Controlling VSMC Proliferation and Migration

控制 VSMC 增殖和迁移

基本信息

批准号：
8203252
负责人：
MING-HUI ZOU
金额：
$ 40.87万
依托单位：
UNIVERSITY OF OKLAHOMA HLTH SCIENCES CTR
依托单位国家：
美国
项目类别：
财政年份：
2011
资助国家：
美国
起止时间：
2011-07-15 至 2016-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8203252
关键词：
5&apos -AMP-activated protein kinase AMP-activated protein kinase kinase Abbreviations Animals Arterial Disorder Atherosclerosis Binding Binding Proteins Biological Assay Blood Vessels Bone Marrow Bone Marrow Transplantation CDK6-associated protein p18 Cardiovascular Diseases Carotid Arteries Cell Cycle Cell Proliferation Cells Clinical Coronary Coronary artery Cyclins Data Development Diabetes Mellitus Disease Event Fluorescence-Activated Cell Sorting Genetic Goals Heart Transplantation Hyperplasia Immigration Immunohistochemistry In Vitro Injury Knock-out Knockout Mice Life Malignant Neoplasms Medial Mediating Messenger RNA Molecular Mus NF-kappa B Pathway interactions Peripheral Vascular Diseases Procedures Proliferating Proliferating Cell Nuclear Antigen Research Ribonucleosides Role S Phase SKP2 gene Serum Skp1-Cullin-F-Box Proteins Skp2 Proteins Smooth Muscle Myocytes Stents Techniques Testing Therapeutic Ubiquitin Up-Regulation cell motility fetal in vivo inhibitor/antagonist insight loss of function migration multicatalytic endopeptidase complex neointima formation novel prevent promoter restenosis sensor therapeutic target tumor ubiquitin-protein ligase vascular smooth muscle cell migration vascular smooth muscle cell proliferation

项目摘要

DESCRIPTION (provided by applicant): Vascular smooth muscle cell (VSMC) proliferation and migration are the major causes of coronary artery in- stent restenosis and accelerated arteriopathy following cardiac transplantation. How VSMC proliferation, migration, and consequent restenosis can be prevented in vivo remains a subject of extensive research in the last decade. Our exciting preliminary data suggest that pharmacological or genetic activation of AMP-activated protein kinase (AMPK) is able to suppress VSMC proliferation and neointimal hyperplasia in vivo. Fluorescence-activated cell sorting (FACS) analysis of VSMC from mice revealed that loss of AMPKa2 increased VSMC transition from G1 to S phase. Consistent with this finding, the cell cycle inhibitor, p27Kip1 (p27), was dramatically down-regulated in AMPKa2-knock out (KO) mouse VSMC but not AMPKa1-KO VSMC. In addition, we found that p27Kip1 deregulation was not due to p27Kip1 mRNA level but due to high Skp2 expression, a subunit of ubiquitin E3 ligase through the STAT binding in the Skp2 promoter. Mechanistically, we found that the S-phase kinase-associated protein 2 (Skp2), an E3 ubiquitin ligase for p27, was elevated in AMPKa2-KO VSMC and was responsible for increased degradation of p27. The most conclusive evidence for AMPK-dependent inhibition of VSMC proliferation and consequent restenosis was that wire injury-induced neointima hyperplasia in the carotid artery was significantly greater in AMPKa2-KO mice than in either AMPKa1-KO or wild type (WT) animals. Thus, the central hypothesis of this application is that loss of AMPKa2 increases Skp2, an E3 ligase for p27, and Skp2-mediated degradation of p27 to produce aberrant VSMC proliferation and migration, critical events in the development of neointimal hyperplasia and restenosis. This hypothesis will be tested in three specific aims: Aim #1 is to establish the central roles of p27 in aberrant VSMC proliferation and migration caused by AMPKa2 inactivation. Aim #2 is to determine if and how Skp2 up-regulation by AMPKa2 deletion causes p27 degradation and enhanced cell proliferation and migration in AMPKa2-KO VSMC. In the last Aim, we will establish a central role for Skp2 and p27 in neointimal hyperplasia in vivo. A combination of in vitro and in vivo techniques, gain-/loss-of-function, and pharmacologic/genetic approaches will used to accomplish the study objectives. The completion of this project will provide novel insights into whether AMPK, p27, and Skp2, are potential therapeutic targets for countering vascular damage associated with common diseases including diabetes, restenosis, atherosclerosis, and cancer. PUBLIC HEALTH RELEVANCE: Aberrant vascular smooth muscle cell (VSMC) proliferation and migration contribute to cardiovascular diseases and are the major causes of coronary artery in-stent restenosis and accelerated arteriopathy following cardiac transplantation. The goal of the proposed studies is to establish a central role for AMPK in Skp2- mediated p27 turnover, VSMC proliferation, VSMC migration, and resultant restenosis. Thus, our studies will provide novel insights into whether AMPK and Skp2 are potential therapeutic targets for neointimal hyperplasia and restenosis which are associated with common diseases including diabetes, restenosis, atherosclerosis, and cancer.

