Role of the mitochondrial LonP1 in myocardial ischemia and reperfusion injury protection

项目总结 缺血-再灌注(IR)损伤是治疗心肌梗死(MI)的重大挑战，心肌梗死(MI)是最主要的心肌梗死 在美国的死因。电子产生的线粒体活性氧(MtROS) 运输链复合体-I是IR损伤的主要介质。在以下过程中生成的mtROS过多 早期IR会触发自由基产生的恶性循环，促进心肌细胞死亡。因此， 了解再灌注早期分子事件将为开发新的靶点提供新的靶点 限制心脏损伤的干预措施。我们发表的研究结果表明，LonP1-a主要的线粒体应激 反应蛋白在早期IR中减轻氧化应激诱导的损伤；因此，LonP1可能是一种 有望成为减轻再灌注损伤的靶点。我们的长期目标是利用线粒体蛋白 LonP1作为制定减轻IR损伤治疗策略的关键点的质量控制机制 以及心肌梗塞后的心力衰竭。我们发表的研究结果表明，心脏中LonP1表达的增加会导致 通过缺血预适应(IPC)或转基因过表达(LonTg)减轻IR损伤并有利于 心脏保护。而LonP1下调(LONP1+/-)可阻断IPC介导的心肌保护作用。 重要的是，LonTg心脏显示出复杂的-I亚基水平降低(但不是复杂的II-V亚单位)和 与正常对照组相比，早期IR期间(再灌流30分钟内)的氧化损伤。相反，我们的 更多的发现表明，LonP1下调心肌细胞中Complex-I的活性， 增加超氧化物歧化水平，并表现出早期再灌流诱导的细胞死亡激活。此外，我们还有 鉴定了一种可显著降低缺氧-复氧(H/R)诱导的LonP1小分子激活剂 体外心肌细胞死亡呈剂量依赖性。关于红外诱导的乙酰化的更多数据 在IR过程中，Complex-I矩阵亚单位和LonP1依赖的Complex-I重构，我们假设LonP1 通过严格调节Complex-I抑制mtROS的过量产生来减轻心肌损伤 在早期IR期间。我们将通过以下具体目标来检验我们的假设：目标1将描述 在IR过程中LonP1调节Complex-I水平、活性和降低氧化应激的机制(S)。 目标2将测试LonP1通过降解IR诱导的分子来重塑复合体-I及其相关的超复合体 翻译后修饰(PTM)复合体-I矩阵亚单位，从而减少早期IR期间的mtROS。目标3 将确定LonP1激活剂在体内治疗心肌缺血再灌注损伤的治疗潜力。通过 LonP1介导的心脏保护的分子机制及其治疗潜力的确定 LonP1激活剂，我们将确定LonP1在心脏保护中的作用，并开发新的治疗工具和 减轻IR损伤的策略。

PROJECT SUMMARY Ischemia-reperfusion (IR) injury is a significant challenge in treating myocardial infarction (MI), the leading cause of death in the United States. Mitochondrial reactive oxygen species (mtROS) generated by electron transport chain (ETC) Complex-I are the principal mediators of IR injury. Excess mtROS generated during early IR triggers vicious cycles of free radical production promoting cardiomyocyte death. Therefore, understanding the early molecular events of reperfusion will provide new targets for developing novel interventions for limiting cardiac injury. Our published findings show that LonP1- a major mitochondrial stress response protease mitigates oxidative stress-induced damage during early IR; therefore, LonP1 could be a promising target for attenuating reperfusion injury. Our long-term goal is to leverage the mitochondrial protein quality control mechanisms of LonP1 as a pivotal point to develop therapeutic strategies for mitigating IR injury and post MI- heart failure. Our published findings show that increased LonP1 expression in the heart induced by ischemic preconditioning (IPC) or transgenic overexpression (LonTg) reduced IR injury and favors cardioprotection. Whereas, LonP1 downregulation (LONP1+/-) abrogated IPC-mediated cardioprotection. Importantly, LonTg hearts showed reduced levels of Complex-I subunits (but not Complex II-V subunit) and oxidative damage during early IR (within 30 min reperfusion) compared to NTg controls. Conversely, our additional findings show that LonP1 downregulation in cardiomyocytes upregulated Complex-I activity, increased superoxide levels, and showed early reperfusion-induced cell death activation. In addition, we have identified a small molecule activator of LonP1 that significantly reduced hypoxia-reoxygenation (H/R) induced myocyte death in a dose-dependent manner in vitro. With additional data on IR-induced acetylation of Complex-I matrix subunits and LonP1 dependent Complex-I remodeling during IR, we hypothesize that LonP1 mitigates myocardial injury by suppressing excess mtROS generation through tight regulation of Complex-I during early IR. We will test our hypothesis by the following specific aims: Aim 1 will delineate the mechanism(s) by which LonP1 modulates Complex-I levels, activity and reduces oxidative stress during IR. Aim 2 will test that LonP1 remodels Complex-I and its associated supercomplexes by degrading IR-induced post-translationally modified (PTM) Complex-I matrix subunits, thereby reduce mtROS during early IR. Aim 3 will determine the therapeutic potential of LonP1 activators in treating myocardial IR injury in vivo. By determining the molecular mechanisms of LonP1-mediated cardioprotection and the therapeutic potential of LonP1 activators, we will define the role of LonP1 in cardioprotection and develop novel therapeutic tools and strategies to mitigate IR injury.