RESTENOSIS AND CONTROL OF VASCULAR MYOCYTE PROLIFERATION

再狭窄和血管心肌细胞增殖的控制

• 批准号: 2771550
• 负责人: VINCENTE ANDRES
• 金额: $ 12.1万
• 依托单位: STEWARD ST. ELIZABETH'S MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 1997
• 资助国家: 美国
• 起止时间: 1997-09-15 至 1999-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2771550
• 关键词: Adenoviridae; animal breeding; artery; atherosclerosis; atherosclerotic plaque; cell cycle; cell proliferation; cyclins; gene expression; genetic regulation; genetic regulatory element; injury; organ culture; restenosis; vascular smooth muscle

DESCRIPTION (Adapted from Investigator's Abstract): The broad objective of this proposal is to elucidate molecular mechanisms underlying the proliferative response of vascular smooth muscle cells (VSMC) to arterial injury, a process that is thought to contribute significantly to restenosis. Recent studies performed in my laboratory have identified positive regulators of cell growth that are induced at early time points following angioplasty in the rat carotid artery, including cdk2, cyclin E, cyclin A, cdk4 and cdc2. Our results show that overexpression of a cdk2 dominant-negative mutant inhibits entry of VSMCs into S-phase. We also have evidence suggesting that inhibition of cdk2 activity by cdk inhibitor p27 may contribute to the reestablishment of the quiescent state at late time points post-angioplasty. To begin to elucidate the mechanisms that control the expression cell cycle regulatory genes in VSMCs, we have demonstrated that elements within the -209 to +79 cyclin E and -79 to +100 cyclin A promoter regions confer cell cycle-dependent transcriptional control of cyclin gene expression in VSMCs. This Proposal represents an extension, as well as a departure from this previous work. The investigations outlined in Specific Aim 1 will further characterize the cell-cycle regulatory networks underlying injury-induced VSMC proliferation. The studies outlined in Specific Aim 2 will identify the cis-elements and cognate binding proteins underlying growth factor- and injury-induced regulation of cyclin E and cyclin A gene expression in VSMCs. Initially, regulatory sequences will be identified by transient transfection of reporter constructs in cultured VSMCs; subsequently, the relevance of cis-elements to injury-induced regulation of cyclin E and A gene expression will be tested in organ cultures of injured arteries. Investigations outlined in Specific Aim 3 will determine the relevance of p27 with respect to intimal thickening in animal models of vascular injury. These studies will include gain-of function experiments using an adenovirus vector directing the overproduction of p27, and loss-of function studies using p27-deficient mice. Finally, to assess the role of p27 on spontaneous atherosclerosis, the kinetics of atheromatous lesion formation will be examined in different genetic backgrounds obtained by crossbreeding p27 null mice and the atherosclerosis susceptible apoE-deficient mice. These studies should yield valuable mechanistic insights into the regulation of VSMC proliferation and may therefore provide a framework for novel therapeutic interventions.

描述(改编自《调查员摘要》)：的总体目标 这一建议是为了阐明 血管平滑肌细胞对动脉的增殖反应 损伤，这一过程被认为是导致再狭窄的重要原因。 最近在我的实验室进行的研究确定了阳性 在下列早期时间点诱导的细胞生长调节因子 大鼠颈动脉血管成形术，包括CDK2，Cyclin E，Cyclin A， CDK4和CDC2。我们的结果表明，CDK2的过度表达 显性负性突变体抑制VSMC进入S期。我们还有 证据表明，CDK抑制剂p27对CDK2活性的抑制 可能有助于在晚些时候重新建立静止状态 血管成形术后积分。开始阐明控制的机制 VSMC中细胞周期调控基因的表达，我们已经证明 -209到+79周期蛋白E和-79到+100周期蛋白A中的元素 启动子区域赋予细胞周期依赖的转录调控 细胞周期蛋白基因在血管平滑肌细胞中的表达这项提议是一种延伸，因为 以及与这项先前工作的背离。中概述的调查 具体目标1将进一步描述细胞周期调控网络的特征 潜在的损伤诱导的VSMC增殖。中概述的研究 特定目标2将确定顺式元件和同源结合蛋白 生长因子和损伤诱导的细胞周期蛋白E和 细胞周期蛋白A基因在血管平滑肌细胞中的表达最初，调控序列将是 瞬时转染培养细胞中报告基因的鉴定 VSMCs；随后，顺式元件与损伤诱导的相关性 将在器官中测试细胞周期蛋白E和A基因表达的调节 损伤动脉的培养。具体目标3中概述的调查 将确定p27与血管内膜增厚的相关性。 血管损伤的动物模型。这些研究将包括收益 用腺病毒载体引导过量生产的功能实验 以及使用p27缺陷小鼠进行的功能丧失研究。最后，为了 评估p27在自发性动脉粥样硬化中的作用，动力学 动脉粥样硬化性病变的形成将在不同的基因中进行检查 P27基因缺失小鼠杂交获得的背景与动脉粥样硬化 易感的载脂蛋白E缺陷小鼠。这些研究应该会产生有价值的 对VSMC增殖调控的机械论见解和可能 因此为新的治疗干预措施提供了一个框架。