Clusterin在大肠癌细胞中的抗凋亡作用

ANTI-APOPTOSIS EFFECT OF CLUSTERIN IN LOVO CELLS

目的:探讨Clusterin及其不同功能区域在大肠癌细胞LOVO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(leader sequence)的Clusterin重组质粒,分别命名为pIRES2-EGFP/cluac、pIRES2-EGFP/clubc。重组质粒转染LOVO细胞后,以Western blot检测LOVO细胞及其培养上清液中Clusterin的表达。应用Na2SeO3诱导LOVO细胞凋亡,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LOVO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LOVO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。FCM及荧光显微镜检测证实,cluac转染对Na2SeO3诱导的LOVO细胞凋亡有明显的抑制作用。结论:Clusterin有明显的抗大肠癌细胞凋亡作用,Clusterin前导序列是其发挥抗凋亡作用的必需功能区域。

Objective：To evaluate the anti-apoptosis effect of clusterin with and without leader sequence in human colonic carcinoma cell line LOVO.Methods： The plasmid pIRES2-EGFP was used as carrier to construct clusterin recombinant with full-length or without the leader sequence （designated as pIRES2-EGFP/cluac, pIRES2-EGFP/clubc, respectively）. Western blot analysis was employed to compare with clusterin expression levels in the lysis and supernatant of clusterin transfected LOVO cells in vitro. The protective role of clusterin against Na2SeO3-induced apoptosis in LOVO cells was examined by flow cytometry （FCM） and fluorescence microscope. Results： Clusterin expression was detected in both the lysis and supernatant of pIRES2-EGFP/cluac transfected LOVO cells,while clusterin was only found in lysis liquid of pIRES2-EGFP/clubc transfected LOVO cells. Its anti-apoptotic property in LOVO cells was proved by FCM and fluorescence microscope. Conclusion： It is apparent that clusterin plays an important role in preventing apoptosis in colonic carcinoma cells, and the presence of the leader sequence is necessary for clusterin＇s anti-apoptotic function.