Mitochondrial pathways in NO induced cardioprotection

描述（由申请人提供）： 在过去的几十年里，研究已经大大提高了我们对心肌缺血损伤的保护和发病机制的信号机制的理解。线粒体功能的保护在心肌缺血再灌注损伤（I/R）的保护中起着重要作用。然而，它仍然几乎是未知的线粒体中的心脏保护的分子靶点是谁;什么特定的分子事件导致线粒体的保护;以及线粒体中是否存在心脏保护信号的系统整合以支持受保护表型的表现。使用一氧化氮（NO）诱导的心脏保护晚期的小鼠模型，我们选择使用一种新的实验策略，通过计算建模并行检查线粒体信号，线粒体蛋白质组和线粒体行为，来检查线粒体的合理内在信号传导特性。建议的研究是基于其他人和我们自己的初步证据，表明激活PKC？Src模块发生在NO供体处理的小鼠中，并且两者都定位于线粒体膜。在这项建议中，我们将测试的创新假设，PKC？- Src模块与短暂的mPTP开口相互作用以保护心肌细胞免受Ca++过载诱导的损伤。工作假设是NO激活PKC？Src模块，导致短暂的mPTP开放，导致瞬时Ca++释放和活性氧（ROS）爆发，从而使Ca++再摄取失活，并进一步激活PKC？Src来形成一个前馈回路。当体内平衡被中断时（例如，钙超载或升高的ROS），短暂的mPTP开放转变为不可逆的、持久的mPTP开放，其反而诱导心脏损伤。在这项应用中，我们建议描绘的功能性影响的短暂开放的mPTP的钙++处理和活性氧的生产，并阐明mPTP调节线粒体蛋白激酶C？Src模块在NO诱导的心脏保护晚期（目的1）的设置。在具体的目标2中，我们将最终确定PKC？线粒体Src信号模块作为NO诱导的心肌缺血保护的强制性信号元件。最后，我们将系统地确定线粒体Src激酶在NO诱导的心脏保护晚期的分子靶点（目的3）。拟议的研究将通过提供新的机制见解来促进我们对心脏生物学的理解，即短暂的mPTP开口与线粒体PKC？Src模块可能在介导NO诱导的晚期心脏保护中是有益的。 (End摘要）

DESCRIPTION (provided by applicant): Investigations in the past decades have significantly advanced our understanding of signaling mechanisms underlying the protection and pathogenesis of myocardial ischemic injury. It is increasingly recognized that preservation of mitochondrial function plays a pivotal role in cardioprotection against ischemia reperfusion injury (I/R). However, it remains virtually unknown as to who the molecular targets of cardioprotection are in the mitochondria; what specific molecular events led to the protection of mitochondria; and whether there is a systems integration of cardioprotective signaling at the mitochondria to support the manifestation of a protected phenotype. Using a murine model of nitric oxide (NO) induced late phase of cardioprotection, we elect to examine the plausible intrinsic signaling properties of mitochondria using a novel experimental strategy enabling a parallel examination of mitochondrial signaling, mitochondrial proteomes, and mitochondrial behavior by computational modeling. The proposed studies are based upon preliminary evidence by others and our own demonstrating that activation of PKC?-Src module occurs in the NO donor treated mice and that both are localized to mitochondrial membranes. In this proposal we will test the innovative hypothesis that the PKC?-Src module interacts with the brief mPTP openings to protect cardiomyocytes from Ca++ overload induced jury. The working hypothesis is that NO activates PKC?-Src module, leading to brief mPTP openings which results in transients Ca++ releases and reactive oxygen species (ROS) bursts, and consequently inactivates Ca++ reuptake and further activates PKC?-Src to form a feed- forward loop. When the homeostasis is interrupted (e.g., calcium overload or elevated ROS), brief mPTP openings transits into irreversible, long-lasting mPTP openings, which instead induce cardiac injury. In this application we propose to delineate the functional effects of brief openings of mPTP on Ca++ handling and ROS production; and to elucidate mPTP regulation by the mitochondrial PKC?-Src module in the setting of NO-induced late phase of cardioprotection (Aim 1). In the specific Aim 2 we will conclusively establish the activation of a PKC?-Src signaling module in the mitochondria as a mandatory signaling element of NO-induced cardioprotection against myocardial ischemic injury. At last we will systematically define the molecular targets of mitochondrial Src-kinase in NO- induced late phase of cardioprotection (Aim 3). The proposed studies will advance our understanding of cardiac biology by providing novel mechanistic insights into how interactions of brief mPTP openings with mitochondrial PKC?-Src module can be beneficial in mediating NO-induced late phase of cardioprotection. (End of Abstract)