小RNA干扰技术沉默Smoothened(Smo)基因表达对胃癌MGC803细胞增殖及凋亡的影响  

Effect of knocking down Smoothened (Smo) expression by specific siRNA in human gastric carcinoma cells

作　　者：戎祯祥[1] 方驰华[1] 朱达坚[1] 刘胜军[1] 

机构地区：[1]南方医科大学附属珠江医院普通外科,广州510282

出　　处：《中华实验外科杂志》2007年第5期531-533,共3页Chinese Journal of Experimental Surgery

摘　　要：目的观察Smo基因在胃癌MGC803细胞中的表达,及其对胃癌MGC803细胞增殖与凋亡的作用。方法半定量逆转录-聚合酶链反应(RT-PCR)及Western blot检测胃癌MGC803细胞中Smo基因的表达;化学合成3条针对Smo mRNA的小干扰RNA(siRNA),阳性脂质体介导转染胃癌MGC803细胞,半定量RT-PCR筛选出降解胃癌MGC803细胞Smo mRNA最有效的siRNA;噻唑蓝(MTT)法和流式细胞仪检测胃癌MGC803细胞Smo mRNA被降解后,对细胞增殖和凋亡率的影响。结果胃癌MGC803细胞内有Smo蛋白及Smo mRNA的强表达,siRNA-1、siRNA-2对胃癌MGC803细胞Smo基因表达均有降解作用,siRNA-1降解作用最明显,达61.7%,与隐性对照组比较,差异有统计学意义(P<0.05)。siRNA-1转染胃癌MGC803细胞24h后,细胞增殖水平较隐性对照组明显下降(P<0.05);转染48h后,细胞凋亡率较隐性对照组明显上升(P<0.05)。结论胃癌MGC803细胞内存在Smo基因的高表达,降低Smo基因表达可抑制胃癌细胞增殖,诱导细胞凋亡,Smo基因具有调控胃癌细胞增殖和凋亡的作用。Objective To study the expression level of Smoothened （Smo） gene in MGC803 gastric cancer cells and the vital role of Smo gene in the proliferation and apoptosis of human gastric carcinoma cells. Methods Using Western blot and semi-quantitative RT-PCR techniques, the expression levels of Smo protein and mRNA in MGC803 gastric cancer cells was detected. Three small fragments of siRNA chemically synthesized in vitro against Smo mRNA were transfected into MGC 803 gastric cancer cells by positive liposomal transfection methods, and intracellular Smo mRNA expression was detected by RT-PCR, which could be interfered by siRNA. The influences on proliferation and apoptosis of gastric cancer MGC cells were studied by flow cytometry and MTr after intracellular Smo mRNA being degraded. Results There was overexpression of Smo protein and mRNA in MGC 803 cells. At 48 h after transfection, the expression level of Smo mRNA in MGC 803 cells of Smo siRNA-1 group was decreased by about 61.7% as compared with other groups. Meanwhile, after 24 h of transfection, the cell proliferation in Smo siRNA-1 group was decreased notably as compared with controls. After 48 h of transfection, the apoptosis rate in Smo siRNA-1 group was 23.6% ,significantly higher than in controls. Conduslon Growth of MGC 803 gastric cancer cells is related to the abnormal expression of Smo gene, Inhibition of Smo gene expression can inhibit proliferation and induce apoptosis in human gastric carcinoma cell- MGC803. Smo gene may play a key role in proliferation and apoptosis of human gastric carcinoma cells.

关 键 词：胃肿瘤 RNA干扰 脱噬作用 

分 类 号：R735.2[医药卫生—肿瘤]