法舒地尔对血管紧张素Ⅱ诱导的足细胞自噬的影响  

Fasudil,a Rho kinase inhibitor,inhibits angiotensinⅡ-induced autophagy in podocytes

作　　者：高雪静[1] 唐莎[1] 尹仕伟[1] 张莹[1] 谢京[1] 施维维[1] 王莉[2] 邹丽云[2] 牟芝蓉[2] 张静波[1] 

机构地区：[1]第三军医大学新桥医院肾内科,重庆400037 [2] 第三军医大学全军免疫研究所

出　　处：《肾脏病与透析肾移植杂志》2014年第1期42-47,共6页Chinese Journal of Nephrology,Dialysis & Transplantation

基　　金：国家自然科学基金(30971366);重庆市国际合作项目(CSTC201110004);第三军医大学临床科研基金(2011XLC37)

摘　　要：目的：探讨Rho激酶抑制剂法舒地尔对血管紧张素Ⅱ（AngⅡ）诱导人足细胞（AB 8/13）自噬的影响. 方法：体外培养人足细胞株,采用不同浓度的AngⅡ（10-8～10-6 mol/L）刺激人足细胞24h和固定浓度AngⅡ（10-7moVL）刺激不同时间（0h、6h、12h、24h、36h）,以及特异性自噬阻断剂3-MA（5 mmol/L）和不同浓度（10-8～1O-6mol/L）法舒地尔进行干预.免疫荧光法和Westem印迹法检测自噬相关蛋白[微管相关蛋白1轻链3B（LC3B）、Beclin-1]的表达变化. 结果：AngⅡ可刺激足细胞LC3B、Beclin-1蛋白表达上调,且呈剂量、时间依赖性（P＜0.05）,在浓度10-7mol/L AngⅡ和时间24h时达高峰.3-甲基腺嘌呤（3-MA）能明显降低AngⅡ引起的LC3B蛋白表达（P＜0.01）.法舒地尔本身对足细胞自噬没有影响,但可以明显下调AngⅡ诱导的足细胞LC3B和Beclin-1蛋白表达（P＜0.05）,10-7mol/L时效果最明显. 结论：AngⅡ可诱导人足细胞自噬增强,3-MA和法舒地尔均可抑制此过程的发生,提示Rho激酶途径可能参与了AngⅡ诱导的足细胞自噬过程.Objective：To investigate the effect of fasudil （a specific Rho-kinase inhibitor） on angiotensin （Ang） Ⅱ-induced autophagy in podocytes. Methodology：The conditionally immortalized human podocytes （AB8/13） were incubated with various concentrations of Ang Ⅱ （ 10-s to 10-6 mol/L） for 2gh or with 10-7 mol/L Ang Ⅱ for different times （0, 6, 12, 24 and 36 hours）, together with or without autophagy inhibitor 3-methyladenine （3-MA） and various concentrations of fasudil （10-8 to 10-6 mol/L） in vitro. The protein expression of LC3B and Beclin-1 were detected by immunofluorescence and western blotting. Results： The proteins expression of LC3B and Beclin-1 were up-regulated significantly by Ang Ⅱ in a dose- and time- dependent manner. The optimal concentration of Ang Ⅱ was 10-7 mol/L and the optimal time was at 24h in which Ang Ⅱ enhanced LC3B and Beclin-1 protein expression to the maximum. The stimulation of podocytes with Ang Ⅱ increased LC3B and Beclin-1 expression that was prevented by 3-MA. The expression enhancement of LC3B and Beclin-1 by Ang Ⅱ were reduced significantly by fasudil in a dose- dependent manner,although fasudil alone does not induce autophagy. The optimal inhibitory concentration of fasudial was 10-7 mol/L. Conclusion： The autophagy of podocytes can be induced by Ang Ⅱ and inhibited by fasudil, which suggests that Rho-kinase pathway may mediate Ang Ⅱ induced the autophagy of podocytes.

关 键 词：血管紧张素Ⅱ 自噬 RHO激酶 足细胞 3-甲基腺嘌呤 ANGIOTENSIN Ⅱ 

分 类 号：R692[医药卫生—泌尿科学]