pEGFP—survivin对GBC—SD细胞生长的抑制及对化疗敏感性的影响  

Effect of survivin shRNA on chemotherapy resistance in GBC-SD cells

作　　者：沈汉斌 吴耀辉[2] 龙浩成 郑启昌[2] 龚建平[3] 

机构地区：[1]武汉市第五医院普外一科,430050 [2]华中科技大学同济医学院附属协和医院外科 [3]华中科技大学同济医学院附属同济医院外科

出　　处：《中华肝胆外科杂志》2010年第1期43-47,共5页Chinese Journal of Hepatobiliary Surgery

摘　　要：目的观察pEGFP—survivin对GBC—SD细胞化疗敏感性的影响。方法以MTT法检测GBC-SD、GBC—SD／EGFP和GBC—SD／survivin三种细胞的增殖活性；用RT—PCR和Western印迹、检测各组细胞中survivin mRNA和蛋白质水平的表达；以合适浓度的DDP作用相同的时间后，用MTT法检测3种细胞存活率，流式细胞仪测细胞凋亡，并用MTT法筛选IC50值，TUNEL法观察细胞核改变；另外检测DDP作用后各组细胞的caspase-3蛋白酶活性的变化。结果GBC—SD和GBC—SD／EGFP细胞的增殖活性大致相同，而GBC-SD／survivin细胞的增殖活性则明显下降；RT—PCR和Western印迹均发现GBC—SD／survivin细胞中的survivin表达水平较其余两种细胞明显下降；在给与DDP作用后，GBC-SD／survivin细胞存活率和IC50明显较低，细胞凋亡率较高，而3种细胞用TUNEL法染色后均可见棕色凋亡细胞核。给与DDP作用后，各组细胞caspase-3活性均呈现先升高后下降的趋势，但GBC-SD／survivin细胞中caspase-3的活性明显高于另外两种细胞。结论pEGFP-survivin表达的survivin shRNA能明显降低GBC—SD细胞中survivin的表达，提高对化疗药物的敏感性。Objective To investingate the effect of survivin shRNA on chemotherapy resistance in GBC-SD cells. Methods They were divided into three groups of GBC-SD, GBC-SD/EGFP and GBC-SD/survivin. MTT assay was used to detect cell viability in the 3groups. mRNA and protein of survivin were tested by RT-PCR and Western blot. Then the cells were treated with proper construction DDP. The cell survival rate and IC50 were determined with MTT, cell apoptosis detected by FACS and the alteration of nucleus observed by TUNEL. Meanwhile, caspase-3 activity was determined u- sing the colorimetric method. Results Cell viability was reduced remarkably in GBC-SD/survivin and survivin expression was decreased obviously. After being treated with DDP, cell survival rate and IC60 were decreased obviously in GBC-SD/survivin, apoptotic rate elevated remarkably compared with other groups. There were brownly nucleuses in three groups. Caspase-3 activity increased first and then decreased, but it exceeded in GBC-SD/survivin than that in other two groups. Conclusion The survivin shRNA can down-regulate the expression of survivin in GBC-SD cells remarkably and improve the sensibility to chemotherapy.

关 键 词：化疗药物 SURVIVIN 短发夹环RNA GBC—SD 

分 类 号：R730.5[医药卫生—肿瘤]