Clusterin在前列腺癌LNCaP细胞中的抗凋亡作用  

Prevention of cell death in LNCaP cells by overexpression of clusterin

作　　者：辛殿祺[1] 朱绪辉[1] 艾军魁[1] 那彦群[1] 郭应禄[1] 

机构地区：[1]北京大学第一医院泌尿外科泌尿外科研究所,北京100034

出　　处：《北京大学学报（医学版）》2004年第2期154-158,共5页Journal of Peking University:Health Sciences

摘　　要：目的 :探讨Clusterin及其不同功能区域在前列腺癌细胞中的抗凋亡作用。方法 :以 pIRES2 EGFP质粒为载体 ,构建含有全长及缺失前导序列 (leadersequence)的Clusterin重组质粒 ,分别命名为 pIRES2 EGFP/cluac、pIRES2 EGFP/clubc。重组质粒转染前列腺癌细胞LNCaP后 ,以Westernblot检测LNCaP细胞及其培养上清液中Clusterin的表达 ,免疫细胞化学法检测不同功能区域Clusterin在细胞中的分布情况。应用Na2 SeO3 诱导LNCaP细胞凋亡 ,用流式细胞术 (FCM )及荧光显微镜检测Clusterin的抗凋亡作用。结果 :在转染 pIRES2 EGFP/cluac的LNCaP细胞培养上清及细胞裂解液中均检测到Clusterin的表达 ,转染 pIRES2 EGFP/clubc的LNCaP细胞 ,仅细胞裂解液中有Clusterin的表达 ,而培养上清液中未检测到。pIRES2 EGFP/cluac转染细胞中Clusterin在胞浆中呈聚集状态 ,而 pIRES2 EGFP/clubc转染细胞中Clusterin呈弥散分布。FCM及荧光显微镜检测证实 ,cluac转染对Na2 SeO3 诱导的LNCaP细胞凋亡有明显的抑制作用。结论 :Clusterin有明显的抗前列腺癌细胞凋亡作用 。Objective: To evaluate the effect of clusterin with and without leader sequence on overexpression preventing apoptosis in human prostate LNCaP cells. Methods:The plasmid pIRES2 EGFP was used to generate the clusterin expression constructs with full length or without the leader sequence (designated as pIRES2 EGFP/cluac, pIRES2 EGFP/clubc, respectively). Western blot analysis was employed to compare clusterin expression levels in the lysis and supernatant fluid of clusterin transfected LNCaP cells in vitro . The distribution of different functional domains of clusterin in cells was detected with Immunocytochemical staining. The clusterin's protective role of Na 2SeO 3 induced apoptosis in LNCaP cells was examined by flow cytometry (FCM) and fluorescence microscope. Results: Clusterin expression was detected in the lysis and supernatant fluid of pIRES2 EGFP/cluac transfected LNCaP cells, while clusterin was found only in lysis liquid of pIRES2 EGFP/clubc transfected LNCaP cells, but not found in their supernatant fluid. The distribution of cluserin in the plasm of pIRES2 EGFP/cluac transfected cells was aggregative, and on the other hand, clusterin distributed dispersedly in pIRES2 EGFP/clubc transfected cells. Its anti apoptotic property in LNCaP cells was proved by FCM and fluorescence microscope.Conclusion: It is apparent that clusterin plays an important role in preventing apoptosis in prostate cancer, and the presence of the leader sequence is necessary for clusterin's anti apoptotic function.

关 键 词：CLUSTERIN 前列腺癌 LNCAP细胞 细胞凋亡 免疫组织化学 流式细胞术 真核表达载体 

分 类 号：R730.23[医药卫生—肿瘤]