肾小球系膜细胞转化生长因子-β_1基因启动子中氧化低密度脂蛋白应答元件的研究  

The cic-element response to oxidized low density lipoprotein of TGF-β_1 promoter in mesangial cell

作　　者：周钦[1] 王远程[2] 兰洋[1] 赵昀[3] 陆长德[3] 吴兆龙[1] 

机构地区：[1]复旦大学中山医院肾内科,上海200032 [2]解放军广州军区武汉总医院肾内科 [3]中国科学院上海生化与细胞研究所

出　　处：《肾脏病与透析肾移植杂志》2002年第3期232-235,共4页Chinese Journal of Nephrology,Dialysis & Transplantation

基　　金：复旦大学首届创新基金暨优秀博士论文培育基金;国家自然科学基金课题资助 (编号 39970 343)

摘　　要：目的 :了解在氧化低密度脂蛋白 (Ox LDL)刺激下 ,转化生长因子 β1(TGF β1)基因启动子中顺式作用元件的应答情况。研究在大鼠肾系膜细胞中 ,Ox LDL对TGF β1基因启动子转录活性的影响作用。　 方法 :用聚合酶链反应 (PCR)、酶切和亚克隆的方法 ,构建出 5个TGF β1启动子缺失体—虫荧光素酶 (luciferase)报告基因 ,利用其瞬时转染系膜细胞并检测luciferase相对活性。　 结果 :在Ox LDL(10 0mg/L)刺激下 ,luciferase的活性 12h即出现 ,并于 36h达到平台期。对Ox LDL刺激的应答 ,TGF β1启动子 15 5 0到 845之间可能存在着增强子 , 6 2 9到 4 2 2之间则可能有抑制子。计算机软件分析表明 ,前者内有一转录激活蛋白 1(AP 1)精确识别位点。　 结论 :在大鼠肾系膜细胞TGF β1基因启动子中 ,AP 1结合序列 (TGAGTCA)可能是应答Ox LDL的顺式作用元件。Ox LDL上调TGF β1的表达至少部分是通过AP 1蛋白来介导的 ,且Ox LDL可能通过蛋白激酶C(PKC)信号通路激活AP 1从而正调控TGF β1基因的转录过程。Objective:To investigate the changes of TGF-β 1 promoter activity in mesangial cell stimulated with oxidized low density lipoprotein(Ox-LDL),and to illustrate the possible cis-elements involved. Methodology:A deletion analysis of TGF-β 1 promoter was applied in SD rat mesangial cells to identify the putative regulatory elements.These deletions were tested for responsiveness to Ox-LDL by fusing them with the luciferase gene,and by measuring the relative enzymatic activity of luciferase after transient transfection of these constructions into SD rat renal mesangial cells. Results:With the stimulation of Ox-LDL,luciferase activity began to rise at the 12th hour and reached the maximum at the 36th hour.Measurement of relative luciferase activity showed that there were two regions that responded to Ox-LDL stimulation in rat mesangial cell.Possibilities existed that enhancer elements locate in the first regions(-1550 to -845,a region containing an accurate activator protein 1 binding sites,AP-1) and inhibitor elements locate in the latter region (-629 to -422). Conclusion:The positive transcriptional regulation of rat TGF-β 1 gene induced by Ox-LDL is probably mediated at least by AP1 transcription.

关 键 词：肾小球系膜细胞 氧化低密度脂蛋白 转化生长因子-Β1 虫荧光素酶报告基因 

分 类 号：R692.02[医药卫生—泌尿科学]