Nexinhib20 inhibits neutrophil adhesion and β2 integrin activation to prevent myocardial ischemia-reperfusion injury

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Abstract Primary percutaneous coronary intervention (PPCI)—a therapy intended to restore blood flow after acute myocardial infarction—can also induce myocardial ischemia-reperfusion (I/R) injury, exacerbating cardiomyocyte death. Neutrophil-mediated cardiac tissue damage contributes prominently to the process of myocardial I/R injury. Therefore, limiting neutrophil recruitment and exocytosis may prevent myocardial I/R injury after PPCI. Here, we found out that a neutrophil exocytosis inhibitor Nexinhib20 inhibits both neutrophil adhesion and exocytosis and prevents myocardial I/R injury in a mouse model. Using a microfluidic chamber, we found that Nexinhib20 inhibited interleukin 8 (IL-8)-induced β2 integrin-dependent human neutrophil adhesion under flow. Using dynamic flow cytometry assay, we discovered that Nexinhib20 suppresses intracellular calcium flux and β2 integrin activation after IL-8 stimulation. Western blots of Rac-1-GTP pull-down assay confirmed that Nexinhib20 inhibited Rac-1 activation in leukocytes. In vitro competing assay showed that Nexinhib20 antagonized the binding of Rac-1 and GTP. Using a mouse model of myocardial I/R injury, Nexinhib20 administration after ischemia and before reperfusion significantly decreased neutrophil recruitment and infarct size. Our results highlight the translational potential of Nexinhib20 as a dual-functional neutrophil inhibitory drug to prevent myocardial I/R injury. This research was supported by grants from the National Institutes of Health, National Heart, Lung, and Blood Institute, USA (R01HL145454, R41HL156322, R44HL152710, and K99HL153678), a Career Development Award from American Heart Association (18CDA34110426), and a startup fund from UConn Health.