miR29c下调S100A4蛋白表达参与人胃癌细胞增殖促进凋亡

MiR-29c downregulated and regulated cell proliferation and apoptosis by suppression S100A4 in human gastric carcinoma cell lines

目的探讨miR-29c对人胃腺癌细胞增殖、凋亡的影响及其与转移相关蛋白S100A4的关系。方法采用细胞培养人胃癌细胞系BGC-823、SGC-7901和人正常胃粘膜上皮GES-1细胞,应用qRT-PCR检测各株细胞miR29c的表达。用hsamiR29c-mimics类似物(miR29c组),hsa-miR29c-inhibitor(阴性对照组)分别转染BGC-823细胞,空白对照组加入转染液,用CCK8及MTT法检测细胞增殖,流式细胞术检测细胞凋亡,Western blot检测S100A4相关肿瘤相关蛋白的表达。结果与正常胃粘膜上皮细胞GES-1表达率(1.131±0.069)相比,MiR-29c在胃癌细胞BGC823(0.254±0.025),SGC-7901(0.466±0.057)(F=57.5,P<0.05)。BGC823细胞转染miR29c模拟物后MiR-29c的表达量为:4.126±0.231,0.405±0.174,0.557±0.102,差异具有统计学意义(F=850.1,P<0.05)。转染24h、48h、72h后,MiR-29c组细胞的OD450值分别为:1.074±0.127、1.609±0.178、2.008±0.104;阴性对照组为:0.419±0.169、0.477±0.092、0.738±0.153,空白对照组为:0.830±0.305。其中,转染48h、72h后,MiR-29c组明显高于对照组(F=59.46,P<0.05)。细胞转染24h后,miR29c组细胞凋亡率分别是(6.900±0.414)%、(4.490±0.316)%、(3.850±0.243)%,,阴性对照组凋亡率分别是(10.830±0.231)%、(11.940±0.279)%、(13.530±0.941)%,空白对照组凋亡率分别是:(10.190±0.188)%、(11.76±0.569)%、(13.260±0.987),(P<0.001).各组S100A4蛋白的表达分别为0.770±0.028、1.109±0.083、1.024±0.042,(P=0.0021)。结论miR29c在胃癌细胞系中表达降低,通过抑制S100A4抑制人胃癌细胞增殖并促进胃癌细胞凋亡。

Objective To explore the effect of MicroRNA-29c(miR29c)on the proliferation and apoptosis of human gastric adenocarcinoma cell lines,and to explore the relationship between miR29c and S100A4 metastasis-related protein.Methods The human gastric carcinoma cell lines BGC-823,SGC-7901 and normal gastric mucosa epithelial GES-1 cell line were cultured in carbon dioxide incubator.The expreesion of miR29c in cell lines were tested by qRT-PCR method.BGC-823 cells were transfected with hsamiR29c-mimics analogs(miR29c group)and hsa-miR29c-inhibitor(negative control group),respectively.In the control group,transfer solution was added,cell proliferation was detected by CCK8 and MTT,cell apoptosis was detected by flow cytometry.The expression of S100A4-associated tumor-associated protein was detected by Western Blot.Results Compared with the expression rate of GES-1 in normal gastric mucosa epithelial cells(1.131±0.069),the expression rate of MiR-29c in gastric cancer cells BGC823(0.254±0.025),SGC-7901(0.466±0.057)(F=57.5,p<0.05).The expression of MiR-29c in BGC823 cells transfected with miR29c mimics was 4.126±0.231,0.405±0.174,0.557±0.102,and the differences were statistically significant(F=850.1,p<0.05).After transfection for 24h,48h and 72h,the OD450 values of miR29c group cells were 1.074±0.127,1.609±0.178 and 2.008±0.104,respectively.The negative control group was 0.419±0.169,0.477±0.092,0.738±0.153,and the control group was 0.830±0.305.After transfection for 48h and 72h,MiR-29c group was significantly higher than control group(F=59.46,p<0.05).After transfection for 24h,the apoptosis rates of miR29c group were(6.900±0.414)%,(4.490±0.316)%,(3.850±0.243)%,respectively.The apoptosis rates of the negative control group were(10.830±0.231)%,(11.940±0.279)%and(13.530±0.941)%,respectively.The apoptosis rates of the blank control group were:(10.190±0.188)%,(11.76±0.569)%,(13.260±0.987),(p<0.001).The expression of S100A4 protein in each group was 0.770±0.028,1.109±0.083,1.024±0.042,respectively(p=0.0021).Conclusion MiR29c can inhibit proliferation and induced apoptosis in human gastric carcinoma cell lines through suppress S100A4 protein expression.