Expression of MicroRNA-1 and MicroRNA-21 in Different Protocols of Ischemic Conditioning in an Isolated Rat Heart Model

Abstract

Background: ‘Conditioning’ [ischemic preconditioning (IPC), ischemic postconditioning (IPO) and remote ischemic preconditioning (RIPC)] the heart to render it more resistant to an episode of acute myocardial ischemia-reperfusion (I/R) injury is an endogenous cardioprotective strategy. There are several mechanisms proposed for ‘conditioning’, such as endogenous mediators or cytoprotective proteins. In recent reports, microRNAs (miRNAs) were involved in controlling the expression of myocardial ischemia-related genes. Some studies have demonstrated that cardiac miRNA-1 and miRNA-21 were significantly increased by late IPC with an increase in their target proteins [endothelial nitric oxide synthase and heat shock protein 70 (HSP70)], but their expression levels in ‘conditioning’ strategies are currently unknown. Methods: In the current study, Langendorff-perfused Sprague-Dawley rat hearts were randomly assigned to one of four groups [control group (CON group, n = 12), IPC group (n = 12), IPO group (n = 12) and RIPC group (n = 12)]. Cardiac function was digitalized and analyzed. The expression of miRNA-1 and miRNA-21 was detected by real-time reverse transcription polymerase chain reaction. The expression of HSP70, programmed cell death protein 4 (PDCD4), B-cell lymphoma/leukemia-2 (Bcl-2) and Bcl-2-associated X protein (Bax) was detected by Western blot. Cardiac infarct size and myocardial apoptosis were determined using the 2,3,5-triphenyltetrazolium chloride assay and terminal deoxynucleotidyl transferase dUTP nick end labeling assay, respectively. Results: The results revealed that miRNA-1 (233 ± 45%) and miRNA-21 (356 ± 33%) expression was up-regulated in the IPC group, but the expression of miRNA-1 was down-regulated in the RIPC (61 ± 16%) group and IPO group (61 ± 13%). The expression of PDCD4 [IPC (74 ± 11%), RIPC (81 ± 16%), IPO (83 ± 12%)], HSP70 [IPC (74 ± 5%), RIPC (81 ± 6%), IPO (67 ± 11%)] and Bax [IPC (27 ± 6%), RIPC (21 ± 3%), IPO (27 ± 4%)] was down-regulated in the conditioning groups compared with the CON group [PDCD4 (130 ± 11%), HSP70 (121 ± 11%) and Bax (63 ± 8%)]. In the conditioning hearts, infarct size [IPC (31.7 ± 4.1%), RIPC (29.6 ± 6.19%) and IPO (32.8 ± 4.71%)] and myocardial apoptosis [IPC (15.2 ± 4.21%), RIPC (17.2 ± 1.92%) and IPO (15.6 ± 4.04%)] were significantly decreased compared with the CON group (infarct size: 51.77 ± 4.3%, myocardial apoptosis: 32.8 ± 3.96%). Conclusion: We concluded that miRNA-1 and miRNA-21 expression differed in IPC, RIPC and IPO groups, and their target proteins were not inversely correlated with the miRNAs in all the conditioning groups, which revealed that the miRNAs were regulated but complicated by the different conditioning protocols.