Modulation of Mitochondrial Function by Pro-Oxidants

描述（由申请人提供）：由心脏供血减少引起的并发症是世界范围内死亡和衰弱的主要原因。然而，冠状动脉血流恢复到看似存活的心肌组织，往往伴随着心功能的丧失，长期来看，会发展为心力衰竭。这种矛盾的现象，被广泛地称为缺血/再灌注损伤，在老年人中表现得更为严重。线粒体可能在心肌缺血/再灌注损伤中起核心作用。线粒体对维持心脏能量状态和功能至关重要，在缺血期间，线粒体的呼吸和氧化磷酸化速率下降，再灌注时进一步出现年龄依赖性缺陷。拟议的研究试图通过统一与缺血/再灌注相关的两个事件来确定导致这种功能丧失的机制：Ca2+过载和促氧化剂产生。心脏缺血导致细胞色素c从线粒体内膜脱落，这是导致电子传递速率下降的一个事件。在再灌注过程中，氧化还原敏感酶复合物I、a-酮戊二酸脱氢酶和乌头酸酶的活性下降。将分离的线粒体暴露于pH和Ca2+浓度的改变中，模拟从缺血到再灌注的转变，导致细胞色素c解离和氧化还原敏感酶的氧化抑制。根据氧化应激的大小和持续时间，可逆抑制可以发展为不可逆失活。假设：心肌缺血/再灌注时线粒体Ca2+浓度的增加导致细胞色素c从线粒体内膜分离，导致自由基产生增加和氧化还原敏感酶的氧化抑制。这暂时降低了线粒体呼吸速率、自由基产生和对不可逆氧化损伤的易感性。衰老增加了线粒体Ca2+超载，增加了线粒体功能从可逆调节到不可逆失活的可能性。利用体内大鼠模型，将使用不同年龄的动物来改变心脏缺血和再灌注的持续时间，以确定导致线粒体促氧化剂产生增加的分子事件（Aim 1），氧化还原依赖性修饰的特定靶点和机制（Aim 2），氧化修饰的生化后果（Aim 2），以及在心脏缺血/再灌注期间促进线粒体功能不可挽回损失的年龄依赖性因素（Aims 1和2）。阐明导致缺血/再灌注损伤的分子事件是优化有利于影响预后的策略的必要条件，尤其是在老年人群中。心脏疾病是导致衰弱和死亡的主要原因，尤其是在老年人群中。我们的研究试图确定与年龄相关的因素，这些因素会提高心脏病的严重程度，从而设计出改善结果的策略。

DESCRIPTION (provided by applicant): Complications arising from reduction of blood supply to the heart are a leading cause of death and debilitation worldwide. Nevertheless, restoration of coronary blood flow to seemingly viable myocardial tissue is often accompanied by loss of cardiac function and, in the long term, development of heart failure. This paradoxical phenomenon, broadly termed ischemia/reperfusion injury, is manifested more severely in the elderly. Mitochondria likely play a central role in myocardial ischemia/reperfusion injury. Critical for the maintenance of cardiac energy status and function, mitochondria exhibit declines in the rate of respiration and oxidative phosphorylation during ischemia, with further age-dependent deficits evident upon reperfusion. The proposed studies seek to define mechanisms responsible for this loss in function by unifying two events associated with ischemia/reperfusion: Ca2+ overload and pro-oxidant production. Cardiac ischemia results in detachment of cytochrome c from the inner mitochondrial membrane, an event responsible for declines in the rate of electron transport. During reperfusion, the redox sensitive enzymes complex I, a-ketoglutarate dehydrogenase, and aconitase exhibit declines in activity. Exposure of isolated mitochondria to alterations in pH and Ca2+ concentration that mimic the transition from ischemia to reperfusion results in cytochrome c dissociation and oxidative inhibition of these redox sensitive enzymes. Depending on the magnitude and duration of oxidative stress, reversible inhibition can progress to irreversible inactivation. It is hypothesized that: Increases in mitochondrial Ca2+ concentration during myocardial ischemia/reperfusion lead to dissociation of cytochrome c from the inner mitochondrial membrane resulting in an increase in free radical production and oxidative inhibition of redox sensitive enzymes. This transiently reduces the rates of mitochondrial respiration, free radical production, and susceptibility to irreversible oxidative damage. Aging augments mitochondrial Ca2+ overload, increasing the likelihood of progression from reversible modulation to irreversible inactivation of mitochondrial function. Utilizing an in vivo rat model, durations of cardiac ischemia and reperfusion will be varied using animals of different ages to identify molecular events that result in increased mitochondrial pro-oxidant production (Aim 1), specific targets and mechanisms of redox- dependent modification (Aim 2), biochemical consequences of oxidative modification (Aim 2), and age- dependent factors that promote irreparable loss in mitochondrial function during cardiac ischemia/reperfusion (Aims 1 and 2). Elucidation of molecular events responsible for ischemia/reperfusion injury is required for optimization of strategies for favorably influencing the outcome particularly in the elderly population. Lay Description: Heart disease is a leading cause of debilitation and death, particularly in the aging population. Our studies seek to define age-related factors that enhance the severity of heart disease in an effort to design strategies to improve the outcome.