds promise as a novel approach for preventing CAV.
GW34-e0977
Ke Gao1,2, Lei Yang1, Yanbo Xue1, Xiaorui Huang1, Yajie Gao1, Ruijuan Shi1, Qian Yin1,2, Hongyan Tian1,2, Xiaohui Zheng3, Xiaopu Zheng1
1Department of Cardiology, The First Affiliated Hospital of Xi’an Jiaotong University, Xi’an, Shaanxi, China
2Department of Peripheral Vascular Diseases, The First Affiliated Hospital of Xi’an Jiaotong University, Xi’an, China
3Key Laboratory of Resources Biology and Biotechnology in Western China, Ministry of Education, College of Life Sciences, Northwest University, Xi’an, China
OBJECTIVES While accelerated arterial stiffening may represent a critical mechanistic factor driving cardiovascular risk in diabetes mellitus (DM), specific therapies to contain the underlying diabetic arterial remodeling have been elusive. The typical pathological changes of vascular remodeling include vascular fibrosis, the destruction of elastic fibers, and the proliferation and migration of vascular smooth muscle cells (VSMC), all of which are the important pathological basis for the occurrence and progression of arteriosclerosis and diabetic macrovascular complications. At present, there is a lack of effective therapeutic drugs to improve diabetic vascular remodeling. Isopropyl 3-(3,4-dihydroxyphenyl)-2-hydroxypropanoate (IDHP) is one of the main bioactive metabolites of the Chinese medicinal herb Danshen, which can be absorbed into blood compounds by oral administration of Compound Danshen dripping pills. The purpose of this study was to investigate whether IDHP can prevent and treat diabetic vascular remodeling, and to explore the possible effects and mechanisms of IDHP on aortic medial fibrosis, elastic fibers destruction, VSMC proliferation and migration during vascular remodeling.
METHODS In vivo, the animal model of diabetic vascular remodeling was established after 8 weeks of diabetes, and IDHP preventive intervention and therapeutic intervention were given respectively. HE, Masson, PAS and Verhoeff’s Van Gieson staining were used to observe the histological morphology of thoracic aorta, the degree of vascular fibrosis, and the destruction of elastic fibers in vascular media. In vitro, VSMCs were pretreated with normal glucose (NG), 25 mM high glucose (HG), IDHP+HG and IDHP for 48 h, respectively. The expressions of Collagen I, p-p38 MAPK, p38 MAPK, Runx2, PCNA, MMP 2, MMP 9, NOX1, NOX4 and ROS levels in VSMC were detected by western blotting, DHE fluorescence probe and flow cytometry. Moreover, VSMCs were pretreated with Runx2 agonist Muramyl dipeptide, p38 MAPK agonist Anisomycin, p38 MAPK inhibitor SB203580, ROS scavenger NAC, the non-specific NOX inhibitor Diphenyleneiodonium chloride, and Mito-TEMPO, a mitochondria-targeted superoxide dismutase mimetic, respectively. Next, VSMCs proliferation was analyzed using CCK-8 method. Cell migration was examined using wound-healing and transwell assay. MitoSOX fluorescent probe was used to detect superoxide levels generated from VSMC mitochondria.
RESULTS Vascular media thickening, VSMC hyperplasia and vascular medial fibrosis were observed in the thoracic aorta of DM rats, and the elastic fibers in the media of thoracic aorta were degraded and fractured obviously. All these changes indicated that there was obvious vascular remodeling in the thoracic aorta of DM rats, but pretreatment with IDHP could prevent DM-induced vascular remodeling. More importantly, post-treatment with IDHP can effectively attenuate vascular remodeling under diabetic conditions. We also demonstrated that IDHP decreased the expression of VSMC Collagen I by inhibiting HG-induced activation of the ROS/p38 MAPK/Runx2 signaling pathway, and thereby improving vascular medial fibrosis. In addition, HG promotes the proliferation and migration of VSMC and enhances the expression of MMP 2 and MMP 9 by inducing the mitochondrial, NOX1 and NOX4-derived ROS-activated p38 MAPK signaling pathway. However, IDHP inhibits VSMC proliferation, migration and MMPs expression by weakening the p38 MAPK signaling pathway mediated by ROS generated from mitochondria, NOX1 and NOX4 in VSMCs.
CONCLUSIONS IDHP can prevent and treat diabetic vascular remodeling, and it attenuates vascular remodeling through ROS/p38 MAPK signaling pathway. These novel findings suggest that IDHP is a potential pharmacologic candidate for treating diabetic vascular remodeling.
GW34-e0996
Dong Guo, Xue Yang, Lang Hu, Yan Li
Department of Cardiology, Tangdu Hospital, Airforce Medical University
OBJECTIVES Iron overload has been cited as an important detrimental factor contributing to the poor prognosis of myocardial infarction (MI). Iron chelating therapy was reported to ameliorate MI-induced cardiac injury. However, the existing iron chelators has many side effects, thereby hindering their application in the treatment of post-MI injury. The current study aimed to explore a biological process-based strategy for iron chelating in MI-induced cardiac injury.
METHODS MI model were constructed by ligation of left anterior descending coronary artery. Cardiomyocytes were subjected to hypoxia treatment to mimic MI in vitro. Extracellular vesicles (EVs) were collected from the culture medium of macrophage by ultracentrifugation and then co-incubated with cardiomyocytes or intramyocardially injected into mice hearts. The iron content, oxidative stress, ferroptosis level, cell survival, infarcted area, and cardiac function were determined to investigate the effect of macrophages-derived EVs on iron overload and cardiac injury in hypoxia cardiomyocytes or post-MI hearts. To further explore the mechanism, multiple approaches including mass spectrometry, immunoelectron microscopy, nano-flow cytometry, and enzyme-linked immunosorbent assay were conducted. Besides, in vivo imaging and fluorescent labeling were employed to elucidate the distribution of macrophages-derived EVs.
RESULTS Cardiac iron level, serum iron level, and cardiac expression of iron-binding proteins, including transferrin and ferritin heavy chain 1 (FTH1), were significantly elevated in MI mice. Moreover, the hearts of MI mice showed obviously increased oxidative stress and ferroptosis level, as evidenced by increased malondialdehyde (MDA) content, elevated 4-hydroxynonenal (4-HNE) expression, decreased glutathione (GSH) expression, and reduced glutathione peroxidase 4 (GPX4) level. EVs were successfully isolated from culture medium of macrophages and showed remarkable iron-chelating capacity in bioactive sample including myocardium lysate, cardiomyocytes lysate, serum and culture medium of cardiomyocytes. Intramyocardially injection of macrophages-derived EVs effectively decreased the iron level of serum and myocardium in post-MI mice. Also, co-incubation with macrophages-derived EVs reduced the iron level of hypoxia-treated cardiomyocytes and their culture medium. Further in vitro and in vivo experiments revealed that iron overload-induced oxidative stress and ferroptosis were also remarkedly alleviated by macrophages-derived EVs. Mechanistically, we found that macrophages-derived EVs inherited TfR from their parent cells and could bind with transferrin. Ablation of TfR on EVs abrogated their capacity in preventing iron overload, oxidative stress, ferroptosis, and cardiac dysfunction in post-MI mice. In addition, we discovered that macrophages-derived EVs were ultimately captured and processed by liver-resident macrophages.
CONCLUSIONS In this study, we identified macrophages-derived EVs as a novel endogenous biological chelator for excess iron in MI hearts. Macrophages-derived EVs inherited TfR from parent cells, based on which macrophages-derived EVs could bind with transferrin, remove excess protein-bound iron, and thus treat iron overload. Our work demonstrated that macrophages-derived EVs were a powerful endogenous tool for iron chelating, which provides a novel and promising therapeutic approach for treating iron overload-induced injury in the pathogenesis of MI and other cardiovascular diseases.
GW34-e0998
Wang Yu, Qin Yanwen
Beijing Anzhen Hospital, Capital Medical University
OBJECTIVES Hypercholesterolemia is an independent risk factor for cardiovascular disease. Studies have shown that effectively lowering circulating levels of low-density lipoprotein cholesterol (LDL-C) can prevent and reduce cardiovascular events. MicroRNA-181d (miR-181d) is a potent inhibitor of liver fat droplets, reducing liver fat droplets by about 60%. Can reduce cell triglycerides and cholesterol esters. The expression of miR-181d was decreased in plasma and adipose tissue of obese people. However, no relationship between miR-181d and LDL-C has been reported.
METHODS Two hyperglycemia animal models were used to analyze the potential relationship between miR-181d-5p and LDL-C. AAV-mediated liver-directed miRNA which over expressed in high cholesterol diet (HCD)-fed mouse model was used to examine the effect of miR-181d-5p on LDL-C and WB/RT-PCR/ELISA/luciferase assays were used to investigate the underlying mechanism.
RESULTS The expressions of miR-181d-5p were obviously lower in hyperglycemia mouse model, decreased 44.2% and had a negative correlation with serum LDL-C level (P<0.001 r=-0.8575); in hyperglycemia hamster model, miR-181d-5p expression was reduced by 48.7% and had a negative correlation with serum LDL-C level (<0.001 r=-0.8811). After AAV-mediated liver-directed miR-181d-5p over expressed, the serum cholesterol was obviously decreased about 15.2%, serum LDL-C decreased about 21.1%. After miR-181d-5p was over expressed, the liver cholesterol was obviously decreased about 25.2%, liver triglyceride decreased about 34.1% than that of control mouse. After confirming the effect of miR-181d-5p on the improvement of serum lipid in vivo, through Target Scan 8.0, we discovered PCSK9 was the possible target genes regulated by miR-181d-5p, which was further proved by in vitro experiments. MiR-181d-5p could directly interact with both the PCSK9 3′-UTR and promoter to inhibit PCSK9 translation and transcription. Furthermore, the result from Dil-LDL uptake assay under the knockdown of PCSK9 demonstrated that miR-181d-5p promoting the absorption of LDL-C in Huh7 cells was dependent on PCSK9. And the result from LDLR−/− mouse model indicated that miR-181d-5p regulating LDL-C was dependent on PCSK9/LDLR pathway.
CONCLUSIONS We discovered miR-181d-5p targets PCSK9 3′-UTR and regulated PCSK9 expression and serum Lipid, suggesting miR-181d-5p might be a new therapeutic target for reducing hypercholesterolemia.
GW34-e1055
Xingdong Ye, Yumei Xue
Guangdong Cardiovascular Institute, Guangdong Provincial People’s Hospital, Guangdong Academy of Medical Sciences
OBJECTIVES Atrial fibrillation (AF) is a common arrhythmia, which is closely related to aging and is a “senile disease”. However, its current treatment effect is not good enough and there is a certain recurrence rate. It is urgent to further explore the specific molecular mechanism of its pathogenesis in order to obtain better treatment. AF is associated with the aging of atrial fibroblasts, but its pathological mechanism is still unclear. The purpose of this study is to observe whether Piezo1/β-catenin pathway participated in the aging of atrial fibroblasts.
METHODS Primary atrial fibroblasts were isolated from male C57BL/6 mice (1 month) by enzyme digestion, and 100 μM tert-butyl hydroperoxide (TBHP) was used to induce the aging of mouse atrial fibroblasts (MAFs). Expression levels of Piezo1, β-catenin and aging-associated proteins p53/p21 in the cells treated with TBHP were detected by Western blot. The aging MAFs were treated with Piezo1 inhibitor GsMTx4 (3 or 10 μM) or Piezo1 siRNA, or β-catenin inhibitor XAV939 (3 or 10 μM) respectively. Changes in expression levels of β-catenin and aging-associated proteins p53/p21 in the MAFs were observed by Western blot.
RESULTS Expression levels of Piezo1, β-catenin and aging-associated proteins p53/p21 in the cells treated with TBHP increased significantly (P<0.05). GsMTx4, Piezo1 siRNA or XAV939 could ameliorate the increased expression levels of β-catenin and aging-associated proteins p53/p21 in the MAFs induced by TBHP (P<0.05).
CONCLUSIONS Piezo1/β-catenin pathway participates in the aging of atrial fibroblasts.
GW34-e1150
Yajun Xue
Beijing Tsinghua Changgung Hospital
OBJECTIVES This study aimed to review the research status and to demonstrate the hotspots and frontiers of chronotherapy.
METHODS Literatures regarding chronotherapy from inception to 2022 were retrieved from the Web of Science Core Collection database. CiteSpace 5.6.R3 was used to generate network maps about the distributions of authors, countries, institutions, references, journals and reveal hot spots and frontiers of chronotherapy.
RESULTS A total of 1059 studies related to chronotherapy were included. The number of literatures regarding chronotherapy was generally increased with some fluctuations. The field of vascular calcification research involves many disciplines, such as pharmacology pharmacy, biology, and physiology. High-yielding countries include the United States and European countries, with China also having a place. Major research institutes include the Hôpital Paul Brousse, Universidade de Vigo, and so on. High-yielding and most cited authors include Levi FA and Hermida RC. Chronobiol Int published the most of research regarding chronotherapy. The first co-cited reference reported the MAPEC trial which analyzed the effect of circadian rhythm time of hypertension treatment on cardiovascular risk. Hot topics focused on the circadian rhythm, hypertension, sleep deprivation, cancer and rheumatoid arthritis. It can be predicted that future correlative studies of chronotherapy will revolve around hypertension, cancers, immune system disorders, and psychiatric disorders.
CONCLUSIONS This study suggested the prosperous research trends and close cooperation. Major ongoing research trends include the timing of antihypertensive medication dosing, sleep deprivation, cancer and rheumatoid arthritis.
GW34-e1170
Ying Wang, Liming Chen, Jinyi Lin, Chancing Huang, Kejia Jin, Hao Wang, Jianguo Jia, Zhiwen Ding, Junbo Ge, Hui Gong, Yunzeng Zou
Zhongshan Hospital of Fudan University
OBJECTIVES Ischemia/reperfusion (I/R) injury causes cardiomyocytes death, including apoptosis and ferroptosis, which remains a significant challenge for reperfusion treatment. This study aimed to investigate the potential protective role of Wnt2 against cardiomyocytes death after I/R injury.
METHODS Serum Wnt2 levels in patients with acute myocardial infarction (AMI) before and after percutaneous coronary intervention (PCI) and in mice subjected to I/R were measured by ELISA. Cardiac function was assessed by echocardiography of M-mode Strain analysis and MRI. Cardiomyocyte death, including apoptosis and ferroptosis, which were examined by TUNEL and thiobarbituric acid reactive substances (TBARs) assay, respectively. TMT6-based proteomics analysis was used to identify critical molecules mediating the effects of Wnt2.
RESULTS Serum Wnt2 levels significantly decreased after 1, 2, 3, and 4 days of PCI in comparison to AMI patients before PCI, and the levels had negative correlation with cTnT and CKMB levels, the cardiac injury biomarkers, respectively, within the first 48 h following PCI. Serum or cardiac Wnt2 also decreased in I/R mice in compared with sham group. Supplement of rbWnt2 significantly improved cardiac function and reduced infarct area and improved the reduced ATP levels in I/R hearts as well. RbWnt2 treatment also significantly inhibits cardiomyocytes apoptosis and ferroptosis by suppressing lipid peroxidation via decreased reactive oxygen species (ROS) production in response to I/R injury. Proteomics analysis revealed that Wnt2 significantly decreased Nap1L1 expression following I/R injury, which was validated by Western Blot. Cardiac specific overexpression of Nap1L1 partly abolished inhibitory effect of rbWnt2 on apoptosis and ferroptosis in vivo and in vitro, and then, attenuated protection effect of rbWnt2 against I/R injury, whereas cardiac specific knockdown of Nap1L1 improved the cardiac function, alleviate infarct area, inhibited apoptosis and ferroptosis and increased ATP levels via suppressing ROS during I/R injury as rbWnt2 treatment. Mechanically, Wnt2 promoted Nap1L1 degradation and subsequently attenuated ROS production and cardiomyocyte death to improve cardiac function following I/R. UbiBrowser database analysis revealed that putative E3 ligases Trim11 and Rapsn interacted with Nap1L1. Western blot analysis indicated rbWnt2 treatment reversed the reduced expression of Trim11 but did not alter the increased expression of Rapsn mediated by H/N. Co-immunoprecipitation (co-IP) analysis demonstrated that the interaction of Trim11 and Nap1L1 was decreased in cardiomyocytes after H/N injury, which was restored by rbWnt2 treatment. Trim11 silence blocked the increased Nap1L1 ubiquitination mediated by Wnt2 in H/N-cardiomyocytes. Further analysis indicated that Wnt2 acted on Lrp6 to enhance Trim11 expression for ubiquitination of Nap1L1 in ischemic cardiomyocytes.
CONCLUSIONS The present study showed that I/R mediated the decreased Wnt2 level in heart and serum, supplement of Wnt2 acts on Lrp6 to attenuate apoptosis and ferroptosis by degradation of Nap1L1 through Trim11 following I/R. Supplement of Wnt2 maybe a novel therapeutic strategy for the clinical treatment of ischemia/reperfusion injury and the present findings also suggest new insights into the regulation of Nap1L1 as a target in the treatment of cardiac ischemia/reperfusion injury.
GW34-e1228
Yishu Song1,2,3, Qiaofeng Jin1,2,3, Cheng Deng1,2,3, Luyang Yi1,2,3, Li Zhang1,2,3, Mingxing Xie1,2,3
1Department of Ultrasound Medicine, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430022, China
2Clinical Research Center for Medical Imaging in Hubei Province, Wuhan 430022, China
3Hubei Province Key Laboratory of Molecular Imaging, Wuhan 430022, China
OBJECTIVES Cardiac patch based therapy has emerged as a promising strategy for the treatment of heart disease, such as severe myocardial infarction. We aimed to develop a novel hydrogel patch for heart disease treatment which can sustained release drugs on the heart surface.
METHODS PLGA microspheres were prepared using a standard water-oil-water double emulsion procedure. Lyophilized PLGA microspheres were doped in GelMA solution containing photoinitiator LAP. To form microspheres hybrid GelMA hydrogel, the precursor solution was illuminated with a 405 nm light source. We used RITC-Dextran as model drug to study the drug release. In vivo, hydrogel precursor solution was spread on the heart surface immediately followed by light crosslinking to prepare the hydrogel cardiac patch.
RESULTS The lyophilized microspheres were white powder with an average particle size of 25.56±0.37 μm. SEM images indicate that the microspheres had a smooth surface and a interconnected porous structure inner it. The PLGA microspheres hybrid GelMA hydrogel was white colloid. SEM images showed that hybrid hydrogel exhibited 3D network structures with an average pore size of 67.96±30.62 μm. Microspheres were encapsulated in the pore wall or distributed in the pore. In vitro drug release study showed that GelMA hydrogel had significant burst release during 0–2 h, with a cumulative release of 96.73±5.38%. The incorporation of microspheres improved the sustained release ability, we demonstrated that RITC-Dextran are released over 28 d from this formulation, with a cumulative release of 30.47±1.57% on 28 d. In vivo experiments demonstrated that the hydrogel precursor solution irradiated by 405 nm light source could be cross-linked in situ to form a hydrogel cardiac patch on the heart surface. The patch also showed wet adhesion behavior and could be attached to the surface of beating heart.
CONCLUSIONS In summary, microspheres hybrid GelMA hydrogel performed a stable sustained drug release behavior and in situ photocuring on the heart surface was also achieved, which suggested a promising approach to the treatment of heart disease such as heart transplantation rejection and myocardial infarction.
GW34-e1290
Guangzhi Cong, Ru Yan, Shaobin Jia
General Hosptital of Ningxia Medical University
OBJECTIVES Promoting angiogenesis in the infarcted area is the key to the treatment of sequelae of AMI. The latest research suggests that monocyte-derived macrophages were recruited to the border zone after MI and secrete vascular endothelial growth factor (VEGFA), promoting angiogenesis. An attractive therapeutic strategy is to stimulate angiogenesis after myocardial infarction.
METHODS We reanalyzed large-scale, publicly available bulk (GSE151834) and single-cell RNA sequencing datasets (GSE163129) from the myocardial infarction mice model. Myeloid-specific TREM2 (Triggering Receptor Expressed On Myeloid Cells 2) knockout mice (TREM2-/-MF) and wild-type mice (TREMflox/flox) underwent MI induced by ligation of the left anterior descending coronary artery. Cardiac functionality was assessed by longitudinal echocardiographic measurements. Histological, fluorescence-activated cell sorting, and in vitro/ex vivo studies were used to assess the myocardial neovascularization, macrophage pro-angiogenesis ability, and activity in response to TREM2.
RESULTS Bioinformatic trajectory analysis indicates TREM2 positive macrophages are the majority monocyte-derived macrophages during the late phase post myocardial infarction and governing VEGFA expression. TREM2-/-MF mice showed a reduced cardiac systolic function with a lower ejection fraction and fractional shortening in a time-dependent manner. Furthermore, the angiogenesis maker CD31 expression decreased in the myocardial infarction border zone. In vitro, when challenged with hypoxia, peritoneal macrophages from TREM2 mice (TREM2-/-MF) have a lower ability to produce VEGFA and NRF2 (Nuclear Factor erythroid 2-related factor 2). When cocultured with HUVEC, angiogenesis was worsened in the peritoneal macrophages from TREM2 mice (TREM2-/-MF) detected by matrigel assay with fewer angiogenesis meshes and junctions. NRF2 inhibitor treatment (MF385) can improve VEGFA production and the pro-angiogenesis ability of TREM2-deficient peritoneal macrophages.
CONCLUSIONS Our findings indicate that cardiac macrophages maintain healing through promoting myocardial angiogenesis by TREM2-NFR2 signaling.
GW34-e1333
You Jieyun1, Li Xuan2, Dai Fangjie2, Liu Jun1, Zhang Qi1, Guo Wei1
1Department of Cardiovascular Medicine, Shanghai East Hospital, School of Medicine, Tongji University, Shanghai, China
2Shanghai Institute of Cardiovascular Diseases, Zhongshan Hospital and Institutes of Biomedical Sciences, Fudan University, Shanghai, China
OBJECTIVES Gasdermin D (GSDMD) forms membrane pores to execute pyroptosis. But the mechanism of how cardiomyocyte pyroptosis induces cardiac remodeling in pressure overload remains unclear. We investigated the role of GSDMD-mediated pyroptosis in the pathogenesis of cardiac remodeling in pressure overload.
METHODS Wild-type (WT) and cardiomyocyte-specific GSDMD-deficient (GSDMD-CKO) mice were subjected to transverse aortic constriction (TAC) to induce pressure overload. Four weeks after surgery, left ventricular structure and function were evaluated by echocardiographic, invasive hemodynamic and histological analysis. Pertinent signaling pathways related to pyroptosis, hypertrophy and fibrosis were investigated by histochemistry, RT-PCR and western blotting. The serum levels of GSDMD and IL-18 collected from healthy volunteers or hypertensive patients were measured by ELISA.
RESULTS We found TAC induced cardiomyocyte pyroptosis and release of pro-inflammatory cytokines IL-18. The serum GSDMD level was significantly higher in hypertensive patients than in healthy volunteers, and induced more dramatic release of mature IL-18. GSDMD deletion remarkably mitigated TAC-induced cardiomyocyte pyroptosis. Furthermore, GSDMD deficiency in cardiomyocytes significantly reduced myocardial hypertrophy and fibrosis. The deterioration of cardiac remodeling by GSDMD-mediated pyroptosis was associated with activating JNK and p38 signaling pathways, but not ERK or Akt signaling pathway.
CONCLUSIONS In conclusion, our results demonstrate that GSDMD serves as a key executioner of pyroptosis in cardiac remodeling induced by pressure overload. GSDMD-mediated pyroptosis activates JNK and p38 signaling pathways, and this may provide a new therapeutic target for cardiac remodeling induced by pressure overload.
GW34-e1361
Qingbo Lv1,2, Yu Xiao1, Guosheng Fu1,2
1Key Laboratory of Cardiovascular Intervention and Regenerative Medicine of Zhejiang Province, Sir Run Run Shaw Hospital, Zhejiang University School of Medicine, Hangzhou 310016, China
2Department of Cardiology, Sir Run Run Shaw Hospital, Zhejiang University School of Medicine, Hangzhou 310016, China
OBJECTIVES Myocardial infarction (MI) is one of the leading cause of global mortality. Ferroptosis, a newly recognized iron-dependent form of cell death, plays a crucial role in progression of MI. The excessive deposition of free irons within cardiomyocytes is a vital cause of severe oxidative stress and lipid peroxidation, directly leading to the occurrence of ferroptosis. Therefore, therapeutic strategies targeting ferroptosis could potentially serve as a novel approach to treat myocardial infarction. In this study, we employed deferoxamine (DFO), a proven effective ferroptosis inhibitor, as our chosen therapeutic agent. However, DFO has an extremely short circulation half-life, making it unfeasible for sustained drug delivery in vivo. In response, we utilized chitosan (CS), a substance with excellent biocompatibility, to self-assemble nanoparticles as carriers. Through nanoscale self-assembly and chemical bonding, DFO was encapsulated within these nanoparticles, forming chitosan-deferoxamine nanosponges (CDNS) capable of adsorbing free iron.
METHODS To construct CDNS, deferoxamine was first dissolved in Dulbecco’s Modified Eagle Medium (DMEM), followed by the addition of CS. The pH was adjusted to 7.4 using 0.1 M Tris solution, and the complex was incubated for 2 hours to form CDNS. After obtaining CDNS, we performed standard material testing. High-performance liquid chromatography (HPLC) experiments were conducted to detect its release curve under different pH solutions and to examine its iron-binding capabilities. Subsequently, we tested the protective effect of CDNS on oxidative stress damage induced by hydrogen peroxide and ferroptosis induced by erastin. We then established a mouse MI model, and delivered CDNS to the margin of the infarcted myocardium via intramyocardial injection. Initially, using the water-soluble fluorescent dye indocyanine green (ICG), we confirmed the CDNS system’s ability for sustained drug release within the heart. Subsequently, with cardiac ultrasound, tissue section staining, and other methods, we focused on whether administration of CDNS improved heart function and reduced infarct size in post-MI mice, and ameliorated adverse myocardial remodeling post-infarction. Furthermore, we closely monitored the indicators of ferroptosis and angiogenesis in the hearts of mice post-MI surgery.
RESULTS First, we observed that CDNS were well-dispersed nanoparticles with diameters ranging from 50 to 200 nm through TEM. HPLC tests confirmed that CDNS were stable in a neutral solution environment, with minimal drug release. In acidic conditions, CDNS could sustainably release DFO and exhibited excellent iron-chelating capabilities. In the in vitro study, we found that CDNS could significantly alleviate oxidative stress damage in cardiomyocytes and inhibit myocardial ferroptosis. In animal experiments, we found that CDNS could achieve in-situ long-term drug release. From echocardiograms, we observed a significant improvement in heart function post-MI in mice treated with CDNS. CDNS significantly reduced apoptosis of myocardial cells and attenuated cardiac inflammation within the acute stage of MI. Meanwhile, CDNS significantly inhibited myocardial fibrosis and promoted angiogenesis in the infarcted area of the heart in the long-term.
CONCLUSIONS Our results suggest that CDNS are the powerful therapeutic approach for the prevention and treatment of MI by inhibiting ferroptosis and promoting angiogensis. Given their high efficacy and good biosafety, CDNS hold great promise for future clinical applications in the management of MI.
CARDIOVASCULAR DISCIPLINARY RESEARCH
PULMONARY VASCULAR DISEASE
GW34-e0131
Xin Li, Zhang Yi, Zhao Zhihui, Liu Zhihong
Fuwai Hospital, National Center for Cardiovascular Diseases, National Clinical Research Center for Cardiovascular Diseases, Chinese Academy of Medical Sciences and Peking Union Medical College
OBJECTIVES The occurrence of complications associated with balloon pulmonary angioplasty (BPA) is frequent and may potentially lead to perioperative mortality. Nevertheless, the current academic literature lacks a predictive model pertaining to the complications related to BPA.
METHODS A retrospective analysis was performed on data obtained from a consecutive series of patients diagnosed with chronic thromboembolic pulmonary hypertension who underwent BPA. The main objective of the study was to investigate complications associated with BPA. The secondary endpoints of the study were mortality rates and hemodynamic changes observed following BPA.
RESULTS The study comprised 207 patients with chronic thromboembolic pulmonary hypertension who underwent a total of 614 sessions of BPA. During the study, complications were observed in 63 BPA sessions (10.26%) among 49 patients. The study findings revealed that hemoptysis or hemosputum (6.51%) was the most frequently encountered complication, whereas the incidence of pulmonary reperfusion edema was relatively low (0.49%). The results of multivariable logistic regression analysis indicated a significant correlation between BPA-related complications and disease duration, mean pulmonary arterial pressure, and the proportion of occlusion lesions. Subsequently, a nomogram was constructed and demonstrated superior predictive accuracy compared to previously reported predictors, as indicated by the area under the curve value of 0.703. Validation and calibration revealed a favorable level of accuracy, as evidenced by a slope of 0.978 and a Brier score of 0.163. Multivariable linear regression analysis that adjusted for the number of BPA sessions revealed that there was no significant correlation detected between the occurrence of complications and alterations in hemodynamics subsequent to BPA. Furthermore, patients who experienced complications exhibited a comparable 3-year survival rate to those who did not encounter such complications (98.0 vs. 94.8%, log-rank P=0.503).
CONCLUSIONS The nomogram, which incorporates multiple parameters such as mean pulmonary artery pressure, occlusion lesions, and disease duration, exhibits superior predictive capabilities for complications associated with BPA compared to individual parameters that have been previously reported. Notably, despite the occurrence of complications, the favorable effects of BPA on hemodynamics and survival were maintained. Thus, the presence of complications should not dissuade patients from pursuing ongoing BPA sessions.
GW34-e0195
Sun Na1,2, Fan Youli1,2, Chen Yansheng1,2, Gao Xuan1,2, Wang Yongbing1, Wu Bingxiang1,2
1Department of Cardiology, The Second Affiliated Hospital of Harbin Medical University, Harbin, China
2Key Laboratory of Myocardial Ischemia, Ministry of Education, Harbin Medical University, Harbin, China
OBJECTIVES The aim of the present study was to evaluate the correlation of plasma DCA levels with clinical and hemodynamic parameters in patients with acute pulmonary embolism patients (APE).
METHODS We prospectively recruited 149 adult patients with APE. Plasma DCA levels were measured by rapid resolution liquid chromatography quadrupole time-of-flight mass spectrometry. Baseline clinical and hemodynamic parameters were evaluated according to different plasma DCA levels.
RESULTS Compared with controls, the plasma DCA levels are significantly lower (0.15 (0.10, 0.31) μg/mL versus 0.08 (0.04, 0.16) μg/mL, P<0.001) in APE patients. Recruited APE patients were divided into two groups according to the media plasma DCA levels (0.08 μg/mL), high DCA group and low DCA group. Compared with high DCA group, low DCA group patients present more adverse cardiac function with higher NT-proBNP levels (2629.0 (752.5, 6225.5) pg/mL versus 1262.0 (158.0, 4444.0) pg/mL, P=0.010), higher WHO functional class levels (P=0.023). Low DCA group also possess serious hemodynamic status, with higher PVR levels (4.3 (1.9, 6.4) Wood units versus 2.5 (1.6, 4.6), P=0.027) and lower CI levels (2.5±0.8 versus 2.8±0.8, P=0.024). For the correlation analysis between DCA and cardiac function and hemodynamic parameters, PVR was negatively correlated with plasma DCA levels (R=-0.198, P=0.027). CI was positively correlated with DCA level (R=0.205, P=0.024). For the cardiac functional parameters, NT-proBNP (R=-0.215, P=0.010) and WHO functional class (R=-0.187, P=0.022) were negatively correlated with plasma DCA levels. Kaplan–Meier survival analysis demonstrated that APE patients with lower plasma DCA levels had significantly higher event rate (P=0.009). In the univariate and multivariate Cox regression analysis, plasma DCA level was an independent predictor to clinical worsening event after adjusted by age, sex, WHO functional class, NT-proBNP, PVR and CI (HR 0.370, 95% CI 0.161, 0.852; P=0.019).
CONCLUSIONS Low plasma DCA levels predict adverse cardiac function and hemodynamic collapse. Low DCA level correlates with higher clinical worsening event rate and could be an independent predictor of clinical outcome in multivariate analysis.
GW34-e0199
Jingxian You1,2,3,4,5, Guili Lian1,2
1Department of Geriatrics, The First Affiliated Hospital of Fujian Medical University, Fuzhou, People’s Republic of China
2Fujian Hypertension Research Institute, The First Affiliated Hospital of Fujian Medical University, Fuzhou, People’s Republic of China
3Clinical Research Center for Geriatric Hypertension Disease of Fujian province, The First Affiliated Hospital of Fujian Medical University, Fuzhou, People’s Republic of China
4Branch of National Clinical Research Center for Aging and Medicine, The First Affiliated Hospital of Fujian Medical University, Fujian Province, Fuzhou, People’s Republic of China
5Department of Geriatrics, National Regional Medical Center, Binhai Campus of the First Affiliated Hospital, Fujian Medical University, Fuzhou, People’s Republic of China
OBJECTIVES Pulmonary arterial hypertension (PAH) is a pathophysiological syndrome, characterized by pulmonary vascular remodeling. Immunity and inflammation are progressively recognized properties of PAH, which are crucial for the initiation and maintenance of pulmonary vascular remodeling. This study explored immune cell infiltration characteristics and potential biomarkers of PAH using comprehensive bioinformatics analysis.
METHODS Microarray data of GSE117261, GSE113439 and GSE53408 datasets were downloaded from Gene Expression Omnibus (GEO) database. The differentially expressed genes (DEGs) were identified in GSE117261 dataset. The proportions of infiltrated immune cells were evaluated by CIBERSORT algorithm. Least absolute shrinkage and selection operator (LASSO) regression analysis was performed to identify feature genes of PAH, and fivefold cross-validation was performed using random forest and logistic regression. The GSE113439 and GSE53408 datasets were used as validation sets and logistic regression and receiver operating characteristic (ROC) curve analysis were performed to evaluate the prediction value of PAH. Weighted gene association network analysis (WGCNA) was used to identify modules associated with PAH. Intersection of genes in the modules screened and DEGs was used to construct protein-protein interaction (PPI) network and identify the core genes. After intersection of feature genes and core genes, the hub genes were identified. Pearson correlation analysis was performed to analyze the correlation between hub genes and immune cell infiltration. Western blotting and immunohistochemistry (IHC) were used to validate the expression of LTBP1 in the lungs and pulmonary arteries in monocrotaline-induced PH rats.
RESULTS A total of 419 DEGs were identified, including 223 upregulated genes and 196 downregulated genes. Functional enrichment analysis revealed that a significantly enrichment in inflammation, immune response, and transformed growth factor β (TGFβ) signaling pathway. CIBERSORT analysis showed that ten significantly different types of immune cells between PAH and control were identified. Seventeen feature genes were identified by LASSO regression for PAH prediction. WGCNA identified 15 co-expression modules. PPI network was constructed and 100 core genes were obtained. Complement C3b/C4b receptor 1 (CR1), thioredoxin reductase 1 (TXNRD1), latent TGFβ binding protein 1 (LTBP1) and toll like receptor 1 (TLR1) were identified as hub genes and LTBP1 has the highest diagnostic efficacy for PAH (AUC=0.968). Pearson correlation analysis showed that LTBP1 was positively correlated with resting memory CD4+ T cells, but negatively correlated with monocytes and neutrophils. We also comfirmed the expression of LTBP1 by Western blotting and immunohistochemistry and the results showed that protein levels of LTBP1 was increased in lungs and pulmonary arteries in PAH rats as compared to control.
