高糖环境对小鼠足细胞骨架结构及相关蛋白的影响

Effects of high glucose on podocyte cytoskeleton and cytoskeleton related proteins

目的探讨高糖培养环境对小鼠足细胞骨架结构及相关蛋白表达及分布的影响。方法体外培养和分化小鼠足细胞,分为正常对照组、高渗对照组和高糖组。免疫荧光法观察足细胞内肌动蛋白纤维骨架(F-actin)、黏着斑蛋白(vinculin)及α-actinin-4的表达及分布。Western blotting半定量检测各组足细胞α-actinin-4和vinculin蛋白的表达。结果正常对照组和高渗对照组足细胞内F-actin以张力纤维丝的形式存在,α-actinin-4主要分布在细胞周边、肌动蛋白微丝以及细胞膜皱褶中;与正常对照组和高渗对照组比较,高糖组足细胞内F-actin形成的张力纤维显著减少,α-actinin-4分布向细胞内部集中;各组vinculin的分布无明显差异。Western blotting检测结果显示:高糖组α-actinin-4蛋白的表达显著低于正常对照组(P<0.001),各组vinculin蛋白的表达量比较差异无统计学意义(P>0.05)。结论高糖可能刺激足细胞肌动蛋白细胞骨架结构重排,引起α-actinin-4的表达和分布改变。

Objective To investigate the effects of high glucose on F-actin cytoskeleton and expression and distribution of cytoskeleton related proteins of podocytes in mice. Methods Mouse podocytes cultured in vitro were divided into normal control group, high permeability control group and high glucose group. The expression and distribution of F-actin, c^-actinin-4 and vinculin in podocytes were determined by immunofluorescence staining. The expression of c^-actinin4 and vinculin protein in podocytes was detected by Western blotting. Results In normal control group and high permeability control group, F-actin existed in the form of tension fibers in podoeytes, and α-actinin-4 mainly located around cells and in actin microfilaments and membrane ruffles. However, F-actin tension fibers decreased, and α-actinin-4 moved into inner cytoplasm in high glucose group. There was no significant difference in the distribution of vinculin among groups. Western blotting revealed that the expression of α-actinin-4 protein in high glucose group was significantly lower than that in normal control group （P 〈 0. 001）, and there was no significant difference in the expression of vinculin protein among groups （P 〉0.05）. Conclusion High glucose may induce F-actin cytoskeleton reorganization and cytoplasmic translocation of α-actinin-4, and may suppress the expression of α-actinin-d in podocytes.