Estrogen Prevents Cardiomyocyte Apoptosis through Inhibition of Reactive Oxygen Species and Differential Regulation of p38 Kinase Isoforms

Mahnaz Razandi,Ali Pedram,Jin Kyung Kim,Ellis R. Levin

doi:10.1074/jbc.m511024200

Abstract

From human and animal studies, estrogen is known to protect the myocardium from an ischemic insult. However, there is limited knowledge regarding mechanisms by which estrogen directly protects cardiomyocytes. In this report, we employed an in vitro model, in which cultured rat cardiomyocytes underwent prolonged hypoxia followed by reoxygenation (H/R), to study the cardioprotective mechanism of estrogen. 17-beta-estradiol (E2) acting via estrogen receptors inhibited H/R-induced apoptosis of cardiomyocytes. Mitochondrial reactive oxygen species (ROS) generated from H/R activated p38alpha MAPK, and inhibition of p38alpha with SB203580 significantly prevented H/R-induced cell death. E2 suppressed ROS formation and p38alpha activation by H/R and concomitantly augmented the activity of p38beta. Unlike p38alpha, p38beta was little affected by H/R. Dominant negative p38beta protein expression decreased E2-mediated cardiomyocyte survival and ROS suppression during H/R stress. The prosurvival signaling molecule, phosphoinositol-3 kinase (PI3K), has previously been linked to cell survival following ischemia-reperfusion injury. Here, E2-activated PI3K was found to inhibit ROS generated from H/R injury, leading to inhibition of downstream p38alpha. We further linked these signaling pathways in that p38beta was activated by E2 stimulation of PI3K. Thus, E2 differentially modulated two major isoforms of p38, leading to cardiomyocyte survival. This was achieved by signaling through PI3K, integrating cell survival mediators.

Highlights

Ischemic heart disease with subsequent myocardial infarction and congestive heart failure is the leading cause of mortality in this country [1]
Using cultured neonatal rat cardiomyocytes and an established in vitro system of hypoxia followed by reoxygenation (H/R) to simulate I/R, we dissected the relationship among E2-activated estrogen receptor (ER), mitochondrial ROS generation, p38␣ and p38␤ MAPK isoforms, and phosphoinositol-3 kinase (PI3K) in the ation; I/R, ischemia/reperfusion; ER, estrogen receptors; ROS, reactive oxygen species; MAPK, mitogen-activated protein kinase; PI3K, phosphatidylinositol 3-kinase; PI, propidium iodide; NAC, N-acetyl-cysteine; DN, dominant negative
We found that E2 modulates two distinct and opposite functions of p38 isoforms in its protection of cardiomyocytes

Summary

Introduction

Ischemic heart disease with subsequent myocardial infarction and congestive heart failure is the leading cause of mortality in this country [1]. One model of cardiac apoptosis is that of I/R injury, associated with both the ischemic and the reperfused heart (14, 19 –21) Perhaps this is related to the bursts of reactive oxygen species (ROS) that are generated during both stages [15]. Additional mechanisms described in other cell models may be involved in I/R-induced apoptosis [11, 13] This includes p38 mitogen-activated protein kinase (p38 MAPK), generally stimulated by heterogeneous forms of cellular stress. Using cultured neonatal rat cardiomyocytes and an established in vitro system of hypoxia followed by reoxygenation (H/R) to simulate I/R, we dissected the relationship among E2-activated estrogen receptor (ER), mitochondrial ROS generation, p38␣ and p38␤ MAPK isoforms, and PI3K in the ation; I/R, ischemia/reperfusion; ER, estrogen receptors; ROS, reactive oxygen species; MAPK, mitogen-activated protein kinase; PI3K, phosphatidylinositol 3-kinase; PI, propidium iodide; NAC, N-acetyl-cysteine; DN, dominant negative

Methods

Results

Conclusion