CONCLUSIONS LTBP1 is upregulated and correlated to immune infiltration in PAH, identified as a new critical biomarker for PAH. Our study suggests that LTBP1 is involved in the development of PAH and serves as a potential diagnostic and therapeutic target for PAH.
GW34-e0208
Jingyuan Chen, Wenjie Chen, Zilu Li, Jun Luo, Li Li
The Second Xiangya Hospital of Central South University
OBJECTIVES Endoplasmic reticulum (ER) stress is involved in the development of pulmonary arterial hypertension (PAH), and glucose-regulated protein-78 (GRP78) is an important marker of ER stress in PAH. Most studies have shown that its expression is increased at the tissue and cellular levels during ER stress. Current studies have shown that its circulating GRP78 levels are also significantly increased in a subset of diseases and can be used as a disease biomarker.
METHODS For in vitro studies, cardiomyocytes and pulmonary arterial smooth muscle cells (PASMCs) were treated by hypoxia and platelet derived growth factor-BB. For in vivo studies, 18 rats were divided equally into three groups: 1) control group received saline injection, 2) MCT group received 60 mg/kg MCT intraperitoneally injection for once, 3) SuHx group received 20 mg/kg Sugeon 5416 once per week and were placed in hypoxia chamber for 3 weeks. Sixty-six PAH participants were recruited from Second Xiangya Hospital from July 2018 to July 2020. A detailed disease history and demographic data, disease related information were collected. Another 40 healthy control participants were recruited from physical examination center at the same period and verified with no cardiopulmonary disease by transthoracic echocardiography. For all plasma or culture medium samples, GRP78 levels are measured by ELISA.
RESULTS Extracellular GRP78 levels were significantly elevated in hypoxia-treated PASMCs and cardiomyocytes compared to the normoxic group (P<0.05 and P<0.001, respectively). Circulating levels of GRP78 were significantly elevated in the MCT and SuHx groups compared to the control group (P<0.05 and P<0.01, respectively). Circulating GRP78 levels were higher in PAH compared to healthy controls (3.20±2.03 vs 2.54±0.66 μg/mL, P<0.001). In subgroup analysis, GRP78 was higher in the IPAH, CHD-PAH, WHO-FC III/IV and medium-high risk groups. The primary endpoint event occurred in 30 patients over a median follow-up time of 20 (15–37) months. Univariate Cox regression analysis showed that GRP78 was a prognostic risk factor HR=1.422 (95% CI: 1.188–1.703) (P<0.001).
CONCLUSIONS In PAH, circulating GRP78 has significant increase and could be a prognostic risk biomarker for disease-free survival.
GW34-e0843
Wanying Xia1, Yuling Qian1, Qing Gu1, Changming Xiong1, Huijun Han2, Jianguo He1
1Department of Pulmonary Vascular Disease, National Center for Cardiovascular Disease, Fuwai Hospital, Chinese Academy of Medical Sciences and Peking Union Medical College, State Key Laboratory of Cardiovascular Disease, Beijing, China
2Institute of Basic Medical Sciences, Chinese Academy of Medical Sciences, Beijing, China
OBJECTIVES Off-label targeted drugs are commonly prescribed for non-operated chronic thromboembolic pulmonary hypertension (CTEPH) in developed and developing countries, but their effect on the long-term prognosis of CTEPH remains unknown. This study aimed to investigate the effect of off-label targeted drugs on the long-term survival of CTEPH patients receiving pharmacotherapies.
METHODS The prospective national registry recruited patients diagnosed with CTEPH between 2009 and 2018 at 18 centers (ClinicalTrials.gov Identifier NCT01417338). Those from the CTEPH registry treated with pharmacotherapies were analyzed. Five-year survival was compared between patients using off-label targeted drugs and patients receiving conventional drugs in the original and propensity score-matched samples.
RESULTS Of 347 enrolled patients, 135 were treated with conventional therapy and 212 with off-label targeted therapy at baseline (173, initial targeted therapy) or during follow-up (39, subsequent targeted therapy). The 1-, 2-, 3-, and 5-year survival of patients with off-label targeted therapy was significantly higher than that of patients with conventional therapy (97.1 vs. 89.4%, 92.3 vs. 82.1%, 83.2 vs. 75.1%, and 71.1 vs. 55.3%, respectively, log-rank test, P=0.005). Initial targeted therapy was correlated with better 5-year survival after excluding patients with subsequent targeted therapy to reduce the immortal time bias (hazard ratio: 0.611; 95% confidence interval: 0.397–0.940; P=0.025). In propensity score-matched samples, patients given initial targeted therapy showed significantly better 5-year survival than those given conventional therapy (68.9 vs. 49.3%, log-rank test, P=0.008).
CONCLUSIONS Off-label targeted drugs contributed to improved long-term survival in CTEPH patients treated with pharmacotherapies.
GW34-e0851
Zhongchao Wang
General Hospital of Northern Theater Command
OBJECTIVES Hemoptysis is a rare but severe complication of advanced pulmonary arterial hypertension (PAH). Yet, no management consensus has been made till present. Importantly, the effect of targeted drug therapy (TDT) still remains unclear concerning its vasodilative effect on the pulmonary circulation, which may aggravate hemoptysis in the context of vascular rupture. Thus, the present study was designed to investigate short-term effects of continued use of TDT on outcomes of advanced PAH complicated by hemoptysis.
METHODS PAH patients associated with congenital heart disease (PAH-CHD) and complicated by hemoptysis from General Hospital of Northern Theater Command since Mar 2011 to Aug 2022 were retrospectively screened and grouped according to in-hospital targeted medication. 1:1 propensity score matching (PSM) analysis was conducted to adjust confounding factors.
RESULTS Seventy-four patients were included and allocated to TDT (n=39) or non-TDT (n=35) group. General comparison showed continued use of TDT was associated with significantly more days of in-hospital stay (P=0.045). No significant difference was found in mortality or other clinical outcomes, including time for hemoptysis termination and other symptom relief, WHO functional class and NT-proBNP at discharge. PSM analysis further selected 21 pairs of cases and showed no significant difference in mortality or other clinical outcomes between groups.
CONCLUSIONS These results have demonstrated no association between continued use of TDT and worse clinical outcomes of PAH-CHD patients complicated by occurrent hemoptysis, supporting continued use of TDT for patients with advanced PAH even when they are complicated by hemoptysis.
GW34-e1005
Gufeng Gao1,2,3,4,5, Ai Chen1,2,3,4,5, Guili Lian1,2,3,4,5, Li Luo1,2,3,4,5
1Department of Geriatrics, The First Affiliated Hospital of Fujian Medical University, Fujian, Fuzhou, People’s Republic of China
2Fujian Hypertension Research Institute, The First Affiliated Hospital of Fujian Medical University, Fujian, Fuzhou, People’s Republic of China
3Clinical Research Center for Geriatric Hypertension Disease of Fujian province, The First Affiliated Hospital of Fujian Medical University, Fujian, Fuzhou, People’s Republic of China
4Branch of National Clinical Research Center for Aging and Medicine, The First Affiliated Hospital of Fujian Medical University, Fuzhou, Fujian, People’s Republic of China
5Department of Geriatrics, National Regional Medical Center, Binhai Campus of the First Affiliated Hospital, Fujian Medical University, Fujian, Fuzhou, People’s Republic of China
OBJECTIVES This study aims to examine the changes in the PKC/IRS-1/ERK in the lung tissue of pulmonary arterial hypertension (PAH) rats induced by monocrotaline (MCT) and to investigate its role in the excessive proliferation and migration of pulmonary arterial smooth muscle cells (PASMCs).
METHODS
(1) 8-week-old male SD rats were injected intraperitoneally with 20 mg/kg MCT twice at a one-week interval to establish the PAH model. We measured mean pulmonary arterial pressure (mPAP) and right ventricular hypertrophy index (RVHI) four weeks after the first treatment. The percentage of wall area (WA%) and the percentage of wall thickness (WT%) are also calculated. The protein levels of protein kinase C (PKC), p-PKC, insulin receptor substrate 1(IRS-1), p-IRS-1, ERK and p-ERK in lung tissue were detected by Western blot.
(2) After starvation for 24 h, platelet-derived growth factor-BB (PDGF-BB), PKC inhibitor Gö 6983 and IRS-1-expressing adenovirus (AdIRS-1) were added for intervention. The expression of PKC, p-PKC, IRS-1, p-IRS-1, ERK, and p-ERK were detected by Western blot. EdU method was used to detect cell proliferation, while the scratch method was performed to observe cell migration.
RESULTS
(1) mPAP, RVHI, WA% and WT% of MCT-PAH rats were significantly increased four weeks after the first MCT administration,
(2) The expression levels of p-PKC, p-IRS-1 and p-ERK in the lung tissue of MCT-PAH rats were significantly elevated compared with control group. IRS-1 was decreased in the lung tissue of PAH rats; however, the difference in the expressions of PKC and ERK between those two groups was insignificant.
(3) The expression levels of p-PKC, p-IRS-1 and p-ERK were significantly increased after PDGF-BB stimulated PASMCs for 15 min; however, there was no statistically significant difference in expression levels of PKC, IRS-1 and ERK between the two groups.
(4) The expression of IRS-1 in PASMCs treated with PDGF-BB for 48 hours was significantly lower than that in the control group; while there was no statistically significant difference in expression levels of p-PKC, p-IRS-1, p-ERK, PKC and ERK between these two groups.
(5) After PKC inhibitor Gö 6983 treatment for 48 h, the expression of IRS-1 in PDGF-BB-PASMCs was significantly increased. The expression levels of p-PKC, p-IRS-1 and p-ERK were significantly lower than those in the control group and the PDGF-BB-PASMCs, while the differences in the expression levels of PKC and ERK among the three groups were not significant.
(6) After being infected with AdIRS-1, the expression of IRS-1 was increased while the level of p-ERK was downregulated compared with AdNull-PASMCs treated by PDGF-BB for 48 h.
(7) The PKC inhibitor Gö6983 and overexpression of IRS-1 inhibited the proliferation and migration levels of PASMCs 48 h after PDGF-BB induction.
CONCLUSIONS
1. Decreased expression of IRS-1 and its enhanced serine phosphorylation were observed in the lung tissue of PAH rats;
2. PDGF-BB promotes the excessive proliferation and migration of PASMCs through abnormal activation of PKC/IRS-1/ERK pathway.
3. Overexpression of IRS-1 can attenuate the proliferation and migration of PASMCs induced by PDGF-BB.
GW34-e1087
Zeying Zhang1, Yunbin Xiao2, Qiming Liu1
1The Second Xiangya Hospital of Central South University
2Hunan Children’s Hospital
OBJECTIVES Congenital malformations are an important cause of death in children whose important component is congenital heart disease (CHD). Mutations in a variety of genes can lead to CHD, including transcription factors, heart and vascular development, and other genes not related to transcription or translation. Pulmonary artery stenosis as a congenital heart disease is attracting increasing attention because of its association with pulmonary hypertension in children.
METHODS Whole exome sequencing (WES) was used to identify clinically relevant gene variants of proband associated with pulmonary artery stenosis. Sanger sequencing is used to identify candidate variants in all individuals of this Chinese family that can be used for familial co-isolation analysis. Bioinformatics tools were used to predict possible pathogenic mutations and multiple sequence alignments were performed. The pathogenicity of this variant was further evaluated by population-based screening of variant frequencies and American Society for Medical Genetics and Genomics (ACMG) scores.
RESULTS We identified a novel heterozygous deletion variant (c.69delC) on the first exon of the elastin gene ELN by WES. This variant resulted in frameshift mutation just before the protein hydrophobic region (p.Ser24Leufs). The deletion variant was present in the proband and his mother and uncle, but not in his father. Bioinformatic analysis predicted that this mutation would cause nonsense-mediated mRNA degradation (NMD) and be pathogenic.
CONCLUSIONS We shown a novel ELN frameshift mutation c.69delC (p.Ser24Leufs*98) in a family, which expands the understanding of the phenotypic characteristics of mutations in the ELN gene. Patients with pulmonary artery stenosis carrying the ELN variant should be given more attention.
GW34-e1090
Sicheng Zhang, Luyang Gao, Zhihua Huang, Zhihui Zhao, Qin Luo, Qing Zhao, Zhihong Liu
Center for Respiratory and Pulmonary Vascular Diseases, Fuwai Hospital, National Center for Cardiovascular Diseases, National Clinical Research Center for Cardiovascular Diseases, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing, China
OBJECTIVES Emerging evidence has showed that outdoor artificial light at night (ALAN) is closely associated with healthy sleep and cardiovascular health. However, few studies have investigated the association between ALAN and sleep disorders. The unique pathophysiologic contributions of obstructive sleep apnea (OSA) toward pulmonary hypertension still represent an understudied area. This study aims to investigate the association of ALAN with OSA and pulmonary hemodynamics.
METHODS From January 1, 2020 to December 31, 2021, a total of 1005 participants were recruited from Fuwai Hospital. Apnea-hypopnea index (AHI) and oxygen desaturation index (ODI) were measured by polysomnography or home sleep test. Participants’ daytime sleepiness scores were evaluated using the Epworth Sleeping Scale (ESS). Right cardiac catheterization was performed in OSA patients with high suspicion of pulmonary hypertension. Outdoor ALAN exposure (in nanowatts per centimeters squared per steradian) within 1000 m of each participant’s residential address obtained from the satellite imagery data. Generalized linear regression models were used to assess the associations of exposure to outdoor ALAN with OSA indexes and right heart catheterization metrics after adjusting for multiple covariates.
RESULTS Of 859 OSA patients (38.7% male), 453 had pulmonary hypertension. In adjusted models, an inter-quartile range (IQR) increase in outdoor ALAN exposure was significantly associated with 11.3% (95% confidence interval [95% CI]: 4.2%, 18.8%) and 10.0% (95% CI: 2.7%, 17.6%) increases in AHI and ODI, respectively. Furthermore, an IQR increase in outdoor ALAN exposure was significantly associated with 4.7% (95% CI: 0.4%, 9.2%) and 7.0% (95% CI: 2.6%, 11.5%) increases in mean pulmonary arterial pressure (mPAP) and pulmonary vascular resistance (PVR), respectively. Mediation analyses suggested that the effect of outdoor ALAN exposure on mPAP could be partly mediated by ODI.
CONCLUSIONS Our study indicates that higher exposure to outdoor ALAN was significantly associated with higher AHI, ODI, mPAP and PVR in patients with OSA in North China. The effect of outdoor ALAN exposure on pulmonary hemodynamics is mediated partly by OSA indexes.
GW34-e1092
Sicheng Zhang, Luyang Gao, Qin Luo, Zhihui Zhao, Qing Zhao, Zhihong Liu
Center for Respiratory and Pulmonary Vascular Diseases, Fuwai Hospital, National Center for Cardiovascular Diseases, National Clinical Research Center for Cardiovascular Diseases, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing, China
OBJECTIVES Emerging evidence has showed that Blood urea nitrogen to serum albumin ratio (BAR) is associated with the severity and prognosis of heart failure. However, its role in pulmonary arterial hypertension (PAH) remains unclear. The aim of this study was to investigate the associations between BAR and functional status, echocardiography, hemodynamics as well as long-term outcomes among patients with PAH.
METHODS Consecutive patients who underwent right heart catheterization and diagnosed with idiopathic or heritable PAH between January, 2014 and December, 2021 in Fuwai hospital were enrolled. The primary outcome was clinical worsening. Spearman correlation coefficients were used to evaluate the association between BAR and established markers of PAH severity. Receiver operating characteristic (ROC) curve analysis was used to determine the optimal cutoff and predictive performance of BAR. Kaplan–Meier analysis and Cox proportional hazard models were used to assess the relationship between BAR and clinical worsening.
RESULTS A total of 472 patients with idiopathic or heritable PAH were included in the study. BAR was correlated with well-validated variables that reflected severity of PAH, such as World Health Organization functional class, 6-min walk distance, N-terminal pro-brain natriuretic peptide (NT-proBNP), mixed venous oxygen saturation and cardiac index. Kaplan–Meier curves indicated that patients with BAR>3.28 had a significantly higher clinical worsening rate (log-rank test, P<0.001) than those with BAR=3.28. Multivariable Cox analysis showed that BAR could independently predict clinical worsening [Hazard ratio (HR): 1.265, 95% confidence interval (CI) 1.072–1.494, P=0.005]. Further, ROC curve analysis showed that BAR provided additional predictive value in addition to the established PAH biomarker NT-proBNP.
CONCLUSIONS BAR could reflect disease severity and was independently associated with prognosis in patients with PAH.
GW34-e1094
Sicheng Zhang, Luyang Gao, Zhihui Zhao, Qing Zhao, Qin Luo, Zhihong Liu
Center for Respiratory and Pulmonary Vascular Diseases, Fuwai Hospital, National Center for Cardiovascular Diseases, National Clinical Research Center for Cardiovascular Diseases, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing, China
OBJECTIVES Greenness has been associated with cardiovascular health, whereas higher air pollution exposure is linked with increased risks of chronic diseases. To date, limited studies have explored the interaction between residential greenness and air pollution on the clinical outcome in patients with pulmonary arterial hypertension (PAH).
METHODS This study analyzed 457 patients with idiopathic or heritable PAH recruited from Fuwai hospital. The normalized difference vegetation index (NDVI) around each participant’s residence was used to characterize greenness exposure. The air pollution data were derived from the China High Air Pollutants (CHAP) dataset, which was used to analyze individual long-term exposure to air pollution, including particulate matter with aerodynamic diameters =2.5 μm (PM2.5) and =10 μm (PM10). Cox proportional hazard models were used to describe the association of residential greenness and air pollutants with the long-term outcome of patients with PAH. Associations of residential greenness and air pollutants with PAH risk stratification and hemodynamic severity at baseline were assessed.
RESULTS Higher estimated exposure to greenness was associated with lower risk of death (hazard ratio (HR)=0.875, 95% confidence interval (CI): 0.773, 0.981 per 0.1-unit increment in NDVI) after adjusted for potential confounding variables. However, long-term exposures to air pollution were associated with higher mortality (HR=1.422, 95% CI: 1.194, 1.692 per 10 μg·m-3 for PM2.5; HR=1.318, 95% CI: 1.160, 1.497 per 10 μg·m-3 for PM10). Mediation analyses suggested that the beneficial effect of residential greenness on long-term outcomes of PAH could be partly mediated by reducing the exposure to PM2.5. In addition, an increase in residential greenness exposure was associated with lower PAH risk score at baseline, reducing the likelihood of being diagnosed within the European Society of Cardiology (ESC)/European Respiratory Society (ERS) intermediate-high risk clinical stratification. Residential greenness correlated with pulmonary hemodynamics at baseline. This association was strongest for cardiac index.
CONCLUSIONS In idiopathic or heritable PAH, higher residential greenness exposure was associated with lower mortality and higher probability of ESC/ERS low-risk categories when diagnosed, while air pollutants were positively associated with long-term mortality. Meanwhile, residential greenness may improve the outcomes of PAH by reducing exposure to PM2.5.
GW34-e1152
Luying Sun1, Xuegang Xie2, Zhenwen Yang3, Bin Jia4, Lan Wang5, Qiguang Wang6, Jiang Li7, Hong Gu8, Weiping Xie9, Nuofu Zhang10, Fenling Fan11
1Johnson and Johnson (China) Investment Ltd, Shanghai, China
2First Affiliated Hospital of Xi’an Jiaotong University, Xi’an, Shanxi, China
3Tianjin Medical University General Hospital, Tianjing, China
4Johnson and Johnson (China) Investment Ltd, Shanghai, China
5Shanghai Pulmonary Hospital, Shanghai, China
6General Hospital of Northern Theater Command, Shenyang, Liaoning, China
7The Second Xiangya Hospital of Central South University, Changsha, Hunan
8Beijing Anzhen Hospital, Capital Medical University, Beijing, China
9Jiangsu Province Hospital, Nanjing, Jiangsu, China
10The First Affiliated Hospital of Guangzhou Medical University, Guangzhou, China
11First Affiliated Hospital of Xi’an Jiaotong University, Xi’an, Shanxi, China
OBJECTIVES Current ESC/ERS Guidelines for the management of pulmonary arterial hypertension (PAH) recommend initial combination therapy with an endothelin receptor antagonist (ERA, e.g. macitentan) and a phosphodiesterase type 5 inhibitor (PDE5i, e.g. tadalafil). For patients without comorbidities and a low/intermediate risk status at diagnosis. A single-tablet fixed dose combination of macitentan 10 mg and tadalafil 40 mg (M/T FDC) would add convenience, increase adherence and ultimately improve clinical outcomes. A DUE evaluated the efficacy and safety of M/T FDC vs monotherapies in PAH. This subgroup analysis aims to explore whether the efficacy and safety of the M/T FDC in Chinese participants are consistent with the overall population.
METHODS A DUE was a multicenter, double-blind Phase 3 adaptive study (NCT03904693) and randomized WHO FC II-III PAH patients: 2:1:1 to M/T FDC, macitentan, or tadalafil, if treatment-naïve; 2:1 to M/T FDC or macitentan, if on prior ERA; and 2:1 to M/T FDC or tadalafil, if on prior PDE5i. The primary endpoint was change in pulmonary vascular resistance (PVR), expressed as the Week 16: baseline ratio of geometric means. Safety/tolerability was also assessed.
RESULTS In the A DUE study, 187 participants were randomized (108 to M/T FDC, 35 to macitentan, and 44 to tadalafil) across 16 countries/territories, including 23 participants in China (13 to M/T FDC, 5 to macitentan, and 5 to tadalafil). Demographics and baseline disease characteristics of Chinese participants were generally consistent with the overall population. As for the overall population, a higher proportion of WHO FC II patients was observed in the M/T FDC arm. In the overall population, PVR reduction with M/T FDC was significantly greater vs macitentan (29%) and vs tadalafil (28%) (Week 16 to baseline PVR ratio 0.71 [95% adjusted repeated confidence limits (RCL): 0.61, 0.82, P<0.0001] for M/T FDC vs macitentan and 0.72 [95% adjusted RCL: 0.64, 0.80, P<0.0001] for M/T FDC vs tadalafil). Consistent results were observed for the primary endpoints in Chinese participants compared with the overall population, with PVR reductions of 50% for M/T FDC vs macitentan (50%) and of 41% for M/T FDC vs tadalafil (41%) (adjusted geometric mean ratios vs. monotherapy were 0.5 [95% CL: 0.35, 0.72, P=0.0017] for M/T FDC vs macitentan and 0.59 [95% CL: 0.43, 0.80, P=0.0040] for M/T FDC vs tadalafil in Chinese participants.) No clinically important differences were observed between the overall population and Chinese participants in the incidence of AEs, SAEs, and AEs leading to premature discontinuation. In overall population, there were 3 deaths (reported as drug-unrelated) in the M/T FDC arm, no deaths were reported in Chinese participants. Adverse events (AE) leading to discontinuation, serious AE and those of special interests (anemia, hypotension, edema) were more frequent in the M/T FDC arm.
CONCLUSIONS M/T FDC significantly improved PVR vs monotherapies in PAH pts, with a safety and tolerability profile consistent with the known profile of the individual components. The observed efficacy results and safety profile for M/T FDC in the Chinese participates were generally consistent with the overall population.
GW34-e1236
Rui Wang1, Jie Wang2, Yun Wu1
1Department of General Medicine, First Affiliated Hospital of Xinjiang Medical University
2Department of Pharmacy, First Affiliated Hospital of Xinjiang Medical University
OBJECTIVES The exact mechanism by which Mfn2 participates in the interaction between the endoplasmic reticulum (ER) and mitochondria remains unclear. Based on our previous studies, we propose that Mfn2 may facilitate the transport of Ca2+ from the ER to the mitochondria by interacting with the calcium channel receptor IP3R in the mitochondria-associated endoplasmic reticulum membranes (MAMs) pathway. This process is crucial for maintaining Ca2+ homeostasis in the cytoplasm, thereby inhibiting the proliferation of pulmonary arterial smooth muscle cells (PASMCs), reducing pulmonary artery stenosis, and lowering pulmonary artery hypertension (PAH).
METHODS (1) The rat model of PAH was established using monocrotaline, and intervention with 4-PBA was conducted. Hemodynamic parameters, including mean pulmonary artery pressure and mean right ventricular pressure, were measured using a biological information acquisition system. Pulmonary vascular remodeling changes were observed through hematoxylin-eosin staining, and relevant indicators of pulmonary vascular remodeling were quantified using ImageJ pathological analysis software. Mitochondrial status in PASMCs was examined under electron microscopy. The mRNA and protein expression levels of key factors involved in the endoplasmic reticulum stress (ERS) pathway were measured using qPCR and Western blotting techniques. (2) In the inflammation-induced injury model of PASMCs stimulated by TNF-α, adenoviral transduction was employed to overexpress or silence Mfn2. Furthermore, intervention with the IP3R3 inhibitor Xestospongin C was performed. Under electron microscopy, the fusion and fission status of mitochondria were observed. Apoptosis and proliferation were assessed using flow cytometry and the CCK-8 assay. The concentrations of Ca2+ in the endoplasmic reticulum, cytoplasm, and mitochondria were measured using fluorescent probes. The mRNA and protein expression levels of key factors involved in the endoplasmic reticulum-mitochondria pathway for Ca2+ transport were determined using qPCR and Western blotting.
RESULTS (1) Following 4-PBA intervention in monocrotaline-induced rat models for PAH, there was a significant decrease in mPAP (53.89±4.69 vs 22.34±1.21) and mRVP (29.61±1.94 vs 17.24±1.59). Additionally, there was a notable improvement in pulmonary arteriole remodeling. The extent of normal mitochondrial structural damage was reduced, and mitochondrial autophagy was decreased. The expression levels of key factors involved in the ERS pathway (Perk/elF2α/Atf4/CHOP) were suppressed, while the expression levels of Mfn2 and IP3R3 were upregulated. (2) In the TNF-α-induced inflammatory injury model of PASMCs, compared to the TNF-α group, overexpression of Mfn2 resulted in an increased number of mitochondria with normal structure and intact mitochondrial cristae. The concentration of Ca2+ within the mitochondria significantly increased, while the cytoplasmic Ca2+ concentration decreased significantly. The expression of key factors involved in ERS pathway, including Perk/elF2α/Atf4/CHOP, was downregulated. Additionally, the expression levels of Drp1, OPA1, and IP3R3 were also decreased. Following the addition of the IP3R3 inhibitor Xestospongin C to the TNF-α group, cell proliferation was suppressed, and apoptosis was increased. The concentration of Ca2+ within the mitochondria increased, while the cytoplasmic Ca2+concentration decreased. The expression of eIF2a, CHOP, and VDAC1 were inhibited, while the mRNA and protein expression of Mfn2 and SEACA were upregulated.
CONCLUSIONS Targeted intervention of Mfn2 can influence intracellular Ca2+ transport by interacting with the key calcium transporter protein IP3R3 on MAMs, thereby reducing calcium overload in PASMCs during PAH and maintaining intracellular Ca2+ homeostasis. This mechanism inhibits excessive proliferation of PASMCs and provides a new perspective for targeted therapy in PAH.
GW34-e1258
Lu Yan, Zhihui Zhang
Department of Cardiovascular Medicine, and the Center for Circadian Metabolism and Cardiovascular Disease, Southwest Hospital, Army Medical University
OBJECTIVES Pulmonary arterial hypertension (PAH) is a life-threatening condition with poor prognosis. Due to its high fatality rate and limited therapies, the pathogenesis of PAH needs to be explored urgently.
METHODS Transcriptome data of patients including PAH from the GSE113439, GSE117261 and GSE15197 were downloaded from the Gene Expression Omnibus (GEO) database. The differentially expressed genes (DEGs) were identified. Hub genes were identified by constructing a protein-protein interaction (PPI) network with the Search Tool for the Retrieval of Interacting Genes/Proteins (STRING) and analyzing in Cytoscape. Further, gene ontology (GO) functional enrichment and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway annotation were performed. CIBERSORT was used to compare immune cells composition in PAH and control groups. Correlations between immune cells and hub genes was analyzed in GSE113439 and validated in GSE117261.
RESULTS Four hundred and fifty-six integrated DEGs with 205 up-regulated and 251 down-regulated were identified, which are mainly enriched in inflammation and immune response GO terms and KEGG pathways, including cell chemotaxis, myeloid leukocyte migration and IL-17 signaling pathway. Forty-four hub genes were selected in top PPI modules. The percentages of resting memory CD4+ T cells and activated dendritic cells were significantly higher in PAH tissues; resting NK cells, monocytes and neutrophils were significantly lower.
CONCLUSIONS Resting memory CD4+ T cells, activated dendritic cells, resting NK cells, monocytes and neutrophils may play important roles in immune responses in the development of PAH. CCL5, ITGAM, FGR, CCL7, S100A8, CCR1, HMOX1 contributing to the pathogenesis of PAH via diverse mechanisms.
GW34-e1282
Jiang Yu, Yongzheng Guo, Suxin Luo
Division of Cardiology, The First Affiliated Hospital of Chongqing Medical University
OBJECTIVES Pulmonary vascular leakage is a prevalent outcome of sepsis, resulting in acute lung injury. We have reported a positive correlation between S100 calcium-binding protein (S100) A8/S100A9 and inflammation-associated damage before. However, limited knowledge exists concerning the biological role of S100A8/A9 in pulmonary vascular endothelial barrier dysfunction, as well as the diagnostic and prognostic value of S100A8/A9 in sepsis.
METHODS Sepsis was induced in C57BL/6J mice and S100A9-knockout (KO) mice lacking the S100A8/A9 heterodimer through the cecal ligation and puncture (CLP) technique. Disease severity was evaluated using the rectal temperature, carotid pressure, and survival rate. Pulmonary vascular leakage was determined by measuring extravasated Evans blue (EB). Reverse transcription polymerase chain reaction and the histological score were used to evaluate inflammation and lung injury, respectively. Immunofluorescence staining and Western blotting were used to evaluate occludin and vascular endothelial (VE)-cadherin expressions in the lungs. Recombinant S100A8/A9 (rhS100A8/A9) was used to identify the effects of S100A8/A9 on endothelial barrier dysfunction in human umbilical vein endothelial cells (HUVECs). Additionally, the diagnostic and prognostic value of S100A8/A9 in sepsis was assessed using receiver operating characteristic and Kaplan–Meier curves.
RESULTS S100A8/A9 expression was up-regulated in the lungs of CLP-operated mice. S100A9 KO in mice significantly reversed CLP-induced hypothermia and hypotension, resulting in an improved survival rate compared with wild-type mice. S100A9 KO also decreased the inflammatory response, EB leakage, and histological scores in the lungs of CLP-operated mice. Occludin and VE-cadherin expressions were decreased in the lungs of CLP-operated mice; however, S100A9 KO attenuated this decrease. Moreover, CLP-induced signal transducer and activator of transcription 3 (STAT3) and p38/extracellular signal-regulated kinase (ERK) signalling activation and apoptosis were mitigated by S100A9 KO in lungs. In addition, rhS100A8/A9 administration significantly decreased occludin and VE-cadherin expressions, increased the phosphorylated (p)-ERK/ERK, p-p38/p38, and B-cell leukaemia/lymphoma 2 protein (Bcl-2)-associated X protein/Bcl-2 ratios in HUVECs. Finally, our findings further support the potential of S100A8/A9 as a diagnostic and prognostic biomarker for human sepsis.
CONCLUSIONS The present study demonstrated a novel role of S100A8/A9 in the progression of sepsis-induced pulmonary inflammation, vascular permeability, and lung injury. This was achieved, at least partially, by activating the P38/STAT3/ERK signalling pathways. Therefore, these findings provide valuable insights into a potential therapeutic target for preventing and treating sepsis-induced pulmonary vascular leakage and lung injury.
DIABETES, CEREBROVASCULAR DISEASES, KIDNEY-DISEASES, CARDIO-ONCOLOGY
GW34-e0053
Houyong Zhu1, Xiaoqun Xu2
1Hangzhou TCM Hospital Affiliated to Zhejiang Chinese Medical University
2Affiliated Hangzhou Chest Hospital, Zhejiang University School of Medicine
OBJECTIVES The purpose of this study was to evaluate the predictive value of the inflammation based Glasgow Outcome Scale (GPS) for cardiovascular death in patients with diabetes.
METHODS This study included 4956 patients (=18 years old) with diabetes in the National Health and Nutrition Survey from 1999 to 2010. The death result is determined by the correlation with the national death index on December 31, 2019. GPS is composed of serum C-reactive protein and albumin. The primary outcome was cardiovascular death and the secondary outcome was all-cause death. The Cox proportional risk model adjusted for demographic factors and traditional cardiovascular risk factors was used to analyze the cumulative risk of outcomes.
RESULTS Among 4956 diabetes patients with a median follow-up of 10.9 years, 601 cardiovascular deaths and 2187 all-cause deaths were recorded. After adequate model adjustment, compared with the low GPS group, the high GPS group [HR, 1.257 (1.007–1.570), P=0.043] had a higher cardiovascular mortality. Compared with the low GPS group, the all-cause mortality of the high GPS group [HR, 1.394 (1.245–1.560), P<0.001] was higher. The trend of subgroup analysis was consistent with that of the overall cohort.
CONCLUSIONS The high level of baseline GPS is closely related to the increased risk of cardiovascular and all-cause death in patients with diabetes. GPS may be a convenient and efficient clinical practical risk assessment tool for diabetes patients.
GW34-e0064
Chan Li1, Zhaoya Liu2
1Xiangya Hospital, Central South University
2Third Xiangya Hospital, Central South University
OBJECTIVES Patients with diabetes mellitus (DM) has a higher mortality and morbidity of ischemic heart disease (IHD). This study aimed to analyze the quantitative analyze the trends and topics of publications related to DM-related IHD in the past three decades.
METHODS Publications indexed under the Medical Subject Headings (MeSH) term “Diabetes Mellitus” and “Myocardial Ischemia” from 1992 to 2021 were obtained from PubMed. Metadata were extracted by R package for bibliometric analyses. MeSH analyses and Latent Dirichlet allocation (LDA) were conducted to speculate research hot spots and establish a topic network to illustrate their relationships.
RESULTS A total of 17,850 publications were included in this study, the quantity of publications constantially increased and reached the peak in 2004. Most of the studies were clinical studies. In MeSH analyses, coronary angiography, body mass index, and biomarkers were recognized as the leading terms of diagnosis. Hypoglycemic agents, coronary balloon angioplasty, and coronary artery bypass were the top terms of treatment. Stroke, life style and recurrence were the top terms of prognosis. Three clusters, including diagnosis and manifestation, epidemiology and mechanism, and treatment and prognosis, were identified by LDA analyses. Most studies focused on the treatment and prognosis of DM-related IHD. Risk factors and biomarkers were the most studied topics.
CONCLUSIONS The annual publication on DM-related IHD scarcely changed during the past two decades. Risk factors and biomarkers attracted most attentions in the field of DM-related IHD. More basic studies should be performed to figure out the associations and specific mechanisms between DM and IHD.
GW34-e0123
Rui Wang1,2,3, Ya-Rong Zhang1,2,3, Yong-Fen Qi1,2,3
1State Key Laboratory of Vascular Homeostasis and Remodeling, Peking University, Beijing 100191, China
2Laboratory of Cardiovascular Bioactive Molecule, School of Basic Medical Sciences, Peking University, Beijing, China
3Department of Pathogen Biology, School of Basic Medical Sciences, Peking University Health Science Center, Beijing, China
OBJECTIVES Vascular calcification is an independent risk factor for cardiovascular events and an important cause of death and disability in patients with type 2 diabetes mellitus (T2DMs). Vascular calcification is common in diabetic patients and multiple factors promote the occurrence and development of diabetic vascular calcification. However, there is still no effective treatment for diabetic vascular calcification. As a new cardiovascular bioactive peptide, intermedin (IMD)/adrenomedullin 2 (ADM2) plays a protective role in cardiovascular disease. Our previous work showed that IMD could inhibit chronic kidney disease- and age-related vascular calcification. However, the role and mechanism of IMD in diabetic vascular calcification remain unknown.
