Regulation of cardiac stress responses by Rho kinase

描述(由申请人提供)：心力衰竭仍然是人类发病率和死亡率的主要原因。Rho激酶(又称ROCK)最近成为治疗心脏疾病的潜在靶点，实验和临床研究表明ROCK抑制剂具有良好的疗效。然而，一个需要解决的重要问题是，ROCK是否真的代表了治疗人类疾病的可行靶点，因为目前可用的ROCK抑制剂具有广泛的特异性。此外，岩石家族的两个成员ROCK1和ROCK2被同等效力的岩石抑制剂抑制，对岩石异构体在体内的功能知之甚少。我们最近发现，全身性ROCK1缺乏症对心脏失代偿有保护作用，而ROCK1缺乏症的抗细胞凋亡作用是一个重要因素。与ROCK1缺失的有益效果相比，我们观察到心脏特异性ROCK2缺失会导致自发性心肌肥大和功能障碍，这表明ROCK2在心脏保护中具有新的作用。我们使用ROCK1或ROCK2缺陷的胚胎成纤维细胞进行的体外研究支持一种新的机制概念，即ROCK1优先通过ROCK1/MYPT/MLC途径介导应激诱导的肌球蛋白收缩，导致细胞死亡增加，而ROCK2优先通过ROCK2/LIMK/Cofilin途径促进肌动蛋白聚合，从而提高应激条件下细胞的存活率。这项应用的目的是剖析ROCK在肥厚性心脏重塑中的异构体功能，并检验一个新的中心假设，即ROCK1和ROCK2在调节心肌细胞死亡和心脏应激反应中的心脏重塑方面功能不同。特定目标1将检验ROCK2促进心肌细胞存活和心脏保护的假设。这些研究将进一步表征心脏特异性ROCK2基因敲除小鼠自发性心肌肥厚的发生和发展，并将确定心肌细胞中有条件的ROCK2缺失是否会加速心力衰竭的进展。具体目标2将决定ROCK1在心脏失代偿中的最终作用。这些研究将确定，当心肌肥厚或扩张性心肌病已经通过慢性压力超负荷发生时，心肌细胞中有条件的ROCK1缺失是否可以限制心力衰竭的进展。具体目标3将验证ROCK1和ROCK2在介导应激诱导的心肌细胞死亡中扮演相反角色的假设，并描述其潜在的机制。这些研究的结果将极大地促进我们对ROCK异构体病理生理学的了解，并为测试ROCK PAN抑制剂以及最终的异构体选择性抑制剂的临床试验提供信息，最终目标是开发防止心肌细胞死亡和减少心力衰竭进展的治疗干预措施。

DESCRIPTION (provided by applicant): Heart failure remains a leading cause of human morbidity and mortality. Rho kinase (also named ROCK) has recently emerged as a potential therapeutic target for the treatment of cardiac diseases with the overall promising studies showing beneficial effects of ROCK inhibitors in experimental and clinical studies. However, one important question needing to be addressed is whether ROCK truly represents a viable target for the treatment of human disease as currently available ROCK inhibitors have broad specificity. In addition, the two members of the ROCK family, ROCK1 and ROCK2, are inhibited by ROCK inhibitors with equal potency, and little is known about ROCK isoform functions in vivo. We recent discovered that systemic ROCK1 deficiency is protective against cardiac decompensation and the anti-apoptotic effect of ROCK1 deficiency is a critical contributor. In contrast to the beneficial effects of ROCK1 deletion, we observed that cardiac-specific ROCK2 deficiency results in spontaneous cardiac hypertrophy and dysfunction, suggesting a novel role for ROCK2 in cardiac protection. Our in vitro studies using ROCK1 or ROCK2 deficient embryo-derived fibroblasts support a novel mechanistic concept that ROCK1 preferentially mediates stress-induced acto-myosin contraction via the ROCK1/MYPT/MLC pathway leading to increased cell death, while ROCK2 preferentially contributes to actin polymerization via the ROCK2/LIMK/cofilin pathway leading to improved cell survival under stress conditions. The goal of this application is to dissect isoform functions of ROCK in hypertrophic cardiac remodeling and to test a novel central hypothesis that ROCK1 and ROCK2 are functionally different in regulating cardiomyocyte death and cardiac remodeling in response to cardiac stress. Specific Aim 1 will test the hypothesis that ROCK2 promotes cardiomyocyte survival and cardiac protection. The studies will further characterize the onset and progression of spontaneous cardiac hypertrophy in cardiac-specific ROCK2 knockout mice, and will determine if conditional ROCK2 deletion in cardiomyocytes accelerates heart failure progression. Specific Aim 2 will determine the ultimate role of ROCK1 in cardiac decompensation. The studies will determine if conditional ROCK1 deletion in cardiomyocytes can limit the progression of heart failure when cardiac hypertrophy or dilated cardiomyopathy has already occurred through chronic pressure overload. Specific Aim 3 will test the hypothesis that ROCK1 and ROCK2 play opposite roles in mediating stress-induced cardiomyocyte death and characterize the underlying mechanisms. Results of these studies will significantly advance our knowledge in ROCK isoform pathophysiology and inform clinical trials testing ROCK pan- inhibitors, and eventually isoform selective inhibitors, with the ultimate goal of developing therapeutic interventions to prevent cardiomyocyte death and reduce heart failure progression.