Mitochondria & NO in Cardiac Ischemia-Reperfusion

描述（由申请人提供）：心肌缺血和再灌注（I-R）后，生物能量功能障碍和钙稳态丧失导致功能恢复不良。最近的研究结果强调了三个潜在的现象，包括活性氮物质（RNS），线粒体扰动和MAP激酶信号的作用。虽然这些机制被认为是相关的，但尚未得到解决。线粒体蛋白质组学的发展和一氧化氮（NO）与线粒体相互作用的新见解允许整合这些机制，并作为本提案的主题。理解线粒体扰动上游事件的顺序和机制在临床上是重要的，因为许多I-R损伤的药理学方法针对调节RNS水平、线粒体和MAP激酶信号传导。此外，RNS在I-R中的治疗作用一直难以解决，因为RNS在I-R中表现出有益和有害的作用。基于本文提供的数据，提出最近对RNS和钙在I-R期间与线粒体的相互作用的了解可以揭示潜在的病理机制。这将使用线粒体功能评估和蛋白质组学方法进行研究。初步数据表明，依赖于线粒体暴露于Ca 2+或RNS，不同的目标被揭示，并确定损伤的结果。此外，MAP激酶在I-R诱导的Ca ~（2+）升高中的新作用被提出。该提案的重点是响应I-R和RNS暴露而改变的两个线粒体参数：这些是呼吸复合物I活性的抑制和线粒体H+泄漏的升高。假设RNS、钙和MAP激酶负责抑制线粒体呼吸复合物I和增加心脏I-R中的H+渗漏。这一假设将通过追求以下目标来检验：1。应用线粒体蛋白质组学技术研究复合物I在缺血再灌注损伤中的抑制机制，以及内源性和外源性NO.2的作用。明确心肌I-R时H+漏出增加的机制及RNS对其的调节作用。3.研究MAP激酶在诱发心肌I-R线粒体功能障碍中的作用。这些目标的实现将极大地增强对I-R中线粒体扰动的基本机制的理解，从而导致新的治疗方法。

DESCRIPTION (provided by applicant): Following myocardial ischemia and reperfusion (I-R), bioenergetic dysfunction and loss of calcium homeostasis contribute to poor functional recovery. Recent findings highlight three underlying phenomena, including the role of reactive nitrogen species (RNS), mitochondrial perturbations, and MAP kinase signaling. Although these mechanisms are thought to be related, this has not been addressed. Developments in mitochondrial proteomics and new insights into nitric oxide (NO) interactions with mitochondria allow integration of these mechanisms, and serve as the theme of this proposal. Understanding the order and mechanism of events upstream of mitochondrial perturbations is clinically important, since many pharmacologic approaches to I-R injury are directed at modulating RNS levels, mitochondria, and MAP kinase signaling. Also, the therapeutic roles of RNS in I-R have been difficult to address, since RNS exhibit both beneficial and damaging effects in I-R. Building upon data presented herein it is proposed that recent insights into the interactions of RNS and calcium with mitochondria during I-R can reveal underlying pathological mechanisms. This will be investigated using assessment of mitochondrial function and a proteomics approach. Preliminary data indicate that dependent upon exposure of mitochondria to Ca2+ or RNS, different targets are revealed, and determine the outcome of injury. In addition, a novel role for MAP kinases in I-R-induced Ca2+ elevation is proposed. The focus of the proposal are two mitochondrial parameters that alter in response to both I-R and RNS exposure: These are the inhibition of respiratory complex I activity, and an elevation in mitochondrial H+ leak. It is hypothesized that RNS, calcium, and MAP kinases are responsible for the inhibition of mitochondrial respiratory complex I and increased H+ leak in cardiac I-R. This hypothesis will be tested by pursuit of the following aims: 1. Apply mitochondrial proteomics to determine the mechanism of complex I inhibition in ischemia-reperfusion, and the effects of endogenous and exogenous NO. 2. Determine the mechanism of increased H+ leak in cardiac I-R and its regulation by RNS. 3. Investigate the role of MAP kinases in eliciting mitochondrial dysfunction in cardiac I-R. Realization of these aims will greatly enhance understanding of the basic mechanisms underlying mitochondrial perturbations in I-R leading to novel therapies.