METHODS Plasma of T2DM patients and healthy controls were used for detecting IMD contents. In vivo, male Sprague-Dawley (SD) rats and Adm2 knockout/transgenic mice were used in the experiment to induce diabetic vascular calcification, IMD was administered with subcutaneous Alzert mini-osmotic pumps in the last 4 weeks of the experiment. In vitro, rat and mouse vascular smooth muscle cells (VSMC) were treated to induce VSMC calcification. Histochemical staining was used to observe vascular morphology and calcium deposition in blood vessels. Enzyme-linked immunosorbent assay, colorimetric method and Western blot were performed to detect the levels of diabetes and vascular calcification-related molecules.
RESULTS IMD level was decreased in the plasma of diabetic patients and rats and calcified rat aortas. Meanwhile, Adm2 mRNA expression was significantly down-regulated in calcified aortas of diabetic rats, and its receptor calcitonin receptor-like receptor (CRLR) and receptor activity modifying protein 1 (RAMP1) were increased, RAMP3 was decreased. Exogenous IMD reduced fructose-induced calcium deposition, calcium contents and expression levels of VSMC osteogenic markers in diabetic rat aortas. Endogenous Adm2 deficiency aggravated and Adm2 overexpression attenuated vascular calcium deposition and alkaline phosphatase (ALP) activity in diabetic mice induced by high-fat diet. In addition, IMD significantly inhibited the high glucose induced rat vascular smooth muscle cell (VSMC) calcium deposition, calcium contents, ALP activity and expression levels of osteogenic markers. Adm2 deficiency exacerbated mouse VSMC calcium deposition and further decreased the expression levels of contractile markers induced by high glucose. Further studies showed that IMD diminished ROS production in diabetic rat calcified aortas and high glucose-treated VSMC. IMD reduced diabetic vascular calcification by inhibiting the rat plasma advanced glycation end products (AGEs) concentration and reducing the receptor for AGEs (RAGE) expression level in calcified aortas and VSMCs.
CONCLUSIONS IMD alleviated diabetic vascular calcification by inhibiting AGEs/RAGE and subsequent oxidative stress.
GW34-e0128
Zidun Wang, Huiyuan Qin, Minglong Chen
Division of Cardiology, The First Affiliated Hospital of Nanjing Medical University, Nanjing, China
OBJECTIVES Previous clinical studies have shown that ibrutinib increases the risk of atrial fibrillation (AF) in chronic lymphocytic leukemia (CLL) patients. However, the mechanism underlying ibrutinib-related AF is not fully understood. Herein, we investigated the proportion of patients with new-onset AF in a CLL cohort treated with ibrutinib.
METHODS Optical mapping was conducted to reveal the proarrhythmic effect of ibrutinib on HL-1 cells. The expression levels of connexins 43 and 40 (Cx43 and Cx40) were further compared between cells that were treated with ibrutinib or the blank control using fluorescence staining and western blotting. Moreover, autophagy-related protein expression was investigated, and an inhibitor of autophagy was administered to reverse the detrimental effect of ibrutinib.
RESULTS As a result, 5% of patients developed atrial arrhythmias after ibrutinib administration. HL-1 cells that were treated with ibrutinib had significantly lower conduction velocity and higher incidence of reentry-like arrhythmia than control cells. In parallel, the expression of Cx43 and Cx40 was significantly decreased and the levels of autophagy markers were increased in HL-1 cells that were treated with ibrutinib. Inhibition of autophagy increased the expression of Cx43 and Cx40. Hence, ibrutinib could induce connexin degradation, thus contributing to atrial arrhythmia, by inducing autophagy via its off-target effect on the PI3K-AKT-mTOR signaling pathway.
CONCLUSIONS Our study provided a new perspective on the mechanisms by which ibrutinib causes atrial arrhythmias and potential intervention targets for treatment.
GW34-e0231
Zhiyue Liu, He Huang
West China hospital, Sichuan University
OBJECTIVES Cancer therapy-related cardiovascular toxicity (CTR-CVT) is a major contributor to poor prognosis in breast cancer (BC) patients undergoing chemotherapy. Left ventricular global longitudinal strain (LV GLS) has predictive value for CTR-CVT, while few researchers take into account late-onset CTR-CVT. This study sought to provide a guide for the prediction of late-onset CTR-CVT in primary BC over the 2 years follow-up via strain and contrast-enhanced echocardiography.
METHODS Anthracycline and anthracycline + targeted medication groups were created from 111 patients with stage I–III primary BC who were prospectively included. The left ventricular diastolic function, LV global long-axis strain (GLS); left ventricular ejection fraction by contrast-enhanced echocardiography (c-LVEF), and electrocardiograms were collected at baseline, 3, 6, 12 and 24 months after the start of cancer treatment. The high-sensitivity troponin-T and NT-pro BNP at baseline and 3 months after chemotherapy were measured.
RESULTS 1) LV GLS decreased in BC patients over time. 2) After 12 months’ follow-up, the LV GLS in the anthracycline+ targeted group was lower than in the anthracycline group. After 24 months’ follow-up, the GLS and c-LVEF in the anthracycline + targeted group declined while the E/e’ increased. 3) Decreased LVEF (56%) and arrhythmia (38%) are the common causes of CTR-CVT. Lower LVEF was a major factor in late-onset CTR-CVT. 4) Combination of LV GLS and c-LVEF at 3 months were used as predictors for CTR-CVT exhibited a higher AUC than either one alone (AUC=0.929, 95% CI: 0.863–0.970). LV GLS at 3 months can predict the late-onset CTR-CVT (AUC=0.745, P<0.001) and the cut-off is 20.32%.
CONCLUSIONS As time went on, the systolic and diastolic dysfunction of BC patients get worsened. The combination of LV GLS and c-LVEF is better in the prediction of CTR-CVT. Only the LV GLS at 3 months can predict the late-onset CTR-CVT.
GW34-e0317
Zhihui Zhu1,2, Chenyu Li1
1Medizinische Klinik und Poliklinik IV, Klinikum der Universität München
2Beijing Anzhen Hospital, Capital Medical University
OBJECTIVES Dual inhibition of the renin-angiotensin system (RAS) and sodium-glucose transporter (SGLT)-2 as well as dual RAS and mineralocorticoid receptor (MR) antagonism have demonstrated renoprotective effects in large clinical trials but whether a triple combination is even more potent is unclear. We hypothesized that triple RAS/SGLT2/MR blockade would be superior to dual RAS/SGLT2 blockade in prolonging kidney lifespan.
METHODS We performed a preclinical randomized controlled trial in Col4a3-deficient mice with spontaneous and progressive CKD (registry ID: PCTE0000266). At 6 weeks of age 50% male and 50% female mice were randomized in a 1:1:1:1 manner to 8 weeks of nil or food admixes of ramipril (exposure dose 10 mg/kg), ramipril plus empagliflozin (30 mg/kg) or ramipril, empagliflozin plus finerenone (10 mg/kg). The prespecified primary endpoint was total lifespan to uremic death. Ancillary studies examined empagliflozin and finerenone monotherapy, baseline histology, and mechanisms.
RESULTS At the time of randomization, Col4a3 −/− mice had albuminuria, elevated serum creatinine, glomerulosclerosis, tubular atrophy, and interstitial fibrosis. Total lifespan was 63.7±10.0 days (vehicle), 77.3±5.3 days (ramipril), 80.3±11.0 days (ramipril+empagliflozin), and 103.1±20.3 days (triple therapy), respectively. Total lifespan was 70.4±9.2 for empagliflozin and 71.1±7.1 for finerenone monotherapy, respectively. Histopathology and RNA sequencing analysis documented a potent anti-sclerotic, anti-inflammatory, and fibrotic effect of the triple combination.
CONCLUSIONS Adding finereone to dual RAS/SGLT2 blockade significantly prolongs uremia-free lifespan even when started at an advanced stage of Alport nephropathy. Triple RAS/SGLT2/MR blockade could be a potent treatment strategy to prolong uremia-free lifespan in patients with CKD related to Alport syndrome and possibly other progressive kidney disorders.
GW34-e0321
Kaiyi Chi1, Tianwang Guan2
1Department of Clinical Medicine, The Second Clinical College of Guangzhou Medical University
2Department of Cardiology, Laboratory of Heart Center, Zhujiang Hospital, Southern Medical University
OBJECTIVES Cardiovascular disease (CVD) is the leading cause of death worldwide. Of concern, older survivors with female-specific cancer (including breast and gynecologic cancers) may have an increased risk of CVD death, owing to estrogen deficiency, anticancer-therapy cardio-toxicities and common risk factors of cancer and CVD. Thus, more attention should be focused on the CVD death risk of older survivors with female-specific cancer. However, there are relatively few studies concerning CVD death risk of older patients with female-specific cancer. Therefore, we performed a comprehensive analysis to evaluate the risk of CVD death in 3 highly prevalent cancers among older female patients with the following two objectives: 1) to quantify the risk of CVD death in the older female-specific cancer compared to the general older population; 2) to identify and characterize age subgroups with a higher CVD death risk. These findings will provide a scientific evidence for CVD death risk management strategies for older survivors with female-specific cancer.
METHODS Older patients (over 65 years) of female-specific cancer diagnosed between 1975 and 2018 were included. The common of cancer in older female, including breast, endometrium and vulva, was defined as female-specific cancer. The proportion of deaths, competing-risk regressions models, standardized mortality ratios (SMRs), and absolute excess risks (AERs) were used to assess the risk of CVD deaths among older patients with female-specific cancer.
RESULTS This study included 230,742 older female patients (195,106 breast cancers, 30,862 endometrium cancers and 4774 vulva cancers). The risk of CVD death exceeded primary neoplasm death in older patients with cancers of the breast, endometrium and vulva over time. Compared to the general older population, increased SMR and AER of CVD death were observed in older patients with breast cancer (SMR 1.24–4.20; AER 36.45–487.77), endometrium cancer (SMR 1.17–3.56; AER 26.03–390.28) and vulva cancer (SMR 2.41–5.76; AER 215.27–725.50). Older patients with female-specific cancer diagnosed after more than 15 years had higher risk of CVD death than those diagnosed after less than 15 years. Compared to the general older population, older breast cancer had increased SMR and AER of CVD death in those diagnosed at 65–74 years (SMR 2.19–14.09; AER 45.80–505.43), those diagnosed at 75–84 years (SMR 1.61–8.85; AER 81.29–1050.73) and those diagnosed at 85+ years (SMR 1.36–3.99; AER 196.11–1634.60). Compared to the general older population, older endometrium cancer had increased SMR and AER of CVD death in those diagnosed at 65–74 years (SMR 1.71–12.01; AER 27.56–424.94), those diagnosed at 75–84 years (SMR 1.46–7.10; AER 61.40–816.55) and those diagnosed at 85+ years (SMR 1.34–4.54; AER 184.91–1934.47). Compared to the general older population, older vulva cancer had increased SMR and AER of CVD death in those diagnosed at 65–74 years (SMR 3.68–18.40; AER 103.65–671.81), those diagnosed at 75–84 years (SMR 2.47–10.63; AER 196.20–1289.73) and those diagnosed at 85+ years (SMR 1.56–5.10; AER 307.24–2243.97).
CONCLUSIONS For older cancers patients, common female-specific cancers including breast, endometrium and vulva were at high risk of CVD death. With the extension of follow-up years after diagnosis, the risk of CVD death may increase correspondingly. Strategies for long-term cardiovascular care in older patients with female-specific cancer are needed.
GW34-e0346
Haowei Li1, Shige Qi2, Zhihui Wang2, Miao Liu3, Yao He1,4
1Institute of Geriatrics, Beijing Key Laboratory of Aging and Geriatrics, National Clinical Research Center for Geriatrics Diseases, Second Medical Center of Chinese PLA General Hospital, Beijing 100853, China
2National Center for Chronic and Non-Communicable Disease Control and Prevention, Chinese Center for Disease Control and Prevention, Beijing 100050, China
3Department of Statistics and Epidemiology, Graduate School of PLA General Hospital, Beijing 100853, China
4State Key Laboratory of Kidney Diseases, Beijing 100853, China
OBJECTIVES Cardiometabolic diseases (CMDs), including diabetes, coronary heart diseases, stroke and hypertension, increases the risk of cognitive decline, but the extent to which this can be offset by adherence to an active integrated lifestyle is unknown. We aimed to associations of CMDs with early cognitive decline and explore the role of integrated lifestyle in this association.
METHODS The participants were from PINDEC Project. A total of 2537 dementia-free older adults≥60 years old who completed the 2015 baseline survey and the 2017 follow-up survey with a cognitive function status assessment were included. Lifestyle factors (including physical exercise, social interaction, leisure activities, sleep quality, smoking status, and alcohol consumption) were collected through questionnaires and the integrated score was calculated. Participants were divided into three groups based on integrated score tertiles (inactive, =3 score; intermediate, 3 score; and active,≥5). Logistic regression was used in data analysis.
RESULTS 35.2% participants had 5–6 healthy components, while only 5.4% of all the participants had all 6 healthy lifestyles. The multiadjusted odds ratios (ORs, 95% confidence interval) of early cognitive decline was 1.223 (0.799–1.871) and 1.832 (1.140–2.943) for participants who had only one CMD and any two or more CMDs, respectively. For integrated lifestyle scores, an inverse dose-response relationship was found between lifestyle scores and early cognitive decline (Ptrend=0.017). In participants with active lifestyle, the OR for early cognitive decline comparing the CMDs status of any two or more CMDs vs. CMDs-free was 0.778 (95% CI: 0.302–2.007). Participants with inactive lifestyle and any two or more CMDs had a near 3.4-fold increased risk of early cognitive decline than those without CMDs who had intermediate to active lifestyle (OR=3.435, 95% CI: 1.772–6.660).
CONCLUSIONS A dose–response relationship exists between CMDs status and risk of early cognitive decline. However, adherence to an active integrated lifestyle may mitigate this risk.
GW34-e0435
Lihua Liao1, Xingmo Dong2, Yani Wang1, Xueqin Lin1, Ying Liao2, Huaijing Luo2, Yu Yi2
1The Third Clinical Medical College, Fujian Medical University
2Longyan First Affiliated Hospital of Fujian Medical University
OBJECTIVES The incidence of kidney stone disease has increased worldwide over the past decades, resulting in high medical costs and social burden. Kidney stone disease shares some common features with the risk factors of cardiovascular diseases (CVDs). Here, we investigated the association between cardiovascular health (CVH) based on the Life’s Essential 8 (LE8) score developed by the American Heart Association and the incidence of kidney stone disease in US adults.
METHODS We analyzed the data of 29,469 US adults aged 20 years or above from the National Health and Nutrition Examination Survey, 2007–2018. The components of LE8 included diet, physical activity, nicotine exposure, sleep health, body mass index, blood lipids, blood glucose, and blood pressure. According to the LE8 score, CVH was divided into three categories: poor, intermediate, and ideal. Logistic regression was used to determine the association between CVH and the incidence of kidney stone disease by estimating odds ratios (ORs) and 95% confidence intervals (CIs).
RESULTS The average age of the participants was 48.6 years, and 50% of the participants were women. The numbers of participants with poor, intermediate, and ideal CVH were 4149, 19,782, and 5538, respectively. After adjusting for related confounding factors, ideal CVH was associated with a reduction in the odds of kidney stone occurrence as compared to poor CVH (adjusted OR [aOR]: 0.45, 95% CI: 0.35–0.57, P<0.001). Moreover, if the ideal CVH score was≥6, the odds of kidney stone occurrence decreased by up to 61% (aOR: 0.39, 95% CI: 0.30–0.51).
CONCLUSIONS In the present study, ideal CVH, a factor indicative of a healthy lifestyle, was associated with lower odds of kidney stone occurrence. These findings suggest that interventions to prevent CVDs are also expected to prevent kidney stone occurrence.
GW34-e0579
Hongzhao You, Heng Zhang, Xiaojue Li, Xinxing Feng, Rong Guo, Hansong Sun, Wei Feng, Guangwei Li, Zhe Zheng, Yanyan Chen
Fuwai Hospital, National Center for Cardiovascular Diseases, Chinese Academy of Medical Sciences and Peking Union Medical College
OBJECTIVES Uncontrolled blood glucose level is a risk factor for postoperative complications after coronary artery bypass grafting (CABG). However, the association between postoperative blood glucose fluctuation and in-hospital adverse events in patients with diabetes remains unknown. The Largest Amplitude of Glycemic Excursions (LAGE), which serves as a reliable indicator of short-term blood glucose fluctuation, is acknowledged as a potent risk factor for diabetes-related complications. We aimed to determine the effect of post-operative blood glucose variability on in-hospital adverse events in diabetic patients undergoing CABG.
METHODS We reviewed medical records for diabetic patients who received CABG at Fuwai Hospital from January 2011 to December 2014. The variability of postoperative glucose during the initial 5 days was gauged by LAGE. Subgroups were divided based on whether the LAGE was equal to or above 4.4 mmol/L. The primary endpoint encompassed a composite endpoint incorporating in-hospital all-cause mortality and major cardiovascular complications. The secondary endpoint encompassed major cardiovascular complications such as acute myocardial infarction, stroke, and acute kidney injury.
RESULTS The study enrolled 3387 eligible patients, among which 241 (7.1%) patients met the composite endpoint. When compared to patients with LAGE below 4.4 mmol/L, the ones with LAGE equal to or above 4.4 mmol/L exhibited a higher rate of both the composite endpoint (6.2 vs 10.2%, P<0.001) and major vascular complications (6.1 vs 10.2%, P<0.001). Moreover, LAGE equal to or above 4.4 mmol/L is an independent risk factor for the composite endpoint (OR=1.69, 95% CI 1.26–2.25, P<0.001) and major vascular complications (OR=1.74, 95% CI 1.30–2.32, P<0.001) after adjusting for the established risk factors (age, sex, BMI, smoking, low density lipoprotein cholesterol, chronic renal failure, congestive heart failure, stroke, previous myocardial infarction and peripheral vascular diseases).
CONCLUSIONS Patients who exhibit a LAGE equal to or above 4.4 mmol/L are subject to an elevated risk of in-hospital adverse events. Relatively large blood glucose fluctuations (LAGE equal to or above 4.4 mmol/L) after CABG is significantly associated with in-hospital adverse events in patients with diabetes who underwent CABG.
GW34-e0640
Cong Liu, Xiaozeng Wang
General Hospital of Northern Theater Command
OBJECTIVES Vascular stiffness increased and vascular compliance decreased on vasculature is a progressive process in diabetics. The pathogenesis underlying diabetic-induced vasculature damages is complex and multifactorial. This paper aim to clarify putative link between aorta abdominal and central retinal artery and the therapeutic implication of antioxidant N-acetylcysteine.
METHODS Forty-two male C57BL/6 mice (20–25 g) were included. Thirty mice were injected intraperitoneally with 150 mg/kg of STZ (STZ, Sigma-Aldrich, St. Louis, MO) in 0.05 M citrate buffer (pH 4.2). As a control group and an only treated with NAC group, 12 mice were injected with equal volume of citrate buffer. Mice with blood glucose levels exceeding 13.5 mmol/L were considered diabetic after five days. The 30 diabetic mice were divided into 5 groups with 6 animals in each group: including DM group (diabetes without NAC treatment), and 4 different NAC treatment groups, namely NAC1, NAC3, NAC5 and NAC7, with the number defining the initiation time of NAC treatment (NAC treatment was done via drinking water starting from 1week, 3weeks, 5 weeks and 7 weeks after STZ injection till the end of the 12 weeks, respectively). The 12 non-DM mice were either untreated (Ctrl) or treated with NAC from 5 weeks (only NAC treated). Vascular function and structure were analyzed by both ultrasound and histology. Body weight and blood glucose level were measured regularly, and the mice were sacrificed at the experimental end points. Aorta abdominal and central retinal artery were collected for Hematoxylin Eosin (HE) and Trichrome staining and ROS staining of all mice after ultrasound.
RESULTS We found that abdominal aorta, retina structure and function were impaired, coupled with endothelial cells, smooth muscle cell and cellular matrix impaired 12 weeks after STZ induction. NAC treatment significantly reduced ROS generation, and suppressed abdominal aorta remodeling and retina damage. Strikingly, the earlier and longer treatment, produced significant improvement of vascular function and structure.
CONCLUSIONS Our study demonstrated that earlier NAC treatment in diabetic effectively defends from diabetic vasculature disease through inhibiting the ROS dependent artery stiffness and restruction, which warrants further clinical trial.
GW34-e0657
Yilan Li, Yao Zhang
Department of Cardiology, The Second Affiliated Hospital of Harbin Medical University,
OBJECTIVES Sorafenib is a standard first-line treatment for advanced hepatocellular carcinoma. However, the serious cardiotoxic effect restricted its clinical application. We previously have illuminated the protective role of canonical Slc7a11/GPX4 signaling in sorafenib-induced cardiotoxicity. Meanwhile, ferroptosis suppressor protein 1 (FSP1) is recently identified as a seminal breakthrough owing to its property of versus ferroptosis. In this study, we investigated the protective effect of FSP1 in sorafenib-induced cardiotoxicity in vitro and in vivo.
METHODS The expression of FSP1 was significantly decreased in rat cardiomyocyte (H9c2), mouse cardiomyocyte (HL-1) (5 μM) cell lines and mice heart (30 mg/kg/injection for 14 days i.p.), which can be partially prevented by treatment with ferroptosis inhibitor Ferrostatin-1(Fer-1).
RESULTS Sorafenib reduced levels of ferroptotic markers involving glutathione peroxidase 4 (GPX4), increased PTGS2, reactive oxygen species (ROS), malonaldehyde (MDA), apart from causing obvious mitochondria damage, which were alleviated by Fer-1. Overexpression of FSP1 protected cells from ferroptosis, while knock-down of FSP1 made cardiomyocytes sensitive to ferroptosis caused by sorafenib. Meanwhile, we found that knockdown for FSP1 showed similar results to GPX4 knockdown, and FSP1 overexpression can largely abrogate GPX4 inhibition-induced ferroptosis. iFSP1, a potent FSP1 inhibitor, was used to induce ferroptosis in GPX4 knockout cells. We found iFSP1 selectively induced ferroptosis in GPX4-knockout cells that overexpressed FSP1.
CONCLUSIONS Based on these findings, we concluded that FSP1-mediated ferroptosis is one of the key mechanisms leading to sorafenib-induced cardiotoxicity. Targeting ferroptosis may be a novel therapeutic approach for preventing sorafenib-induced cardiotoxicity in the future.
GW34-e0800
Yemin Li1,2, Tianwang Guan3,4, Caiyun Ou5
1Department of Clinical Medicine, The First Clinical College of Guangzhou Medical University, Guangzhou 510182, China
2Cardiovascular Medicine and Cardio-Oncology Group, Medical Exploration and Translation Team, Guangzhou 510000, China
3Department of Cardiology, Laboratory of Heart Center, Zhujiang Hospital, Southern Medical University, Guangzhou 510280, China
4Dongguan Hospital of Southern Medical University (Dongguan People’s Hospital), Southern Medical University, Guangdong Provincial Key Laboratory of Cardiac Function and Microcirculation, Guangzhou 510280, China
5The 10th Affiliated Hospital of Southern Medical University (Dongguan People’s Hospital), Southern Medical University, Guangdong Provincial Key Laboratory of Cardiac Function and Microcirculation, Guangzhou 510280, China
OBJECTIVES Penile cancer is a rare malignancy, with almost 35,000 new cases worldwide every year. The incidence peak is around 60 years of age and increases with increasing age. Recently, Cardiovascular disease (CVD) has become the leading cause of death for cancer patients, which should receive more attention in older patient with penile cancer. Neoadjuvant chemotherapy as the strategy of treatment before surgical consolidation may exposes patients to cardiac toxicity, such as paclitaxel, ifosfamide and cisplatin. In addition, shared risk factors between penile cancer and CVD suggest CVD as a frequent co-morbidity of penile cancer, including chronic inflammation and smoking. Therefore, reinforcing our understanding and management of CVD may help improve survival in older patients with penile cancer. However, due to the rarity of this disease, there is still a lack of research to explore the risk of CVD death in older patients with penile cancer. Our study aimed to assess the risk of CVD death in older patients with penile cancer.
METHODS Older patients with penile cancers aged over 65 years were included in the study. The proportions of deaths, competing-risk regressions models, standardized mortality ratios (SMRs), and absolute excess risks (AERs) were utilized to evaluate the risk of CVD death.
RESULTS This study included 1283 older patients with a median follow-up of 13.5 years. Older patients with penile cancer were at a higher risk of CVD death. The proportions of CVD death increased witha peak in 5 to 10 years following diagnosis (except subgroup age 85+) and the proportions of primary neoplasms death tended to decrease. The cumulative mortality from CVD death exceeded primary neoplasm death 5–10 years after the cancer diagnosis. Compared with the general older population, older patients with penile cancer had higher risk of CVD death with increased SMR and AER (SMR 1.70–9.13; AER 133.01–1145.74).
CONCLUSIONS For older patients, penile cancer was at a higher risk of CVD death which exceeded primary neoplasm death over the survival time and became the major cause of death. Long-term strategies for cardiovascular care and a multidisciplinary approach in older patients with penile cancer are imperative.
GW34-e0964
Yan Wei, Xiaoping Wang, Guanjing Ling, Yong Wang
Beijing University of Chinese Medicine
OBJECTIVES Doxorubicin (DOX), as a first-line chemotherapy agent, plays an irreplaceable role in anti-tumor therapy. However, cumulative doses of DOX can cause severe cardiotoxicity even put cancer patients at risk for developing dilated cardiomyopathy. Up to now, the precise molecular mechanisms underlying the progression of DOX-induced cardiac remodeling (DICR) remain puzzling. Mitochondrial bioenergy is critical for maintaining healthy cardiac function. Previously, we have demonstrated that Traditional Chinese medicine compound of Qishen granule (QSG), which is composed of Radix Astragali mongolici, Radix Salvia miltiorrhizabunge, Flos Lonicerae, Radix Scrophulariae, Radix Aconiti Lateralis Preparata, and Radix Glycyrrhizae, possesses beneficial effects on mitochondrial ecosystem. Here, we aim to evaluate whether QSG exerts protective effects on DICR and identify the deep regulatory mechanism.
METHODS C57BL/6 mice were injected with DOX (5 mg/kg) through the tail vein once every 7 days for 4 weeks to establish the DIC model. In vivo, the mice were divided into four groups, control group, model group, QSG group and Enalapril (positive drug) group. The protective effect of QSG on the heart was evaluated by echocardiographic assessment, histological examination, myocardial hypertrophy and energy metabolism gene (qPCR) and pathway protein (Western blots) analysis. Mechanistically, H9c2 cells were induced with 1 μmol/L DOX, combined with Rev-erbα siRNA, Cpt1b siRNA model, exerted Seahorse analysis and Mito-SOX immunofluorescence for mitochondrial function, and Phalloidin immunofluorescence for evaluate cell hypertrophy to further investigate pharmacodynamic mechanism of QSG.
RESULTS In vivo, QSG effectively improved the cardiac structure and function of DICR mice. Further, QSG promoted mitochondrial structural and functional integrity, reduced ROS production and enhanced oxidative metabolism, of which reversed DICR. Moreover, the protein levels of Rev-erbα and Cpt1b were up-regulated, and E4bp4 was down-regulated in the hearts of DICR mice treated with QSG. Mechanistically, the protective effects conferred by QSG against mitochondrial oxidative stress, cardiomyocyte hypertrophy and apoptosis were counteracted by Rev-erbα gene silencing in vitro. Meanwhile, the expression of E4bp4 was increased and Cpt1b was decreased. Of note, after interfering with Cpt1b, the protective effect of QSG was also weakened, but the expression of Rev-erbα and E4bp4 were not affected.
CONCLUSIONS In summary, the results demonstrated QSG could prevent from DICR by regulating the Rev-erbα-E4bp4-Cpt1b axis, which plays an indispensable role in cardiac energy metabolism This study will provide a potential approach for the treatment of DOX induced clinical cardiotoxicity.
GW34-e1012
Mekhman Mamedov, Azamat Karimov, Ksenia Badeynikova, Bahodir Mardanov
National Research Center for Therapy and Preventive Medicine
OBJECTIVES The chronic noncommunicable diseases remain the leading cause of disability and death among adults in the developed world. Their total share in mortality is about 77%. Cancers, along with cardiovascular diseases (CVD) associated with atherosclerosis, are among the top three causes of death in the adult population. According to the forecast of WHO experts, in the next 5–10 years, CVD will account for 26.5% in the structure of mortality, and 8.5% for cancers of various localization. Determination of the comorbidity of somatic diseases in patients with coronary artery disease and cancer of various localization plays an important role in the development of treatment tactics in order to improve the prognosis and quality of life.
METHODS The cross-sectional clinical study included 220 men and women aged 40–65 years. Depending on the presence of coronary artery disease and cancer of various localization, patients were divided into two group. The first group with ischemic heart disease and lung cancer (n=110), the second group with ischemic heart disease and colon cancer (n=110). In stationary conditions, with the help of clinical and instrumental studies, other somatic diseases were recorded in patients.
RESULTS According to results among patients with lung cancer and colon cancer, the most common disease was AH. In patients with cancer of both localizations, its frequency was comparable, 76 and 75%, respectively. The comorbidity of somatic diseases is also associated with the cancer’s localization. Type 2 diabetes in the group with lung cancer was diagnosed in 13.6% of cases, while in the group with bowel cancer it occurred twice as often, in 26% of cases. In the group with lung cancer, stomach diseases were detected in almost every second patient (44.5%), among people with bowel cancer this figure reached 70% of cases (P=0.034). Concomitant chronic obstructive pulmonary disease in patients with lung cancer was diagnosed in 62% of cases, and in the group of patients with bowel cancer it was 3.5 times less and amounted to 18% (P<0.001).
CONCLUSIONS Thus, in patients with coronary artery disease and cancer, additional somatic diseases are identified that are associated with cancer localization. When developing secondary prevention, it is necessary to take into account the presence of additional diseases that play an important role in the prognosis and quality of life of patients.
GW34-e1034
Fei Liu, Peiling Mi, Chenglin Li, Xiaolei Yang, Yunlong Xia
First Affiliated Hospital of Dalian Medical University
OBJECTIVES The aim of this study is to analyze and compare the distribution of cardiovascular (CV) comorbidities, risk factors and Electrocardiograph (ECG) parameters and other baseline characteristics in patients with primary Lung cancer (LC) and Thyroid cancer (TC), with the aim of providing a reference for the monitoring, early prevention and treatment of CV toxicity in these patients.
METHODS This study retrospectively included 15,145 patients who were hospitalized at the First Hospital of Dalian Medical University with a primary diagnosis of LC and TC between January 1, 2011 and May 31, 2022. Patients were finally included. General information, cardiovascular comorbidities, risk factors and electrocardiographic parameters were collected and statistically analyzed for both groups.
RESULTS 1. LC and TC patients carry a high burden of CV comorbidities at the time of initial diagnosis. The most common comorbidity in both groups was hypertension (20.2%), and the occurrence rate of hypertension was significantly higher in the LC group than in the TC group (21.8 vs. 16.6%, P<0.001), followed by coronary heart disease (8.1%), myocardial infarction (2.0%), atrial fibrillation (1.0%), atrial premature beats (0.7%), atrial tachycardia (0.4%), ventricular tachycardia (0.4%), and atrial flutter (0.3%), which were all significantly higher in the LC group than in the TC group (P<0.001); 2. The most common abnormal ECG parameter was prolonged QTc interval (15.4%), followed by increased P-wave duration (5.4%) and increased Sokolow-Lyon index (4.1%). The occurrence rates of prolonged QTc interval (16.8 vs. 12.3%, P<0.001), prolonged P-wave duration (5.5 vs. 4.1%, P<0.001), and increased Sokolow-Lyon index (4.9 vs. 2.7%, P<0.001) were significantly higher in the LC group. 3. After adjusting for age and sex, binary logistic regression analysis revealed that LC patients with concomitant low hemoglobin had a significantly increased risk of prolonged QTc interval (OR=1.01, 95% CI: 1.0–1.01, P<0.001), while TC patients with concomitant high systolic blood pressure (OR=3.3, 95% CI: 2.02–5.36, P<0.001), low hemoglobin (OR=0.98, 95% CI: 0.96–1, P=0.037), and low potassium (OR=0.38, 95% CI: 0.17–0.8, P=0.012) had a significantly increased risk of prolonged QTc interval. 4. In this study, a total of 142 patients (6.2%) with prolonged QTc interval and concomitant low potassium were identified. Among these patients, 25 (44%) in the LC group and 38 (45%) in the TC group did not undergo dynamic monitoring of potassium levels, and 28 (49%) in the LC group and 45 (53%) in the TC group did not receive potassium supplementation.
CONCLUSIONS Patients with initial diagnosis of LC and TC often have comorbid CV diseases or multiple CV risk factors even before initiating anti-tumor treatment. A considerable proportion of patients have abnormal ECG parameters, with prolonged QTc interval being the most common. Patients with prolonged QTc interval and concomitant hypokalemia account for a certain proportion in both the LC and TC groups, but the rate of close monitoring and potassium supplementation for these patients during hospitalization is relatively low, indicating inadequate management of CV risk factors in cancer patients in current clinical practice.
GW34-e1047
Zeya Li1, Shanshan Wu2, Dan Li3, Gang Wang1, Na Zhang1, Feng Zhao3, Jing Hao3, Chunlei Yang3, Jiashu Song3, Xianzhong Gu3, Rongchong Huang1
1Department of Cardiology, Beijing Friendship Hospital, Capital Medical University
2National Clinical Research Center for Digestive Diseases, Beijing Friendship Hospital, Capital Medical University
3Beijing Tongzhou Yongshun Community Health Service
OBJECTIVES The role of uric acid (UA) in chronic kidney disease (CKD) remains unclear. In this study, the effects of UA levels on kidney function (KF) in patients with normal versus impaired KF at baseline were investigated.
METHODS Data were extracted from the community-based Tongzhou Cohort Study (ClinicalTrials.gov Identifier: NCT05156580). The cohort of 4246 patients was divided into the normal KF group (estimated glomerular filtration rate [eGFR]≥90 mL/min/1.73 m2) and impaired KF group (60=eGFR<90 mL/min/1.73 m2). The primary endpoint was the composite of a decrease in eGFR of≥30% from baseline or new onset of CKD diagnosed during the annual checkups. Multivariate logistic and restricted cubic splines models were employed to evaluate potential associations.
RESULTS A total of 284 participants reached the primary endpoint during the 5-year follow-up period. There was a significant association between UA levels and baseline eGFR (P<0.001). Restricted cubic spline analysis revealed a U-shaped association between baseline UA levels and the risk of decreased KF for participants with normal KF (cut-off value=5.0 mg/dL; pnonlinear=0.025) and a linear association for participants with impaired KF (pnonlinear=0.384). The odds ratio of an increase in UA levels by 1 mg/dL was 1.15 [95% confidence interval (CI) 1.02–1.31, P=0.028] for the primary endpoint and 1.15 (95% CI=1.01–1.31, P=0.037) for new onset of CKD.
CONCLUSIONS Baseline KF is predictive of the association of increased UA levels and decreased KF. Patients with 60=eGFR<90 mL/min/1.73 m2 could benefit from UA-lowering therapy.
