大鼠系膜细胞增殖时S期激酶相关蛋白2表达的变化及机制  

Changes of Kinase-Associated Protein 2 During S Phase of Rat Mesangial Cells Proliferation

作　　者：刘书馨[1] 常明[1] 刘焱[1] 王志宏[1] 付瑶[1] 

机构地区：[1]辽宁省大连市中心医院肾内科,大连116033

出　　处：《中国中西医结合肾病杂志》2009年第7期595-598,共4页Chinese Journal of Integrated Traditional and Western Nephrology

基　　金：大连市科技局优秀青年科技人才基金资助项目(No.2008J23JH034)

摘　　要：目的:探讨系膜细胞增殖时细胞周期蛋白依赖性蛋白激酶抑制物p27kip1的降解限速蛋白——S期激酶相关蛋白2(Skp2)表达的变化及其机制。方法:原代培养的大鼠系膜细胞同步后分为无血清组、20%胎牛血清(FCS)刺激组。MTT法检测细胞增殖;实时定量PCR和Western Blot检测p27kip1和Skp2的mRNA、蛋白表达。结果:p27kip1在静止期系膜细胞高丰度表达,20%FCS刺激诱导细胞增殖后p27kip1蛋白表达显著下调;实时定量PCR研究结果显示p27kip1的mRNA表达水平差异无统计学差异,p27kip1蛋白和mRNA表达不一致。蛋白酶体抑制剂MG-132可显著改善系膜细胞增殖时p27kip1蛋白水平的下降。Skp2表达在静止期系膜细胞几乎检测不出,20%FCS刺激诱导细胞增殖后Skp2表达量显著上调。结论:系膜细胞Skp2表达显著上调导致p27kip1降解增加,可能是p27kip1蛋白水平下降、系膜细胞增殖的重要原因,为进一步阐明疾病状态下系膜细胞增殖的机制提供了新的研究靶点。Objective： To explore the changes of the rate - limiting enzyme in p27^kip1 degradation - S phase kinase - associated protein 2 during rat mesangial cells proliferation. Methods： Primary cultured rat mesangial cells were growth arrested by withdrawal FCS for 24 hours and then stimulated by 20 % FCS. Expression of p27^kip1 and Skp2 which played a crucial role in p27^kip1 degradation through ubiqutinproteasome proteolytic pathway were detected by western blot and real - time quantified PCR. Resuits： The p27^kip1 protein was highly expressed in quiescent mesangial cells and significantly decreased after 20% FCS stimulated. However, real time quantified PCR revealed no changes on the abundance of the p27^kip1 mRNA level during mesangial cells proliferation. Proteasome inhibitor MG 132 could significantly alleviated p27^kip1 protein decrease induced by FCS. Contrast to p27^kip1 protein, Skp2 was rarely detected in quiescent cells while dramatically increased after stimulation of 20 % FCS. Skp2 mRNA level was similar to protein. Conclusion：High expression of Skp2 may be the reason of low p27^kip1 protein level during mesangial cell prolifera- tion, and Skp2 will become a new study target for clarifying the mechanism of mesangial cell proliferation.

关 键 词：系膜细胞 细胞周期调节蛋白 S期激酶相关蛋白2 

分 类 号：R692.6[医药卫生—泌尿科学]