Egr-1 functions as a master switch regulator of remote ischemic preconditioning-induced cardioprotection

Abstract

Despite improved treatment options myocardial infarction (MI) is still a leading cause of mortality and morbidity worldwide. Remote ischemic preconditioning (RIPC) is a mechanistic process that reduces myocardial infarction size and protects against ischemia reperfusion (I/R) injury. The zinc finger transcription factor early growth response-1 (Egr-1) is integral to the biological response to I/R, as its upregulation mediates the increased expression of inflammatory and prothrombotic processes. We aimed to determine the association and/or role of Egr-1 expression with the molecular mechanisms controlling the cardioprotective effects of RIPC. This study used H9C2 cells in vitro and a rat model of cardiac ischemia reperfusion (I/R) injury. We silenced Egr-1 with DNAzyme (ED5) in vitro and in vivo, before three cycles of RIPC consisting of alternating 5 min hypoxia and normoxia in cells or hind-limb ligation and release in the rat, followed by hypoxic challenge in vitro and I/R injury in vivo. Post-procedure, ED5 administration led to a significant increase in infarct size compared to controls (65.90\u2009±\u20092.38% vs. 41.00\u2009±\u20092.83%, p\u2009<\u20090.0001) following administration prior to RIPC in vivo, concurrent with decreased plasma IL-6 levels (118.30\u2009±\u20094.30 pg/ml vs. 130.50\u2009±\u20091.29 pg/ml, p\u2009<\u20090.05), downregulation of the cardioprotective JAK–STAT pathway, and elevated myocardial endothelial dysfunction. In vitro, ED5 administration abrogated IL-6 mRNA expression in H9C2 cells subjected to RIPC (0.95\u2009±\u20090.20 vs. 6.08\u2009±\u20091.40-fold relative to the control group, p\u2009<\u20090.05), resulting in increase in apoptosis (4.76\u2009±\u20090.70% vs. 2.23\u2009±\u20090.34%, p\u2009<\u20090.05) and loss of mitochondrial membrane potential (0.57\u2009±\u20090.11% vs. 1.0\u2009±\u20090.14%-fold relative to control, p\u2009<\u20090.05) in recipient cells receiving preconditioned media from the DNAzyme treated donor cells. This study suggests that Egr-1 functions as a master regulator of remote preconditioning inducing a protective effect against myocardial I/R injury through IL-6-dependent JAK–STAT signaling.