GW34-e1128
Junzhen Li
Zhongnan Hospital of Wuhan University
OBJECTIVES The objective of this study was to examine the correlation between fluctuations in metabolic syndrome (MetS) status and the occurrence of cardiovascular diseases (CVD), stroke, and all-cause mortality.
METHODS This prospective cohort study consisted of 4840 individuals who participated in the China Longitudinal Study of Health and Retirement (CHARLS) form 2011 to 2018. Participants were classified according to the change in their MetS status as follows: MetS-free at both time point (n=2169), MetS-recovery (n=744), MetS-developed (n=534), and MetS-chronic (n=1393). CVD and stroke were defined as the presence of physician-diagnosed heart disease and stroke. Logistic regression models were done to calculate the odd ratio (ORs) and confidence intervals (95% CIs).
RESULTS These participants were followed up in 2018. During follow-up period, 346 (7.1%) participants developed CVD, 246 (5.1%) participants developed stroke in 4840 participants in the longitudinal cohort. Compared with the MetS-free group, MetS-chronic increased the risk of CVD (adjusted OR, 1.40 [CI, 1.07 to 1.83]) and stroke (adjusted OR, 2.56 [CI, 1.83 to 3.58]). However, the increased risk in all MetS status were not significant for all-cause mortality. When stratified by age of MetS status changes, compared with the MetS-free group, MetS-chronic (<65 years) had higher risks of CVD (adjusted OR, 1.45 [CI, 1.03 to 2.05]), MetS-chronic (=65 years) had higher risks of stroke (adjusted OR, 2.81 [CI, 1.66 to 4.75]). In both genders, the MetS-chronic group had the highest risk of stroke, respectively, in women (adjusted OR, 3.32 [CI, 1.90 to 5.75]) and men (adjusted OR, 2.28 [CI, 1.46 to 3.58]), compared with the MetS-free group.
CONCLUSIONS Dynamic changes of MetS altered the risks of CVD and stroke, MetS-chronic increased the risk of CVD and stroke.
GW34-e1202
Xinhai Cui1, Yuanlong Hu2, Mengkai Lu3, Zhiyuan Zhang3, Yunlun Li3, Chao Li3
1College of Traditional Chinese Medicine, Shandong University of Traditional Chinese Medicine
2First Clinical Medical College, Shandong University of Traditional Chinese Medicine
3Innovation Research Institute of traditional Chinese Medicine, Shandong University of Traditional Chinese Medicine
OBJECTIVES Carotid femoral pulse wave velocity has been found to be an independent predictor of both cardiovascular mortality and kidney injury, which can be estimated non-invasively using estimated pulse wave velocity (ePWV). The objective of this study was to investigate the association between ePWV and in-hospital mortality of critically ill patients diagnosed with acute kidney injury (AKI).
METHODS This study included 5960 AKI patients from the Medical Information Mart for Intensive Care IV (MIMIC-IV) database. The low and high ePWV groups were compared using a Kaplan–Meier survival curve to evaluate differences in survival status. Cox proportional hazards models were utilized to explore the association between ePWV and in-hospital mortality among critically ill patients with AKI. To examine the dose-response relationship further, we used a restricted cubic splines (RCS) model. Additionally, stratification analyses were conducted to investigate the effect of ePWV on hospital mortality across various subgroups.
RESULTS Survival analysis results indicated that high ePWV had lower survival rate compared with low ePWV. Following adjustment, high ePWV demonstrated a statistically significant association with increased risk of in-hospital mortality among AMI patients (HR=1.84, 95% CI=1.62–2.08, P<0.001). Analysis using RCS model confirmed a linear increase in the risk of hospital mortality as ePWV values increased (P for nonlinearity=0.602).
CONCLUSIONS High ePWV was significantly associated with increased risk of in-hospital mortality among AKI patients. Furthermore, ePWV proved to be an independent predictor of hospital mortality in critically ill individuals with AKI.
GW34-e1214
Xixiang Tang1, Jiafu Wang2, Yesheng Ling2, Xiaolan Ouyang2, Qian Chen2, Long Peng2, Suhua Li2
1VIP medical service center, The Third Affiliated Hospital, Sun Yat-sen University
2Department of Cardiovascular Medicine, The Third Affiliated Hospital, Sun Yat-sen University
OBJECTIVES Risk of contrast-induced nephropathy (CIN) is markedly increased in patients with type 2 diabetes mellitus (T2DM). This study aimed to investigate the relationship between the adipose tissue thickness and the CIN in T2DM patients following coronary catheterization.
METHODS A total of 603 T2DM patients undergoing percutaneous coronary angiography or angioplasty were enrolled in this study. The thicknesses of perirenal fat (PRF), subcutaneous fat (SAT), intraperitoneal fat (IPT) and epicardial fat (ECF) were measured by Color Doppler Ultrasound respectively. The association of various adipose tissues with CIN were analyzed.
RESULTS Out of the 603 patients, 77 (12.8%) developed CIN. Patients who developed CIN had significantly thicker PRF (13.7±4.0 mm vs. 8.9±3.6 mm, P<0.001), slightly thicker IPT (P=0.046), and similar thicknesses of SAT (P=0.782) and ECT (P=0.749) compared to those who did not develop CIN. Correlation analysis showed that only PRF was positively associated with maximal sCr postoperatively (r=0.18, P=0.012), maximal absolute change (r=0.33, P<0.001) and maximal percentage of change in sCr (r=0.36, P<0.001). In ROC analysis, the area under the curve of PRF (0.809) for CIN was significantly higher than those of SAT (0.490), IPT (0.594) and ECT (0.512). Multivariate logistic regression analysis further confirmed that the thickness of PRF, rather than other adipose tissues, was independently associated with the development of CIN after adjusted for confounding factors (OR=1.53, 95% CI: 1.38–1.71, P<0.001).
CONCLUSIONS PRF is independently associated with the development of CIN in T2DM patients undergoing coronary catheterization.
GW34-e1260
Yan Ma, Dong Han, Shasha Sun, Feng Cao
Second Medical Centre & National Clinical Research Center for Geriatric Diseases, Chinese PLA General Hospital, Beijing, China
OBJECTIVES Despite the irreversible cardiotoxicity, anthracyclines, represented by doxorubicin, are still widely used to treat various tumors, which necessitates more effective strategies to lessen their cardiotoxicity. The role of cancer-heart crosstalk in DOX-induced cardiotoxicity (DOXIC) remains unidentified.
METHODS A co-culture system was used to observe the effects of DOX-injured BCCs on DOXIC in cardiomyocytes. EXOs derived from DOX-treated BCCs (4T1 cell, D-BCC-EXOs) were extracted and purified to examine their effects on adult murine ventricular cardiomyocytes (AMVC) injury and pyroptosis. An orthotopic breast cancer mouse model was used to explore whether EXOs could aggravate DOXIC in vivo. Exosomal miRNA sequencing and mechanistic studies were used to identify the functional miRNAs encapsulated in D-BCC-EXOs. Bioinformatics analysis combined with experimental validations were used to explore the upregulation and selective packaging mechanism of the functional miRNAs in D-BCC-EXOs as well as the downstream targets of the identified miRNAs.
RESULTS DOX-induced AMVCs injury and pyroptosis can be worsened by co-culturing with 4T1 BCCs as well as D-BCC-EXOs treatment in vivo and in vitro. Depletion of BCCs’ miRNA cargo by Dicer knockout blocked the effect of D-BCC-EXOs to aggravate DOXIC. Exosomal miRNA sequencing and mechanistic investigations identified that miR-216a-5p was highly expressed in DOX-induced EXOs from both DOX-treated tumor-bearing mouse serum and 4T1 BCCs and functioned as a cardiomyocytes pyroptosis aggravator both in vivo and in vitro. Mechanistically, DOX-induced ATF3 promotes miR-216a-5p transcription in BCCs, and ATF3 knockdown attenuated the pro-pyroptotic effects of D-BCC-EXOs. RNA binding protein-associated assays showed that SF3B4 bound to miR-216a-5p and facilitated the selective packaging and delivery of miR-216a-5p to BCC-D-EXOs. ITCH was identified as a bona fide target mRNA of miR-216a-5p, and miR-216a-5p-mediated ITCH decay reduced TXNIP ubiquitination and degradation to activate the TXNIP/NLRP3 inflammasome pathway to incite pyroptosis in cardiomyocytes, thereby aggravating DOXIC.
CONCLUSIONS Taken together, our study found that EXO-mediated pathological communication between BCCs and cardiomyocytes caused additional cardiac injury in the setting of DOXIC. This effect is achieved through the exosomal transfer of miR-216a-5p from DOX-induced BCCs to cardiomyocytes and activation of the ITCH/TXNIP/NLRP3 inflammasome pathway to predispose cardiomyocytes to DOX-induced pyroptosis. Targeting the pathological EXOs releasing or blocking miR-216a-5p may be an effective strategy to mitigate additional cardiac injury in DOXIC.
GW34-e1287
Lichuan Chen, Jilang Zeng, Kaiyang Lin, Yansong Guo
Shengli Clinical Medical College of Fujian Medical University, Department of Cardiology, Fuzhou, China
OBJECTIVES Contrast-associated acute kidney injury (CA-AKI), a common cause of hospital-acquired AKI, is related to poor outcome. Recent studies suggested that a novel biomarker described as the derived neutrophil-to-lymphocyte ratio (dNLR) can reflect systemic inflammation and is related to the adverse outcome of coronary heart disease, chronic inflammatory diseases, and cancers. However, it is unclear whether dNLR has the capacity to predict CA-AKI and long-term mortality in patients receiving elective percutaneous coronary intervention (PCI). The aim of the current study was to assess the predictive value of dNLR for the occurrence of CA-AKI and long-term mortality in patients undergoing elective percutaneous coronary intervention.
METHODS This was a retrospective analysis of 5807 consenting patients from January 2012 to December 2018. The dNLR is calculated by the following formula: neutrophil count/(leukocyte count - neutrophil count). CA-AKI was defined as an increase in serum creatinine≥50% or 0.3 mg/dL within 48 hours after contrast medium exposure.
RESULTS Overall, the incidence of CA-AKI after PCI was 6.1% (n=352). Based on receiver operating characteristic (ROC) curves, a baseline dNLR value of 1.77 was used as the best cut-off for predicting CA-AKI. The patients were divided into low dNLR group (dNLR<1.77) and high dNLR group (dNLR=1.77) according to the cut-off value. The dNLR showed an area under the curve (AUC) of 0.618 [95% confidence interval (CI): 0.589–0.649] in predicting CA-AKI. Multivariate logistic analysis revealed that the dNLR=1.77 was independently associated with the incidence of CA-AKI (odds ratio=1.45, 95% CI: 1.12–1.88, P=0.004). There were 499 (8.6%) deaths during the median follow-up of 2.85 years. Kaplan–Meier survival analysis suggested dNLR=1.77 was significantly associated with worse outcomes (P<0.001). Furthermore, the patients in the high dNLR group had a significantly higher risk of all-cause death than those in the low dNLR group (hazard ratio=1.35, 95% CI: 1.11–1.65, P=0.003) in the fully adjusted Cox proportional hazards model.
CONCLUSIONS In patients treated with elective PCI, dNLR is an independent predictor of CA-AKI and long-term mortality.
GW34-e1289
Qiaorui Wen1,2, Shengfeng Wang1,2
1Department of Epidemiology and Biostatistics, School of Public Health, Peking University Health Science Center, Beijing 100191, China
2Key Laboratory of Epidemiology of Major Diseases (Peking University), Ministry of Education, Beijing 100191, China
OBJECTIVES To investigate whether Life’s Essential 8 (LE8) could attenuate the association between cancer and incident cardiovascular diseases (CVD).
METHODS The baseline survey of UK Biobank study (ethics approval No. 16/NW/0274) was conducted in 2006–2010, and followed up till 2021. Exposure and covariates information were collected through baseline questionnaires. Cardiovascular health (CVH) was divided into low (<50), moderate (50–79), and high (=80) according to LE8 score, which included diet, physical activity, smoke, sleep, BMI, non-HDL cholesterol, blood glucose, and blood pressure. CVD (I20–I25, I60–I69) diagnosis were ascertained through electronic linkage. Hazard ratios (HR) and 95% confidence intervals (CI) for the associations of cancer diagnosis and chemotherapy with incident CVD were estimated by Cox regression, with adjustment for sociodemographic characteristics, lifestyles and family history. Joint analysis was conducted to investigate the effect of cancer diagnosis and CVH on CVD risk.
RESULTS Among 270,807 participants free of CVD at baseline and with complete data on LE8, 10.91% were cancer survivors. The mean±SD age was 55.9±8.1 years. Over a median 12.5-year follow-up, 25,353 CVD cases were recorded. Compared with the general population, cancer survivors had a 21% (95% CI 1.17–1.25) higher risk of incident CVD. While considering the CVD subtypes, cancer survivors had higher risks of incident coronary heart disease and stroke, with HR (95% CI) of 1.18 (1.13, 1.23) and 1.26 (1.19, 1.34). Compared with non-cancer patient (despite of their CVH), cancer survivors with low or moderate CVH had 100% (1.75–2.29) and 23% (1.18–1.28) higher CVD risks respectively, while those with high CVH had no increased risk (0.95, 0.86–1.05). Compared with those free of cancer but with low CVH, cancer survivors with low CVH had were associated 19% (1.03–1.37) higher risk of CVD, while those with moderate or high CVH were associated with lower risks of CVD, with HR (95% CI) of 0.72 (0.68, 0.77) and 0.56 (0.50, 0.62), respectively.
CONCLUSIONS Achieving higher CVH could attenuate or even counteract the increased CVD risk in cancer survivors.
GW34-e1359
Meizhen WU1,2, Jiayi Huang1,2, Qingwen Ren1,2, Kai-Hang Yiu1,2
1The University of Hong Kong, Queen Mary Hospital
2The University of Hong Kong-Shenzhen Hospital
OBJECTIVES Sodium-glucose cotransporter-2 (SGLT2) inhibitors are increasingly prescribed due to its potential cardiorenal protective effect, while it was demonstrated to be associated with increased risk of urinary tract infection (UTI). Whether to continue or stop SGLT2 inhibitors after experiencing a UTI remains unclear. The aim of present study is to explore the association of incident UTI with subsequent cardiovascular and renal events. Additionally, the impact of continuing versus stopping SGLT2 inhibitors after UTI on subsequent cardiovascular and renal events.
METHODS We recruited patients with T2DM prescribed SGLT2 inhibitors from 2015 to 2022 from Clinical Data Analysis and Reporting System in Hong Kong. The primary outcomes were major adverse cardiovascular events (MACE), a composite of renal outcome, all-cause mortality, and recurrent UTI. UTI was modelled as time-varying covariates. A Fine-Gary model was applied to adjust for competing risk. Multivariable Cox proportional hazards model was used to estimate the risk of UTI versus non-UTI and continuing versus stopping SGLT2 inhibitors after UTI on subsequent adverse events.
RESULTS Among 63,799 SGLT2 inhibitors users, 4769 (7.48%) patients developed incident UTI during a median follow-up of 1.80 years. After UTI, 1836 (38.50%) patients stopped using SGLT2 inhibitors. UTI was associated with higher risk of MACE [hard ratio (HR) 2.02, 95% confidence interval (CI) 1.83–2.23], a composite of renal outcome (HR 2.01, 95% CI 1.83–2.19), and all-cause mortality (HR 3.90, 95% CI 3.58–4.25). Stopping SGLT2 inhibitors after incident UTI was associated with lower risk of recurrent UTI (HR 0.70, 95% CI 0.61–0.85), but higher risk of a composite of renal outcome (HR 1.61, 95% CI 1.35–1.91), MACE (HR 1.25, 95% CI 1.04–1.51) and all-cause mortality (HR 2.54, 95% CI 2.21–2.92).
CONCLUSIONS UTI has been linked to increased risk of cardiovascular and renal outcomes in patients with T2DM compared to no incident UTI. Moreover, stopping SGLT2 inhibitors was associated with reduced risk of recurrent UTI, but increased risk of adverse cardiovascular and renal events.
PERIPHERAL VASCULAR DISEASE
GW34-e0164
Peibiao Mai1, Junping Li2, Chunhong Wang2, Shuwan Xu1, Xiaofang Wang3, Kun Zhang2,4
1Department of Cardiology, The Eighth Affiliated Hospital, Sun Yat-sen University
2Department of Cardiology, Sun Yat-sen Memorial Hospital, Sun Yat-sen University
3Department of Dermatology and Venerology, University of Chinese Academy of Sciences Shenzhen Hospital
4Guangdong Provincial Key Laboratory of Malignant Tumor Epigenetics and Gene Regulation, Department of Cardiology, RNA Biomedical Institute, Sun Yat-sen Memorial Hospital, Sun Yat-sen University
OBJECTIVES Peripheral arterial calcification is a prevalent condition in patients with type 2 diabetes mellitus (T2DM), resulting in lower-limb amputation and reduced life quality. Nonalcoholic fatty liver disease (NAFLD) which can be simply evaluated by fatty liver index (FLI) is closely associated with T2DM development. In this study, we aimed to explore the relationship between FLI and lower limb arterial calcification (LLAC) in T2DM patients.
METHODS This was a retrospective, observational, single-center study between January 2018 and January 2019. Seventy-seven T2DM patients with suspiciously symptomatic lower limb PAD were enrolled in this study. Clinical and laboratory data were collected. FLI was calculated by body mass index, waist circumference, triglycerides, and γ-glutamyltransferase. LLAC was evaluated by computed tomography with Agatston scoring algorithm.
RESULTS T2DM patients were divided based on the FLI values: low risk (FLI<30, n=29), intermediate risk (30=FLI<60, n=32) and high risk (FLI=60, n=16) groups. Compared with low risk group, T2DM patients in high risk group had significantly lower LLAC scores [270.0 (3.3, 1020.3) vs. 2024.0 (481.5–7317.5); P=0.018], accompanied with higher triglyceride and glucose (TyG) index [(9.89±0.70) vs. (8.86±0.57) vs. (8.34±0.61); P<0.001]. The correlation analysis showed FLI was negatively associated with LLAC scores (r=-0.311, P=0.006).
CONCLUSIONS Major finding of the present study shows that FLI, an indicator used to diagnose NAFLD, is negatively associated with LLAC in T2DM patients, which implies that NAFLD may not be a risk factor for LLAC in T2DM patients.
GW34-e0609
Jiaxin Cheng1,2
1College of Medicine and Biological Information Engineering, Northeastern University, Shenyang, Liaoning, China
2National Key Laboratory of Frigid Zone Cardiovascular Disease, Cardiovascular Research Institute and Department of Cardiology, General Hospital of Northern Theater Command, Liaoning, China
OBJECTIVES This study aims to investigate the effects of statin therapy on patients with acute Stanford type B aortic dissection after undergoing TEVAR procedure.
METHODS This retrospective analysis reviewed the medical records of 742 patients who had acute type B aortic dissection and received TEVAR treatment at Department of Cardiology of General Hospital of Northern Theater Command from April 2002 to March 2022. The patients were categorized into two groups: the statin group (S-group, n=366), and non-statin group (NS-group, n=376) based on wheater they accepted postoperative statin therapy or not. The main observation outcomes were all-cause mortality and aortic-related adverse events. Kaplan–Meier survival curves analyzed survival between the two groups, and COX regression analysis corrected for confounding factors.
RESULTS There were significant differences between the S-group and NS-group in terms of coronary heart disease history (S-group 20.77%, NS-group 9.57%), postoperative Calcium channel blockers (S-group 87.43%, NS-group 77.66%), and postoperative β-blockers (S-group 86.25%, NS-group 79.79%) (P<0.05). There were no significant differences between the two groups in other baseline data (P>0.05). Significant differences between the S-group and NS-group were observed for all-cause mortality (S-group n=26, NS-group n=46, P=0.018), stent graft-induced distal redissection (S-group n=6, NS-group n=23, P=0.001), and re-intervention (S-group n=4, NS-group n=17, P=0.005). While Kaplan–Meier survival analysis showed that statin therapy had differences for stent graft-induced distal redissection (P=0.01), re-intervention (P=0.03), it had no significant impact on all-cause mortality over time. COX regression analysis showed that statin therapy was an independent protective factor, reducing stent graft-induced distal redissection [HR (95% CI): 0.241 (0.081, 0.723), P=0.011] and re-intervention [HR (95% CI): 0.210 (0.052, 0.840), P=0.027].
CONCLUSIONS Statin therapy after TEVAR treatment for patients with acute Stanford type B aortic dissection can help reduce aortic adverse events and the risk of both stent graft-induced distal redissection and re-intervention. Nevertheless, larger samples and multi-center studies are needed to validate these findings.
GW34-e0715
Jiaxin Cheng1,2, Zhiqiang Zhang2, Yasong Wang2, Xiaozeng Wang2
1College of Medicine and Biological Information Engineering, Northeastern University, Shenyang, Liaoning, China
2The Department of Cardiology of General Hospital of Northern Theater Command of Chinese People’s Liberation Army, Shenyang, China
OBJECTIVES This study aims to investigate the effects of statin therapy on patients with acute Stanford type B aortic dissection after undergoing TEVAR procedure.
METHODS This retrospective analysis reviewed the medical records of 742 patients who had acute type B aortic dissection and received TEVAR treatment at Department of Cardiology of General Hospital of Northern Theater Command from April 2002 to March 2022. The patients were categorized into two groups: the statin group (S-group, n=366), and non-statin group (NS-group, n=376) based on wheater they accepted postoperative statin therapy or not. The main observation outcomes were all-cause mortality and aortic-related adverse events. Kaplan–Meier survival curves analyzed survival between the two groups, and COX regression analysis corrected for confounding factors.
RESULTS There were significant differences between the S-group and NS-group in terms of coronary heart disease history (S-group 20.77%, NS-group 9.57%), postoperative Calcium channel blockers (S-group 87.43%, NS-group 77.66%), and postoperative β-blockers (S-group 86.25%, NS-group 79.79%) (P<0.05). There were no significant differences between the two groups in other baseline data (P>0.05). Significant differences between the S-group and NS-group were observed for all-cause mortality (S-group n=26, NS-group n=46, P=0.018), stent graft-induced distal redissection (S-group n=6, NS-group n=23, P=0.001), and re-intervention (S-group n=4, NS-group n=17, P=0.005). While Kaplan–Meier survival analysis showed that statin therapy had differences for stent graft-induced distal redissection (P=0.01), re-intervention (P=0.03), it had no significant impact on all-cause mortality over time. COX regression analysis showed that statin therapy was an independent protective factor, reducing stent graft-induced distal redissection [HR (95% CI): 0.241 (0.081, 0.723), P=0.011] and re-intervention [HR (95% CI):0.210 (0.052, 0.840), P=0.027].
CONCLUSIONS Statin therapy after TEVAR treatment for patients with acute Stanford type B aortic dissection can help reduce aortic adverse events and the risk of both stent graft-induced distal redissection and re-intervention. Nevertheless, larger samples and multi-center studies are needed to validate these findings.
GW34-e0795
Liyuan Chen1, Xue Wang2, Lina Pan1, Lingling Yang2, Xutao Zhang1, Chang Liu3, Houyuan Hu1
1Department of Cardiology, Southwest Hospital, Third Military Medical University
2Department of Occupational Health (Key Laboratory of Electromagnetic Radiation Protection, Ministry of Education), Third Military Medical University
3Department of Neurology, The Second Affiliated Hospital of Chongqing Medical University
OBJECTIVES Even if endovascular treatment (EVT) getting to be the first-line therapeutic modality for large vessel occlusion in the anterior circulation (ALVO), nearly one in three of these patients obtained poor outcome. With the facts that valvular diseases were widely detected in these patients, present study aimed to evaluate the roles of valvular diseases in determining outcomes of these individuals to explore novel risk factors for poor clinical consequences among these individuals.
METHODS Among 798 LVO patients receiving EVT across 32 medical centers in China who underwent transthoracic echocardiography (TTE) were included in analysis. After grading the regurgitation and stenosis of the mitral, aortic, and tricuspid valves, logistics regression analysis was engaged in evaluating their relationship with outcomes. Outcomes includes acute heart failure after EVT, modified Rankin Scale score (mRS) of 0–2 and mortality at 3 months. Causal mediation analyses were used to estimate the effects of valvular diseases and the potential mediators.
RESULTS The presence of MR was found in 27.2% of patients. Compared to non-significant MR (nMR), patients with significant mitral regurgitation (sMR) were older (P=0.005), and they also had more comorbidities, such as diabetes mellitus (30.61 vs. 19.85%; P<0.080) and long-standing atrial fibrillation (29.39 vs. 6.12%; P<0.001). sMR was independently associated with functional independence after 3 months [adjusted odds ratio: 0.41, 95% confidence interval: 0.21 to 0.77; P=0.006] and mortality [adjusted odds ratio: 1.92, 95% confidence interval: 1.21 to 2.47; P=0.012]. Mediation analysis showed sMR-increased acute heart failure explained 20.0% (95% CI 4.1–76.0%) of the effect of sMR on functional independence. In addition to MR, we did not detect significant relationship between outcomes and the other valvular diseases among enrolled patients.
CONCLUSIONS sMR constitutes an underrated high-risk group in LVO stroke patients who received EVT, with an increase in the overall high mortality and significant reduced functional independence. Our study also stressed the importance of echocardiography monitoring of the leaflet of MR in high-risk stroke patients.
PSYCHO-CARDIOLOGY
GW34-e0757
Li Feng, Li Hui
The Second Affiliated Hospital of Soochow University, Suzhou, Jiangsu 215004, China
OBJECTIVES Accumulated studies have indicated that depression was significantly associated with the atrial fibrillation (AF) risk. However, the causal effect of major depressive disorder (MDD) on the risk of AF was elusive.
METHODS We performed a two-sample Mendelian randomization (MR) analysis using two genetic instrument tools for MDD to assess the potential causal effect of MDD on the AF risk, including a training dataset (37 single-nucleotide polymorphisms (SNPs) significantly associated with MDD) and a validation dataset (11 SNPs robustly associated with MDD). Inverse-variance weighted method (IVW), weighted median method, MR-Egger, MR-Pleiotropy REsidual Sum and Outlier (MR-PRESSO) test, pleiotropy test, and leave-one-out sensitivity analysis were performed for MR analyses.
RESULTS We first performed MR analysis for a training data using multiple data processing steps, including removing SNPs related with AF risk factors, removing one outlier with MR-PRESSO, and tighten instrument P value threshold. The final MR estimates suggested that MDD had no causality on AF incidence (IVW method, OR=1.04, 95% CI=0.79–1.36, P value=0.785; weighted median, OR=1.15, 95% CI=0.90–1.46, P value=0.268; MR-Egger, OR=1.00, 95% CI=0.35–2.85, P value=0.999). Moreover, a consistent result was displayed in MR estimates using a validation data. Pleiotropy test and leave-one-out sensitivity analysis revealed that our results were robust.
CONCLUSIONS Our MR analysis suggested that MDD may have no causality on the AF risk.
GW34-e0999
Shenghui Zhang, Siqi Hu, Yao You, Jiake Tang, Chen Chen, Mingwei Wang
Department of Cardiology, Affiliated Hospital of Hangzhou Normal University, Hangzhou Institute of Cardiovascular Diseases, Zhejiang Key Laboratory for Research in Assessment of Cognitive Impairments, Hangzhou Normal University, Hangzhou 310015, China
OBJECTIVES Previous studies have found that amygdala activity independently and robustly predicts cardiovascular disease events. The purpose of this study was to calculate functional connectivity (FC) using resting-state functional MRI (rs-fMRI). To explore the characteristics of brain functional network in patients with coronary heart disease (CHD), and to better understand the relationship between the changes of amygdala-based FC and the clinical characteristics and cognition of patients with CHD.
METHODS A total of 14 CHD patients and 10 healthy controls were enrolled in this study. All subjects underwent resting-state fMRI scan in the Affiliated Hospital of Hangzhou Normal University, and administered the biochemical testing of blood. Neuropsychological assessment was performed by 2 trained physicians: the assessment scale includes Mini-Mental State Scale (MMSE), Hamilton Rating Scale for Depression (HAMD-17).
RESULTS Compared with healthy controls, patients with CHD had increased functional connectivity between the amygdala and the medial superior frontal gyrus (BA11, P<0.05, FDR corrected, Q<0.05). The FC between amygdala and medial superior frontal gyrus was significantly associated with HDL-C (r=−0.53, P<0.05), HAMD score (r=0.66, P<0.05) and MMSE scores (r=−0.60, P<0.05).
CONCLUSIONS This study found that the whole brain functional connectivity based on amygdalae in CHD patients is different from that in healthy controls, and this functional connectivity pattern is correlated with laboratory indicators and cognitive scales, which may help us to further understand the relationship between brain function and diseases.
GW34-e1165
Xing Liang, Qiaoling Ye, Yisong Pei, Guozhu Chen
The Second Affiliated Hospital of Chongqing Medical University
OBJECTIVES Anxiety, often accompanied by somatic symptoms and sleep disorder, is commonly observed in populations infected with COVID-19. Patients with pre-existing cardiac conditions are considered a high-risk demographic for fatality following COVID-19 infection. This study aims to evaluate the psychological conditions, including the severity of anxiety, somatic symptoms, and sleep quality in patients with ACS (acute coronary syndrome) during the COVID-19 pandemic.
METHODS The subjects of the study were 336 patients diagnosed with ACS who sought medical consultation between February 2022 and February 2023 at the Second Affiliated Hospital of Chongqing Medical University and the Wushan County People’s Hospital of Chongqing. Among these, nine patients were not infected with COVID-19. All patients underwent assessment using the following scales: Short Health Anxiety Inventory (SHAI), Patient Health Questionnaire-15 (PHQ-15), and the Pittsburgh Sleep Quality Index (PSQI). Assessments were conducted through online questionnaires.
RESULTS In the SHAI scale, 44.3% of the patients with ACS displayed symptoms of anxiety, with an average score of 21.56±7.377. The ACS onset time showed a significant correlation with the state of anxiety (P<0.05), the earlier the onset of ACS, the longer the time from COVID-19 infection, and the lower rates of anxiety. In the PHQ-15 scale, 24.71% of the patients had somatic symptoms. Patients with mild somatic symptoms were 9 times more likely to moderate ones, no severe cases. The earlier the ACS onset, the longer the time from COVID-19 infection, the longer follow-up interval, and the lower incidence of somatic symptoms (P<0.05). STEMI and NSTEACS differed markedly in terms of somatic symptoms measured by PHQ-15, the incidence in STEMI group is notable higher than the NSTEACS. A significant positive correlation was found between SHAI and PHQ-15 scores (P<0.001). The PSQI scale indicated a sleep disorder incidence of 22.9%, somatic symptoms had a obvious impact on sleep quality (P<0.05), and the more severe the somatic symptoms, the more likely to have sleep disorders. In the subgroup analysis, remarkable differences were observed in PHQ-15 scores between patients with stents [3.0 (2.0–5.0)] and those without stents [5.0 (2.0–6.0)] (P<0.05). The analysis indicated that they were independent of factors such as age, gender, ischemic time, severity of COVID-19 infection, and the presence of residual symptoms (P>0.05).
CONCLUSIONS After the recent COVID-19 pandemic, psychological problems such as anxiety, somatic symptoms and sleep disorders were common in ACS patients. It was found that the incidence of anxiety and somatic symptoms is related to the time from ACS onset to COVID-19 infection, the longer the time, the lower the proportion of patients with psychological condition. The somatic symptoms are related to the severity of ACS, meanwhile they may decrease sleep quality. These demonstrated that more severe ACS and shorter interval between ACS onset and COVID-19 infection may increase psychological problems of patients. Those patients require more active psychological attention and intervention.
CARDIOVASCULAR IMAGING
GW34-e0037
Yao Lu, Xiaoli Zhang
Beijing Anzhen Hospital Affiliated Capital Medical University
OBJECTIVES To evaluate the predictive value of the proportion of hibernating myocardium (HM) in total perfusion defect (TPD) on reverse left ventricle remodeling (RR) after coronary artery bypass graft (CABG) in patients with heart failure with reduced ejection fraction (HFrEF) by 99mTc-methoxyisobutylisonitrile (MIBI) single photon emission computed tomography (SPECT) myocardial perfusion imaging (MPI) combined with 18F-flurodeoxyglucose (FDG) gated myocardial imaging positron emission computed tomography (PET).
METHODS Inpatients diagnosed with HFrEF at the Cardiac Surgery Center, Anzhen Hospital of Capital Medical University from January 2016 to January 2022 were prospectively recruited. MPI combined with 18F-FDG gated PET was performed before surgery for viability assessment and the patients received follow-up MPI and 18F-FDG gated PET at different stages (3∼12 months) after surgery. Δ indicated changes (post-pre). Left ventricular end-systolic volume (ESV) reduced at least 10% was defined as RR, patients were divided into reverse remodeling (RR+) group and the non-reverse group (RR−). Binary logistic regression analysis was used to identify predictors of RR. Receiver operating characteristic (ROC) curve analysis was performed and the area under the curve (AUC) was calculated to assess the cut-off value for predicting RR. Additionally, we retrospectively enrolled inpatients with HFrEF at the Cardiac Surgery Center, Anzhen Hospital of Capital Medical University from January 2021 to January 2022 as the validation group, who underwent MPI and 18F-FDG gated PET before surgery. Echocardiography was performed before CABG and after CABG (3∼12 months). In the validation group, the reliability of obtaining the cut-off value for the ROC curve was verified.
RESULTS A total of 28 patients with HFrEF (26 males; age (56.9±8.7) years) were included in the prospective cohort. HM/TPD was significantly higher in the RR+ group than in the RR-group ((51.8±17.9%) vs. (35.7±13.9%), P=0.016). Binary logistic regression analysis revealed that HM/TPD was an independent predictor of RR [Odds ratio=1.073, 95% Confidence interval: 1.005–1.145, P=0.035]. ROC curve analysis revealed that HM/TPD=38.3% yielded the highest sensitivity, specificity, and accuracy (all 75%) for predicting RR and the AUC was 0.786 (P=0.011). Meanwhile, a total of 100 patients with HFrEF (90 males; age (59.7±9.6) years) were included in the validation group. In the validation group, HM/TPD=38.3% predicted RR in HFrEF patients after CABG with the highest sensitivity, specificity and accuracy (82, 60 and 73% respectively). Compared with the HFrEF patients in the HM/TPD<38.3% group (n=36), RR and cardiac function improved more significantly in the HM/TPD=38.3% group (n=64) (all P<0.05).
CONCLUSIONS Preoperative HM/TPD ratio is an independent factor for predicting RR in patients with HFrEF after CABG, and HM/TPD=38.3% can accurately predict RR and the improvement of cardiac function after CABG.
GW34-e0039
Yankai Mao1, Yuan Yang1, Jürgen Duchenne2, Christophe Garweg2, Xia Sheng3, Jiefang Zhang3, Yang Ye3, Min Wang3, Ying Yang3, Gabor Vöros2, Yaxun Sun3, Mingming Ma1, Guosheng Fu3, Jens-Uwe Voigt2
1Department of Diagnostic Ultrasound & Echocardiography, Sir Run Run Shaw hospital, Zhejiang University
2Department of Cardiovascular Sciences, Catholic University Leuven, Belgium
3Department of Cardiology, Key Laboratory of Cardiovascular Intervention and Regenerative Medicine of Zhejiang Province, Sir Run Run Shaw Hospital, School of Medicine, Zhejiang University
OBJECTIVES Left bundle branch pacing (LBBP) has emerged as a novel pacing modality. Although it has been proved to maintain electrical synchrony better than right ventricular pacing (RVP), little is known about the impact on mechanical synchrony. This study investigates if LBBP preserves mechanical synchrony and cardiac function better compared to conventional (CRVP) and leadless (LRVP) RVP.
