Mechanisms of Hypothermic Protection from Ischemia/Reperfusion Cardiac Injury

描述（由申请人提供）：治疗性低温可以显著减轻复苏后的损伤，提高一些患者的生存率。在我们的细胞和动物全脑缺血模型中，有明显的复苏后心脏损伤。该建议的中心假设是，在缺血期间启动的低温通过相关途径保护心肌细胞并保持心功能，涉及一氧化氮（NO）的产生和p53的下调，它们作用于线粒体以防止凋亡级联的启动。利用已建立的恒温和低温细胞和动物模型，我们提出了两个主要的具体目标：1)确定低温诱导的一氧化氮，活性氧（ROS）和线粒体的调节在保护心肌细胞和心脏免受缺血/再灌注（l/R）诱导的损伤中的相互作用。我们假设缺血低温保护涉及ROS和活性氮（RNS）谱改变的产生，允许缺血减少的线粒体膜电位恢复，并阻止线粒体释放细胞色素c （cyt c）。在这些研究中，我们将研究通过低温诱导或调节特定氧化剂，如超氧化物和一氧化氮，以及这些对细胞凋亡线粒体途径启动的影响。2)确定线粒体Bcl-2家族蛋白（特别是Bax和Bak）与其他“线粒体靶向”蛋白（特别是p53）之间的相互作用，通过低温诱导的NO生成提供保护并阻止促凋亡线粒体蛋白cyt c的释放和半胱天冬酶的激活。我们假设在I/R过程中p53的激活导致Bax和/或Bak的激活，而低温阻断p53的激活、与Bax/Bak的相互作用和/或易位将抑制这些相互作用，阻断cyt c的释放并保护细胞免于凋亡。拟议的研究将阐明低温诱导心肌细胞保护的分子机制，包括“适应性”或保护性低温氧化剂的细节。了解特定线粒体凋亡途径受低温信号影响的机制将有助于制定低温、药物和基因治疗方法的最佳保护方案。

DESCRIPTION (provided by applicant): Therapeutic hypothermia can significantly attenuate post-resuscitation injury and improve survival in some patients. In our cellular and animal models of global ischemia, there is significant post-resuscitation cardiac injury. The central hypothesis of this proposal is that hypothermia initiated during ischemia protects cardiomyocytes and preserves cardiac function through interrelated pathways, involving the production of nitric oxide (NO) and p53 down-regulation, which act at the mitochondria to prevent initiation of the apoptotic cascade. Using a combination of established normothermic and hypothermic cellular and animal models, we propose two major specific aims: 1) Determine the interacting roles of hypothermia-induced nitric oxide, modulation of reactive oxygen species (ROS) and the mitochondrion in protecting cardiomyocytes and the heart from ischemia/reperfusion (l/R)-induced injury. We hypothesize that intra-ischemic hypothermic protection involves the generation of an altered ROS and reactive nitrogen species (RNS) profile, allowing recovery of ischemia-reduced mitochondrial membrane potential and blocking the release of cytochrome c (cyt c) from the mitochondria. In these studies, we will examine the induction or modulation of specific oxidants, such as superoxide and nitric oxide, by hypothermia and these effects on the initiation of the mitochondrial pathway of apoptosis. 2) Determine the interactions between mitochondrial Bcl-2 family proteins, specifically Bax and Bak, and other "mitochondria-targeted" proteins, specifically p53, through which hypothermia-induced NO generation provides protection and prevents release of pro-apoptotic mitochondrial proteins cyt c and activation of caspases. We hypothesize that activation of p53 during I/R leads to activation of Bax and/or Bak and hypothermic blockade of p53 activation, interactions with Bax/Bak, and/or translocation will inhibit these interactions, blocking the release of cyt c and protecting the cells from apoptosis. The proposed studies will elucidate the molecular mechanisms involved in the hypothermic induction of protection of cardiomyocytes, including details of the "adaptive" or protective hypothermic oxidant profile. An understanding of mechanisms by which specific mitochondrial pathways of apoptosis are affected by hypothermic signaling will aid in developing optimal protective protocols in hypothermic, pharmacological and genetic therapeutic approaches.