TGF-β1/ILK/FSP1信号通路在环孢素诱导肾小管上皮细胞转分化中的作用  

Role of TGF-β1/ILK/FSP1 signaling pathway in cyclosporin A-induced epithelial-mesenchymal transition in cultured renal tubular epithelial cells

作　　者：白志勋 陆静[2] 杨亦彬 BAI Zhixun;LU Jing;YANG Yibin(Department of Nephrology and Rheumatology, Second Affiliated Hospital of Zunyi Medical University, Zunyi 563000,China;Zunyi Medical and Pharmaceutical College, Zunyi 563006, China;Department of Nephrology, Affiliated Hospital of Zunyi Medical University, Zunyi 563006, China)

机构地区：[1]遵义医科大学第二附属医院肾病风湿科,贵州遵义563000 [2]遵义医药高等专科学校,贵州遵义563006 [3]遵义医科大学附属医院肾病风湿科,贵州遵义563006

出　　处：《南方医科大学学报》2019年第7期804-809,共6页Journal of Southern Medical University

基　　金：国家自然科学基金(81260118);贵州省教育厅自然科学基金(黔教科2010044);遵义市科技计划项目[遵市科合社字(2012)42号]~~

摘　　要：目的通过体外特异阻断试验,探讨转化生长因子-β1/整合素连接激酶/成纤维细胞特异蛋白1(TGF-β1/ILK/FSP1)信号通路在环孢素A致肾小管上皮细胞转分化中的作用。方法构建并鉴定ILK-RNAi慢病毒表达载体ILKshRNA。将NRK52E细胞培养分为5组,分别为空白对照组;环孢素诱导组:细胞培养液加环孢素A 1 mg/L;TGF-β1干预组:阻断剂(SB431542)10μmol/L加环孢素A 1 mg/L;ILK干预组:阳性转染ILKshRNA后加入环孢素A 1 mg/L;阴性对照组:阴性转染ILKshRNA后加入环孢素A 1 mg/L。实时荧光定量PCR检测TGF-β1、ILK和FSP-1 mRNA的表达,Western blot检测TGF-β1、ILK、FSP-1的蛋白表达。免疫组化检测细胞爬片α-SMA阳性表达细胞数。结果与空白对照组比较,环孢素诱导组细胞TGF-β1、ILK、FSP-1基因及蛋白表达量均显著增加(P<0.05);TGF-β1干预组经SB431542处理后,TGF-β1、ILK和FSP1水平较环孢素诱导组降低(P<0.05);经ILKshRNA沉默后,ILK和FSP1水平较环孢素诱导组降低(P<0.05)。α-SMA阳性细胞数经SB431542和ILKshRNA处理后较环孢素诱导组降低(P<0.05)。结论 TGF-β1/ILK/FSP1信号通路激活是环孢素A诱导大鼠肾小管上皮细胞转分化的重要机制之一,ILK参与环孢素A诱导大鼠肾小管上皮细胞间充质转分化过程。Objective To explore the role of transforming growth factor-β1/integrin- linked kinase/fibroblast-specific protein 1 (TGF-β1/ILK/FSP1) signaling pathway in cyclosporine A (CsA)-induced renal tubular epithelial cell transdifferentiation. Methods Rat renal tubular epithelial NRK-52E cells were induced with 1 mg/L CsA, treated with TGF-β1 inhibitor (SB431542, 10 μmol/L), or transfected with the ILK-RNAi lentiviral expression vector (ILKshRNA) or a negative control vector before CsA induction. The expressions of TGF-β1, ILK and FSP-1 mRNAs and proteins in the cells were detected using real-time PCR and Western blotting. The positive cells for α-SMA expression were detected by immunohistochemistry. Results Compared with the blank control cells, the cells treated with CsA showed significantly increased levels of TGF-β1, ILK and FSP-1 mRNAs and proteins (P<0.05). The expressions of TGF-β1, ILK and FSP-1 were significantly lower in TGF-β1 inhibitor group than in CsA group (P<0.05). The levels of ILK and FSP-1 were significantly decreased after shRNA-mediated ILK silencing (P<0.05). The number of positive cells for α-SMA was significantly lower in cells treated with SB431542 and in cells with ILK silencing than in the cells treated with CsA alone (P<0.05). Conclusion The activation of TGF-β1/ILK/FSP-1 signaling pathway is an important mechanism for CsA-induced transdifferentiation in rat renal tubular epithelial cells. ILK participates in CsA-induced epithelialmesenchymal transition of renal tubular epithelial cells.

关 键 词：环孢素肾病 上皮细胞转分化 转化生长因子-Β1 整合素连接激酶 成纤维细胞特异蛋白1 

分 类 号：R692[医药卫生—泌尿科学]