METHODS Sixty-eight patients with pacing indication for bradycardia were prospectively enrolled: Twenty-five were treated with LBBP, 23 with CRVP and 20 with LRVP. All patients underwent echocardiography before and after implantation and at one-year follow-up. Left ventricular (LV) volumes, ejection fraction (EF) and global longitudinal strain (GLS) were measured. Regional septal (SW) and lateral wall work (LW) was calculated as the average from the respective basal and mid-ventricular segments in the apical four-chamber and three-chamber view. The lateral-septal (LW-SW) work difference was used as a measure of mechanical dyssynchrony.
RESULTS At baseline, LW-SW work difference were similar in all three groups. SW was markedly decreased in CRVP and LRVP while LW work remained unchanged, resulting in a larger LW-SW work difference compared to LBBP (1308.4±732.9 mmHg*% and 1451.3±606.1 mmHg*% vs. 286.0±479.9 mmHg*%, both P<0.001). During one year follow-up, LVEF and LV GLS decreased more in CRVP compared to LBBP (both P<0.05).
CONCLUSIONS LBBP causes less LV dyssynchrony than CRVP and LRVP as it preserves a more physiologic conduction pattern. With this, LBBP appears to preserve LV function better than CRVP. CRVP and LRVP did not differ in mechanical dyssynchrony or LV remodelling.
GW34-e0044
Yankai Mao1, Yuan Yang1, Chan Yu1, Yunhe Wang2, Chenyang Jiang2
1Department of Diagnostic Ultrasound & Echocardiography, Sir Run Run Shaw Hospital, Zhejiang University School of Medicine
2Department of Cardiology, Key Laboratory of Cardiovascular Intervention and Regenerative Medicine of Zhejiang Province, Sir Run Run Shaw Hospital, School of Medicine, Zhejiang University
OBJECTIVES Atrial fibrillation (AF) is associated with left atrial (LA) mechanical dysfunction, which predispose to stasis of blood within the LA and thus increase the risk of stroke. However, limited is known about the mechanical functional alterations of left atrial appendage (LAA) in relation to stages of AF. Here, we sought to analyse LA and LAA mechanics during different stages of AF in detail.
METHODS A total of 92 patients with AF referring for radiofrequency ablation were consecutively enrolled. Patients were categorized according to rhythm and stages of AF: sustained AF (SAF) (n=31), paroxysmal AF with sinus rhythm (PAF-SR) (n=30) and AF rhythm (PAF-AF) (n=31) at the time of echocardiography. Twenty-seven patients without a history of AF were included as controls. Using speckle-tracking echocardiography, LA reservoir strain (LASr), LAA global longitudinal strain (GLS) and mechanical dispersion (MD) of LA and LAA were measured in all patients. Mechanical dispersion was defined as the standard deviation of the time to peak of the regional strain corrected by R-R interval.
RESULTS Patients with PAF-AF and SAF displayed prominently impaired LA and LAA mechanics (all P<0.01 vs. PAF-SR and no AF). In the presence of SR, only LAA mechanical function were lower in PAF patients than patients without AF (LAA GLS 20.5±5.7% (no AF) vs 15.9±5.4% (PAF-SR); LAA MD 6.9±3.8% (no AF) vs 9.4±3.9 (PAF-SR), both P<0.01). AF stages were independent determinants of LA and LAA mechanics after adjusting for age and comorbidities.
CONCLUSIONS AF affected mechanical function of LA and LAA on their own, and more so if stage of AF has advanced to more persistent forms. In addition, LAA mechanical dysfunction occurred earlier than that of LA.
GW34-e0203
Yining Wang1, Xuejing Duan2, Leyi Zhu1, Jian He1, Baiyan Zhuang1, Jing Xu1, Di Zhou1, Wenjing Yang1, Arlene Sirajuddin3, Andrew E. Arai3, Hongyue Wang2, Shihua Zhao1, Minjie Lu1
1Department of Magnetic Resonance Imaging, Fuwai Hospital, State Key Laboratory of Cardiovascular Disease, National Center for Cardiovascular Diseases, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing, China
2Department of Pathology, Fuwai Hospital, State Key Laboratory of Cardiovascular Disease, National Center for Cardiovascular Diseases, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing, China
3Department of Health and Human Services, National Heart, Lung and Blood Institute (NHLBI), National Institutes of Health (NIH), Bethesda, USA
OBJECTIVES The diagnostic performance of cardiovascular magnetic resonance (CMR) compared with endomyocardial biopsy (EMB) in patients with suspected acute myocarditis (AMC) has not been extensively evaluated, and characteristics of patients with false-negative EMB results are not well understood. This study aimed to evaluate the diagnostic performance of CMR compared with EMB in patients with suspected AMC and identify characteristics of patients with false-negative EMB results.
METHODS A total of 311 patients with clinically suspected AMC who underwent CMR with 3.0 T. The CMR protocol consisted of cine-SSFP, T2-weighted STIR, late gadolinium enhancement (LGE), T1 and T2 mapping. Patients were considered to have confirmed myocarditis when both ESC statement and 2018 Lake Louise criteria (LLC) supported the diagnosis.
RESULTS The 2018 LLC yielded a sensitivity of 96.3% and a specificity of 84.6%, while the sensitivity of EMB was 62.8% and the specificity was 85.7%. Of the patients who underwent EMB, 55 (59.1%) were diagnosed with myocarditis, three (3.2%) were diagnosed with dilated cardiomyopathy and no specific myocardial injury was found in the remaining 35 (37.6%). Based on predetermined classification criteria, 27 patients (29.0%) were classified in the false-negative group, 53 (57.0%) in the true-positive group, and 11 (11.8%) in the true-negative group. Patients prone to have false-negative EMB results may have clinical symptoms present as infarct-like or arrhythmias and normal cardiac function, with CMR performance including preserved left ventricular ejection fraction, atypical LGE patterns and elevated T1 or T2 relaxation times.
CONCLUSIONS CMR shows superior diagnostic performance compared with EMB in the diagnosis of AMC. False-negative EMB results may occur more frequently in patients with specific clinical and CMR characteristics. Our findings suggest that all patients with clinically suspected myocarditis should undergo CMR to optimize diagnosis and clinical management.
GW34-e0215
Jinwei Tian1, Chao Li2, Zhifeng Qin1, Taishi Yonetsu3, Rocco Vergallo4, Bo Zhang5, Zhao Wang2, Bo Yu1
1Department of Cardiology, Second Affiliated Hospital of Harbin Medical University, Harbin, China
2School of Electronic Science and Engineering, University of Electronic Science and Technology of China, Chengdu, China
3Department of Cardiovascular Medicine, Tokyo Medical and Dental University, Tokyo, Japan
4Department of Cardiovascular Medicine, Catholic University of the Sacred Heart, Rome, Italy
5Department of Cardiology, First Affiliated Hospital of Dalian Medical University, Dalian, China
OBJECTIVES Coronary artery calcification (CAC) is a surrogate of atherosclerotic disease burden and has been associated with worse stent expansion and clinical outcomes. However, evidence from large-scale high-resolution imaging studies is lacking, and the 3D morphological significance of CAC is yet to be elucidated.
METHODS We proposed a novel deep learning method that can automatically identify and quantify CAC in massive intravascular OCT data. 1,106,347 OCT images from 1048 patients with ischaemic heart disease were collected and utilized to train and evaluate the method. All possible CACs were segmented from OCT pullbacks using the proposed method (Ca-Net) trained with sparsely annotated data, which were then screened and classified by a 3D CNN network. After validating the performance of the AI-model by external evaluation, we applied the method to 1259 patients with ST-segment elevation myocardial infarction (STEMI) and evaluated the predictive value and clinical significance of CAC.
RESULTS The Dice similarity coefficient for CAC segmentation and the accuracy for CAC classification in internal evaluation were 0.693 and 0.932 respectively, reaching human-level performance. We found that patients with greater extent of calcification in the culprit vessels were significantly more likely to have major adverse cardiovascular and cerebrovascular events (MACCE), and calcification is more severe in culprit vessels than in non-culprit vessels.
CONCLUSIONS The proposed AI method trained with massive clinical data has achieved human-level performance in CAC segmentation and quantification. Severe CACs in culprit vessels of STEMI patients identified by the AI algorithm were associated with MACCE.
GW34-e0240
Nuo Xu, Yingjie Zhao, Haiyan Chen, Xianhong Shu
Zhongshan Hospital, Fudan University
OBJECTIVES Transjugular intrahepatic portosystemic stent shunt (TIPS) is a non-surgical treatment for portal hypertension. Right heart overloading is one of the main complications of the procedure, leading to heart failure. However, research on right heart function after TIPS is limited. This study aims to assess the efficiency of speckle tracking echocardiography for evaluating cardiac function before and after TIPS and to examine the short-term impact of TIPS on right heart function.
METHODS This prospective study included 51 patients (32 males, 19 females; 56±9 yrs) with a clinical history of intractable esophageal varices or refractory ascites. Transthoracic echocardiograph data were collected at baseline and 7 days, 90 days after TIPS. Conventional and advanced echocardiographic parameters were analyzed, including right atrium (RA) and right ventricle (RV) structure, function and 2D strain. The Model for End-stage Liver Disease (MELD) score was also calculated.
RESULTS
• RASr showed a significant increase at day 7, and although it decreased after day 7, it was still remarkably higher at day 90 than before therapy. (PRE vs Day7: 42.35±6.90% vs 47.45±5.56%, P<0.01; PRE vs Day90: 42.35±6.90% vs 46.12±4.26%, P<0.05). RAScd and RASct showed the same trends.
• RVFWSL decreased significantly at day 7 but increased significantly at day 90. (PRE vs Day7: −33.61±4.70% vs −30.35±3.35%, P<0.01; Day7 vs Day90: −30.35±3.35% vs −34.78±3.18%, P<0.01).
• ΔRASr was remarkably linearly correlated with ΔRAA (PRE vs Day7: P=0.014, r=0.484; PRE vs Day90: P=0.016, r=0.529).
• ΔRVFWSL was linearly related to ΔFAC (PRE vs Day7: P=0.042, r=-0.414).
• The MELD score correlated with RVFWSL, indicating that RV strain is somewhat predictive of liver cirrhosis outcome. (PRE: P=0.039, r=-0.378, Day90: P=0.030, r=-0.424).
CONCLUSIONS
• Speckle tracking echocardiography is more sensitive than conventional echocardiography in detecting changes in right heart function.
• TIPS may cause RA overload and RV function decrease shortly after the procedure, but right heart strains and function ultimately increase 90 days after therapy.
• The MELD score correlates with RV longitudinal strain, indicating its potential as a predictor of liver cirrhosis outcome.
GW34-e0287
Ran Xin1,2, Guanhua Dou3, Yundai Chen1, Junjie Yang1
1Senior Department of Cardiology, The Sixth Medical Center, Chinese PLA General Hospital
2School of Medicine, Nankai University
3Department of Cardiology, The Second Medical Center & National Clinical Research Center for Geriatric Diseases, Chinese PLA General Hospital
OBJECTIVES To explore the value of CCTA-based radiomics technology in improving the identification efficiency of left atrial appendage (LAA) thrombus in patients with nonvalvular atrial fibrillation (NVAF).
METHODS Six hundred and seventy patients with NVAF who underwent CCTA and transesophageal echocardiography (TEE) from May 2015 to May 2020 were retrospectively enrolled and classified as thrombus group and non-thrombus group by TEE, dividing the training dataset (N=404) and the validation dataset (N=266) by a ratio of 6:4. Clinical baseline data and CCTA parameters were collected and the ratio of minimum Hounsfield units (HU) in the thrombus (LAAmin) to those in the ascending aorta (AAmin) was also calculated (LAAmin/AAmin). Mean value interpolation was used to interpolate the missing values. Radiomics features were automatically extracted from the region of interest (ROI) of LAA. Random Forest (RF) were used for feature selection, and multivariate logistic regression was used for modeling. Receiver operator characteristic (ROC) curve, calibration curve and decision curve analysis (DCA) were plotted and the performance parameters, the net reclassification index (NRI) and C-index were calculated to evaluate the efficacy in terms of discrimination, calibration and clinical net benefit. The radiomics nomogram was plotted to describe the fraction of components in the predictive model. Optimal radiomics was compared between different groups.
RESULTS The AF type, CHA2DS2-VAS score, HAS-BLED score, LAA length (LAA-L), LAA diameter (LAA-D) and LAAmin/AAmin were significantly different between two groups (P<0.05). 1232 radiomics features were extracted and 20 features were finally selected for modeling. The composite model had a higher area under curve (AUC) than the other three models (Training dataset: 0.949 vs. 0.757, 0.829, 0.913; Validation dataset: 0.900 vs. 0.832, 0.825, 0.822, respectively). The accuracy (ACC), specificity (SPE), positive likelihood ratio (PLR), Positive predictive value (PPV), Negative percent Agreement (NPA) and Kappa value of composite model were significantly higher (Training dataset: 0.874, 0.870, 7.296, 0.275, 0.997, 0.874; Validation dataset: 0.917, 0.924, 10.38, 0.367, 0.917, 0.461, respectively). The composite model had a positive NRI compared to the other three models (0.421, 0.059, 0.113, respectively), and a C-index greater than 0.5 (0.802, 0.662, 0.648, respectively). The calibration curve and decision curve suggested that the composite model had better calibration and clinical net benefits. The radiomics nomogram showed that the efficiency ratio of radiomics features in the composite model. The value of optimal radiomics feature in thrombus group was significantly higher than that in non-thrombus group (P<0.05), which had no significant difference in the subgroups divided by LAA morphology and CHA2DS2-VAS score (P>0.05).
CONCLUSIONS CCTA-based radiomics can improve the predictive efficiency of traditional predictive models for LAA thrombosis in NVAF patients, and it can also assess factors associated with thrombosis in multiple dimensions.
GW34-e0288
Ran Xin1,2, Yipu Ding1,2, Xi Wang1, Zinuan Liu1,3, Yundai Chen1, Junjie Yang1, Dongkai Shan1
1Senior Department of Cardiology, The Sixth Medical Center, Chinese PLA General Hospital
2School of Medicine, Nankai University
3Medical School of Chinese PLA
OBJECTIVES The study aims to investigate the association of Perivascular adipose tissue (PVAT) volume and attenuation with the risk of coronary artery disease (CAD) in patients with type 2 diabetes mellitus (T2DM).
METHODS T2DM patients with suspected CAD were retrospectively included in this study, and non-diabetic patients were selected as the control group according to the 1:1 propensity matching score. Demographic data and clinical risk factors were collected, and the PVAT volume and attenuation measured by CCTA were calculated. The PVAT volume and attenuation differences between T2DM patients and non-diabetic patients were compared. And the correlation between RCA, LAD, and LCX volume and metabolic syndrome-related factors, univariate and multivariate regression analysis of RCA-PVAT volume and attenuation, metabolic syndrome-related factors, and diabetes status. Continuous normal data were represented by mean±standard deviation (mean±SD), while non-normal data were represented by quartile and median. The student’s T-test was used for the comparison of normal distribution and the Mann-Whitney U test was used for that of non-normal distribution. The categorical data were expressed as percentages and compared using the Chi-square test or Fisher’s exact test. Pearson or Spearman r correlation method was used for correlation analysis. Odds ratios (OR) and 95% confidence intervals (CI) were calculated.
RESULTS One hundred and thirty patients with T2DM complicated with CAD were enrolled in this study, and a total of 260 subjects were enrolled after 1:1 propensity score matching (PSM). A comparison of baseline data found that waist circumference, BMI, total serum cholesterol, triglycerides, LDL-C, and HbA1C were significantly higher in T2DM patients compared with those without diabetes. Compared with the non-diabetic population, the PVAT volume in T2DM patients increased significantly, while the attenuation did not change statistically. Multivariate regression analysis indicated that RCA-PVAT volume, waist circumference, and HbA1C were independent correlation factors of T2DM.
CONCLUSIONS PVAT volume, rather than attenuation, was associated with an increased risk of CAD in T2DM patients.
GW34-e0291
Wang Yi
Sichuan Provincial People’s Hospital
OBJECTIVES Heart failure remains the main cause for cardiac death. Lots of heart failure with reduced ejection fraction (HFrEF) patients often have concomitant significant mitral regurgitation. It’s not quite clear who would benefit from transcatheter edge-to-edge repair (TEER). Some evidence showed that right ventricular function was an important predictor for heart failure. We therefore try to refine the evaluation of the association between right ventricular function evaluated by pressure-strain loops (PSL) and the clinical improvement after TEER. We sought to (1) analyze the change of different right ventricular myocardial work (RVMW) parameters by PSL in HFrEF patients undergoing TEER; (2) evaluate the association between immediately change of RVMW parameters and clinical improvement among HFrEF patients who underwent TEER treatment for secondary mitral regurgitation (SMR).
METHODS Eligible patients had ischemic or nonischemic cardiomyopathy with a left ventricular ejection fraction of 20 to 50%, had moderate-to-severe (grade 3+) or severe (grade 4+) SMR that was confirmed at the echocardiographic laboratory before enrollment, and remained symptomatic (NYHA functional class II, III, or IVa [ambulatory]) despite the use of optimal doses of guideline-directed medical therapy and cardiac resynchronization therapy (if appropriate), which were administered in accordance with guidelines of professional societies. If the interventional cardiologist confirmed that the patient was anatomically eligible for device implantation, and the cardiothoracic surgeon determined that mitral valve surgery was not appropriate, patients would undergo TEER procedure. From May, 2021 to January, 2023, a total of 48 HFrEF patients (median age 68 (57–84) years) with moderate-to-severe or severe SMR were enrolled. Patients were followed up for 6 months. One patient died during follow up. Non-invasive analysis of LVMW and RVMW was performed before and immediately after MitraClip treatment.
RESULTS There’s no significant change in RVEDV (128.5±20.7 mL vs 136.7±28.5 mL), RVESV (82.9±17.6 mL vs 73.5±20.6 mL) after TEER. While RVSV (44.5±9.8 mL vs 58.8±14.5 mL) and RVEF (36.6±3.6% vs 45.4±3.9%) increased. TAPSE, RV GLS, RV S’and RV FAC were not significantly changed immediately after MitraClip treatment (16.5±5.1 mm, −9.7±3.8%, 6.7±2.2 m/s and 37±14% before vs 17.1±2.1 mm, −10.5±3.5%, 6.9±1.8 m/s and 39±12% after MitraClip treatment). While RVGWI, RVGCW, RVGWE were significantly increased after MitraClip treatment (452.4±112.5 mmHg%, 596.3±127.5 mmHg% before and 85.7±15.6 mmHg% vs 589.4±119.6 mmHg%, 778.8±135.3 mmHg% and 80.9±22.4% after MitraClip treatment). And RVGWW was significantly decreased after MitraClip treatment (113.8±19.7 mmHg% vs 91.2±22.4 mmHg%). Mean KCCQ-OS and 6MWD were increase after MitraClip treatment (23.7±9.8 points and 103 (79–121) m). On multivariable linear regression analysis, RVGWI and RVGCW immediate change was independently associated with KCCQ-OS (ΔRVGWI: β=0.40, P<0.001; ΔRVGCW: β=0.39, P=0.003), RVGWI, RVGCW and RVGLS immediate change were independently associated with 6MWD improvement (ΔRVGWI: β=0.31, P=0.029; ΔRVGCW: β=0.30, P=0.039; ΔRVGLS: β=0.35, P=0.041).
CONCLUSIONS RVMW was significantly increased after MitraClip treatment. And RVGWI and RVGCW increase were independently associated with clinical improvement among HFrEF patients who underwent MitraClip treatment for SMR. The application of the proposed RVMW analysis is easy to perform and RV reserve function is an important predictor of clinical improvement in HFrEF patients with TEER.
GW34-e0302
Fei Yu1, Yan Huang2, Han Zhang1, Xuepin Hu1, Shanshan Qin1
1Shanghai Tenth People’s Hospital
2Anhui University of Science and Technology
OBJECTIVES Gate myocardial perfusion single-photon emission computer tomography (GSPECT) overestimated left ventricle (LV) ejection fraction (EF) in patients with small LV volumes, the SH reconstructed algorithm dedicated for small LV volumes provides an enhanced reconstructed spatial resolution. This study aimed to explore the clinical application value of SH.
METHODS We retrospectively analyzed patients who had both underwent GSPECT and Echocardiography (Echo) within one week. Small LV volume was defined as rest end-systolic volume (rESV)=25 mL which was reconstructed using the Standard (SD) algorithm. The LVEF difference >5% between the two algorithms was considered successfully corrected, and the ROC curve was used to calculate the optimal cut-off value of the SH algorithm. Echo being the gold standard for LV function parameters, the differences between the three methods were evaluated in comparison.
RESULTS The final study included 209 patients (73.21% female, age 67.34±7.85 years). In the overall population, SH significantly decreased LVEF (67.43±7.38% vs 71.30±7.61%, P<0.001) compared with SD, but still had statistical difference compared with Echo in the overall patients (67.43±7.38% vs 63.15±2.65%, P<0.001). The optimal cut-off value for using SH was rESV>17 mL (AUC=0.651, sensitivity=78.43%, specificity=45.57%, P=0.001). In the subgroup of rESV>17 mL, there was no statistical difference of LVEF (61.84±4.67% vs 62.83±2.85%, P=0.481) between SH and Echo.
CONCLUSIONS The pilot study demonstrated the SH algorithm could effectively correct the overestimation of LVEF in patients with small LV volumes, which yield more clinical value in the subgroup of rESV>17 mL.
GW34-e0337
Zinuan Liu1,2, Yipu Ding2,3, Guanhua Dou4, Xi Wang2, Dongkai Shan2, Bai He2, Jing Jing2, Tao Li5, Junjie Yang2, Yundai Chen2
1Medical School of Chinese PLA, Beijing, China
2Senior Department of Cardiology, The Sixth Medical Center, Chinese PLA General Hospital, Beijing, China
3School of Medicine, Nankai University, Tianjin, China
4Department of Cardiology, The Second Medical Center & National Clinical Research Center for Geriatric Diseases, Chinese PLA General Hospital, Beijing, China
5Department of Radiology, The First Medical Center of PLA General Hospital, Beijing, China
OBJECTIVES Coronary computed tomography angiography (CCTA)-derived fractional flow reserve (CT-FFR) enables physiological assessment and risk stratification, which is of significance in diabetic patients with nonobstructive coronary artery disease (CAD). We aim to evaluate prognostic value of the global trans-lesional CT-FFR gradient (GΔCT-FFR), a novel metric, in patients with diabetes without flow-limiting stenosis.
METHODS Patients with diabetes suspected of having CAD were prospectively enrolled. GΔCT-FFR was calculated as the sum of trans-lesional CT-FFR gradient in all epicardial vessels greater than 2 mm. Patients were stratified into low-gradient without flow-limiting group (CT-FFR >0.75 and GΔCT-FFR <0.20), high-gradient without flow-limiting group (CT-FFR >0.75 and GΔCT-FFR≥0.20), and flow-limiting group (CT-FFR =0.75). Discriminant ability for major adverse cardiovascular events (MACE) prediction was compared among 4 models [model 1: Framingham risk score; model 2: model 1+Leiden score; model 3: model 2+ high-risk plaques (HRP); model 4: model 3+Global ΔCT-FFR] to determine incremental prognostic value of GΔCT-FFR.
RESULTS Of 1215 patients (60.1±10.3 years, 53.7% male), 11.3% suffered from MACE after a median follow-up of 57.3 months. GΔCT-FFR (HR: 2.88, 95% CI: 1.76–4.70, P<0.001) remained independent risk factors of MACE in multivariable analysis. Compared with the low-gradient without flow-limiting group, the high-gradient without flow-limiting group (HR: 2.86, 95% CI: 1.75–4.68, P<0.001) was associated with higher risk of MACE. Among the 4 risk models, model 4, which included GΔCT-FFR, showed the highest C-statistics (C-statistics: 0.75, P=0.002) as well as a significant net reclassification improvement (NRI) beyond model 3 (NRI: 0.605, P<0.001).
CONCLUSIONS In diabetic patients with non-obstructive CAD, GΔCT-FFR was associated with clinical outcomes at 5-year follow-up, which illuminates a novel and feasible approach to improved risk stratification for a global hemodynamic assessment of coronary artery in diabetic patients.
(Approval S2020-255-01).
GW34-e0558
Shuang Li, Baiyan Zhuang, Yue Ren, Hui Wang, Lei Xu
Department of Radiology, Beijing Anzhen Hospital, Capital Medical University, Beijing, China
OBJECTIVES Myocardial fibrosis is a significant pathological characteristic observed in various cardiovascular diseases and is closely associated with prognosis. The noninvasive assessment of myocardial fibrosis using cardiac magnetic resonance (CMR) has emerged as a promising method to improve risk stratification in patients. However, limited knowledge exists regarding the myocardial fibrosis in individuals with alcoholic cardiomyopathy (ACM). The study aims to investigate the prognostic importance of myocardial fibrosis detected through CMR in patients diagnosed with ACM.
METHODS In this retrospective study, we included a consecutive series of patients with ACM who underwent enhanced CMR with T1 mapping between September 2015 and January 2023. Myocardial fibrosis was assessed by late gadolinium enhancement (LGE), as well as native T1 and extracellular volume (ECV) fraction. The primary endpoint was a composite of cardiac-related death, heart transplantation, hospitalization for heart failure, life-threatening arrhythmias, and implantable cardioverter-defibrillator or cardiac resynchronization therapy implantation. Cox proportional hazards models were used to identify independent predictors of these endpoints.
RESULTS A total of 127 patients with ACM (127 men, 51±10 years) were enrolled. Over a median follow-up period of 23.0 months (interquartile range (IQR) of 10.8–34.3), 35 patients experienced the primary endpoints. Multivariate analyses revealed that the LGE presence (hazard ratio [HR]: 1.86; 95% confidence interval [CI]: 1.08, 4.25; P=0.01), native T1 (per 10 ms increase, HR: 1.06; 95% CI: 1.02, 1.11; P=0.002), and ECV (per 3% increase, HR: 1.74; 95% CI: 1.31, 2.31; P<0.001) were identified as independent predictors of the primary endpoints. In a series of prognostic models, the addition of myocardial fibrosis (LGE, native T1 and ECV) improved the predictive capability compared to models comprising only clinical and functional variables. We found a nonlinear correlation between the total lifetime ethanol dosage and myocardial fibrosis, with a slightly increased or relatively stable fibrosis level at lower total lifetime doses but a more pronounced increase at higher doses of ethanol exposure.
CONCLUSIONS CMR-detected myocardial fibrosis in patients with ACM demonstrated predictive and incremental value beyond clinical and functional variables for major adverse events, suggesting its potential as a tool for risk stratification in this population.
GW34-e0627
Yan-qiu Wang, Wei-wei Zhou
General Hospital of Northern Theater Command
OBJECTIVES To investigate the clinical application value of instant bedside echocardiography in etiology differential diagnosis of patients with acute chest pain.
METHODS A total of 114 patients with acute chest pain admitted to the General Hospital of Northern Theater Command from November 2022 to February 2023 were selected as the study subjects. All patients were admitted to the chest pain center and underwent bedside echocardiography within 30 minutes. The results of bedside echocardiography and misdiagnosed cases were recorded. Bedside echocardiography was recorded and compared with clinical diagnosis.
RESULTS A total of 114 patients with acute chest pain admitted to the General Hospital of Northern Theater Command from November 2022 to February 2023 were selected as the study subjects. All patients were admitted to the chest pain center and underwent bedside echocardiography within 30 minutes. The results of bedside echocardiography and misdiagnosed cases were recorded. Bedside echocardiography was recorded and compared with clinical diagnosis.
CONCLUSIONS Instant bedside echocardiography can quickly and effectively assist clinical diagnosis of common acute chest pain causes, and can effectively differentiate the diagnosis of common acute chest pain diseases such as acute myocardial infarction, aortic dissection, acute pulmonary embolism, etc.
KEY WORDS: Bedside echocardiography; Acute chest pain; Chest pain center; Cause of disease; Differential diagnosis.
GW34-e0644
Wenwen Yang1,2,3, Yueqi Wang3, Changgeng Fu1,2, Changjian Li4, Feng Feng5, Hongzheng Li1,2,6, Ling Tan1,2, Hua Qu1,2, Hui Hui3, Jingjing Wang3,7, Jie Tian3, Linzi Long1,2,8
1Department of Cardiology, Xiyuan Hospital of China Academy of Chinese Medical Sciences, Beijing 100091, People’s Republic of China
2National Clinical Research Center for Cardiovascular Diseases of Traditional Chinese Medicine, Beijing 100091, People’s Republic of China
3CAS Key Laboratory of Molecular Imaging, Beijing Key Laboratory of Molecular Imaging, the State Key Laboratory of Management and Control for Complex Systems, Institute of Automation, Chinese Academy of Sciences, Beijing 100190, People’s Republic of China
4School of Engineering Medicine, Beihang University, Beijing 100191, People’s Republic of China
5College of energy engineering, Zhejiang University, Zhejiang 310058, People’s Republic of China
6Beijing University of Traditional Chinese Medicine Graduate School, Beijing University of Chinese Medicine, Beijing 100105, People’s Republic of China
7Department of Cardiovascular Medicine, First Medical Center, General Hospital of the People’s Liberation Army of China, Beijing 100853, People’s Republic of China
8Academy of Integrative Medicine, Fujian University of Traditional Chinese Medicine, Fuzhou 350122, People’s Republic of China
OBJECTIVES The most effective therapy for limiting infarct size and improving clinical outcomes following acute myocardial infarction (MI) is the rapid restoration of blood flow through the occluded coronary artery via primary percutaneous coronary intervention. Compared with thrombolytic therapy, primary percutaneous coronary intervention exhibits reduced morbidity and mortality; however, MI/R lesion, a complication of vascular reperfusion therapy for acute myocardial infarction, is estimated to occur in≥60% of patients. Specifically, reperfusion injury may be independently associated with adverse left ventricular (LV) remodeling, an increased risk of fatal arrhythmias, and hospitalization for heart failure. Ferroptosis, an iron-dependent form of regulated cell death, has been increasingly recognized as an important process that mediates the pathogenesis and progression of numerous cardiovascular diseases, including MI/R injury, sepsis-induced cardiomyopathy, arrhythmia, etc. The absorption of iron by cells is mediated by iron-binding transferrin (Tf) and its receptor 1 (TfR1), which together can induce the endocytosis of reticulin-dependent complexes. The changes in TfR1 expression and alterations in iron transport from cytosol to mitochondria result in the increased mitochondrial iron associated with ferroptosis during heart injury. To this end, TfR1 represents a promising biomarker for specifically detecting MI/R-induced ferroptosis, as it corresponds to MI/R changes injury. The early stages of MI/R lesions are associated with ferroptosis, however, in vivo, noninvasive visualization of ferroptosis in MI/R using molecular imaging methods remains difficult.
METHODS Seven-week-old C57BL/6N mice were purchased from Beijing Charles River Experimental Animal Technology Company, China. In the MI/R mouse model, using 2% isoflurane to anesthetize 60 mouse. Following this, the skin was cut between the third and fourth ribs on the left side of the mouse, and the muscles were passively separated layer-by-layer, quickly extruding the myocardium and ligating the anterior descending branch of the coronary artery of the heart ∼2 mm below the junction of the left atrial appendage and conus arteriosus. The heart was quickly placed back into the heart cavity and the skin was sutured. After 30 min of ligation, the ligature thread was removed to restore cardiac perfusion. When the electrocardiogram showed obvious ischemia, the MI/R mice were randomly divided into two groups, as follows: (1) CON NPs and (2) CCI NPs. Each group was injected with 100 μL of nanoparticles at a concentration of 1 mg·mL−1 for 120 h of multimodal imaging, including ex vivo NIR and MPI imaging for 48 h.
RESULTS The feMPI, based on the TfR1-targeting and cell-penetrating peptides (CPPs) dual-targeted probe, detects cardiac injury ∼48 h in advance and quantitatively determines damage degree during post-MI/R, as compared to existing clinical imaging detection methods.
CONCLUSIONS This new imaging strategy compensated for the difficulty in detecting I/R damage during cardiac remodeling. These findings are notably consistent with the commonly used clinical biochemical indicators in the early stage of MI/R. In addition, optical and MI/R imaging was integrated as a multimodal to precisely monitor the occurrence and development of MI/R-induced cardiac injury. This study proposes a powerful imaging-effect-based feMPI strategy for the precise assessment of MI/R-induced cardiac injury, which may help elucidate diagnostic methods for ferroptosis-related heart diseases.
GW34-e0661
Yu-Ting Tan1,2,3, Yu-Man Li1,2,3, Li Zhang1,2,3, Ming-Xing Xie1,2,3, Jing Wang1,2,3
1Department of Ultrasound, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei Province, China
2Clinical Research Center for Medical Imaging in Hubei Province, Wuhan, Hubei Province, China
3Hubei Province Key Laboratory of Molecular Imaging, Wuhan, Hubei Province, China
OBJECTIVES Left atrial reservoir strain (LARS) is a sensitive imaging biomarker of left ventricular (LV) diastolic dysfunction. However, the prognostic implications of LARS in aortic regurgitation (AR) remains unknown. The purpose of this study was to examine the prognostic value of LARS in bicuspid aortic valve (BAV) patients with significant (=moderate) AR.
METHODS Two hundred and twenty BAV patients with significant AR were enrolled in our study. LARS and left ventricular global longitudinal strain (LVGLS) were derived from speckle-tracking echocardiography. The endpoint was a composite of all-cause mortality, heart failure hospitalization, and aortic valve repair or replacement. The Cox regression was used to evaluate the independent association between LARS and the endpoint.
RESULTS During a median follow-up of 364 days, 46 (20.9%) patients reached the composite endpoint. On multivariable Cox analysis, LARS (adjusted hazard ratio [aHR]: 0.85, 95% confidence interval [CI]: 0.80–0.90, P<0.001) was independently associated with adverse outcomes after adjustment for other clinical and echocardiographic indices. As dichotomized variables, patients with LARS<24% showed increased adverse outcomes risk (aHR: 2.22, 95% CI: 1.01–4.47, P=0.025). Furthermore, Model 2 incorporating LARS exhibited the best performance in model discrimination and reclassification for predicting adverse outcomes (Net reclassification index [NRI]=0.78, Integrated discrimination improvement [IDI]=0.17, all P<0.001 compared with Model 1 [i.e. Basic model plus LVGLS (%)]; NRI=0.62, IDI=0.10, all P<0.001 compared with Basic model [i.e. age, New York Heart Association≥ III, and LV end-systolic dimension]).
CONCLUSIONS In BAV patients with significant AR, LARS is an independent predictor of adverse outcomes and confers incremental prognostic utility for adverse outcomes. These findings suggest that LARS could be considered in clinical decision-making processes for such populations.
GW34-e0662
Yuting Tan1,2,3, Yuman Li1,2,3, Zhang Li1,2,3, Mingxing Xie1,2,3, Jing Wang1,2,3
1Department of Ultrasound, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei Province, China
2Clinical Research Center for Medical Imaging in Hubei Province, Wuhan, Hubei Province, China
3Hubei Province Key Laboratory of Molecular Imaging, Wuhan, Hubei Province, China
OBJECTIVES Noninvasive left ventricular (LV) pressure-strain myocardial work (MW) is a novel method for evaluating LV function by integrating myocardial deformation and afterload, with advantages over global longitudinal strain (GLS). The study aims to analyze MW in patients with well-functioning bicuspid aortic valve (BAV) and explore the influences of aortic dilation and arterial stiffness on LV function.
