线粒体细胞色素c氧化酶在心肌组织损伤与修复中的作用

Sustained pressure overload causes myocardial tissue injury leading to cardiac hypertrophy. Suppressed cytochrome c oxidase (CCO) activity and mitochondriopathy are involved in the pathogenesis. We have observed that copper (Cu) loss in the heart due to pressure overload is highly responsible for the depressed CCO activity because Cu is an essential component of CCO enzyme complex. Increasing dietary Cu intake from 6 mg Cu/kg to 20 mg/kg diet can recover CCO activity, repair mitochondriopathy, and reverse the established cardiac hypertrophy in a mouse model. To provide mechanistic insights, we propose to test the hypothesis that increase in homocysteine and oxidative stress associated with sustained pressure overload cause Cu mobilization and redistribution leading to Cu restriction to CCO, thus suppressed CCO activity and the pathological consequences, and Cu supplementation reverses these adverse effects. We will carry out the following aims: Aim 1 is to explore the molecular mechanism by which sustained pressure overload causes Cu restriction to CCO. Sustained pressure overload leads to increase in homocysteine and oxidative stress. Homocysteine would form complexes with Cu leading to Cu exclusion from cardiomyocytes. Oxidative stress causes Zn release from metallothionein and Zn replacement of Cu-binding site leading to Cu mobilization and redistribution. Both restrict the availability of Cu to CCO. We will use cardiac-specific catalase overexpressing transgenic mice to prevent oxidative stress and siRNA interfering homocysteine synthesis to address these two scenarios. Aim 2 is to determine why Cu excluded from the heart cannot be reused by the heart and how to make Cu from unavailable to available form to the heart. We will determine changes in Cu speciation in the blood of cardiac hypertrophic mice, then test in primary cultures of neonatal rat cardiomyocytes the uptake of different Cu-speciation and differentiate the unavailable from available forms of Cu speciation. Finally, we will attempt to use compounds that can manipulate Cu speciation change in the blood based on differential Cu-binding affinities to make Cu available to the heart. Aim 3 is to determine the essential role of recovery of suppressed CCO activity in the regression of cardiac hypertrophy. We will use a cardiac COX10 conditional knockout mouse model, in which treatment with tamoxifen induces cardiac-specific COX10 deletion and CCO suppression independent of Cu status. We have observed that without the recovery of CCO activity, regression of cardiac hypertrophy did not occur. We will examine the effects of CCO activity and mitochondrial functional recovery on cardiac angiogenesis, regression signaling transduction, and inhibition of cardiac hypertrophic pathways. This study will generate novel insights into the suppression of CCO activity and the affected mitochondrial dysfunction and their recovery in prevention of heart failure induced by pressure overload.

在压力负荷持续增高的情况下，心肌细胞线粒体损伤、功能异常导致能量代谢紊乱和活性氧自由基增加，细胞凋亡、坏死，进而引起心肌组织损伤。已有研究证明，保护线粒体功能可抑制心肌组织损伤的发生与发展。细胞色素c氧化酶活性下降是引起线粒体损伤及功能异常的主要原因。我们的前期研究发现压力负荷持续增高引起心肌组织铜的含量减少是细胞色素c氧化酶活性下降的主要原因。我们将在本研究中检验"在压力负荷增加诱导的心肌损伤中，通过补充铜可以促进细胞色素c氧化酶活性和线粒体功能恢复，最终实现受损心肌组织的修复"这一科学假说。利用转基因和基因敲除小鼠，我们将深入探索（1）压力负荷增加诱导的心肌组织铜含量下降以及心肌组织摄取铜的机制；（2）在众多铜-结合蛋白中，细胞色素c氧化酶对缺铜的特殊敏感性和活性下降的机制；（3）细胞色素c氧化酶活性在心肌损伤恢复中所发挥的关键作用机制。本项研究结果将为心肌组织损伤与修复提供新的认识。

在压力负荷持续增高的情况下，心肌细胞线粒体损伤、功能异常导致能量代谢紊乱和活性氧自由基增加，细胞凋亡、坏死，进而引起心肌组织损伤。已有研究证明，保护线粒体功能可抑制心肌组织损伤的发生与发展。细胞色素c氧化酶（CCO）活性下降是引起线粒体损伤及功能异常的主要原因。我们的前期研究发现压力负荷持续增高引起心肌组织铜的含量减少是细胞色素c氧化酶活性下降的主要原因。本研究中，我们用细胞模型和动物模型都证明了在压力负荷增加诱导的心肌损伤中，铜含量下降。通过补充铜可以促进细胞色素c氧化酶活性和线粒体功能恢复，最终实现受损心肌组织的修复。我们使用心肌特异性COX10条件下敲除小鼠研究发现，CCO活性的恢复对补铜治疗修复心肌组织是必须的，PGK1通路参与了此过程。同时我们深入探讨了压力负荷增加情况下，心肌组织铜丢失和摄取铜的机制，发现同型半胱氨酸能与铜形成复合物，引起铜流失。此外，我们找到了可以将血液中不可利用的铜转化为可利用的铜的方式，实现体内铜的重分布，受损心肌靶向补铜治疗。本项研究结果除了以科研论文发表以外，还将为临床上心肌组织损伤与修复提供新的治疗方式，有临床应用前景。