针对CTGF的siRNA对高糖诱导小鼠足细胞nephrin、podocin减少的影响  

Effect of siRNA Targeting CTGF on the Decreased Expression of Nephrin and Podocin Induced by High Glucose in Mouse Podocyte

作　　者：杨蕾[1] 汤珣[1] 李萍华[1] 杜庆生[1] 郭婷婷[1] 章俊[1] 

机构地区：[1]南方医科大学珠江医院肾内科,广州510280

出　　处：《中国医科大学学报》2011年第6期512-515,共4页Journal of China Medical University

基　　金：广东省科技计划项目(2007B031503001);广东省医学科学技术研究基金(B2008115)

摘　　要：目的探讨结缔组织生长因子(CTGF)在高糖诱导小鼠足细胞nephrin、podocin表达减少中的作用,及针对CTGF基因的siRNA对其影响。方法将体外培养的小鼠足细胞分为6组:(1)正常对照组,1640培养基含D-葡萄糖1 g/L;(2)等渗对照组,1640培养基含D-葡萄糖1g/L,甘露醇3.5 g/L;(3)高糖组,1640培养基含D-葡萄糖4.5 g/L;(4)高糖+空白对照组:细胞转染空白质粒后于含D-葡萄糖4.5 g/L中的1640培养基中培养;(5)高糖+阴性对照组:细胞转染含无关序列的重组质粒后于含D-葡萄糖4.5 g/L中的1640培养基中培养;(6)高糖+干扰组:细胞转染针对CTGF的siRNA表达质粒后于含D-葡萄糖4.5 g/L中的1640培养基中培养。应用Realtime PCR检测nephrin、podocin、CTGF mRNA水平,Western blot检测CTGF、nephrin、podocin蛋白表达水平。结果高糖可诱导CTGF mRNA及蛋白表达增加,下调足细胞的nephrin、podocin mRNA及蛋白水平,而通过特异性siRNA抑制CTGF mRNA表达后,CTGF表达减少,同时伴有细胞nephrin、podocin表达升高。结论 CTGF是高糖诱导小鼠足细胞损伤的重要介质,高糖可通过CTGF诱导nephrin,podocin表达减少。针对CTGF的siRNA能明显改善这种损伤,为DN发病机制的研究提供新的实验依据。Objective To study the effect of connective tissue growth factor （CTGF） on the expression of nephrin and podocin in mouse potocyte induced by high glucose in vitro, and the role of small interfering RNA （siRNA） targeting CTGF. Methods Cultured mouse poto- cytes were divided into 6 groups： （ 1 ） normal control group：RPMI1640 medium containing 1 g/L glucose; （2） iso-osmolar control group： RPMI1640＋ 1 g/L glucose ＋ 3.5 g/L mannitol; （3） high glucose group： ,RPMI1640＋4.5 g/L glucose; The cells were transfected with pGenesil-1 ,pGenesil/neg,or pGenesil/siRNA-CTGF and then cultured in RPMI1640 medium containing 4.5 g/L glucose as the （4） high glucose ＋ blank control group, （5） high glucose ＋negative control group and （6） high glucose＋ interference group, respectively. Real-time PCR and Western blotting were apphed respectively to detect the mRNA levels and the protein levels of CTGF, nephrin and podocin. Results High glucose induced the expression of CTGF mRNA and protein, but down regulated mRNA and protein of nephrin and podocin. Transfection with siRNA targeting CTGF significantly decreased the expression of CTGF, and this was accompanied by increase in the expression of nephrin and podocin. Conclusion CTGF is an important mediator of high glucose-induced potocyte damage. High glucose decreases the level of nephrin and podocin through CTGF. Transfeetion with siRNA targeting CTGF can alleviate the damage, which provides new experimental basis for the research of DN pathogenesis.

关 键 词：结缔组织生长因子 小分子干扰RNA 糖尿病肾病 足细胞 

分 类 号：R587.1[医药卫生—内分泌]