METHODS A total of 104 patients with well-functioning BAV and 50 controls were enrolled in our study. Global work index (GWI), global constructive work (GCW), global wasted work (GWW), global work efficiency (GWE), global longitudinal strain (GLS), and aortic stiffness index were measured. Based on the ascending aorta diameters, BAV patients were divided into three subgroups (nondilated, mild dilated, and moderate dilated).
RESULTS GWI, GCW, GWW, and aortic stiffness index were increased (all P<0.05), while GWE and GLS were significantly reduced (all P<0.001) in total BAV patients compared with controls. BAV patients with mild dilated aorta and moderate dilated aorta had increased GWW and aortic stiffness index, and a decreased GWE compared with BAV patients with nondilated aorta (all P<0.05), whereas GCW and GLS did not differ among BAV subgroups (all P>0.05). GWI was elevated in BAV patients with moderate dilated aorta compared with BAV patients with nondilated aorta (P<0.05).
CONCLUSIONS Myocardial work could assess myocardial impairment early in patients with well-functioning BAV. MW may help to differentiate the detrimental effect of aortic dilation on LV function, whereas GLS may not.
GW34-e0663
Yu-Ting Tan1,2,3, Ming-Xing Xie1,2,3, Jing Wang1,2,3
1Department of Ultrasound, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei Province, China
2Clinical Research Center for Medical Imaging in Hubei Province, Wuhan, Hubei Province, China
3Hubei Province Key Laboratory of Molecular Imaging, Wuhan, Hubei Province, China
OBJECTIVES Chronic aortic regurgitation (AR) is a cardiac lesion combining left ventricular (LV) pressure and volume overload characteristics, eventually detrimental to the myocardium. Noninvasive myocardial work offers a promising method to evaluate LV myocardial performance as it integrates myocardial deformation and afterload. The study aimed to investigate the value of noninvasive LV myocardial work in LV dysfunction in patients with chronic AR after aortic valve surgery.
METHODS Pre-operative LV ejection fraction (LVEF), global longitudinal strain (GLS), and noninvasive LV myocardial work parameters (work index [GWI], constructive [GCW] and wasted [GWW] work, and work efficiency [GWE]) were measured. Post-operative LV impairment defined as LVEF<50% at 6 to 12 months after surgery. The logistic regression model was used to determine the independent association between noninvasive LV myocardial work indices and post-operative LV dysfunction.
RESULTS The study included one hundred and twenty-five (50.02±10.16 years; 83.2% men) with severe chronic AR and preserved LVEF who underwent aortic valve surgery. Seventeen percent developed LV impairment after aortic valve surgery. Patients with post-operative LV dysfunction displayed significantly higher E/e′ ratio, lower LVEF and LV GLS. Compared with patients without post-operative LV dysfunction, patients with post-operative LV dysfunction showed lower GWI (1932.85±136.55 mmHg% vs 1795.43±143.83 mmHg%, P<0.001), GCW (2223.96±164.53 mmHg% vs 2120.57±162.57 mmHg%, P=0.010) and GWE (92.68±1.39% vs 90.38±2.13%, P<0.001), higher GWW (165.88±37.83 mmHg% vs 207.48±57.74 mmHg%, P<0.001). Moreover, a multivariate logistic regression model with GWE (Odd ratio [OR]: 0.47; 95% confidence interval [CI]: 0.34 to 0.67; P<0.001; Akaike information criterion [AIC]=86.234, C-index=0.836) showed the best capability in predicting post-operative LV dysfunction.
CONCLUSIONS Noninvasive LV myocardial work may be paramount in predicting post-operative LV impairment with advantages over afterload-dependent echocardiographic parameters in patients with chronic AR.
GW34-e0666
Su Yuxin
The Department of Cardiology, General Hospital of Northern Theater Command, No. 83 Wenhua Road, Shenyang 110016, Liaoning Province, China
OBJECTIVES Using transthoracic echocardiography (TTE) and transesophageal echocardiography (TEE) to investigate the occurrence and related causes of iatrogenic atrial septal defect (iASD) after catheter ablation combined with left atrial appendage closure (LAAC) for atrial fibrillation (AF).
METHODS We retrospectively analyzed 330 patients that underwent combined procedure of catheter ablation for AF and LAAC at General Hospital of Northern Theater Command from January 2018 to March 2022. There were 179 males and 151 females, with an average age of 65.81±4.35 years. These patients underwent TEE at 3 month and TTE at 3, 6 and 12 months after operation and were divided into persistent iASD group and spontaneous closure group according to whether there was persistent iASD at 3-months follow-up. CHA2DS2-VASc score, HAS-BLED score, atrial fibrillation type, preoperative anticoagulation, type/size of device, type/size of sheath, number/position of TSP, procedure time of left atrium. TTE was used to measure the left atrial diameter (LAd), left atrial ejection fraction (LAEF), mitral and tricuspid valve regurgitation area and regurgitation velocity. TEE was used to assess the LAA. The data from TTE and TEE before and after surgery were collected to evaluate the relevant causes of iASD after combined operation of catheter ablation and LAAC.
RESULTS
1. Comparison of baseline information and operative data between the two groups:
Preoperative left atrial volume was larger in the persistent iASD group (76.5±35.2 mL vs 61.1±21.9 mL, P=0.036).
The persistent iASD group has longer procedure time within left atrium (92.1±36.3 vs 69.1±17.8 min, P=0.003) and larger size of sheath (4.1±1.2 mm vs 3.3±1.2 mm, P=0.022). We found that the use of ICE was associated with iASD (P=0.048). In addition, we also observed that the puncture site was relatively low, about 1.85±2.23 cm above the ostium of the inferior vena cava.
2. Comparison on Echocardiographic parameters 3 months after operation between the two groups:
The persistent iASD group had higher occurrence of postoperative MR (32.3 vs 12.8%, P=0.018) and postoperative TR (27.1 vs 7.3%, P=0.002).
Results of Logistic regression analysis:
The risk factors for the closure of iASD were larger left atrium before operation, higher pressure of left atrium, longer operation time within left atrium, larger size of sheath and the use of ICE during operation (P<0.05). The risk of non-closure of iASD increased by 20.8% for every 1 mm increase in the size of left atrium before operation. The more severe the MR, the more difficult the closure of iASD (OR 3.936).
CONCLUSIONS
1. The occurrence of iASD 3 months after combined atrial fibrillation ablation and LAAC was high.
2. The size of left atrium, the pressure of left atrium, the manipulation time within left atrium, the size of sheath, and the use of ICE during operation was related to the closure of iASD.
GW34-e0674
Yu Xie1,2, Li Zhang1,2, Mingxing Xie1,2
1Department of Ultrasound Medicine, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology
2Hubei Province Key Laboratory of Molecular Imaging
OBJECTIVES Acute cardiac allograft rejection (ACAR) is one of the main reasons for mortality after heart transplantation. Its precise diagnosis has always to be a big obstacle in the clinic. Lymphocyte infiltration is essential in ACAR, and CD4+ lymphocytes were crucial. We hypothesized that ultrasound molecular imaging with nanobubbles targeted to CD4+ lymphocytes (NBCD4) could achieve the precise diagnosis of ACAR.
METHODS NBCD4 were fabricated by conjugating anti-CD4 antibodies onto nanobubbles, and isotype-conjugated nanobubbles (NBIso) were the isotype control. The characteristics and target abilities were assayed. Rat heart transplantation models were established. Ultrasound molecular imaging was performed in the syngeneic group, the allogeneic groups on post-operative day (POD) 1 and 3, and cyclosporin A (CsA)-treated allogeneic group. Histology was further analyzed for ACAR grading and infiltrating CD4+ cell quantification.
RESULTS NBCD4 adhered to CD4+ lymphocytes more efficiently than NBIso did. In the allogeneic POD3 group and CsA-treated group, the ultrasound molecular imaging signals of NBCD4 were significantly higher than those of NBIso. In the syngeneic group and allogeneic POD1 group, the signals of NBCD4 and NBIso were comparable. Furthermore, the signal of NBCD4 on POD3 allografts was significantly higher than that on CsA-treated allografts, which was still significantly higher than that on POD1 allografts. The signals of NBIso were not conspicuously different in different groups. Histology confirmed no rejection in isografts, 1R rejection in POD1 allografts, 2R rejection in CsA-treated allografts, and 3R rejection in the untreated POD3 allografts. Moreover, the signals of NBCD4 were strongly correlated with the rejection grades, as well as the number of infiltrated CD4+ lymphocytes.
CONCLUSIONS Ultrasound molecular imaging with NBCD4 might be a new approach to achieve the precise diagnosis of ACAR.
GW34-e0802
Zhuxin Wei1, Hongchuang Xu2, Xing Yang2, Shihua Zhao1
1Department of MRI, Fuwai Hospital, National Center for Cardiovascular Diseases, Chinese Academy of Medical Sciences and Peking Union Medical College
2Department of Nuclear MedicinePeking, University First Hospital
OBJECTIVES Ischemic myocardial tissue often causes an unsuitable local inflammatory response resulting in structural and biochemical changes that cause adverse cardiac remodeling and the subsequent heart failure after MI. Thus, accurate evaluation of harmful inflammation and apoptosis after MI can help guide preventive treatment in the clinic. The role of inflammation mediated by monocyte-macrophage cell system and molecular imaging targeted these have been the subject of experimental research in the field of myocardial injury remodeling for decades, whereas T lymphocytes cells have only recently come into focus. Recently, the role of cytotoxic CD8+ T lymphocytes has been revealed that the CD8+ T lymphocytes are recruited in the ischemic heart and release of Granzyme B (GzmB) leading to enhanced apoptosis and myocardial inflammation. Studies on GzmB indicated that it contributes to tissue remodeling and fibrosis by augmenting inflammation and promoting fibroblast activation. Therefore, we investigated for the first time (to our knowledge) the effects of GzmB on inflammation, repair and functional outcomes in myocardial infarction rats model using 68Ga-grazytracer PET/CT imaging.
METHODS Forty-six male Sprague-Dawley rats underwent permanent ligation and a 30-min transient occlusion of left coronary artery. The rats underwent 68Ga-grazytracer PET/CT on days 1, 3, 6, 14 and 28 after myocardial infarction. The percentage injected tracer dose per gram of within the infarct and remote tissue were assessed. Autoradiography was used to verify the effect of in vitro imaging. The Masson stain and immunohistochemical of GzmB and CD8+ were tested to assess CD8+ T cells recruited, GzmB released and fibrous repairment in infarct and remote tissues. Hearts were harvested on days 1, 3, 6, 14 and 28 after myocardial infarction for ex vivo analyses (enzyme-linked immunosorbent assay) of inflammation and repair related markers. The ischemia reperfusion model rats were tested PET/CT to analyze the relationship between infarct size and GzmB signal. The treatment group received GzmB inhibitor carbobenzoxy-IETD-fluoromethylketone (Z-IETD-FMK) early and late induce CD8+ T cells functional depletion and then underwent 68Ga-grazytracer PET/CT to test inhibitory effect. Cardiac functional tests were performed by echocardiograph to assess cardiac function 6 weeks after myocardial infarction.
RESULTS There were significantly increased 68Ga-grazytracer uptake within the infarct on days 1, 3, 6, 14 after myocardial infarction (P<0.01). Immunohistochemical showed significantly higher expression of GzmB and CD8, in line with PET/CT imaging result. And small amount of GzmB release was also present in the remote myocardial tissue. Autoradiography showed that 68Ga-grazytracer accumulation was observed in the infarcted myocardium defined by histology. The 68Ga-grazytracer uptake of ischemia reperfusion rats and GzmB inhibitor treated rats were significantly reduced compared with myocardial infarction groups (P=0.015 and P=0.013). Echocardiographic cardiac function showed that left ventricular end-diastolic volume (EDV) and end-systolic volume (ESV) were larger and left ventricular ejection fraction (LVEF) was lower in myocardial infarction than in ischemia reperfusion group. Late GzmB inhibitor treatment was shown to be effective in improving cardiac function without significant shortening of infarct size.
CONCLUSIONS This study highlighted the potential of 68Ga-grazytracer imaging to delineate adverse inflammatory response and pathological cardiac remodeling, predict heart function. PET/CT imaging-guided therapy can limit myocardial injury and improve heart function after short-term GzmB inhibition in myocardial infarction.
GW34-e1116
Yifan Chen1, Leilei Cheng2, Junbo Ge1
1Department of Cardiology, Zhongshan Hospital, Fudan University, Shanghai Institute of Cardiovascular Diseases, China
2Department of Echocardiography, Zhongshan Hospital, Fudan University, Shanghai Institute of Cardiovascular Diseases, Shanghai Institute of Medical Imaging
OBJECTIVES Transcatheter duct closure (TCDC) of patent ductus arteriosus (PDA) can be guided by aortoangiography (AAG) or transthoracic echocardiography (TTE), but feasibility of TTE-guided TCDC for all duct types and sizes remains uncertain. This study sought to investigate the comparative outcomes of TTE-guided versus AGG-guided TCDC for all duct types with various sizes.
METHODS AAG and TTE was used as a major procedural guidance of AGG-guided and TTE-guided TCDC, respectively. Clinical and echocardiographic follow-up were conducted. The primary endpoints were the intraprocedural device occluding success (DOS) defined as successfully delivering, positioning, fixing and releasing ADO without ADO-dislodgement/embolism; and one-year clinical occluding success (COS) as no residual shunting or procedure-associated complications (PACs). The secondary endpoint was PACs and its component of ADO-migration/embolism, ADO-caused obstruction, hemolysis, any cardiac surgery due to PACs or death.
RESULTS Of 842 included patients, 308 were treated by AAG-guided TCDC and 534 by TTE-guided TCDC. There was insignificant difference in baseline characteristics between the treatments. Compared AAG-guided versus TTE-guided TCDC, the final DOS was similar (100.0 vs. 99.8%, P>0.05) with more ADO replacing-reoccluding per DOS (0.12 vs. 0.06, P<0.05) in AAG-guided TCDC; COS was also similar (95.3 vs. 97.1%, P>0.05) with comparable PACs (2.5 vs. 1.2%, P>0.05) at one-year. There were 2 intraprocedural ADO-dislodgement/embolization (1 in each treatment), and 1 acute hemolysis (leading to surgical ADO extraction and duct ligation) in TTE-guide TCDC. No death occurred in one-year follow-up.
CONCLUSIONS For closure of all duct types with various sizes, TTE-guided and AAG-guided TCDC are associated similarly with high intraprocedural device success, one-year clinical success and few procedure-associated complications.
GW34-e1118
Xiaorui Xiang, Shihua Zhao
Fuwai Hospital
OBJECTIVES Trabecular complexity is a unique biometric marker like fingerprint, but its prognostic impact in patients with dilated cardiomyopathy (DCM) remains unclear. This study aimed to explore the prognostic value of trabecular complexity by using fractal analysis in patients with DCM.
METHODS A total of 276 consecutive patients with DCM from January 2011 to December 2012 were enrolled in this study. Comprehensive clinical evaluation and cardiovascular magnetic resonance (CMR) imaging investigation were obtained. Trabecular complexity was quantified with fractal analysis of cine images to estimate the global, basal, and apical fractal dimensions (FD). All patients were followed up for major adverse cardiac events (MACE) of all-cause mortality, aborted sudden cardiac death, and heart transplantation. Univariable and multivariable Cox regression analyses were applied to identify the predictors. Patients’ survival was illustrated by Kaplan-Meier curves and differences were evaluated by log-rank test. The reproducibility of FD assessment was evaluated by intraclass correlation coefficients.
RESULTS Over a 5.37-year median follow-up, 103 (37.32%) patients experienced MACE. All left ventricular FD parameters were higher in patients with events than those without events (all P<0.05). Max Basal FD emerged as the strongest MACE prognosticator among FD parameters (area under the curve [AUC], 0.84 [95% CI, 0.78–0.88]), and the optimal cutoff value was 1.27. Furthermore, Cox proportional hazards analysis revealed that Max Basal FD was independently associated with MACE (hazard ratio [HR]:1.07 per %, P=0.002) after adjustment for clinical and imaging risk factors including NT-proBNP, left ventricular ejection fraction (LVEF), and late gadolinium enhancement (LGE) presence. By Kaplan-Meier analysis, the risk of MACE increased significantly with increased Global FD, Mean Basal FD, and Max Basal FD (all log-rank P<0.001).
CONCLUSIONS Left ventricular max basal FD was an independent predictor of adverse outcomes, and fractal analysis may contribute to improving the risk stratification for patients with DCM.
GW34-e1158
Ruohan Zhao1,2, Jing Zhang1,2, Yuting Tan1,2, Siyi Zhang1,2, Li Zhang1,2, Mingxing Xie1,2, Qing Lv1,2
1Department of Ultrasound, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430022, China
2Hubei Province Key Laboratory of Molecular Imaging
OBJECTIVES Coronary artery disease (CAD) without wall motion abnormality is difficult to diagnose clinically before coronary angiography. The object of this study is to investigate the diagnostic value of echocardiographic non-invasive pressure/volume loop (P/V loop) for CAD without wall motion abnormality and to establish a novel diagnostic model.
METHODS A total of 204 patients with suspected CAD without wall motion abnormality were enrolled. According to random allocation at the ratio of 7:3, they were divided into the training set (144 cases) and the validation set (60 cases). All patients underwent coronary angiography to confirm diagnosis. Conventional echocardiographic parameters such as left atrial diameter, left ventricular diastolic volume index, etc. were obtained. The noninvasive P/V loop was obtained by left ventricular diastolic volume, left ventricular systolic volume, systolic blood pressure, diastolic blood pressure, early diastolic mitral flow velocity E, late diastolic mitral flow velocity A, and early diastolic mitral lateral annulus velocity e′. Stroke work (SW), filling energy (FE), total energy (TE), end systolic energy (ESE) and energy efficiency (EE) were inferred from the P/V loop. Logistic regression was used to develop a multi-parameter model including gender, diabetes, and EE for the diagnosis of CAD without wall motion abnormality. The diagnostic performance of the model was tested in the validation set.
RESULTS There were 101 (70.1%) patients with CAD in the training set. TE (10186.77±2888.34J vs. 9017.12±2531.66J, P<0.05) and ESE (4942.10± 1623.74J vs. 4126.67±1250.82J, P<0.05) were significantly higher in CHD patients than in non-CHD patients. EE (51.31±4.68% vs. 53.80±5.57%, P<0.05) was significantly lower than that in patients without CAD (P<0.05). SW, TE, FE, ESE were weakly correlated with Gensini score, and the correlation coefficients were r=0.18 (P<0.05), r=0.22 (P<0.05), r=0.20 (P<0.05), r=0.25 (P<0.05), respectively. Multivariate stepwise Logistic regression analysis showed that gender (OR, 9.83; 95% CI, 3.66–26.39; P<0.001), diabetes (OR, 9.00; 95% CI, 1.85–43.74; P<0.05), A (OR, 30.52; 95% CI, 2.10–443.41; P<0.05), EE (OR, 0.91; 95% CI, 0.83–0.99; P<0.05) had greater diagnostic value (AUC in training set: 0.84; AUC in validation set: 0.76).
CONCLUSIONS The non-invasive P/V loop based on echocardiography has an independent diagnostic value for coronary artery disease without wall motion abnormality. The diagnostic model in this study included gender, diabetes, A and EE, which could provide a noninvasive and simple pretest probability estimation for CAD without wall motion abnormality before coronary angiography.
GW34-e1175
Tian Xu1, Jia-Cong Nong1, Yi Xu1, Wei You1, Yan-Qing Wang2, De-Feng Pan3, Hai-Bo Jia1, Fei Ye1
1Nanjing First Hospital, Nanjing Medical University
2Jinling Hospital, Clinical Medicine School of Nanjing University
3The Affiliated Hospital of Xuzhou Medical University
OBJECTIVES Negative remodelling (NR) often contributes to stenosis of side branch ostium (SBO), which cannot be recognized by only angiography in bifurcation lesions. Whether NR affects the efficacy of percutaneous coronary intervention (PCI) for SBO in bifurcation lesions remains unclear. To investigate the influence of severe NR (sNR) of SBO for in-stent neointimal hyperplasia (NIH) and clinical outcome for patients who underwent PCI with a 2-stent strategy for distal left main bifurcation (LMb) lesions.
METHODS A total of 328 patients from four Chinese heart centres with true LMb lesions treated with a 2-stent strategy guided by intravascular ultrasound (IVUS) were enrolled into analysis retrospectively. Among them, 48 patients with completed IVUS data pre-, post-PCI and at the 1-year follow-up were considered in phase I analysis with the endpoint of in-stent NIH, then the cut-off value of remodelling index (RI) predicting percent NIH≥50% was used as the grouping basis for sNR and its correlation with target lesion failure (TLF) within 5 years was analyzed in all populations by Kaplan-Meier analysis (in phase II).
RESULTS In phase I analysis, only NR of the left circumflex ostium (LCxO) showed correlation with in-stent NIH (P=0.044), but not for the left anterior descending ostium (LADO). The predictive cut-off values of the RI of the LCxO for percent NIH≥50% was 0.854 by receiver operating characteristic curve analysis (P=0.002). In phase II analysis, the incidence of TLF related to LMb at 5-year follow-up was higher in the sNR (defined as RI =0.85) group than the non-sNR group (32.0 vs 8.1%, P<0.001).
CONCLUSIONS NR of LCxO is associated with more in-stent NIH for distal LMb lesions with a 2-stent strategy, and sNR is linked to percent NIH≥50% at the 1-year follow-up and more TLF at the 5-year follow-up.
GW34-e1199
Ying Han, Wei Tong, Yundai Chen
Department of Cardiology, the Sixth Medical Centre, Chinese PLA General Hospital, Beijing, China
OBJECTIVES Inflammation contributes to progression of atherosclerotic plaques, and reflects the disease activity and severity of atherosclerosis. However, detecting and evaluating inflammatory reaction of the unstable plaque by 18F-FDG-PET imaging may put human body in danger due to its radioactivity. Magnetic particle imaging (MPI) is an emerging, highly sensitive, radiation-free, and no-tissue-background tomographic technique that uses superparamagnetic iron oxide nanoparticles (SPIONs) as its contrast agent. Theoretically, SPIONs can be taken up by accumulated macrophages via surface scavenger at sites of inflammation. Therefore, We aimed to investigate: 1) whether SPIONs-enhanced MPI could assess inflammation in unstable plaques by imaging macrophages; 2) compare the efficacy of four types of commercially available SPIONs in atherosclerotic inflammation imaging using MPI.
METHODS We fed six-week-old male ApoE−/− mice a high-fat diet and induced unstable plaques by cast placement over the left carotid artery. After twenty weeks, mice were divided in four groups (N=3–4) and intravenously administered with four types of SPIONs (Fe concentration, 8 mg/kg), including Nanoeast (Nanjing Nanoeast Biotech Co., China), Vivotrax (Magnetic Insight Inc., USA), Synomag (Micromod Partikeltechnologie GmbH, Germany) and Ferumoxytol (AMAG pharmaceuticals, USA). MPI scanning was performed at Pre, 4 h, 12 h, 24 h, 48 h, and 72 h post-injection. Signal-to-noise ratio (SNR) value of unstable plaques on MPI images was calculated. Plaque specimens were analyzed histologically.
RESULTS For the ApoE−/− mice injected with Nanoeast, the MPI SNR values of unstable plaques at multiple time points were 1.20±0.12 (Pre), 7.11±0.79 (4 h), 9.86±1.94 (12 h), 12.20±1.23 (24 h), 6.70±1.15 (48 h), 6.01±1.57 (72 h). These revealed that MPI SNR values reached the peak at 24 h after intravenous injection of Nanoeast. While the MPI SNR values reached the peak at 12 h after intravenous injection of Synomag and Vivotrax, and were both significantly lower than the peak SNR values of the Nanoeast group (Synomag 6.91±0.52 vs. Nanoeast 12.20±1.23, P<0.05; Vivotrax 3.14±0.36 vs. Nanoeast 12.20±1.23, P<0.05). Of interest, the ferumoxytol group showed no significant differences in MPI SNR values between different time points. Histological analysis confirmed that more SPIONs accumulated in the ustable plaques of Nanoeast group than in those of the three other groups.
CONCLUSIONS The radiation-free MPI enabled macrophage detection in atherosclerotic plaques via imaging of SPIONs, which allows in vivo visualization of inflammation in atherosclerosis. Among the commercial MPI contrast agents, Nanoeast shows the highest MPI SNR values and may represent the most promising contrast agent in MPI imaging of atherosclerotic inflammation.
GW34-e1356
Shukun He1,2,3, Qiaofeng Jin1,2,3, Tianshu Liu1,2,3, Mingxing Xie1,2,3, Jing Wang1,2,3
1Department of Ultrasound Medicine, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430022, China
2Clinical Research Center for Medical Imaging in Hubei Province, Wuhan 430022, China
3Hubei Province Key Laboratory of Molecular Imaging, Wuhan 430022, China
OBJECTIVES Pressure overload leads to myocardial fibrosis formation and reduction of myocardial compliance, further leads to left ventricular dysfunction, and eventually heart failure (HF). Early assessment of left ventricular remodeling and dysfunction in HF will have implications for the clinical management of HF patients. First-phase ejection fraction (EF1) has been proved to be a sensitive parameter for detection early left ventricular systolic dysfunction. Here we aimed to investigate the diagnostic value of EF1 in the assessment of left ventricular remodeling and dysfunction in HF.
METHODS Six-week-old SD rats were randomly divided into blank control group (n=6), sham-operated group (n=6) and modeling group (n=7). Pressure overload-induced HF model was established by minimally invasive transverse aortic constriction. Serial echocardiography was performed weekly to evaluate EF1, myocardial strain and conventional parameters in baseline and 4 consecutive weeks after operation. Rat hearts were examined weekly for haematoxylin and eosin staining and Masson’s trichrome staining analysis. The correlation between EF1 and left ventricular fibrosis and hypertrophy were analyzed.
RESULTS EF1 was decreased at 2 weeks after operation, and further reduced with the progress of observation time (P<0.05). Left ventricular global longitudinal strain (GLS) and ejection fraction (EF) impaired at 3 weeks after operation (P<0.05). Left ventricular fibrosis and hypertrophy presented at 2 weeks after operation, and continued to show an upward trend (P<0.05). Correlation analysis showed that EF1 was strongly correlated with myocardial fibrosis and hypertrophy (r=-0.80, P<0.001 and r=-0.75, P<0.001, respectively).
CONCLUSIONS EF1, the early left ventricular systolic dysfunction predictor, is a sensitive and dynamically accessible biomarker for detecting primitive left ventricular remodeling and dysfunction of HF induced by pressure overload.
CARDIOVASCULAR DISEASE NURSING
GW34-e0016
HuiMin Zhang
Second Hospital of the Army Medical University
OBJECTIVES Growing evidence indicates that nurse staffing affects the hospital mortality. but little is known about whether the educational composition of registered nurses (RNs) in hospitals is related to patient outcomes. To examine whether the proportion of hospital RNs educated at the baccalaureate level or higher is associated with risk-adjusted mortality and failure to rescue (deaths in cardiac patients with serious complications).
METHODS Cross-sectional analyses of outcomes data for 1825 cardiovascular medical, Cardiovascular surgery and CCU patients discharged from our hospitals between March 1, 2022, and December 31, 2022, linked to administrative and hospital data providing information on educational composition, staffing, and other characteristics. Risk-adjusted patient mortality and failure to rescue within 30 days of admission associated with nurse educational level.
RESULTS The proportion of RNs holding a bachelor’s degree or higher ranged from 77 to 97% across the hospital. After adjusting for patient characteristics and wards structural characteristics (size, teaching status, level of technology), as well as for nurse staffing, nurse experience, a 10% increase in the proportion of nurses holding a bachelor’s degree was associated with a 5.6% decrease in both the likelihood of patients dying within 30 days of admission and the odds of failure to rescue (odds ratio, 0.95; 95% confidence interval, 0.91–0.99 in both cases).
CONCLUSIONS In wards with higher proportions of nurses educated at the baccalaureate level or higher, CCU patients experienced lower mortality and failure-to-rescue rates.
GW34-e0220
Zhipeng Bao, Ying He
The First Affiliated Hospital of Nanjing Medical University
OBJECTIVES Radiofrequency catheter ablation (RFCA) for patients with atrial fibrillation (AF) can generate considerable physical and psychological discomfort under conscious sedation. App-based mindfulness meditation combined with an electroencephalography (EEG)-based brain-computer interface (BCI) shows promise as effective and accessible adjuncts in medical practice.
METHODS This single-center pilot randomized controlled trial involved 84 eligible patients with AF scheduled for RFCA, who were randomized 1:1 to the intervention and control groups. Both groups received a standardized RFCA procedure and a conscious sedative regimen. Patients in the control group were administered conventional care, while those in the intervention group received BCI-based app–delivered mindfulness meditation from a research nurse. The primary outcomes were the changes in the numeric rating scale, State Anxiety Inventory, and Brief Fatigue Inventory scores. Secondary outcomes were the differences in hemodynamic parameters (heart rate, blood pressure, and peripheral oxygen saturation), adverse events, patient-reported pain, and the doses of sedative drugs used in ablation.
RESULTS BCI-based app–delivered mindfulness meditation, compared to conventional care, resulted in a significantly lower mean numeric rating scale (mean 4.6, SD 1.7 [app-based mindfulness meditation] vs mean 5.7, SD 2.1 [conventional care]; P=0.008), State Anxiety Inventory (mean 36.7, SD 5.5 vs mean 42.3, SD 7.2; P<0.001), and Brief Fatigue Inventory (mean 3.4, SD 2.3 vs mean 4.7, SD 2.2; P=0.01) scores. No significant differences were observed in hemodynamic parameters or the amounts of parecoxib and dexmedetomidine used in RFCA between the 2 groups. The intervention group exhibited a significant decrease in fentanyl use compared to the control group, with a mean dose of 3.96 (SD 1.37) mcg/kg versus 4.85 (SD 1.25) mcg/kg in the control group (P=0.003). The incidence of adverse events was lower in the intervention group (5/40) than in the control group (10/40), though this difference was not significant (P=0.15).
CONCLUSIONS BCI-based app–delivered mindfulness meditation effectively relieved physical and psychological discomfort and may reduce the doses of sedative medication used in RFCA for patients with AF.
GW34-e0223
Tianxi Yu, Guozhen Sun
School of Nursing, Nanjing Medical University
OBJECTIVES Chronic Heart Failure (CHF) still affects millions of people worldwide despite great advances in therapeutic approaches in the cardiovascular field. Cardiac rehabilitation (CR) is known to improve disease-related symptoms, quality of life and clinical outcomes, yet implementation was suboptimal, a frequently low engagement in rehabilitation programs has been found globally. To quantify diverse CR-engaged processes and elucidate predictors of the various levels of CR engagement in CHF patients.
METHODS CHF patients admitted or discharged from cardiology departments between May 1 2022 to November 1 2022 were enrolled. Individuals who met the inclusion criteria filled the questionnaires, including the generalized anxiety disorders scale, patient health questionnaire, cardiac rehabilitation inventory, patient activation measure, Tampa scale for kinesiophobia heart, social frailty, Patient Health Engagement Scale (PHE-s®) We obtained sociodemographic characteristics and clinical data from medical records. The survey was distributed via mobile phone text messaging or face-to-face completed. Chi-square tests and multivariable logistic regression analyses were performed to examine the factors associated with CR engagement phases.
RESULTS A total of 684 patients were included in the study. Univariate analysis showed that only kinesiophobia had no correlation with engagement phases besides socio-demographic and clinical variables. Multivariate logistic regression analysis revealed that compared with the blackout phase process anxiety (Arousal: OR 0.829, 95% CI: 0.73∼0.94; Adhesion: OR 0.725, 95% CI: 0.64∼0.82; Eudaimonic Project: OR 0.674, 95% CI: 0.59∼0.77), monthly income (RMB yuan) equal to or more than 5000 (Arousal: OR 6.342, 95% CI: 1.30∼31.01; Adhesion: OR 5.226, 95% CI: 1.09∼24.96; Eudaimonic Project: OR 6.658, 95% CI 1.26∼34.76) were the most important factor impacting CHF patients CR engagement. In the Arousal phase, versus the Blackout phase, regular exercise or not (OR 3.29, 95% CI: 1.19∼9.10), severe depression (OR 0.019, 95% CI: 0.00∼0.813), previous cardiac-related hospitalizations 1 or 2 times (OR 3.75, 95% CI: 1.19∼11.86), Age (OR 0.958, 95% CI: 0.92∼0.998) influenced patient CR engagement. Besides, compared to the Blackout phase, outcome anxiety (OR 1.269, 95% CI: 1.11∼1.46) and activation level (level 2: OR 9.357, 95% CI: 1.44∼60.68; level 3: OR 29.96, 95% CI: 3.67∼244.92; level 4: OR 29.71, 95% CI: 3.62∼243.61) were independent factors predicting the Eudaimonic Project phase.
CONCLUSIONS This study characterized CR engagement, and explored demographic, medical, and psychological factors-with the most important being process anxiety, monthly income, patient activation, severe depression, and previous cardiac-related hospitalizations. The predictor factors of CR engagement were not identical among different phases which strongly indicates a significant role in quantifying CR engagement. Our findings suggested that factors could potentially be targeted in clinical practice to identify low CR engagement patients, and strategies implemented to strengthen or overcome these associations to address low CR engagement in CHF patients.
GW34-e0264
Zhijie Tang, Guozhen Sun
Nanjing Medical University School of Nursing
OBJECTIVES The sleep quality following radiofrequency ablation in patients with atrial fibrillation is subject to variation due to numerous factors. However, the specific trajectories of sleep quality remain unmapped, and the predictors of such trajectories remain unclear. The objective of this study is to identify the trajectories of sleep quality within six months post-radiofrequency ablation in patients with atrial fibrillation and to determine the multivariate predictors thereof.
METHODS Three hundred patients with atrial fibrillation undergoing radiofrequency ablation were followed up four times over a six-month period. We used patient self-report questionnaire and clinical information system to collect patient related information. The latent class growth analysis (LCGA) was used to identify trajectories of sleep quality. Multinomial Logical Regression was used to identify the influence of socio-demographic, psychological, and clinical factors on trajectories of symptom burden for patients.
RESULTS The study’s findings indicate that patients with atrial fibrillation who underwent ablation experienced a gradual improvement in sleep quality across four assessments, with four distinct trajectories observed: Mild-Gradual Improvement 25.0%, Mild-Stability 32.7%, High-Stability 32.0%, and High-Deterioration 10.3%. Predictive factors for poor trajectories included hypertension, Type D personality, history of atrial fibrillation>3 years, frailty, female, monthly income <3000¥.
CONCLUSIONS There are different trajectory subgroups of sleep quality in patients after atrial fibrillation ablation. Clinical and community health care workers can give patients targeted sleep support measures based on the characteristics of patients after radiofrequency ablation, in order to improve sleep quality.
GW34-e0413
Shuyi Sun1, Min Gao2, Guozhen Sun1,2, Tianxi Yu1, Shenxinyu Liu1, Zhijie Tang1
1The Nursing School, Nanjing Medical University, Nanjing, China
2Department of Cardiology Medicine, The First Affiliated Hospital of Nanjing Medical University, Nanjing, Jiangsu, China
OBJECTIVES Patients with heart failure require multiple self-control to handle physical, psychological, and social challenges, which may result in ego-depletion. However, there is a lack of evidence about ego-depletion in patients with heart failure in China. Hence, this study aimed to explore the level of ego-depletion in patients with heart failure and the associated factors.
METHODS In this cross-sectional study, we screened patients with heart failure treated in the Department of Cardiology Medicine, People’s Hospital of Jiangsu Province between February 2022 and October 2022. T-test, analysis of variance, Pearson correlation analysis, and multiple regression analysis were applied to explore the factors associated with ego-depletion in patients with heart failure.
