Activation of NADPH oxidase mediates increased endoplasmic reticulum stress and left ventricular remodeling after myocardial infarction in rabbits

Abstract

Nicotinamide adenine dinucleotide 3-phosphate (NADPH) oxidase activity and endoplasmic reticulum (ER) stress are increased after myocardial infarction (MI). In this study, we proposed to test whether activation of the NADPH oxidase in the remote non-infarcted myocardium mediates ER stress and left ventricular (LV) remodeling after MI. Rabbits with MI or sham operation were randomly assigned to orally receive an NADPH oxidase inhibitor apocynin or placebo for 30 days. The agents were administered beginning at 1 week after surgery. MI rabbits exhibited decreases in LV fractional shortening, LV ejection fraction and the first derivative of the LV pressure rise, which were abolished by apocynin treatment. NADPH oxidase Nox2 protein and mRNA expressions were increased in the remote non-infarcted myocardium after MI. Immunolabeling further revealed that Nox2 was increased in cardiac myocytes in the remote myocardium. The apocynin treatment prevented increases in the Nox2 expression, NADPH oxidase activity, oxidative stress, myocyte apoptosis and GRP78, CHOP and cleaved caspase 12 protein expression in the remote myocardium. The apocynin treatment also attenuated increases in myocyte diameter and cardiac fibrosis. In cultured H9C2 cardiomyocytes exposed to angiotensin II, an important stimulus for post-MI remodeling, Nox2 knockdown with siRNA significantly inhibited angiotensin II-induced NADPH oxidase activation, reactive oxygen species and GRP78 and CHOP protein expression. We conclude that NADPH oxidase inhibition attenuates increased ER stress in the remote non-infarcted myocardium and LV remodeling late after MI in rabbits. These findings suggest that the activation of NADPH oxidase in the remote non-infarcted myocardium mediates increased ER stress, contributing to myocyte apoptosis and LV remodeling after MI.

Endoplasmic reticulum stress: a novel mechanism and therapeutic target for cardiovascular diseases

“…Luo et al [110] reported that MI induced ERS and provoked cardiac apoptosis and fibrosis by increasing the expression of ERS markers GRP78 and CHOP, pro-apoptotic proteins Bax and caspase 3, and pro-fibrotic proteins transforming growth factor-beta 1 (TGF-β1) and Smad 2/3, culminating in cardiac rupture and remodeling. Li et al [111] reported that ERS contributed to MI through the activation of NADPH oxidase. NADPH oxidase inhibition attenuated an increase in ERS in the remote non-infarcted myocardium and LV remodeling after MI by preventing the increase of GRP78, CHOP and cleaved caspase 12 protein expression in rabbits.…”

Progressive Reduction in Myocyte Autophagy After Myocardial Infarction in Rabbits: Association with Oxidative Stress and Left Ventricular Remodeling

“…Healthy adult male New Zealand White rabbits (2-2.5 kg) obtained from the Academy of Military Medical Sciences were prepared for MI produced by coronary artery ligation as described before [13]. The rabbits were anesthetized by intraperitoneal injection of pentobarbital sodium (50mg/kg) and mechanically ventilated with a respirator (HX-101E, Taimeng Technology Co., Ltd., Chengdu, China).…”

“…The H9C2 rat cardiomyocyte line was obtained from the Chinese Academy of Sciences Cell Bank (Shanghai, China; The American Type Culture Collection, Manassas, VA, USA) [13]. The H9C2 cardiomyocytes were cultured in 25 cm 2 flask in Dulbecco's modified Earle's medium (DMEM) (Gibco, Life Technologies, USA) with 3.7 g/L sodium bicarbonate, 4.5 g/L D-glucose, and 110 mg/L sodium pyruvate, supplemented with 10% fetal bovine serum (Australia) and penicillin (100 units/mL) and streptomycin (100 μg/mL) in a humidified incubator with 95% air and 5% CO 2 at 37 °C.…”

“…The two-dimensional and M-mode echocardiograms were conducted before surgery and 1, 4 and 8 weeks after surgery to assess LV dimension and function using a UCG machine (AE33, Philips) equipped with a 3-7 MHz transducer (S5-1, Philips) [13]. Maximal LV end-diastolic dimension (EDD) and end-systolic dimension (ESD) were used to calculate LV fractional shortening (FS) by the following equation: FS =[(EDD− ESD) × 100] / EDD.…”

“…Myocytes with nuclei in the middle or near the middle were selected for the analysis. The quantification of myocyte diameter was determined using NIH ImageJ software as described previously [13]. Measurements of cardiac fibrosis LV tissue sections were stained with Masson's trichrome kit (Sigma) and examined under a light microscope (AX 10, Zeiss).…”