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Analysis of the Beta2-chimaerin signaling in phenotypiC modulation of vascular smooth muscle cells

血管平滑肌细胞表型调节中 Beta2-chimaerin 信号传导的分析

基本信息

批准号：
13832007
负责人：
KATO Seiya
金额：
$ 2.3万
依托单位：
Kurume University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
2001
资助国家：
日本
起止时间：
2001 至 2003
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-13832007/
关键词：
smooth muscle, vascular beta2-chimaerin protein kinase C phenotypes atherosclerosis プロテインキナーゼ C

项目摘要

Phenotypic change of vascular smooth muscle cells (SMC) is an important aspect of atherogenesis. Activated cytokinegrowth factor network in the injured vessels subsequently induces a number of intracellular events resulting in the phenotypic modulation of SMC. Here, we investigated the role of beta2-chimaerin, a novel phorbol ester receptor with Rac GTPase-activating protein activity, in growth factor-stimulated SMC. Endogenous expression of beta2-chimaerin was detected in cultured human SMC by RT-PCR and immunohistochemistiy. Next, an overexpression of HA-tagged wild type human beta2-chimaerin was attempted in cultured rat SMC with a recombinant adenovirus (Adv-Ch). Confocal microscopy revealed 12-O-tetradecanoyl phorbol 13-acetate (TPA)-induced translocation of exogenous beta2-chimaerin from cytoplasm to plasma membrane in the presence of the PKC inhibitor (GF109203X). Proliferation of SMC stimulated by 10% FCS, bFGF (25ng/ml), and PDGF (10ng/ml) were inhibited by the infection with Adv-Ch (10-200MOI), but not with control viruses (Adv-LacZ), which was measured by cell counting and BrdU incorporation assays. PDGF-induced SMC migration was inhibited by the infection with Adv-Ch (200MOI) by about 25% in a modified Boyden chamber assay with the fibronectin-coated membrane. These data suggested that beta2-chimaerin might regulate proliferation and migration of SMC at the down-stream of the receptor tyrosine kinases. Beta2-chimaerin system may be involved in human atherogenesis, as is a potential therapeutic target.

血管平滑肌细胞（SMC）的表型改变是动脉粥样硬化发生的一个重要方面。受损血管中活化的细胞因子网络随后诱导了许多细胞内事件，导致SMC的表型调节。在这里，我们研究了β 2-嵌合蛋白在生长因子刺激的SMC中的作用，β 2-嵌合蛋白是一种具有Rac gtpase激活蛋白活性的新型磷酯受体。采用RT-PCR和免疫组织化学方法检测体外培养的人SMC内源性β -嵌合蛋白的表达。接下来，用重组腺病毒（Adv-Ch）在培养的大鼠SMC中过表达ha标记的野生型人β -嵌合蛋白。共聚焦显微镜显示，在PKC抑制剂（GF109203X）存在的情况下，12- o -十四烷醇phorbol 13-乙酸（TPA）诱导外源性β -嵌合蛋白从细胞质转移到质膜。通过细胞计数和BrdU掺入实验检测，10% FCS、bFGF （25ng/ml）和PDGF （10ng/ml）刺激的SMC增殖被Adv-Ch （10-200MOI）感染所抑制，而不被对照病毒（Adv-LacZ）所抑制。在改良的Boyden室实验中，pdgf诱导的SMC迁移被Adv-Ch （200MOI）感染抑制了约25%。这些数据提示β -嵌合蛋白可能在受体酪氨酸激酶下游调控SMC的增殖和迁移。β 2-嵌合蛋白系统可能参与人类动脉粥样硬化的形成，是一个潜在的治疗靶点。