RESULTS A total of 327 patients with HF were included in the study. The mean and SD of ego-depletion was 44.50±8.18. Univariate analysis showed that education level, household per capita monthly income, exercise habits, sleep status, food intake status, Cardiac function grade of NYHA, age, the number of comorbidities, LAD, BNP, self-care ability, anxiety, depression, and psychological distress were associated with ego-depletion (P<0.05). The multiple linear regression analysis showed that psychological distress, depression and exercise habits were found to be independently associated with the ego-depletion in patients with HF (P<0.01).
CONCLUSIONS Patients with HF experience high level of ego-depletion. Depression, psychological distress, and lack of exercise were considered as independent risk factors of ego-depletion. Health workers should pay enough attention to ego-depletion, improve the understanding of ego-depletion, and apply targeted interventions to alleviate ego-depletion in patients with heart failure.
GW34-e0980
Sisi Wu, Daqiong Wei, Chunhua Long, Xuemei Tu, Shasha Li
Department of Cardiovascular Medicine, Chongqing University Three Gorges Hospital
OBJECTIVES Low physical activity has long been identified as a risk factor for coronary heart disease. Risk factors for physical activity after percutaneous coronary intervention need to be determined. To investigate the level of physical activity in patients after percutaneous coronary intervention at 3 months after discharge, and to explore the influencing factors of physical activity in patients after percutaneous coronary intervention.
METHODS A longitudinal research design was adopted. From April 2021 to December 2021, the patients with acute myocardial infarction in the cardiovascular medicine ward of a tertiary hospital in China were selected by convenience sampling. A general information questionnaire, anxiety screening scale (GAD-7), depression screening scale (PHQ-9), Pittsburgh Sleep Quality Index (PSQI), and the Mini-Nutrition Assessment (MNA) were used to investigate influencing factors of physical activity in patients after percutaneous coronary intervention before discharge. Two hundred and forty-one patients were investigated with the International Physical Activity Questionnaire at 3 months after discharge.
RESULTS 72.2% of patients with acute myocardial infarction reported low levels of physical activity 3 months after discharge. Logistic regression analysis showed age (OR=2.060, P<0.001) and sleep quality (OR=21.732, P=0.044) were risk factors for physical activity.
CONCLUSIONS The level of physical activity of patients with acute myocardial infarction 3 months after PCI is low, and age and sleep quality are important predictors of physical activity in patients with acute myocardial infarction. It is recommended to develop a physical activity strategy suitable for the rehabilitation of patients with acute myocardial infarction and to carry out continuous health promotion behaviors as soon as possible.
GW34-e1209
Mengqi Liu1, Pingping He2
1School of Nursing, University of South China
2Medical College, Hunan Normal University
OBJECTIVES This study aimed to describe social isolation in elderly patients with coronary heart disease and examine the relationship between Type D personality, chronic disease resources, experiential avoidance, and social isolation.
METHODS A total of 322 elderly patients with coronary heart disease were recruited at 3 general hospitals from Hengyang, China using convenience sampling method. The Chinese version of the General Alienation Scale (GAS), the Chronic Illness Resource survey (CIRS), the type D personality scale (DS-14) and the Acceptance and Action Questionnaire-II (AAQ-II) were used to collect data from September 2022 to April 2023. The structural equation modelling was used to analyze the serial mediation model.
RESULTS Results from 322 study subjects indicated that social isolation in elderly patients with coronary heart disease is at a moderate or severe levels. Multiple mediation analysis showed that Type D personality had a direct positive effect on social isolation, and the relationship between them can be mediated by chronic disease resources and experiential avoidance, respectively. Moreover, there are serial mediating effect between chronic disease resources and experiential avoidance in this relationship.
CONCLUSIONS Social isolation in elderly patients with coronary heart disease needs attention., especially in patients with type D personality. Improving chronic disease resource and reducing experiential avoidance may be effective strategies to mitigate the adverse impact of type D personality and social isolation. This study extends our understanding of the relationships between Type D personality, chronic illness resources, experiential avoidance, and social isolation among older patients with coronary heart disease. Effective interventions by well-trained healthcare providers are needed to alleviate patients’ social isolation.
CARDIOVASCULAR LAB MED
GW34-e0245
Umida Kamilova, Gulnosa Zakirova, Dylafruz Masharipova, Nuriddin Nuritdinov, Bobur Utemuradov
Republican Specialized Scientific and Practical Medical Center of Therapy and Medical Rehabilitation
OBJECTIVES Assessment of hemostasis parametrs and endothelial dysfunction in COVID-19 convalescents.
METHODS One hundred and five COVID-19 convalescents were examined. Patients were included in the study 4–6 months after infection with COVID-19. The average age of the participants was 51.8±6.7 years. The following laboratory studies were performed: general blood analysis, coagulogram, detection of D-dimer by immunoenzyme method (Vector-Best, Russia), high-sensitivity C reactive protein (CRP) (Demeditec Diagnostics, Germany), endothelin-1, von Willebrand factor (VWF), thrombomodulin was determined by immunoenzymatic method on the Humareader HS (“HUMAN”, Germany) analyzer with the use of Elabscience (USA) reagents.
RESULTS The main markers of endothelial dysfunction - endothelin-1, VWF and thrombomodulin were analyzed depending on hemostasis indicators. In patients with a normal level of fibrinogen (less than 400 mg/dL), endothelin-1 was 82.81±3.12 pg/mL, VWF - 122.69±4.8%, thrombomodulin - 1384±19.2 pg/mL did, and in the group of patients with elevated fibrinogen level (more than 400 mg/dL) compared to patients with normal fibrinogen level, these indicators were 28.5% (r<0.01), 23.5% (r<0.05) and 15.7% (r<0.05) was correspondingly high. In patients with a normal level of D-dimer (less than 0.6 mg/L), the endothelin-1 index was 85.44±2.82 pg/mL, VWF - 116.78±5.95% and thrombomodulin 1215.9±24, 7 pg/mL, and compared to patients with normal D-dimer levels, in patients with high D-dimer levels (more than 0.6 mg/L), these values were 30.8% for endothelin-1 (r<0.01), 28.7% (r<0.01) for VWF and 18.6% (r<0.05) for thrombomodulin were convincingly higher, respectively. Studies have shown that D-dimer levels in patients with COVID-19 persisted long into the post-COVID period and were associated with impaired endothelial function.
CONCLUSIONS In COVID-19 convalescents, coagulation hemostasis indicators - fibrinogen and D-dimer levels were significantly increased, which were associated with signs of endothelial dysfunction. The combined use of three humoral markers characterizing endothelial dysfunction - endothelin-1, VWF and thrombomodulin - opens new perspectives for monitoring in the post-Covid period.
GW34-e0985
Ryuya Naruta1, Sari Nurmali1, Yasufumi Katanasaka1,2,3, Yoichi Sunagawa1,2,3, Toshihide Hamabe-Horiike1,2,3, Masafumi Funamoto1,2, Satoshi Shimizu1, Kana Shimizu1, Akira Murakami4, Koji Hasegawa2, Tatsuya Morimoto1,2,3
1Division of Molecular Medicine, School of Pharmaceutical Sciences, University of Shizuoka
2Division of Translational Research, Kyoto Medical Center
3Shizuoka General Hospital
4Department of Food Science and Nutrition, School of Human Science and Environment, University of Hyogo
OBJECTIVES Heart failure (HF) is one of the leading causes of death in developed countries. Cardiac hypertrophy and fibrosis are known to be essential factors in cardiac remodeling and the progression of HF, but highly effective therapeutic agents targeting these processes have not yet been developed. We have previously shown that natural compounds such as Curcumin inhibit HF by suppressing cardiac hypertrophy and fibrosis in vivo, suggesting that these compounds are possible candidates for HF therapy and prevention. Zerumbone (Zer) is a major active terpene found in the endemic wild ginger species of Southeast Asia. This wild ginger is usually used not only as a spice and fragrance but also as a medicine, mainly in Indonesia and Malaysia. The purpose of this study is to investigate whether the natural compound Zer inhibits cardiomyocyte hypertrophy, fibrosis, and the development of HF.
METHODS To investigate the effect of Zer on cardiac hypertrophy and cardiac fibrotic responses, primary cultured cardiomyocytes and cardiac fibrosis prepared from neonatal rats were pretreated with Zer and then stimulated with phenylephrine (PE) or transforming growth factor beta (TGF-β), respectively. Immunofluorescent staining, quantitative PCR (qPCR) analysis, and western blotting (WB) were performed. To determine whether Zer prevents the development of pressure overload-induced HF in vivo, a transverse aortic constriction (TAC) mouse model was utilized. Cardiac function was evaluated by echocardiography. The cardiomyocyte surface area was observed using histological analysis (HE and WGA staining). Fibrosis formation was measured by picrosirius staining. The total mRNA levels of genes associated with hypertrophy (ANF and BNP) and fibrosis (Postn and α-SMA) were examined by qRT-PCR. The protein expressions (Akt and α-SMA) were assessed by western blotting.
RESULTS To investigate the effect of Zer on cardiac hypertrophy and cardiac fibrotic responses, primary cultured cardiomyocytes and cardiac fibrosis prepared from neonatal rats were pretreated with Zer and then stimulated with phenylephrine (PE) or transforming growth factor beta (TGF-β), respectively. Immunofluorescent staining, quantitative PCR (qPCR) analysis, and western blotting (WB) were performed. To determine whether Zer prevents the development of pressure overload-induced HF in vivo, a transverse aortic constriction (TAC) mouse model was utilized. Cardiac function was evaluated by echocardiography. The cardiomyocyte surface area was observed using histological analysis (HE and WGA staining). Fibrosis formation was measured by picrosirius staining. The total mRNA levels of genes associated with hypertrophy (ANF and BNP) and fibrosis (Postn and α-SMA) were examined by qRT-PCR. The protein expressions (Akt and α-SMA) were assessed by western blotting.
CONCLUSIONS The present study has shown for the first time the protective effect of Zer against PE-induced hypertrophy in cultured cardiomyocytes and TGF-β-induced fibrotic phenotypes in cardiac fibroblasts. Moreover, Zer ameliorates TAC-induced cardiac dysfunction. These suggest that Zer may be a prospective drug candidate for HF prevention in humans.
TRADITIONAL CHINESE MEDICINE
GW34-e0427
Yuxuan Liu1,2, Ping Li2
1Third Clinical Medical College, Beijing University of Traditional Chinese Medicine
2Department of Cardiology, Beijing University of Chinese Medicine Third Affiliated Hospital
OBJECTIVES Visually analyze the scientific achievements of Curcumin in the field of cardiovascular disease, and clarify its research status, hot spots and frontiers.
METHODS The data comes from the Web of Science (WoS) core collection database. The search strategies was TS=(curcumin or “Turmeric Yellow” or “Yellow Turmeric” or “Curcumin Phytosome” or “Phytosome Curcumin” or Diferuloylmethane or Mervia) AND TS=(Cardiovascular or Cardiology or circulation or heart). The language is English. The search scope is from January 1, 2004 to June 13, 2023. The selected literature types are research papers and reviews. All records (including title, author, source code, keywords, abstract, doi) and references in search results are exported in plain text format and download– * * Name in txt format. The search was completed within 1 day (June 13, 2023) to avoid changes caused by daily database updates. Use visual analysis software Cite Space 5.8.R3 for country, institutional, and keyword analysis.
RESULTS A total of 1235 articles related to Curcumin and cardiovascular diseases were included, including 843 research papers and 392 reviews. The global research of Curcumin in the field of cardiovascular disease is on the rise. A total of 82 countries participated in the study, with China being the most productive country and publishing a total of 345 papers. Germany is the country with the highest between centrality (BC). Mashhad University Medical Science not only has the highest BC (0.2), but also has the highest publication volume (66 articles). Sahebkar Amirhossein devoted himself to the research of Curcumin, and published 47 articles in total. Keywords that occur more than 100 times include oxidative stress, curvature, nf kappa b, in vivo, and expression. The top 5 keywords with BC include cancer, activation, disease, apoptosis, and advertisement issue. Lipid peroxidation is the keyword with the strongest explosive intensity.
CONCLUSIONS The research on Curcumin and cardiovascular disease is on the rise as a whole, and the antioxidant mechanism of Curcumin is the focus of researchers.
GW34-e0963
Guanjing Ling, Xiaoping Wang, Yan Wei, Yong Wang
Beijing University of Chinese Medicine
OBJECTIVES Although the development of therapies for heart failure (HF) continues apace, clinical outcomes are often far from ideal. Current studies suggest that mitophagy defects and accumulation of damaged mitochondria are linked to the progression of HF. Developing molecularly targeted agents to enhance mitophagy and characterizing more in-depth mechanisms are of great research value and clinical development prospects for curing HF. Of note, Unc51-like-kinase 1 (ULK1)-mediated mitophagy prevents pathological cardiac remodeling and HF. However, molecularly ULK1-targeted agent to enhance mitophagy is scanty. Ginsenoside Rg3 (Rg3), a major active ingredient of the root of Panax ginseng or Panax notoginseng, exerts a wide range of pharmacological protective effects on cardiovascular diseases. However, whether Rg3 could regulate mitophagy in the heart has never been reported up to now. This study aimed to investigate whether Ginsenoside Rg3 (Rg3) can activate ULK1 to trigger FUNDC1-mediated mitophagy for protecting HF.
METHODS An established HF rat model and transcriptome sequencing were used to evaluate the therapeutic effect and regulatory mechanism of Rg3. Surface plasmon resonance was used to detect the ULK1 binding behavior of Rg3. Loss-of-function approaches in vivo and in vitro were performed to determine the intrinsic mechanism of ULK1 in Rg3-elicited myocardial protection against HF. Adenovirus expressing the pH-sensitive fluorescent protein mt-Keima was used to visualize and quantify mitophagy. Fundc1 siRNA, Fundc1-S17A mutant and Fundc1-S17A mutant of plasmid were applied for mechanistic research.
RESULTS In vivo, Rg3 improved cardiac dysfunction, adverse remodeling, and mitochondrial damage in HF rats. Transcriptomics profiling revealed that Rg3 promoted Ulk1-Fundc1 axis-mediated mitophagy in the heart tissues of HF rats. Consistent with this observation, Rg3 treatment increased the phosphorylation level of Ulk1, which in turn induced the colocalization of Fundc1 and Lc3 to trigger mitophagy both in vitro and in vivo. Structurally, a good binding mode was unveiled between ULK1 and Rg3. Of note, the protective effects conferred by Rg3 against mitophagy defects, the collapse of mitochondrial potential, oxidative stress, and cardiac dysfunction were compromised by Ulk1 gene silencing both in vitro and in vivo, consolidating a pivotal role of ULK1 in Rg3-elicited myocardial protection against HF. Mechanistically, we demonstrated that Rg3 activated mitophagy through inducing ULK1-mediated phosphorylation of FUNDC1 at Ser17 site, not Ser13 site.
CONCLUSIONS Together these observations demonstrated that Rg3 acts as a ULK1 activator for the precise treatment of HF, which binds to ULK1 to activate FUNDC1-mediated mitophagy. Since mitophagy downregulation are observed in various pathological conditions in the heart, pharmacological intervention of Rg3 that activates mitophagy may contribute to curing a broad spectrum of heart diseases via maintaining mitochondria homeostasis and energy metabolism.
GW34-e1147
Meiqi Miao
Heilongjiang University of Traditional Chinese Medicine
OBJECTIVES In order to advance the research progress of combining Chinese and Western medicine in the prevention and treatment of MetS and to provide new ideas for it.
METHODS By outlining the process and mechanism of cellular autophagy, focusing on describing its relationship with the main pathological mechanisms of MetS, and based on modern medical theory and the direction of Chinese medicine understanding, we review the preventive and curative effects of Chinese medicine whole formula and its single active ingredient on MetS through the cellular autophagy pathway.
RESULTS MetS leads to an increased risk of cardiovascular disease and is now a global problem. Autophagy is a dynamic process necessary for the maintenance of energy metabolism in the body. Defects in autophagic homeostasis in the body are closely related to metabolic abnormalities, and the imbalance of energy metabolism caused by autophagy dysregulation is closely related to the main pathological mechanisms of MetS. The whole formula of TCM and the main chemical active ingredients of its single herbs can regulate cellular autophagy and intervene in the development of MetS through anti-inflammation, anti-oxidative stress and regulation of glucolipid metabolism, fully demonstrating its multi-pathway, multi-target and multi-mechanism synergistic regulation. Therefore, the addition of Chinese medicine in clinical practice is expected to bring into full play the interventional effects of Chinese medicine on MetS while Western medicine achieves significant efficacy in a single disease, and the combined treatment of MetS with Chinese and Western medicine will be a promising development direction. The understanding of autophagy in modern medicine is still in progress, and the current treatment methods based on autophagic pathways (such as autophagy enhancers) to improve the metabolic symptoms of MetS have certain clinical effects, but there are still some questions and side effects. The research of autophagy mechanism in Chinese medicine to prevent and treat MetS is also in its initial stage, and there are still many questions that need to be addressed by researchers as far as the current situation is concerned. Therefore, how to regulate autophagy homeostasis, determine its threshold value, and prevent the deterioration of MetS due to defective or excessive autophagy becomes the focus of future research in this field.
CONCLUSIONS In conclusion, this paper summarizes the relationship between cellular autophagy pathway and MetS, and the current status of research on the prevention and treatment of MetS based on autophagy pathway in Chinese medicine, which provides a new idea for the prevention and treatment of MetS in Chinese medicine, and provides a scientific basis for advancing the experimental research and clinical treatment of MetS by combining Chinese and Western medicine.
GW34-e1188
Zhiyuan Zhang, Yuanlong Hu, Xinhai Cui, Mengkai Lu, Chao Li
Shandong University of Traditional Chinese Medicine
OBJECTIVES Astragaloside IV (ASIV) is the primary pharmacologically active compound found in Astragalus propinquus Schischkin, which has potential protective effects on cardiac function. However, there are almost no systematic evaluations of ASIV for the treatment of heart failure (HF).
METHODS Preclinical studies published before December 27th, 2022, were retrieved from PubMed, Web of Science, MEDLINE, SinoMed, Chinese National Knowledge Infrastructure (CNKI), VIP information database, and Wanfang Data information site. The quality of included research was evaluated using SYRCLE’s RoB tool. Review Manager 5.4.1 was used to perform meta-analyses of cardiac function parameters and other indicators. Regression analysis was conducted to observe the dose-efficacy relationship.
RESULTS Nineteen studies involving a total of 489 animals were analyzed in this study. The findings revealed that ASIV exhibited positive effects on various indicators of cardiac function when compared to the control group. These indicators included left ventricular ejection fraction (LVEF), left ventricular fractional shortening (LVFS), left ventricular pressure change rate (±dp/dtmax), left ventricular end-diastolic pressure (LVEDP), left ventricular systolic pressure (LVSP), heart weight/body weight (HW/BW), and left ventricular weight/body weight (LVW/BW). Furthermore, the regression analysis conducted demonstrated that the effectiveness of ASIV in treating heart failure was dependent on the dosage administered.
CONCLUSIONS This is the first meta-analysis evaluating ASIV for HF treatment. Findings suggest that ASIV can inhibit cardiac hypertrophy by reducing cardiac preload and afterload, thereby protecting cardiac function.
GW34-e1294
Menghui Du1, Xiujuan Yan1, Yu Bai1, Jiong Zhang1, Chunying Si2, Tan Wang1, Jie Sun1, Xianghua Liu1, Aishe Gao1, Fang Chen1
1Department of Medical School, Henan University of Traditional Chinese Medicine, Zhengzhou, Henan, P.R. China
2The First Affiliated Hospital of Henan University of Traditional Chinese Medicine, Zhengzhou, Henan, P.R. China
OBJECTIVES We have already established a novel herbal screening system for selecting special herbs on calcified aortic valve calcification (CAVD) by Constructing a stable pGL4.10-RLUC-RAGE/CDH11 dual fluorophorence reporter gene vector. The test results show that Puerarin may have effect on aortic valve calcification. In this study, we further investigated the effects of Puerarin against aortic valve calcification via animal experiments.
METHODS Fifty ApoE−/− mice were randomly divided into 5 groups: Control, Model (normal saline of 0.1 mL/10g/d), the puerarin groups (200 mg/kg/d), FPS-ZM1 group (FPS, 1 mg/kg/d of FPS-ZM1) and Rosuvastatin group (ST, 1.6 mg/kg/d of Rosuvastatin calcium). Except for the Control, all mice in the other 4 groups were given free consumption of high-cholesterol diet for 36 weeks to construct calcified aortic valve disease (CAVD) model. Mental state and body weight were observed daily. Cardiac ultrasonography was performed to test Transvalvular pressure difference, peak flow velocity and so on. Blood glucose, blood lipids, blood calcium and phosphorus, alkaline phosphatase (ALP), alanine aminotransferase (ALT), and blood urea nitrogen (BUN) were monitored. HE staining and Alizarin red staining of AV have proceeded. The expression of calcification-related osteocalcin, Runx2, RAGE and CDH11 in the valve was measured by western blot. The expressions of IL-1β, IL-6, and TNF-α in serum were detected by ELISA.
RESULTS The body weight of mice consuming high-cholesterol diet was generally higher than that of the control group (P<0.05). The levels of blood lipids, blood glucose and blood calcium were generally increased in all experimental groups compared to the control (P<0.05). Blood ALP was significantly higher in the model group (P<0.05) and puerarin could reduce the level of ALP. The ultrasonic examination of the heart in the experimental group exhibited symptoms in different degrees while the control was more normal and the symptoms were relieved in puerarin group and ST group (P<0.05). HE staining and Alizarin Red staining showed the points of calcification and lipid deposition in the model group, accompanied by a large number of neutrophils, lymphocytes and other inflammatory infiltrates in the experimental groups. The calcification was reduced in puerarin group, ST group and FPS group (P<0.05). The expressions of calcification related protein osteocalcin, Runx2, RAGE and CDH11 were decreased in puerarin group, ST group and FPS group by western blot (P<0.05). IL-1β, IL-6, and TNF-α were decreased in puerarin group, ST group and FPS group (P<0.05).
CONCLUSIONS High-cholesterol diet and aging promote calcification of the aortic valve. Our novel herbal screening system can effectively select special herbs for CAVD. Puerarin can improve the symptoms of CAVD by reducing inflammatory response and partially by inhibiting RAGE and CDH11.
GW34-e1297
Fang Chen1, Yu Bai1, Menghui Du1, Chunying Si2, Tan Wang1, Jie Sun1, Xianghua Liu1, Aishe Gao1
1Department of medical school, Henan University of Traditional Chinese Medicine, Zhengzhou, Henan, P.R. China
2The First Affiliated Hospital of He’nan University of Chinese Medicine, Zhengzhou, Henan, P.R. China
OBJECTIVES Decotion of Huatan Huoxuejiedu is the classical clinic treatment of chest Bi syndrome at the First Affiliated Hospital of Henan University of Traditional Chinese Medicine. This decotion can effectively improved the symptoms of chest Bi by clinical observation. In this study, we investigated the effects and possible mechanisms of Huatan Huoxuejiedu Decotion against Aortic valve calcification via network pharmacological analysis and animal experiment.
METHODS Bioinformatic methods were used to find the effective ingrendients of Huatan Huoxuejiedu Decotion and genes related to the drug targets. 70 ApoE−/− mice were randomly divided into 7 groups: Control, Model (normal saline of 0.1 mL/10g/d), the low, medium and high dose groups of decotion (JDF-L, M, H, 2 g/kg, 4 g/kg and 8 g/kg of the decotion per day), FPS-ZM1 group (FPS, 1 mg/kg/d of FPS-ZM1) and Rosuvastatin group (ST, 1.6 mg/kg/d of Rosuvastatin calcium). Except for the Control, all mice in the other 6 groups were given free consumption of high-fat diet for 36 weeks. Mental state and body weight were observed daily. At the end of 36 week, Cardiac ultrasonography was performed to test Transvalvular pressure difference, peak flow velocity and so on. Blood glucose (Glu), blood lipids, blood calcium (Ca) and phosphorus (P), alkaline phosphatase (ALP), alanine aminotransferase (ALT), blood urea nitrogen (BUN) were monitored. HE staining and Alizarin red staining of AV were proceeded. The expression of osteocalcin, Runx2, and RAGE, MAPK/NF-κB, JAK/STAT expression in valve were measured by western blot. The expressions of inflammatory cytokines in serum were detected by ELISA.
RESULTS Network pharmacology showed that Decotion of Huatan Huoxuejiedu could retard CAVD progress by AGE/RAGE axis and subsequent MAPK/NF-κB, JAK/STAT pathways. Animal experiment showed the body weight of mice consuming high-fat diet was generally higher than that of the control group (P<0.05). Compared with the Control group, the levels of HDL and P in all the drug administration group and the model group were decreased, and the levels of TG, TC, LDL, Ca, ALT and BUN were generally increased. Blood ALP was significantly higher in the model group and the JDF-L group (P<0.05), while ALP levels were significantly lower in the remaining groups (P<0.05). The ultrasonic examination of the heart in the experimental group exhibited symptoms in different degrees while control was more normal and the symptoms relieved in JDF-L, M, H groups (P<0.05). HE staining and Alizarin Red staining showed the points of calcification and lipid deposition in the model group, accompanied by a large number of neutrophils, lymphocytes and other inflammatory infiltrates. The calcification was reduced most in JDF-H group (P<0.01). The expressions of calcification related protein osteocalcin, Runx2 and pathway related protein RAGE, p38MAPK, JAK, p-STAT3, STAT3, STAT5, p-STAT5, NF-κB (p65), ERK and Jun in JDF-H, statin and FPS group were decreased by western-blot (P<0.05). IL-1β, IL-6, and TNF-α were decreased in JDF-L, M, H groups (P<0.05), especially for the JDF-H group (P<0.01).
CONCLUSIONS High-fat diet and aging promote calcification of the aortic valve. Huatan Huoxuejiedu decotion, FPS-ZM1 and statin can improve the symptoms of CAVD especially for the JDF-H group. Huatan Huoxuejiedu decotion may exhibit anti-CAVD effect via RAGE related pathways and decreasing of inflammatory cytokine release.
CARDIOVASCULAR PREVENTION AND REHABILITATION
EPIDEMIOLOGY AND EVIDENCE-BASED MEDICINE
GW34-e0075
Zhenhong Ou1, Xiaoqing Liu2, Yunlin Chen2, Fangchao Wang1, Boli Ran1, Kui Cui1, Yuehui Yin2
1Chongqing General Hospital
2The Second Affiliated Hospital of Chongqing Medical University
OBJECTIVES Intensive lipid-lowering using proprotein convertase subtilisin/kexin 9 (PCSK9) inhibitors reduces the risk of major adverse cardiovascular events (MACEs) in patients with high-risk or established atherosclerotic cardiovascular disease (ASCVD). But time to benefit (TTB) for the addition of PCSK9 inhibitors is not to be investigated. We performed this meta-analysis to evaluate the TTB for prevention of a MACE with the addition of PCSK9 inhibitors upon standard therapy.
METHODS MEDLINE, Embase, CENTRAL and Web of Science were searched for Randomized controlled trials that compared the addition of PCSK9 inhibitors with placebo upon standard treatment in adults with high-risk or established ASCVD until November 18, 2022. Kaplan-Meier curves were extracted to reconstruct individual survival data. Individual TTB and confidence intervals under the specific absolute risk reduction (ARR) threshold at each trial were calculated by Weibull curve fitting and Markov chain Monte Carlo methods. A random-effects meta-analysis was conducted to estimate the overall TTB.
RESULTS Seven trials randomizing 56954 adults (mean age, 60.9 years; 27.8% were woman) were included in the analysis. Similar MACEs commonly including myocardial infarct, stroke and cardiovascular death were defined in each trial. The addition of PCSK9 inhibitors to standard treatments significantly reduced the hazard of MACE by 19% (hazard ratio, 0.81; 95% CI, 0.77–0.85; P<0.001). Overall, 2.6 (95% CI, 1.2–6.3) months was the average needed to prevent 1 MACE for 1000 patients (ARR, 0.001) treated with PCSK9 inhibitors. Similarly, 4.3 (95% CI, 2.4–8.4), 9.1 (95% CI, 5.4–15.8) and 16.6 (95% CI, 10.4–27.8) months were estimated to avoid 1 MACE for 500 (ARR, 0.002), 200 (ARR, 0.005) and 100 (ARR, 0.01) patients, respectively.
CONCLUSIONS In patients with high-risk or established ASCVD, the addition of PCSK9 inhibitors upon standard therapy may need approximately 3 months at least to produce additional cardiovascular benefits and are more likely to benefit individuals with a life expectancy of longer than 1.5 years.
GW34-e0091
Zhen Hu1, Xin Wang1, Congyi Zheng1, Linfeng Zhang1, Zuo Chen1, Haoqi Zhou2, Xue Cao1, Yixin Tian1, Runqing Gu1,3, Ye Tian1, Lan Shao1, Zengwu Wang1
1Division of Prevention and Community Health, National Center for Cardiovascular Disease, National Clinical Research Center of Cardiovascular Disease, State Key Laboratory of Cardiovascular Disease, Fuwai Hospital, Peking Union Medical College & Chinese Academy of Medical Sciences, Beijing 102308, China
2Department of Biostatistics, Peking University, Beijing 100191, China
3School of Population Medicine and Public Health, Chinese Academy of Medical Sciences & Peking Union Medical College, Beijing 100730, China
OBJECTIVES Both medication and non-medication therapies are effective approaches to control blood pressure (BP) in hypertension patients. However, the association of joint changes in antihypertensive medication use and healthy lifestyle index (HLI) with BP control among hypertension patients is not reported, which needs to be validated by prospective intervention studies. We examined the association of antihypertensive medication use and HLI with BP control among employees with hypertension in China based on a workplace-based multicomponent intervention program.
METHODS Between January 2013 and December 2014, a cluster randomized clinical trial of a workplace-based multicomponent intervention program was conducted in 60 workplaces across 20 urban areas in China. Workplaces were randomly divided into intervention (n=40) and control (n=20) groups. Basic information on employees at each workplace was collected by trained professionals, including sociodemographic characteristics, medical history, family history, lifestyle behaviors, medication status and physical measurements. After baseline, the intervention group received a 2-year intervention to achieve BP control, which included: (1) a workplace wellness program for all employees; (2) a guidelines-oriented hypertension management protocol. HLI including nonsmoking, nondrinking, adequate physical activity, weight within reference range and balanced diet, were coded on a 5-point scale (range, 0–5, with higher score indicating a healthier lifestyle). Antihypertensive medication use was defined as taking drug within the last 2 weeks. Changes in HLI, antihypertensive medication use and BP control from baseline to 24 months were measured after the intervention.
RESULTS Overall, 4655 employees were included (mean [standard deviation] age, 46.3 [7.6] years; 3547 men [82.3%]). After 24 months of the intervention, there was a significant improvement in lifestyle [smoking (0.65, 95% CI, 0.43∼0.99; P=0.045), drinking (OR=0.52, 95% CI, 0.40∼0.68; P<0.001), regular exercise (OR=3.10, 95% CI, 2.53∼3.78; P<0.001), excessive intake of fatty food (OR=0.17, 95% CI, 0.06∼0.52; P=0.002), restrictive use of salt (OR=0.26, 95% CI, 0.12∼0.56; P=0.001)]. Compare to employees with a deteriorating lifestyle after the intervention, those with an improved lifestyle had a higher BP control. In the intervention group, compared with employees not using antihypertensive medication, those who consistent used (OR, 2.34; 95% CI, 1.16∼4.72; P=0.017) or changed from not use to use antihypertensive medication (OR, 2.24; 95% CI, 1.08∼4.62; P=0.030) had higher BP control. Compared with those having lower HLI, participants with a same (OR, 1.38; 95% CI, 0.99∼1.93; P=0.056) or high (OR, 1.79; 95% CI, 1.27∼2.53; P<0.001) HLI had higher BP control. Those who used antihypertensive medication and had a high HLI had the highest BP control (OR, 1.88; 95% CI, 1.32∼2.67, P<0.001). Subgroup analysis also showed the consistent effect as the above.
CONCLUSIONS These findings suggest that adherence to antihypertensive medication treatment and healthy lifestyle were associated with a significant improvement in BP control among employees with hypertension.
GW34-e0122
Xue Cao, Yixin Tian, Zengwu Wang
National Center for Cardiovascular Disease, National Clinical Research Center of Cardiovascular Disease, State Key Laboratory of Cardiovascular Disease, Fuwai Hospital, Peking Union Medical College & Chinese Academy of Medical Sciences
OBJECTIVES Elaborating and understanding disparities in the burden of cardiovasular disease (CVD) attributable to high fasting plasma glucose (FPG) is important to improve diabetes prevention and cardiovascular health. In this study, we estimate the CVD burden attributable to high FPG at national and provincial levels in China.
METHODS We pooled data on people aged 25 years and older from population-based studied that had collected data on FPG levels through measurement of its biomarkers. The temporal-spatial Bayesian hierarchical model was used to estimate age-specific, sex-specific, province-specific, and year-specific average FPG levels from 2010 to 2018. Then we estimate population attributable fraction for CVDs by age and sex for 31 provinces. The mortality data for CVD at the provincial level were obtained from surveillance systems, which were used to compute the portion of deaths and years of life lost (YLLs) due to CVD that could be attributed to high FPG using the counterfactual scenario of theoretical minimum risk level. We also decomposed trends in attributable burdens into four explanatory components, including population growth, population aging, risk exposure to FPG, and in risk-deleted mortality rates for CVD.
RESULTS In 2018, an estimated total of 512.29 thousand (95% UI 488.60 to 538.65) adults aged 25 or older in China were attributable to high FPG, with 281.54 thousand (95% UI 267.89 to 295.05) of these deaths occurring in men and 229.75 thousand (95% UI 210.98 to 251.89) in women. High FPG accounted for 1093.45 crude YLLs per 100,000 people (95% UI 1059.13 to 1131.73) due to FPG-related CVD in 2018, with substantial variation across provinces. The age-standardised CVD mortality rate attributable to high FPG increased by 3.99%, from 41.93 per 100,000 people (95% UI 40.07 to 43.88) in 2010 to 43.60 per 100,000 (95% UI 41.77 to 45.62) in 2018. For specific causes, ischaemic heart disease was the most important contributor to FPG-related CVD deaths (254.13 thousand [95% UI 235.63 to 274.51]) in 2018, followed by ischemic stroke (139.95 thousand [95% UI 128.14 to 152.81]) and hemorrhagic stroke (117.56 thousand [95% UI 112.65 to 123.23]). In 2018, the highest age-standardised CVD mortality rate attributable to high FPG was observed in the high-middle SDI region and the middle SDI region. By province, the age-standardised mortality rate due to FPG-related CVD in both sexes combined varied from 21.31 per 100,000 people in Tibet to 77.66 per 100,000 people in Heilongjiang in 2018. Nationally, compared to 2010, exposure to high FPG and population aging in 2018 were the primary drivers of increased FPG-related deaths due to CVD, and the proportion of FPG-related CVD death attributed to population ageing in 31 provinces ranged from −3.10 in Hainan to 58.36% in Shanghai.
CONCLUSIONS The CVD burden attributable to high FPG is still large in China, and varied markedly across regions and provinces. Population ageing was associated with substantial changes in the number of FPG-related CVD deaths between 2010 and 2018. Findings in this study call for renewed efforts to implement population-specific tailored measures for diabetes prevention and cardiovascular health.
