hsMAD2基因及Nocodazole对人肝癌细胞生长和细胞周期的影响  

Effects of hsMAD2 gene and Nocodazole on the growth and cell cycle of human primary hepatocellular carcinoma cells

作　　者：李擒龙[1] 王文亮[1] 张晓晖[1] 晏伟[1] 成胜权[2] 郭双平[1] 王文勇[1] 张志培[1] 王丽[1] 严晓昱[1] 张静[1] 

机构地区：[1]第四军医大学西京医院病理科,肿瘤生物学国家重点实验室,西安710033 [2]第四军医大学西京医院小儿科,西安710033

出　　处：《临床与实验病理学杂志》2006年第5期590-594,共5页Chinese Journal of Clinical and Experimental Pathology

基　　金：国家自然科学基金(No39900175)

摘　　要：目的观察hsMAD2过表达以及联合Nocodazole处理对HepG2细胞的生长和细胞周期的影响。方法将人全长hs-MAD2基因经脂质体包裹转染肝细胞肝癌HepG2细胞,经G418筛选期获得转基因癌细胞克隆,采用免疫细胞化学法、Westernblot方法检测hsMAD2的表达情况。观察细胞形态、测量生长曲线和分裂指数曲线,观察hsMAD2基因对HepG2细胞生长和细胞增殖能力的影响,流式细胞仪检测HepG2细胞周期的变化。然后观察在纺锤体阻滞剂50nmol/L Nocodazole作用下,hs-MAD2对HepG2细胞的影响。结果稳定表达外源性hsMAD2蛋白的HepG2细胞生长速度和细胞增殖能力并无变化;流式细胞仪检测结果显示HepG2-hsMAD2细胞周期G2、S期比例增加,而G1期细胞比例下降,但差异无显著性。经过nocodazole处理后,HepG2-hsMAD2细胞生长速度下降,差异具有显著性,细胞周期出现G2期阻滞。相反,HepG2在受到Nocodazole处理后无明显变化。结论单纯过表达外源性hsMAD2对细胞无影响。由于hsMAD2蛋白发挥了细胞周期关卡作用,使受到No-codazole破坏作用的细胞出现细胞周期的阻滞。Purpose To investigate the effects of over expression of hsMAD2 and Nocodazole on the cell growth and cell cycle of HepG2 cells. Methods A full human hsMAD2 gene was cloned into expression vector pcDNA3. 1 and transfected into human hepatocellular carcinoma cells HepG2 by lipofectamine,and the positive clone was screened by G418. The expression of hsMAD2 protein in HepG2 cells were examined by immunocytochemistry and Western blotting, and its effect on cell growth and cell cycle were observed by electron microscope and flow eytometer（FCM）. Finally,the effects on cell growth and cell cycle were detected after be treated with 50 nmol/L Nocodazole. Results There was no remarkable change in the growth rate and proliferation of HepG2-hsMAD2 cells. Cell cycle of HepG2-hsMAD2 cells was measured with Flow cytometry （FCM） results showed that ratio of G2 and S stage increased while ratio of G1 stage decreased. But there was no significant difference. It was revealed that there were no obvious effects on cells which have been transferred into hsMAD2 gene. Nocodazole＇s action on cells：the growth rate of HepG2-hsMAD2 which was treated with 50 nmol/L Nocodazole,a spindle retarder,dropped significantly and G2 stage block occurred,while there was no obvious change in HepG2 processed by Nocodazole. Therefore,it was shown that hsMAD2 played a role as checkpoint and induced a cycle block in the cell treated by No- codazole. Conclusions No obvious effects on the cells are be found which have been transferred into hsMAD2 gene. As to HepG2-hsMAD2 treated with Nocodazole,G2 stage block occurred,and cell＇s growth is inhibited,here hsMAD2 played a role as checkpoint.

关 键 词：肝肿瘤 hsMAD2基因 NOCODAZOLE 基因转染 细胞周期 

分 类 号：R735.7[医药卫生—肿瘤]