Abstract 115: Post-ischemic Estrogen Administration Protects the Myocardium Against Ischemia/Reperfusion Injury by Reducing Mitochondrial Protein Ubiquitination, Acetylation and Calpain10 Levels and Inhibition of the mPTP Opening via GPER1 Activation

Abstract

Introduction: We recently found in isolated perfused hearts that acute pre-ischemic estrogen-induced cardioprotection against ischemia/reperfusion injury is mainly mediated via G protein-coupled estrogen receptor1 (GPER1) activation but not through classical estrogen receptors: alpha (ERα) and beta (ERβ). Hypothesis: We investigated whether acute post-ischemic estrogen (PI-E2) treatment can also induce cardioprotective effects via GPER1 activation in the intact animal subjected to ischemia/reperfusion injury. Methods: Male and ovariectomized female Sprague-Dawley rats were anesthetized with ketamine (80 mg/kg i.p.) and xylazine (8 mg/kg i.p.). Hearts were subjected to 35 min of the left anterior descending (LAD) artery occlusion, followed by 180 min reperfusion. An E2 bolus (50 mg/kg body weight) or PBS (same volume) was applied via the femoral vein 5 min before reperfusion and a GPER1 antagonist, G15, was given 10 min before E2. Area at risk (AAR) was identified using Evans Blue dye and myocardial infarct size assessed by TTC staining method. Mitochondria calcium retention capacity (CRC) required to induce mitochondrial permeability transition pore (mPTP) opening was assessed after 10 min reperfusion. Expression of ubiquitinated, acetylated, as well as calpains 1 and 10 proteins was measured by Western Blot in mitochondrial and cytosolic fractions. Results: We found that PI-E2 treatment reduced myocardial infarct size normalized to the AAR or the whole LV and improved mitochondrial CRC. PI-E2 treatment reduced mitochondrial protein acetylation, ubiquitination, and decreased calpain10 levels in mitochondrial but not in cytosolic fractions as compared to control, respectively. Interestingly, all these effects of E2 were abolished by addition of G15. Conclusion: Acute post-ischemic GPER1 activation by E2 induces cardioprotection against ischemia/reperfusion injury. PI-E2 effects through GPER1 involve the reduction of the levels of mitochondrial protein acetylation, ubiquitination, and calpain10. These PI-E2-GPER1 effects lead to inhibition of the mPTP opening.