Cell cycle activation for cardiac repair

描述（由申请人提供）：尽管成年哺乳动物心肌表现出有限的再生生长能力，但内在的更新速度不足以逆转病理生理性心肌细胞损失。因此，在受伤的心脏中重建失去的心脏质量的能力可能具有相当大的治疗价值。实现这一目标的一种方法是在存活的心肌细胞中诱导细胞周期活动。初步数据表明，cyclin D2的靶向表达足以诱导成人心脏的心肌细胞周期活性。cyclin D2是驱动转运通过G1/S细胞周期检查点的调节复合体的关键成员。此外，细胞周期蛋白d2诱导的细胞周期活性可以逆转心肌损伤后的结构损伤和恢复功能。本实验将进一步阐明cyclin d介导的心肌细胞周期调控机制。特异性目的1将验证d型细胞周期蛋白的翻译后修饰调节其介导心肌损伤后再生生长的能力的假设。这些实验将利用新产生的表达d型细胞周期蛋白的转基因小鼠，携带相关的模拟磷酸化和不可磷酸化的氨基酸残基取代。Aim 1中的其他研究将验证特定磷酸化事件介导d型细胞周期蛋白与p193/Cul7 E3泛素连接酶相互作用的假设，并且阻断这种相互作用可增强损伤心脏的细胞周期活性。在Specific Aim 2中提出的实验将进一步验证cyclin d2介导的心肌细胞周期激活可用于促进成人心脏心肌修复的假设。初始实验将利用条件转基因小鼠模型来确定细胞周期蛋白D2是否可以诱导成人心脏新生心肌细胞增殖。其他的研究被提出来确定限制不良损伤后重塑的药物干预能够在多大程度上有利于长期的心肌细胞周期诱导的再生。综上所述，这些研究将有助于确立d型细胞周期蛋白调控心肌细胞周期进入的机制，以及周期蛋白D2靶向表达对心肌再生的促进程度。总体目标是了解如何操纵细胞周期调节途径来促进损伤心脏的修复。这些分子靶点的鉴定可能最终导致药物的发展，以促进病变心脏的再生生长。

DESCRIPTION (provided by applicant): Although the adult mammalian myocardium exhibits a limited ability to undergo regenerative growth, the intrinsic renewal rate is insufficient to reverse pathophysiologic cardiomyocyte loss. The ability to reconstitute lost cardiac mass in injured hearts could thus be of considerable therapeutic value. One approach to accomplish this entails inducing cell cycle activity in the surviving cardiomyocytes. Preliminary data indicate that targeted expression of cyclin D2, a key member of the regulatory complex which drives transit through the G1/S cell cycle check-point, is sufficient to induce cardiomyocyte cell cycle activity in adult hearts. Moreover, cyclin D2-induced cell cycle activity can reverse structural damage and restore function following myocardial injury. The experiments proposed in this application will further elucidate the mechanism of cyclin D-mediated cardiomyocyte cell cycle regulation. Specific Aim 1 will test the hypothesis that post-translational modification of the D-type cyclins regulates their ability to mediate regenerative growth following myocardial injury. These experiments will utilize newly generated transgenic mice expressing D-type cyclins carrying the relevant phospho-mimetic and non-phosphorylatable amino acid residue substitutions. Other studies in Aim 1 will test the hypothesis that specific phosphorylation events mediate the interaction of D-type cyclins and the p193/Cul7 E3 ubiquitin ligase, and that blocking this interaction enhances cell cycle activity in injured hearts. Experiments proposed in Specific Aim 2 will further test the hypothesis that cyclin D2-mediated cardiomyocyte cell cycle activation can be used to promote myocardial repair in the adult heart. Initial experiments will utilize a conditional transgenic mouse model to determine if cyclin D2 can induce de novo cardiomyocyte proliferation in adult hearts. Other studies are proposed to determine the degree to which pharmacologic interventions which limit adverse post-injury remodeling are able to benefit long-term, cardiomyocyte cell cycle-induced regeneration. Collectively, these studies will help establish the mechanism by which D-type cyclins regulate cardiomyocyte cell cycle entry, and the degree to which targeted expression of cyclin D2 is able to promote myocardial regeneration. The overall goal is to gain an understanding of how cell cycle regulatory pathways can be manipulation to promote the repair of injured hearts. Identification of such molecular targets may ultimately lead to the development of pharmacologic agents to promote regenerative growth in diseased hearts.