Oxidative Stress and Nitric Oxide in Aged Myocardium

本先导研究资助计划的重点是研究目标2：心脑血管老化。衰老与许多组织中氧化应激和线粒体功能障碍的增加有关。这些因素被认为在与年龄相关的心室功能、冠状动脉血管储备和心肌缺血再灌注耐受性下降中发挥作用，所有这些因素都可能导致心血管疾病随年龄增长的增加。越来越多的证据表明，线粒体除了维持心室心肌高工作需求的能量储存外，还在决定细胞从可逆损伤到凋亡和坏死的转变中发挥作用。然而，关于老龄动物心肌缺血再灌注损伤的研究很少，对老龄动物完整心室肌细胞线粒体功能的系统研究也很少。这一提议将验证衰老心肌对缺血再灌注损伤和氧化应激的耐受性降低的假设，部分原因是一氧化氮（NO）代谢和信号传导的改变。特异性目的1将确定老年大鼠体内心肌和离体心室肌细胞是否表现出缺血再灌注和心肌缺血再灌注的耐受性较差

This pilot research grant proposal focuses on research objective 2: Cardiovascular and Cerebrovascular Aging. Aging is associated with increased oxidative stress and mitochondrial dysfunction in numerous tissues. These factors are thought to play a role in aging-associated decreases in ventricular function, coronary vascular reserve, and tolerance to myocardial ischemia-reperfusion all of which may contribute to the increase in cardiovascular disease with aging. Significant evidence has accumulated that mitochondria, in addition to maintaining the energy stores for the high work demands of ventricular myocardium, play a role in determining the cell's transition from reversible injury to apoptosis and necrosis. However there have been limited studies of ischemia-reperfusion injury in in vivo aged myocardium and few systematic investigations of mitochondrial function in intact ventricular myocytes of aged animals. This proposal will test the hypothesis that aged myocardittm exhibits less tolerance to ischemia-reperfusion injury and oxidative stress in part due to altered nitric oxide (NO) metabolism and signaling. Specific Aim 1 will determine whether in vivo myocardium and isolated ventricular myocytes from aged rats exhibit less tolerance to ischemia-reperfusion and hypoxia-reoxygenation, respectively. Specific Aim 2 will test the hypothesis that aging associated oxidative stress and mitochondrial dysfunction is due to increased formation of reactive nitrogen species. In Aim 1 studies adult (6 month) and aged (24 month) Fischer 344 x Brown Norway F1 hybrid (F344xBN) rats will be submitted to in vivo coronary artery occlusion and reperfusion and infarct size will be measured. Left ventricular myocytes isolated from adult and aged rats will be submitted to hypoxia-reoxygenation and twitch amplitude and intracellular oxidative stress will be measured. In Aim 2 studies isolated ventricular myocytes will be exposed to/-/202 tO induce oxidative stress. Mitochondrial ([Ca2+]), mitochondrial redox state, and intracellular oxidative stress will be measured under baseline conditions and during and following exposure to H202. The role of NO in modulating aged myocyte response to oxidative stress will be determined by modulating NOS activity and arginine levels and with a NO donor. The expression and subcellular compartmentation of anti-oxidant enzymes and NOS isoforms will be determined by Western blotting of subcellular fractions. The proposed studies will provide significant insight into the role of acute and chronic oxidative stress in aged myocardium and its modulation by NO. The results obtained may lead to the development of new therapies for treating cardiovascular disease in the elderly.