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      Cathepsin B-Mediated Activation of Trypsinogen in Endocytosing Macrophages Increases Severity of Pancreatitis in Mice


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          BACKGROUND & AIMS:

          Acute pancreatitis is characterized by premature intracellular activation of digestive proteases within pancreatic acini and a consecutive systemic inflammatory response. We investigated how these processes interact during severe pancreatitis in mice.


          Pancreatitis was induced in C57Bl/6 wild-type (control), cathepsin B (CTSB)- knockout, and cathepsin L-knockout mice by partial pancreatic duct ligation with supramaximal caerulein injection, or by repetitive supramaximal caerulein injections alone. Immune cells that infiltrated the pancreas were characterized by immunofluorescence detection of Ly6g, CD206, and CD68. Macrophages were isolated from bone marrow and incubated with bovine trypsinogen or isolated acinar cells; the macrophages were then transferred into pancreatitis control or cathepsin-knockout mice. Activities of proteases and nuclear factor (NF)- κB were determined using fluorogenic substrates and trypsin activity was blocked by nafamostat. Cytokine levels were measured using a cytometric bead array. We performed immunohistochemical analyses to detect trypsinogen, CD206, and CD68 in human chronic pancreatitis (n = 13) and acute necrotizing pancreatitis (n = 15) specimens.


          Macrophages were the predominant immune cell population that migrated into the pancreas during induction of pancreatitis in control mice. CD68-positive macrophages were found to phagocytose acinar cell components, including zymogen-containing vesicles, in pancreata from mice with pancreatitis, as well as human necrotic pancreatic tissues. Trypsinogen became activated in macrophages cultured with purified trypsinogen or co-cultured with pancreatic acini and in pancreata of mice with pancreatitis; trypsinogen activation required macrophage endocytosis and expression and activity of CTSB, and was sensitive to pH. Activation of trypsinogen in macrophages resulted in translocation of NF-kB and production of inflammatory cytokines; mice without trypsinogen activation (CTSB-knockout mice) in macrophages developed less severe pancreatitis compared with control mice. Transfer of macrophage from control mice to CTSB-knockout mice increased the severity of pancreatitis. Inhibition of trypsin activity in macrophages prevented translocation of NF- κB and production of inflammatory cytokines.


          Studying pancreatitis in mice, we found activation of digestive proteases to occur not only in acinar cells but also in macrophages that infiltrate pancreatic tissue. Activation of the proteases in macrophage occurs during endocytosis of zymogen-containing vesicles, and depends on pH and CTSB. This process involves macrophage activation via NF- κB-translocation, and contributes to systemic inflammation and severity of pancreatitis.

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          Most cited references43

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          Lactate reduces liver and pancreatic injury in Toll-like receptor- and inflammasome-mediated inflammation via GPR81-mediated suppression of innate immunity.

          The NACHT, LRR, and pyrin domain-containing protein 3 (NLRP3) inflammasome induces inflammation in response to organ injury, but little is known about its regulation. Toll-like receptors (TLRs) provide the first signal required for activation of the inflammasome and stimulate aerobic glycolysis to generate lactate. We examined whether lactate and the lactate receptor, Gi-protein-coupled receptor 81 (GPR81), regulate TLR induction of signal 1 and limit inflammasome activation and organ injury.
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            Anti-inflammatory lipid mediators and insights into the resolution of inflammation.

            The pro-inflammatory signalling pathways and cellular mechanisms that initiate the inflammatory response have become increasingly well characterized. However, little is known about the mediators and mechanisms that switch off inflammation. Recent data indicate that the resolution of inflammation is an active process controlled by endogenous mediators that suppress pro-inflammatory gene expression and cell trafficking, as well as induce inflammatory-cell apoptosis and phagocytosis, which are crucial determinants of successful resolution. This review focuses on this emerging area of inflammation research and describes the mediators and mechanisms that are currently stealing the headlines.
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              Priming of the neutrophil respiratory burst: role in host defense and inflammation.

              Neutrophils are the major circulating white blood cells in humans. They play an essential role in host defense against pathogens. In healthy individuals, circulating neutrophils are in a dormant state with very low efficiency of capture and arrest on the quiescent endothelium. Upon infection and subsequent release of pro-inflammatory mediators, the vascular endothelium signals to circulating neutrophils to roll, adhere, and cross the endothelial barrier. Neutrophils migrate toward the infection site along a gradient of chemo-attractants, then recognize and engulf the pathogen. To kill this pathogen entrapped inside the vacuole, neutrophils produce and release high quantities of antibacterial peptides, proteases, and reactive oxygen species (ROS). The robust ROS production is also called 'the respiratory burst', and the NADPH oxidase or NOX2 is the enzyme responsible for the production of superoxide anion, leading to other ROS. In vitro, several soluble and particulate agonists induce neutrophil ROS production. This process can be enhanced by prior neutrophil treatment with 'priming' agents, which alone do not induce a respiratory burst. In this review, we will describe the priming process and discuss the beneficial role of controlled neutrophil priming in host defense and the detrimental effect of excessive neutrophil priming in inflammatory diseases.

                Author and article information

                4 July 2019
                10 January 2018
                February 2018
                29 July 2019
                : 154
                : 3
                : 704-718.e10
                [1 ]Department of Medicine A, University Medicine Greifswald, Greifswald, Germany;
                [2 ]Interfacuity Institute for Genetics and Functional Genomics, University Medicine Greifswald, Greifswald, Germany;
                [3 ]Department of Surgery, University Medicine Greifswald, Greifswald, Germany;
                [4 ]Institute of Pathology, University Medicine Greifswald, Greifswald, Germany;
                [5 ]VA Greater Los Angeles Healthcare System; David Geffen School of Medicine, University of California at Los Angeles, California;
                [6 ]Medizinische Klinik und Poliklinik II, Universitätsklinikum der Ludwig-Maximilians-Universität, Klinikum Grosshadern, Munich, Germany
                Author notes

                Authors share co-senior authorship.

                Reprint requests: Address requests for reprints to: Julia Mayerle, MD, Medizinische Klinik und Poliklinik II, Klinikum der LMU München-Grosshadern, Anstalt des öffentlichen Rechts, Marchioninistr. 15, D-81377 München, Germany. julia.mayerle@ 123456med.uni-muenchen.de ; fax: +49 (0) 89 4400-78887.

                This is an open access article under the CC BY-NC-ND license ( http://creativecommons.org/licenses/by-nc-nd/4.0/).


                Gastroenterology & Hepatology
                pancreatic inflammation,mechanisms,immune response,mouse model
                Gastroenterology & Hepatology
                pancreatic inflammation, mechanisms, immune response, mouse model


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