Controlling VSMC Proliferation and Migration

控制 VSMC 增殖和迁移

基本信息

批准号：
9059320
负责人：
MING-HUI ZOU
金额：
$ 40.94万
依托单位：
GEORGIA STATE UNIVERSITY
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 2016-12-14
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9059320
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 genetic approach 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.

描述（由申请人提供）：血管平滑肌细胞（VSMC）增殖和迁移是心脏移植后冠状动脉内部再狭窄和加速动脉炎的主要原因。在过去十年中，可以防止VSMC增殖，迁移和随之而来的再狭窄。我们令人兴奋的初步数据表明，AMP激活蛋白激酶（AMPK）的药理或遗传激活能够抑制VSMC增殖和体内新内膜增生。对小鼠VSMC的荧光激活细胞分选（FACS）分析表明，AMPKA2的损失增加了VSMC从G1到S相的过渡。与这一发现一致的是，在AMPKA2-KNOCK OUT（KO）小鼠VSMC中，细胞周期抑制剂P27KIP1（P27）急剧下调，而不是AMPKA1-KO VSMC。此外，我们发现P27KIP1放松管制不是归因于P27KIP1 mRNA水平，而是由于SKP2表达高，这是泛素E3连接酶通过SKP2启动子中的STAT结合的亚基。从机械上讲，我们发现S期激酶相关蛋白2（SKP2）是P27的E3泛素连接酶，在AMPKA2-KO VSMC中升高，并负责增加P27的降解。 AMPKA2-KO小鼠中丝损伤引起的颈动脉动脉中线损伤诱导的新内膜增生的抑制作用和随之而来的最终证据的证据要比AMPKA1-KO或野生型动物（WT）动物明显大于AMPKA2-KO小鼠。因此，该应用的中心假设是AMPKA2的损失增加了SKP2，P27的E3连接酶以及SKP2介导的p27降解以产生异常的VSMC增殖和迁移，对新内在增生和再生病的发展中的关键事件。该假设将以三个具体的目的进行检验：目标＃1是确定P27在异常VSMC增殖和由AMPKA2失活引起的迁移中的核心作用。 AIM＃2是确定AMPKA2缺失的SKP2是否以及如何在AMPKA2-KO VSMC中导致P27降解并增强细胞增殖和迁移。在最后一个目标中，我们将在体内新内膜增生中确立SKP2和P27的核心作用。体外和体内技术，功能丧失和药理学/遗传方法的结合将用于实现研究目标。该项目的完成将为AMPK，P27和SKP2提供新的见解，是针对与包括糖尿病，再狭窄，动脉粥样硬化和癌症在内的常见疾病相关的血管损害的潜在治疗靶标。