Abstract 15664: Conditional Overexpression of Neuronal Nitric Oxide Synthase is Cardioprotective in Ischemia-Reperfusion
Introduction: We previously demonstrated that conditional overexpression of nNOS inhibited L-type Ca2+-channels and decreased myocardial contractility. However, nNOS has multiple targets within the cardiac myocyte. We now hypothesized that nNOS overexpression acts cardioprotective after ischemia-reperfusion because of inhibition of mitochondrial function and a reduction in ROS generation.
Methods and results: We assessed the effect of conditional nNOS overexpression in cardiac myocytes in ischemia-reperfusion injury. NOS-activity (22 ± 1.5 vs. 29 ± 1μM/sec, n=18, p<0.05) was significantly enhanced after nNOS overexpression. Electron microscopy with immunogold labeling, immunprecipitation experiments and co-immunfluorescence-staining of adult cardiac myocytes indicate an interaction of nNOS at the mitochondria in nNOS overexpressing animals. In vitro ischemia-reperfusion experiments with isolated hearts showed a cardioprotective effect of nNOS overexpression (30 min post-ischemia, LVDP 20±6 in non-induced animals vs. 60±11 mmHg in nNOS overexpressing animals, n=6, p<0.05). In vivo ischemia-reperfusion experiments showed a significantly reduced infarct size in nNOS overexpressing mice (14.4±0.8 % vs. 19.4±1%, n=8, p<0.05). O2-consumption in isolated heart muscle stripes was decreased in nNOS overexpressing mice already under resting conditions (0.018±0.01 vs. 0.025±0.007ml [O2] × mm−3 × min−1, n=11, p<0.05). We also investigated the function of mitochondrial nNOS. nNOS overexpression caused a significant decrease of cytochrome c oxidase activity (OD 0.21±0.03 vs. OD 0.24±0.02, n=7,p<0.05). Additionally, we found that the ROS concentration was significantly decreased in hearts of nNOS overexpressing mice (6.14±0.685μM vs. 14.53±1.7μM, n=8, p<0.01) which was caused by inhibition of XOR activity by abundant nNOS expression. We also demonstrated interaction of nNOS with HSP90. HSP90 acts as a carrier of nNOS across the mitochondrial membrane Discussion: We demonstrated that conditional transgenic overexpression of nNOS resulted in myocardial protection after ischemia-reperfusion injury by suppression of ROS generation and inhibition of myocardial oxygen consumption in the mitochondria.
- © 2010 by American Heart Association, Inc.