描述（申请人提供）：血管平滑肌细胞（VSMC）增殖和迁移是心脏移植后冠状动脉支架内再狭窄和加速动脉病变的主要原因。如何在体内预防 VSMC 增殖、迁移和随后的再狭窄仍然是过去十年中广泛研究的主题。我们令人兴奋的初步数据表明，AMP 激活蛋白激酶 (AMPK) 的药理学或基因激活能够抑制体内 VSMC 增殖和新内膜增生。对小鼠 VSMC 的荧光激活细胞分选 (FACS) 分析表明，AMPKa2 的缺失会增加 VSMC 从 G1 期向 S 期的转变。与这一发现一致的是，细胞周期抑制剂 p27Kip1 (p27) 在 AMPKa2 敲除 (KO) 小鼠 VSMC 中显着下调，但在 AMPKa1-KO VSMC 中则不然。此外，我们发现 p27Kip1 失调并不是由于 p27Kip1 mRNA 水平，而是由于 Skp2 的高表达，这是泛素 E3 连接酶的亚基，通过 Skp2 启动子中的 STAT 结合。从机制上讲，我们发现 S 期激酶相关蛋白 2 (Skp2)（一种 p27 的 E3 泛素连接酶）在 AMPKa2-KO VSMC 中升高，并导致 p27 降解增加。 AMPK 依赖性抑制 VSMC 增殖和随后的再狭窄的最确凿证据是，AMPKa2-KO 小鼠中线损伤诱导的颈动脉新内膜增生明显大于 AMPKa1-KO 或野生型 (WT) 动物。因此，本申请的中心假设是，AMPKa2 的缺失会增加 Skp2（p27 的一种 E3 连接酶）和 Skp2 介导的 p27 降解，从而产生异常的 VSMC 增殖和迁移，这是新内膜增生和再狭窄发展中的关键事件。该假设将在三个具体目标中得到检验：目标#1 是确定 p27 在 AMPKa2 失活引起的异常 VSMC 增殖和迁移中的核心作用。目标 #2 是确定 AMPKa2 缺失导致的 Skp2 上调是否以及如何导致 p27 降解以及 AMPKa2-KO VSMC 中细胞增殖和迁移的增强。在最后一个目标中，我们将确定 Skp2 和 p27 在体内新生内膜增生中的核心作用。将结合体外和体内技术、功能获得/丧失以及药理学/遗传学方法来实现研究目标。该项目的完成将为 AMPK、p27 和 Skp2 是否是对抗与糖尿病、再狭窄、动脉粥样硬化和癌症等常见疾病相关的血管损伤的潜在治疗靶点提供新的见解。公众健康相关性：异常的血管平滑肌细胞 (VSMC) 增殖和迁移会导致心血管疾病，也是心脏移植后冠状动脉支架内再狭窄和加速动脉病变的主要原因。拟议研究的目标是确定 AMPK 在 Skp2 介导的 p27 更新、VSMC 增殖、VSMC 迁移和由此产生的再狭窄中的核心作用。因此，我们的研究将为 AMPK 和 Skp2 是否是与糖尿病、再狭窄、动脉粥样硬化和癌症等常见疾病相关的新内膜增生和再狭窄的潜在治疗靶点提供新的见解。