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RhoA-Rho激酶信号转导通路参与TGF-β1诱导的血管外膜成纤维细胞表型转化为肌成纤维细胞

陈闻东, 楚玉峰, 刘建军, 洪墨纳, 高平进

上海交通大学医学院附属瑞金医院，上海市高血压研究所，医学基因组学国家重点实验室，上海 200025；中国科学院上海生命科学研究院健康科学研究所血管生物学实验室，上海 200025；山东大学附属山东省立医院，济南 250021

摘要

血管外膜成纤维细胞表型转化为肌成纤维细胞是血管重塑的重要病理特征。本研究旨在探讨小分子G蛋白RhoA及其下游Rho激酶信号通路在转化生长因子β1 (transforming growth factor β1, TGF-β1)诱导的血管外膜成纤维细胞/肌成纤维细胞表型转化中的作用。用10 ng/mL TGF-β1诱导体外培养的大鼠胸主动脉外膜成纤维细胞表型转化为肌成纤维细胞，使用亲和沉淀法检测RhoA活性、使用免疫印迹检测RhoA、Rho激酶蛋白表达和Rho激酶活性；使用免疫印迹和免疫细胞化学检测肌成纤维细胞标记蛋白的表达。结果显示，TGF-β1上调体外培养的血管外膜成纤维细胞RhoA蛋白表达和RhoA活性。TGF-β1增加Rho激酶下游底物肌球蛋白磷酸酶目标亚单位的磷酸化，但不改变Rho激酶的蛋白表达，提示TGF-β1增加Rho激酶活性。腺病毒Ad-N19RhoA-hrGFP感染和Rho激酶特异性抑制剂Y27632都呈剂量依赖性地抑制TGF-β1诱导的肌成纤维细胞标记分子α平滑肌肌动蛋白和钙结合蛋白Calponin的蛋白表达。本研究证明RhoA-Rho激酶信号通路参与了TGF-β1诱导的血管外膜成纤维细胞/肌成纤维细胞表型转化。

关键词： 血管外膜成纤维细胞; 肌成纤维细胞; 转化生长因子β1; RhoA; Rho激酶

分类号：R331.3+2

RhoA-Rho kinase signaling pathway mediates adventitial fibroblasts differentiation to myofibroblasts induced by TGF-β1

Chen Wen-Dong, Chu Yu-Feng, Liu Jian-Jun, Hong Mo-Na, Gao Ping-Jin

State Key Laboratory of Medical Genomics and Shanghai Key Laboratory of Vascular Biology at Ruijin Hospital and Shanghai Institute of Hypertension, Shanghai Jiaotong University School of Medicine, Shanghai 200025, China； Laboratory of Vascular Biology, Institute of Health Science, Shanghai Institute for Biological Sciences (SIBS), Chinese Academy of Sciences (CAS), Shanghai 200025, China； Provincial Hospital Affiliated to Shandong University, Jinan 250021, China.

Abstract

Vascular adventitial fibroblasts (AF) differentiation to myofibroblasts (MF) is the critical physiopathologic feature of vascular remodeling. This study was to investigate the role of RhoA-Rho kinase signaling pathway in AF differentiation to MF induced by transforming growth factor β1 (TGF-β1). The results showed that TGF-β1 up-regulated total RhoA protein expression and RhoA activity in cultured AF by Western blotting and Rho pull-down assay, respectively. TGF-β1 up-regulated phospho-Myosin phosphatase target subunit (MYPT1, a downstream substrate of Rho kinase) expression without altering Rho kinase protein expression, indicating TGF-β1 induced the enhancement of activity of Rho kinase. Ad-N19RhoA-hrGFP virus infection and Y27632, a specific inhibitor of Rho kinase, dose-dependently inhibited TGF-β1-induced α-SM-actin and Calponin expression, as markers of MF differentiation. In conclusion, the RhoA-Rho kinase pathway is involved in AF differentiation to MF induced by TGF-β1.

Key words: adventitial fibroblast; myofibroblast; transforming growth factor β1; RhoA; Rho kinase

收稿日期：2012-07-31　　录用日期：2012-09-21

通讯作者：高平进　　E-mail: gaopingjin@yahoo.com.cn

引用本文：

陈闻东, 楚玉峰, 刘建军, 洪墨纳, 高平进. RhoA-Rho激酶信号转导通路参与TGF-β1诱导的血管外膜成纤维细胞表型转化为肌成纤维细胞[J]. 生理学报 2013; 65 (2): 113-121.

Chen Wen-Dong, Chu Yu-Feng, Liu Jian-Jun, Hong Mo-Na, Gao Ping-Jin . RhoA-Rho kinase signaling pathway mediates adventitial fibroblasts differentiation to myofibroblasts induced by TGF-β1. Acta Physiol Sin 2013; 65 (2): 113-121 (in Chinese with English abstract).