Nephrin分子细胞内吞转运途径的研究  

A Study on Endocytosis Pathway of Nephrin

作　　者：秦晓松[1] 刘勇[1] 吴丽娜[1] 刘建华[1] 王丹丹[1] 

机构地区：[1]中国医科大学附属盛京医院检验科,沈阳110004

出　　处：《中国细胞生物学学报》2010年第4期527-532,共6页Chinese Journal of Cell Biology

基　　金：辽宁省自然科学基金(No.20082095);辽宁省高校科研计划(No.L20100610)资助项目~~

摘　　要：为研究nephrin分子在细胞内的转运途径,探讨其在维持肾小球裂孔膜完整性上的作用,构建了nephrin真核表达载体,并转染至COS-7细胞内,应用免疫荧光三重标记的方法,分别进行细胞内及细胞表面的荧光标记,联合笼型蛋白介导的内吞(clathrin-mediated endocytosis,CME)和脂筏介导的内吞(raft-mediated endocytosis,RME)标记物,通过共聚焦显微镜对裂隙膜分子nephrin在不同时间点细胞内的内吞转运特点进行研究。结果发现在转运启动2 min时,68.44%±4.65%的nephrin通过CME途径进行细胞内转运;在20 min时,65.24%±4.02%的nephrin以RME途径进行转运,并且两种转运途径均可被相关途径抑制物所阻断。表明nephrin通过两种途径进行细胞内转运,提示不同的转运途径可能与其实现不同功能有关。To study the intracellular endocytic pathway of nephrin and the possible effects on maintaining the integrate of slit diaphgram. An eukaryotic expression vector of nephrin was constructed and was transfeeted into COS-7 cells. Triple immunofluorescence was used to stain membrane and plasma expressing nephrin molecules. By using markers of clathrin- and raft-mediated pathways, a laser scanning confocal microscopy was applied to study the endocytic character of nephrin in different time points. 68.44%±4.6% of nephrin was endocytosed via clathrin pathway at 2min, 65.24%±4.02% of nephrin was endocytosed via a raft associated pathway at 20 min. Both routes can be inhibited by the relative inhibitors. The data indicated that both CME and RME pathways were involved in nephrin endocytosis suggesting that nephrin perform its function via distinct pathways.

关 键 词：NEPHRIN 免疫荧光技术 蛋白质转运 

分 类 号：R329[医药卫生—人体解剖和组织胚胎学]