GW34-e0125
Yi Ren
Institute of Microcirculation, Chinese Academy of Medical Sciences (CAMS) & Peking Union Medical College (PUMC), Beijing 100005, China
OBJECTIVES High blood pressure is a key pathogenetic factor that contributes to the deterioration of kidney function. However, the incidence trend of hypertension-related chronic kidney disease (CKD) has rarely been studied; therefore, we aimed to analyze the global, regional, and national patterns, temporal trends as well as burden of hypertension-related CKD.
METHODS We extracted data on hypertension-related CKD from the Global Burden of Disease (GBD) study database, including the incidence, prevalence, disability-adjusted life years (DALYs), and mortality numbers and rates (per 100,000 population) and further described according to year, location, sex, age, and socio-demographic index (SDI). The estimated annual percentage changes (EAPCs) were calculated to assess the variation in incidence, DALYs, and mortality. We used an age-period-cohort (APC) model framework to analyze the underlying trends in prevalence by age, period, and birth cohort. Nordpred APC analysis was performed to predict the future morbidity and mortality of hypertension-related CKD.
RESULTS In 2019, a total of over 1.57 million new hypertension-related CKD cases were reported worldwide, a 161.97% increase from 1990. Compared to 1990, the age-standardized incidence rates (ASIR) increased in all 21 regions in 2019. In all countries and territories except Iceland, the EAPC in ASIR and the lower boundary of its 95% confidence interval (CI) were higher than 0. From 1990 to 2019, the global age-standardized mortality rates (ASMR) from hypertensive heart disease, ischemic heart disease, and stroke attributed to high systolic blood pressure decreased by 21.49% (95% UI: 10.13 to 35.18%), 32.64% (95% UI: 28.74 to 36.81%) and 34.89% (95% UI: 28.69 to 40.87%), respectively, but a similar decrease was not observed for hypertension-related CKD, which actually increased by 17.56% (95% UI: 7.59 to 24.87%). ASIR, age-standardized prevalence rates (ASPR), age-standardized DALYs rates (ASDR), and ASMR were not identical among countries with different SDI regions in 2019; additionally, ASIR and ASMR were significantly different among sexes in all SDI regions in 2019. The predicted incidence and mortality counts globally continue to increase to 2044, and there is an upward trend in ASIR for both men and women.
CONCLUSIONS Between 1990 and 2019, the ASIR of hypertension-related CKD demonstrated an ascending trend, and according to our projections by Norpred APC model, it would remain on the rise for the next 25 years and reach 21.28 (per 100,000 population) in 2044. Blood pressure lowering treatment significantly reduced the risk of major adverse cardiovascular events and stroke in various populations of hypertensive patients; however, there was a lack of overall benefit of blood pressure lowering for CKD. With remarkable global population growth, aging, and an increasing number of patients with hypertension, the burden of disease caused by hypertension-related CKD continues to increase.
GW34-e0158
Yunfei Wu1, Tianshu Han2, Jun Lyu3, Yidan Wang1, Wenbo Qu1, Shanjie Wang1, Feiyuan Han1, Shiqi Yuan4, Xiangwen Xi1, Wenjun Ni1, Yanwen Zhang1, Xueyu Wang1, Shan Zhong1, Minghui Piao1, Bo Yu1, Jinwei Tian1
1Department of Cardiology, The Second Affiliated Hospital of Harbin Medical University; The Key Laboratory of Myocardial Ischemia, Chinese Ministry of Education, 246 Xuefu Road, Harbin 150081, China
2National Key Discipline, Department of Nutrition and Food Hygiene, School of Public Health, Harbin Medical University, 157 Baojian Road, Harbin 150081, China
3Department of Clinical Research, The First Affiliated Hospital of Jinan University, 613 West Huangpu Avenue, Guangzhou 510630, China
4Department of Neurology, The First Affiliated Hospital of Jinan University, 613 West Huangpu Avenue, Guangzhou 510630, China
OBJECTIVES The risk of cardiovascular disease (CVD) in cancer survivors is an important area of concern, and there is a lack of relevant prospective studies. A reliable evaluation of CVD risk for sitespecific cancers is therefore crucial to guide the treatment and improve the prognosis of survivors with various site-specific cancers. This study aimed to analyze the risk of a wide range of specific CVDs in survivors with site-specific cancers compared with cancer-free controls.
METHODS Cancer diagnosis was determined by linked data from the Medical Research Information Service and The Information Services Division, while CVD diagnosis was ascertained via record linkage from health episode statistics records and Scottish morbidity records. We used Cox proportional hazards regression to assess the association between cancer sites and CVD outcomes after adjustment for age, sex, Townsend deprivation index, ethnicity, college degree, drinking status, smoking status, physical activity, healthy diet, family history (stroke, heart diseases), and 11 comorbidities (type 2 diabetes, lipidemia, hypertension, chronic obstructive pulmonary disease, chronic kidney disease, thyroid gland disorders, obesity, depression, peptic ulcer, rheumatoid arthritis, gout).
RESULTS Of the 404,727 participants, 62,009 (15.3%) survivors with a diagnosis of cancer and 342,718 (84.7%) controls were followed up for a median of 12.7 years. Risk of pulmonary circulation disease was elevated in survivors with any cancer sites compared with cancer-free controls, especially in thoracic cancer survivors [hazard ratio: 12.89, 95% confidence interval: 11.13–14.94]. Both heart failure and atrial fibrillation were at increased risks at 6 cancer sites, as were acute myocardial infarction and angina at 3 cancer sites. Survivors with digestive, thoracic or hematological cancers had elevated risks of all CVD outcomes.
CONCLUSIONS With risk patterns ranging by cancer sites and particular CVD outcomes, survivors of most site-specific cancers had greater risks of specific CVD than those who had not been diagnosed with cancer, especially digestive, thoracic and hematological cancers. For the expanding population of cancer survivors, strategies to monitor and control cardiovascular risk are needed.
GW34-e0269
Yingjian Deng, Qiang Li, Faguang Zhou, Dong Chang
Xiamen Cardiovascular Hospital of Xiamen University
OBJECTIVES Observational studies suggested that physical activity (PA) and sedentary behavior are associated with the risk of atrial fibrillation (AF). Thus, we performed this Mendelian randomization (MR) study to assess the causal effects of PA and sedentary behavior on the risk of AF.
METHODS A two-sample MR analysis was performed to investigate the associations between PA and sedentary behavior and AF. The summary statistics for PA and sedentary behavior were obtained from different genome-wide association studies (GWASs), and cohorts of vigorous PA (n=251,501), moderate PA (n=343,827), Light PA (n=64,949), television watching (n=437,887), driving (n=310,555), mobile phone use (n=456,972), computer use (n=261,987), and playing computer games (n=462,433) were obtained. Summary data regarding AF were gathered from a meta-analysis on GWASs involving a total of 1,030,836 individuals.
RESULTS The results suggested that vigorous PA (OR 1.64, 95% CI [1.13, 2.40], P=0.01), driving (OR 1.92, 95% CI [1.12, 3.29], P=0.02), mobile phone use (OR 1.21, 95% CI [1.04, 1.40], P=0.01), and playing computer games (OR 1.61, 95% CI [1.16, 2.24], P<0.01) were associated with an increased risk of AF, while no causal associations between other PA or sedentary behavior and AF were found.
CONCLUSIONS The present study demonstrated that vigorous PA, driving, mobile phone use, and playing computer games were associated with an increased risk of AF. The relationship between moderate PA, Light PA, television watching, and computer use with AF needs further investigation in future studies.
GW34-e0270
Xiong Gao, Qiuxia Zhang, Miaomiao Yang, Wei Luo, Yuegang Wang, Jianchen Xiu
Nanfang Hospital of Southern Medical University
OBJECTIVES Observational studies have shown that atrial fibrillation (AF) can lead to cognitive impairment ranging from mild cognitive dysfunction to dementia. Whether this association reflects a causal relationship is still unclear. This study aims to explore the causal relationship between AF and cognitive impairment.
METHODS A two-sample Mendelian randomization (TSMR) analysis is used to assess the potential causality of AF on cognitive dysfunction. Single nucleotide polymorphisms (SNPs) strongly associated with atrial fibrillation were extracted as instrumental variables using a large genome-wide association study (GWAS) pooled dataset of atrial fibrillation (AF). The association of SNPs with Alzheimer’s disease dementia, Parkinson’s disease dementia, vascular dementia, Lewy body dementia, frontotemporal dementia, undefined dementia, and overall cognitive function assessment was then extracted separately from publicly available GWAS data on cognitive dysfunction. The inverse variance-weighted method (IVW) was used for the main analysis, and sensitivity analyses were conducted using Cochran’s Q test, MR-Egger regression, and leave-one-out method. To verify the robustness of the results, replicate analyses and meta-analyses were performed using different GWAS data.
RESULTS In the initial analysis, 101 SNPs were extracted as instrumental variables from a recent meta-analysis of a genome-wide association study involving up to 1,030,836 individuals. The IVW analysis showed no evidence for causal associations between AF and dementia [Dementia (OR, 1.03; 95% CI, 0.97–1.09; P=0.29), Parkinson’s disease dementia (OR, 1.00; 95% CI, 0.78–1.29; P=0.97), vascular dementia (OR, 1.12; 95% CI, 0.96–1.30; P=0.12), or unspecified dementia (OR, 1.01; 95% CI, 0.91–1.12; P=0.80)]. In the replication analysis, 27 SNPs were extracted as instrumental variables from the FinnGen alliance’s AF GWAS data, the IVW analysis are consistent with the initial analysis [Cognitive function (OR, 0.99; 95% CI, 0.98–1.01; P=0.87), Alzheimer’s disease dementia (OR, 0.97; 95% CI, 0.94–1.01; P=0.19), Lewy body dementia (OR, 1.01; 95% CI, 0.89–1.14; P=0.82), or frontotemporal dementia (OR, 0.99; 95% CI, 0.74–1.33; P=0.98)]. Both Mendelian randomization (MR) analyses and meta-analyses showed no evidence of association between genetically predicted AF and different types of dementia or overall cognitive function assessment. MR-Egger regression suggested no horizontal pleiotropy, and leave-one-out analysis showed stable results after individually removing each SNP.
CONCLUSIONS No evidence of a causal relationship between AF and cognitive impairment was found. The associations observed in observational studies can be partially attributed to confounding factors such as shared biology or co-morbidities.
GW34-e0275
Hui Zhang1, Qi Guo1,2
1Jiangwan Hospital of Shanghai Hongkou District
2Shanghai University of Medicine and Health Sciences
OBJECTIVES Cardiovascular disease (CVD) is the leading cause of mortality in patients with end-stage renal disease on hemodialysis (HD), and Ideal cardiovascular health (CVH) metrics is commonly used to assess cardiovascular health status. Epidemiological studies have found that sleep complaints such as obstructive sleep apnea (OSA) and abnormal sleep duration, which are common in HD patients, may increase the risk of cardiovascular disease. Therefore, the purpose of this study was to explore the separated and combined associations of OSA risk and sleep duration with ideal cardiovascular health metrics in HD patients.
METHODS We recruited 880 patients over 18 years old who underwent hemodialysis at least twice a week and obtained informed consent from all participants. After excluding participants with a history of cardiovascular disease and taking sleep medications, 470 HD participants (average: 59.48±12.89 years, 281 men) were included in this study. Sleep duration was measured as self-reported average sleep time during the previous month. OSA risk was evaluated by using STOP-BANG questionnaire. Ideal CVH metrics was defined by the American Heart Association, including 7 modifiable health behaviors and factors, and participants were divided the into three groups by the number of ideal CVH metrics: 0–2 (poor), 3–4 (intermediate), and 5–7 (ideal). Ordinal logistic regression was performed to model the associations of CVH metrics with sleep duration, OSA risk, and their combined effects by adjusting for certain covariates.
RESULTS Compared with patients with ideal CVH, poor and intermediate CVH individuals had poorer CVH indicators including BMI, smoking, physical activity, salty diet, glucose, blood lipids and blood pressure. HD patients with poor and moderate CVH had higher age, spkt/v and potassium, they were also more likely to be male, drinker, fluid overload, shorter sleepers and at higher OSA risk. After adjusting for age, sex, job category, drinker, IPAQ, fluid overload, spkt/v, HDL, and potassium, short sleep duration (<7 h) (OR=0.53; 95% CI [0.30, 0.92]) and OSA risk (OR=0.58; 95% CI [0.32, 0.83]) were negative associated with better CVH (ideal vs. intermediate; intermediate vs. poor), respectively. For HD patients with both short sleep duration (<7 h) and OSA risk, the proportional odds of having better CVH were 72% lower (OR=0.28; [95% CI 0.13, 0.60]).
CONCLUSIONS Short sleep duration (<7 h) and OSA risk were negative associated independently with ideal CVH metrics in HD patients, the combination of short sleep duration (<7 h) and OSA risk significantly further reduced the likelihood of ideal CVH metrics. This study supports population-based prevention programmes in HD patients, improving sleep to minimize the risk of subsequent cardiovascular disease.
GW34-e0283
Zhiyuan Wu1,2, Jinqi Wang1, Haiping Zhang1, Lixin Tao1, Xiuhua Guo1
1Capital Medical University
2Edith Cowan University
OBJECTIVES Arteriosclerosis and atherosclerosis are closely related with cardiovascular disease (CVD) risk. Remnant cholesterol (RC) could predict CVD. However, its effect on joint arteriosclerosis and atherosclerosis progression remains unclear. This study aims to evaluate the association of RC with joint arteriosclerosis and atherosclerosis progression trajectories in the general population.
METHODS This study collected data across five biennial surveys of the Beijing Health Management Cohort from 2010 to 2019. Multi-trajectory model was used to determine the joint arteriosclerosis and atherosclerosis progression patterns by brachial-ankle pulse wave velocity (baPWV) and ankle brachial index (ABI). We also performed discordance analyses for RC vs. low density lipoprotein cholesterol (LDL-C) using ordinal logistics model.
RESULTS A total of 3186 participants were included, with three clusters following distinct arteriosclerosis and atherosclerosis progression patterns identified using a multi-trajectory model. In the multivariable-adjusted ordinal logistics analyses, RC was significantly associated with baPWV and ABI progression (OR: 1.20; 95% CI: 1.13–1.28, per 10 mg/dL). For the discordance analyses, the discordant low RC group was associated with decreased risk compared to the concordant group (OR: 0.73; 95% CI: 0.60–0.89). People with a high RC level were at an increased risk of joint arteriosclerosis and atherosclerosis progression, even with optimal LDL-C.
CONCLUSIONS RC is independently associated with joint arteriosclerosis and atherosclerosis progression beyond LDL-C. RC could be an earlier risk factor than LDL-C of arteriosclerosis and atherosclerosis in the general population.
GW34-e0382
Juanjuan Zheng, Shuangshuang Song, Jiamin Zou, Cheng Liang, Wenyou Wang, Yangyang Zhou, Yuming Zhang, Hong Peng, Wei Eric Wang
Department of Geriatrics, Southwest Hospital, Army Military Medical University
OBJECTIVES Omicron virus infection has been spreading rapidly in China since December 2022, and a part of the infected patients are complicated with myocardial injury. Myocardial injury affects the quality of life and prognosis of patients, and is an important factor in the occurrence of “Long-COVID”. However, the risk factors of myocardial injury caused by Omicron virus infection are still unclear. To explore the risk factors of patients with Omicron virus infection complicated with myocardial injury, thereby providing scientific basis to prevent and treat myocardial injury in clinic.
METHODS Demographic information, clinical signs and symptoms, laboratory examination and other medical records of 691 hospitalized patients with Omicron virus infection in Southwest Hospital from December 1, 2022 to March 3, 2023 were retrospectively collected. Myocardial injury was defined according to whether the hypersensitive troponin was elevated in the first serum examination. The risk factors of myocardial injury were analyzed by multivariate Logistic regression model.
RESULTS Among the 691 Omicron virus infected patients who were tested for hypersensitive troponin, the average age was 50 (40, 59) years, males were 418 (60.49%), and 235 (34.01%) patients were with myocardial injury. Compared with control group, myocardial injury group had a higher median age (53 (45, 63) vs 48 (38, 57), P<0.05); Among them, 160 (68.09%) were males. There were statistically significant differences in admission temperature, maximum body temperature during hospitalization, symptoms on admission (fever, fatigue, poor appetite, cough), cardiovascular diseases (hypertension, diabetes, coronary heart disease), COPD, blood gas analysis, left ventricular ejection fraction (LVEF) and other clinical indicators between the two groups (all P<0.05). There were no statistically significant differences in BMI, vaccination or palpitation between the two groups (P>0.05). NT-proBNP, blood urea, creatinine and glomerular filtration rate were strongly correlated with the increase of troponin (r>0.5, all P<0.001). Multivariate Logistic regression analysis showed that age (OR=1.056, 95% CI=1.040–1.072), male (OR=1.738, 95% CI=1.136–2.658, P<0.05), renal insufficiency (OR=2.318, 95% CI=1.318∼4.076, P<0.05), renal dialysis history (OR=6.566, 95% CI=2.954–14.594, P<0.001), glomerular filtration rate decreased (OR=0.984, 95% CI=0.970∼0.998, P<0.05), anemia (OR=2.512, 95% CI=1.354–4.662, P<0.05), platelet level decreased (OR=0.990, 95% CI=0.982∼0.998, P<0.05) and ST-T changes in ECG (OR=2.308, 95% CI=1.224∼3.395, P<0.05) were independent risk factors for myocardial injury of patients with Omicron virus infection.
CONCLUSIONS Age, male, renal insufficiency, renal dialysis history, decreased glomerular filtration rate, NT-proBNP, anemia, decreased platelet level and ST-T changes in ECG are independent risk factors for Omicron virus infection caused myocardial injury.
GW34-e0555
Zheming Yang1,2, Xiaodong Jia2, Jiayin Li1,2, Zhu Mei1,2, Chenghui Yan2, Yaling Han1,2
1College of Medicine and Biological Information Engineering, Northeastern University, Shenyang, Liaoning 110167, China
2National Key Laboratory of Frigid Zone Cardiovascular Disease, Cardiovascular Research Institute and Department of Cardiology, General Hospital of Northern Theater Command, Shenyang 110016, China
OBJECTIVES Cardiovascular disease (CVD) is a serious condition that poses threats to patients’ quality of life and life expectancy. Cardiac rehabilitation is a crucial treatment option that can improve outcomes for CVD patients. Hybrid comprehensive telerehabilitation (HCTR) is a relatively new approach. In the context of pandemics, HCTR can minimize the risk of cluster infections by reducing hospital visits, while delivering effective rehabilitation care. This study aimed to assess the efficacy and safety of HCTR as a secondary prevention measure for CVD patients compared to usual rehabilitation care.
METHODS We searched Pubmed, Embase, The web of science, The Cochrane Library, and PsychINFO for all related studies up to January 20, 2023. Two reviewers independently screened the titles and abstracts of potentially eligible articles based on the predefined search criteria. Data were analyzed using Comprehensive Meta-Analysis software (RevMan5.3).
RESULTS Eight trials, involving 1578 participants, were included. HCTR and usual rehabilitation care provide similar effects on readmission rates (odds ratio [OR]=0.90 [95% CI 0.69–1.17], P=0.43) and mortality (odds ratio [OR] = 1.06 [95% CI 0.72–1.57], P=0.76). Effects on Short Form-36 Health Status Questionnaire (SF-36) score were also similar (SMD: 1.32 [95% CI −0.48–3.11], P=0.15). Compared with usual rehabilitation care, HCTR can improve peak oxygen uptake (VO2 peak) (SMD: 0.99 [95% CI 0.23–1.74], P=0.01) and 6-minute walking test (6MWT) (SMD: 10.02 [95% CI 5.44–14.60], P<0.001) of patients.
CONCLUSIONS Our findings indicate that HCTR is as effective as traditional rehabilitation care in reducing readmission rates, mortality, and improving quality of life in patients with CVD. However, HCTR offers the added advantage of improving VO2 peak and 6MWT, measurements of cardiorespiratory fitness and functional capacity, respectively. These results suggest that HCTR can be a safe and effective alternative to traditional rehabilitation care, offering numerous benefits for CVD patients.
GW34-e0824
Hualin Wang, Bowen Du, Zhuoyan Li, Yujian Wu, Qianchuo Wang, Jian Wang, Kun Sun
Department of Pediatric Cardiology, Xinhua Hospital, School of Medicine, Shanghai Jiao Tong University
OBJECTIVES Studies have reported that prenatal exposure to air pollution is associated with elevated blood pressure and impaired cardiometabolic health in children. However, the effects of prenatal air pollution exposure on children’s cardiac structure and function, as well as the susceptible windows for these effects, remain unknown. The objective of this study is to investigate the association between prenatal air pollution exposure with cardiac structure and function in preschool-aged children.
METHODS A total of 1270 mother-offspring pairs from the Shanghai Birth Cohort study were included. Anthropometric assessment and detailed echocardiography examination were conducted in these children at 4 years old. Weekly averaged concentrations of air pollutants including PM2.5, PM10, SO2, NO2, CO and ozone over gestation were assessed based on data from the National Urban Air Quality Real-Time Publishing Platform. Distributed lag non-linear models (DLNMs) were fitted by incorporating multiple linear models and logistic regression models to evaluate weekly exposure-lag-response associations between prenatal air pollution levels and cardiac structure and function, as well as left ventricular hypertrophy (LVH).
RESULTS In this study, we found that higher levels of prenatal exposure to PM2.5, PM10, SO2, and carbon monoxide were associated with increased cardiac wall thickness and decreased cardiac diameter in 4-year-old children. These associations were particularly pronounced during early and mid-pregnancy, with no apparent threshold observed in the concentration-response curve for these pollutants. Furthermore, prenatal exposure to PM2.5, PM10, SO2, NO2, and carbon monoxide during early pregnancy was associated with a higher risk of LVH, with critical windows identified as 1st–12th weeks, 2nd–11th weeks, 3rd–14th weeks, 4th–11th weeks and 1st–15th weeks of gestation, respectively. The strongest effect of each 10 μg/m3 per increment in prenatal exposure to PM2.5, PM10, SO2, NO2 on the risk of LVH displayed in the 1st week (HR:1.10, 95% CI: 1.03–1.17), 8th week (HR:1.07, 95% CI: 1.04–1.10), 7th week (HR:1.21, 95% CI: 1.14–1.29), and 9th week (HR:1.14, 95% CI: 1.05–1.25), respectively. And each 50 μg/m3 increase in exposure to carbon monoxide showed strongest effect on the risk of LVH in 8th–9th weeks (HR:1.07, 95% CI: 1.04–1.09).
CONCLUSIONS Our findings suggest that prenatal exposure to air pollution is associated with cardiac structural changes in preschool-aged children and exposure during early pregnancy is associated with a higher risk of LVH.
GW34-e0955
Chaoqun Ma, Dingyuan Tu
National Key Laboratory of Frigid Zone Cardiovascular Diseases (NKLFZCD), Cardiovascular Research Institute and Department of Cardiology, General Hospital of Northern Theater Command, Shenyang, Liaoning 110000, P.R. China
OBJECTIVES Emerging evidence have reported that lifestyle behaviors play critical roles in the pathogenesis of various cardiovascular disorders. The joint associations of anti-inflammatory diet and vigorous leisure-time physical activity (LTPA) with all-cause mortality and cardiovascular disease (CVD) mortality are incompletely understood.
METHODS This secondary data analysis included 16,068 adults from the National Health and Nutrition Examination Survey (2007–2014). Mortality outcomes were ascertained by linkage to National Death Index records through December 31, 2019. Participants were categorized into four lifestyle patterns based on the inflammatory properties of the diet and the degree of vigorous LTPA: pro-inflammatory diet and insufficient vigorous LTPA, anti-inflammatory diet and insufficient vigorous LTPA, pro-inflammatory diet and sufficient vigorous LTPA, anti-inflammatory diet and sufficient vigorous LTPA. Multivariable Cox proportional hazards models were used to estimate the hazards ratio (HR) and 95% confidence intervals (CI) for all-cause and CVD mortality. Two-sided P value <0.05 was considered statistically significant.
RESULTS During a median follow-up of 8.5 years, 1904 overall deaths and 484 confirmed CVD deaths were reported. Compared with the pro-inflammatory diet and insufficient vigorous LTPA group (reference), those in the anti-inflammatory diet and sufficient vigorous LTPA group had a significantly lower risk of all-cause (HR, 0.51; 95% CI, 0.32–0.81; P=0.004) and CVD (HR, 0.31; 95% CI, 0.12–0.80; P=0.016) mortality. Similarly, the lifestyle pattern of anti-inflammatory diet and insufficient vigorous LTPA also decreased all-cause (HR, 0.80; 95% CI, 0.69–0.92; P=0.002) and CVD (HR, 0.71; 95% CI, 0.53–0.95; P=0.020) mortality risk. In contrast, all-cause mortality was not significantly reduced in pro-inflammatory diet and sufficient vigorous LTPA group (HR: 0.75, 95% CI: 0.54–1.06, P=0.101), as well as CVD mortality (HR: 0.60, 95% CI: 0.32–1.13, P=0.114). Consistent results were obtained in subgroup and sensitivity analyses.
CONCLUSIONS Adhering to the anti-inflammatory diet and sufficient vigorous LTPA was associated with lower all-cause and CVD mortality. Furthermore, an anti-inflammatory diet can be instrumental in counteracting the harms of insufficient vigorous LTPA, while sufficient vigorous LTPA fails to offset the detrimental effect of pro-inflammatory diet.
GW34-e0991
Hailin Du1, Hongtu Qiao1, Chunxian Lin1, Bin Xue1, Tao Chen1, Shengping Wang1, Wenyong Zhang1,2
1Chengdu Qingbaijiang District People’s Hospital
2Chengdu Second People’s Hospital
OBJECTIVES The aging population, high-altitude terrain, and unique population characteristics in the Garze region have led to a significantly higher prevalence of hypertension compared to other areas. Previous studies have shown that the prevalence of hypertension in the Garze region is 22.9%, higher than 17.9% in Tibetan farming and pastoral areas, and 11.64% in the Han ethnic population. Moreover, the Garze region exhibits characteristics such as low awareness, low control rate, low treatment rate, poor medication adherence, and high morbidity and mortality rates for hypertension. This study aimed to investigate the current status of blood pressure control (achievement rate and non-achievement rate) and medication adherence among hypertensive patients in the multi-ethnic region of Garze, Sichuan, and analyze the main influencing factors among hypertensive patients of different ethnicities. The findings aim to provide data support and theoretical basis for developing intervention measures for hypertensive patients in the multi-ethnic region of Garze.
METHODS A multi-stage stratified cluster random sampling method was employed in this study. Based on the latest population census data in Garze in 2020, 2100 hypertensive patients from Kangding, Jiulong, Dege, and Batang counties were selected for questionnaire surveys, and 2009 valid responses were collected, resulting in a response rate of 95.6%. According to whether blood pressure met the standard (blood pressure standard: systolic pressure <140 mmHg, diastolic pressure <90 mmHg), the patients were divided into an achievement group (n=1033) and a non-achievement group (n=976). The gender, age, ethnicity, education level, marital status, medical history (diabetes, stroke, chronic obstructive pulmonary disease), lifestyle habits (drinking, smoking), BMI index, and medication adherence were analyzed and compared between the two groups. Multivariate logistic regression analysis was conducted to identify potential risk factors.
RESULTS The overall achievement rate of blood pressure control among hypertensive patients was 51.42%. There were no statistically significant differences between the two groups in terms of ethnicity, gender, education level, stroke, or chronic obstructive pulmonary disease (P>0.05). However, there were significant differences (P<0.05) in terms of age, diabetes, BMI index, drinking, smoking, and medication adherence. Regression analysis indicated that age, diabetes, drinking, smoking, BMI index, and medication adherence were independent risk factors for uncontrolled blood pressure in hypertensive patients. Receiver operating characteristic (ROC) analysis showed that age, BMI index, diabetes, and smoking had significantly larger areas under the curve in predicting uncontrolled blood pressure compared to drinking and medication adherence, with statistical significance (P<0.05).
CONCLUSIONS When patients with hypertension have comorbidities such as diabetes, an increased BMI index, older age, smoking, and non-adherence to medication, the risk of inadequate blood pressure control increases. Clinical doctors and public health workers should intervene early for the above-mentioned risk factors to improve the rate of achieving blood pressure control targets in patients.
GW34-e1048
Xin Yin, Yu-Tao Li, Ying Pan, Ting-Ting Wu, Yi-Tong Ma, Xiang Xie
Xinjiang Medical University
OBJECTIVES To further refine the association between sleep duration with all-cause mortality (ACM), cardiovascular events, mortality and stroke, and make it possible to personalize risk prediction.
METHODS A systematic review and meta-analysis of prospective cohort studies that estimate the relation between sleep duration and adverse outcomes was conducted. We searched PubMed, Embase, Web of Science, and Cochrane Library databases for these studies, and extracted data from identified studies up to 1 January 2023. Prospective cohort study with sleep duration as exposure and presenting clear data on adverse outcomes were included in; We took 7–7.9 hours as reference when calculating effect size.
RESULTS A total of 110 research projects involving about 2.28 million people were evaluated. Consistent with previous studies, the risk was lowest at 7–8 hours of sleep. But too much sleep is more harmful than sleep deprivation, showing a “J” shape rather than a “U” shape between the risk and sleep duration. When less than 5 hours or more than 9 hours, the magnitude of the increased risk changed significantly. When sleep duration =6 h, the risk in people aged 40–49.9 years (123% risk for ACM and 125.1% risk for CVD Mortality) was much higher than people aged over 50 years. When sleep duration≥10 h, the risk in people aged 50–54.9 years (153% risk for ACM and 235.6% risk for CVD Mortality) was higher than others. When sleep duration =5 hours and≥9 hours, people aged 40 to 49.9 years unexpectedly had the highest risk of stroke (141.2% at 5 h and 181.1% at 9 h). As the duration of sleep increased, the risk was much higher in women (Sleep duration≥10 h, 178% risk for ACM and 188% risk for CVD Mortality) than in men (Sleep duration≥10 h, 150% risk for ACM and 158% risk for CVD Mortality). We find that when sleep duration was more than 7 hours, the risk of yellow people (Sleep duration≥10 h, 164% risk for ACM and 171% risk for CVD Mortality) was significantly higher than that of white people (Sleep duration≥10 h, 128% risk for ACM and 135.8% risk for CVD Mortality). However, when sleep duration =5 hours and≥9 hours, white people have a higher risk of Stroke (138.7% at 5 h and 146.4% at 9 h) than yellow people (113.4% at 5 h and 131.4% at 9 h). The results of studies with more than 10 years of follow-up were more “conservative” than those with less than 10 years of follow-up. In terms of numbers, the results for the 10,000-plus cohort were more “conservative”. We recommend that even if staying up late is inevitable, you should also ensure at least 5 hours of sleep, and do not sleep in more than 9 hours, especially for yellow women.
CONCLUSIONS More attention should be paid to sleep in middle-aged people. People aged 40–49.9 years, during extreme sleep, has the highest risk of stroke. People aged 50–54.9 years also suffer the highest risk of all-cause and cardiovascular mortality when they experience extreme sleep.
GW34-e1070
Binglu Wang1, Jun Wang2
1Xiang Ya Nursing School of Central South University, Changsha, Hunan 410013, China
2China CDC Key Laboratory of Environment and Population Health, National Institute of Environmental Health, Chinese Center for Disease Control and Prevention, Beijing 100021, China
OBJECTIVES The impact of adherence to plant-based dietary patterns on life expectancy among older adults remains unclear. We aimed to quantify the associations of plant-based dietary patterns with life expectancy among Chinese older adults.
METHODS We used data from Chinese Longitudinal Healthy Longevity Survey (CLHLS, 2008–2018) of 15,706 adults aged 65 years and older (57.48% female). The overall plant-based diet index (PDI), healthful plant-based diet index (hPDI) and unhealthful plant-based diet index (uPDI) were assessed using dietary data collected by a simplified and validated food frequency questionnaire (FFQ), which consists of 16 food components: whole grains, vegetable oils, fresh fruit, fresh vegetables, legumes, garlic, nuts, tea, refined grains, preserved vegetables, sugar, meat, fish and aquatic products, eggs, milk or dairy products, and animal fat. Hazard ratios (HRs) of all-cause mortality and the life expectancy by levels of three versions of plant-based diet indices (overall, healthful, and unhealthful) were estimated.
RESULTS During a follow-up of 69,700 person-years, 10,573 deaths were recorded. Compared with the lowest tertile, participants in the highest tertile of PDI and hPDI had lower risk of all-cause mortality (HR=0.92; 95% CI 0.87–0.96 for PDI; HR=0.93, 95% CI, 0.88–0.97 for hPDI), whereas participants with the highest uPDI scores had a 11% (HR=1.11, 95% CI 1.06–1.17) increased mortality risk. We observed gradual gains in life expectancy toward PDI or hPDI dietary patterns, such as, compared with lowest tertile of PDI/hPDI, participants with highest tertile of PDI and hPDI had an average 0.82 (95% CI 0.43–1.21) and 0.68 (95% CI 0.28–1.01) more years of life expectancy at age 65 years, respectively. Compared with lowest tertile of uPDI, those with highest uPDI lost 1.16 (95% CI 0.81–1.50) years in life expectancy. Several sensitivity analyses suggested these results were relatively stable.
CONCLUSIONS Adopting to a high level of plant-based dietary pattern, especially for healthful plant-based dietary pattern, was related to lower risk of all-cause mortality and longer life expectancy, supporting current recommendations to increase intake of healthy plant foods, and reducing intake of unhealthy plant foods for prevention of all-cause mortality.
GW34-e1074
Binglu Wang1, Jun Wang2
1Xiang Ya Nursing School of Central South University, Changsha, Hunan 410013, China
2China CDC Key Laboratory of Environment and Population Health, National Institute of Environmental Health, Chinese Center for Disease Control and Prevention, Beijing 100021, China
OBJECTIVES The effect of plant-based dietary patterns on annual change in cognitive performance is uncertain among Chinese older adults. This study examined the association between plant-based dietary patterns and annual change in cognitive performance.
METHODS Data were from Chinese Longitudinal Healthy Longevity Survey (CLHLS, 2008–2018), a population based, prospective cohort study. Individuals aged 65 years or older who had normal cognition at baseline were included, and followed up until death, discontinuation, or December 2018. A simplified and validated food frequency questionnaire (FFQ) was used to assess the dietary information. Three versions of graded plant-based diets were constructed: an overall plant-based diet index (PDI), healthful plant-based diet index (hPDI), and unhealthful plant-based diet index (uPDI). Cognitive performance was measured via the Chinese version of Mini-Mental State Examination. Multivariable generalized linear regression models with repeated measures analyses were used to examine the associations of three versions of plant-base diets with cognitive performance annual change.
RESULTS 8865 participants were included (mean age of 82.63 years), 53.38% were women. Higher healthful-PDI was associated with slower cognitive decline. The annual changes in cognitive performance were −0.67 (95% CI: −0.74, −0.60), −0.61 (95% CI: −0.67, −0.55), and −0.57 (95% CI: −0.63, −0.51) across tertiles of overall PDI (Ptrend=0.0313); −0.71 (95% CI: −0.79, −0.64), −0.60 (95% CI: −0.66, −0.54), and −0.54 (95% CI: −0.61, −0.48) across tertiles of healthful PDI (Ptrend=0.0003). The uPDI was positively associated with cognitive decline, with −0.53 (95% CI: −0.59, −0.47), −0.65 (95% CI: −0.72, −0.58), and −0.68 (95% CI: −0.75, −0.62) across tertiles of unhealthful PDI (Ptrend=0.0022). For sub-domains of cognitive performance, similar results were observed for the following two dimensions: orientation in time and space, and calculation.
CONCLUSIONS Individuals with higher healthful PDI were associated with slower cognitive decline. This study might offer important information to protect older adults against cognitive decline.