饰胶蛋白聚糖基因转染对大鼠肾系膜细胞转化生长因子βⅠ、βⅡ型受体表达的影响  

Influence on the expression of type I and type II transforming growth factor-β receptors in cultured rat mesangial cells transfected by decorin gene

作　　者：冯秀艳[1] 张志刚[1] 赵仲华[1] 陈琦[1] 郭慕依[1] 

机构地区：[1]复旦大学上海医学院病理学系,上海200032

出　　处：《中华肾脏病杂志》2005年第5期265-269,共5页Chinese Journal of Nephrology

基　　金：国家自然科学基金(30170431)

摘　　要：目的探讨饰胶蛋白聚糖(DCN)拮抗肾小球硬化进展的作用是否与其抑制系膜细胞(MsC)转化生长因子βⅠ、βⅡ型受体(TGF—βRⅠ、TGF—βRⅡ)表达有关。方法经外源性TGF—β1刺激培养的大鼠肾MsC,采用RT—PCR、Western印迹法检测其TGF—βRⅠ、TGF—βRⅡ的表达。用脂质体介导法将DCN基因转染MsC,经筛选和鉴定,用RT—PCR、Western印迹法和免疫荧光法分别观察DCN基因转染对TGF—βRⅠ、TGF—βRⅡ表达的影响。结果经外源性TGF—β1刺激的正常MsC,其TGF—βRⅠ、TGF—βRⅡ基因转录及蛋白表达均明显升高,且呈时间依赖性,于作用24h达高峰。与转染空载体的MsC(1P-1)相比,转染DCN基因的MsC克隆(3D-5,7D-1)的TGF-βRⅠmRNA表达分别为对照组的20%和32%,TCF-βRⅡmRNA表达分别为对照组的64%和59%;TGF—βRⅠ蛋白表达分别为对照组的34%和25%,TGF—βRⅡ蛋白表达分别为对照组的49%和57%。同时转染DCN基因也能阻止MsC对外源性TGF-β1刺激所致的两种受体表达升高作用。结论过表达DCN基因的MsC,其TGF-βRⅠ、TGF—βRⅡmRNA和蛋白表达水平均明显降低,这可能是DCN拮抗由TGF-β介导的肾小球硬化发生的重要机制之一。Objective To explore whether the antagonistic effect of decorin (DCN)on progression of glomeruloselerosis is associated with the inhibition of the expression of type I and type II transforming growth factor-β receptors (TGF-βR I and TGF-βR II )in mesangial cells (MsC). Methods RT-RCR and Western blot analysis were used to detect the expression of TGF-βR I and TGF-βR II mRNA and their proteins on cultured rat MsC stimulated by exogenous TGF-β1. Lipofectin-mediated method was used to transfect DCN vector into MsC. After screening and identifying of transfected MsC, RT-PCR and Western blot analysis were adapted to detect the changes of TGF-βR I and TGF-βR II expression respectively. Results The expression of TGF-βR I and TGF-βR II mRNA and their proteins on normal MsC stimulated by exogenous TGF-β1 increased in time-dependent manner and reached the peak at 24th hour. Compared with normal and untransfected MsC (1P-1), the mRNA and protein expression of TGF-βR I and TGF-βR II on MsC (3D-5, 7D-1) transfecled by DCN gene decreased significantly, and DCN gene transfection could antagonize the increase of mRNA and protein expression of both receptors caused by exogenous TGF-β1. Conclusions The expression of both TGF-βR I and TGF-βR II decreases obviously in MsC overexpressing DCN gene, which may be one of the importan antagonistic mechanisms of decorin involved in the development of glomeruloselerosis mediated by TGF-β.

关 键 词：饰胶蛋白聚糖 细胞转化生长因子β 受体表达 WESTERN印迹法 TGF-βRⅠ TGF-βRⅡ 大鼠 DCN基因转染 mRNA表达 肾小球硬化 蛋白表达 脂质体介导法 MsC 免疫荧光法 时间依赖性 对照组 外源性 系膜细胞 Ⅱ型受体 基因转录 

分 类 号：R346[医药卫生—基础医学]