幽门螺杆菌上调miR-7致胃上皮细胞GES-1凋亡

Helicobacter pylori induces apoptosis of gastric epithelial cells GES-1 by upregulating miR-7 expression

目的探讨miR-7对幽门螺杆菌感染胃上皮细胞凋亡的影响。方法以Hp感染GES-1细胞,感染24 h的细胞分为1∶50(细胞∶细菌)组和1∶100组;当感染比率为1∶100(细胞∶细菌)时,按感染时间分为24 h组和48 h组,同时设未感染细胞为空白对照组,观察不同感染菌量和感染时间对GES-1细胞miR-7和RELA表达的影响。以miR-7-5p类似物转染GES-1细胞设为miR-7 mimics组,miR-阴性序列转染GES-1为阴性对照组(NC组),未转染细胞作为空白对照组,观察各组细胞增殖凋亡及RELA蛋白表达情况。实时定量PCR法检测miR-7表达量,CCK-8法检测细胞增殖能力,流式细胞仪检测细胞凋亡率,Western blot检测RELA蛋白表达量。结果与空白对照相比较,感染24 h时1∶50组及1∶100组的miR-7表达量上调,差异有统计学意义(P<0.05)。当感染率1∶100时,与空白对照组比较,感染24 h组和48 h组miR-7表达量均显著上调(P<0.05),48 h组RELA蛋白显著下降(P<0.05),但24 h组差异无统计学意义。转染实验中发现,与空白对照组和NC组相比较,mimics组细胞增殖能力明显下降(P<0.05),细胞凋亡率明显增加(P<0.05),RELA蛋白表达显著降低(P<0.05)。结论Hp通过上调感染细胞的miR-7分子调控其靶基因RELA的表达,抑制细胞增殖,促进细胞凋亡,这可能是Hp导致胃上皮细胞损伤致病的机制之一。

Objective To investigate the effect of miR-7 on apoptosis of gastric epithelial cells GES-1 infected by Hp.Methods GES-1 cells were divided into 1∶50 group and 1∶100 group according to the ratio of cell and bacteria when infected with Hp for 24 h,and GES-1 cells were divided into 24 h group and 48 h group according to the infection time when the cell/bacteria ratio of 1∶100,and the uninfected cells were chosen as control group.GES-1 cells was transfected with miR-7-5p mimics in mimics group and miR-negative sequences in NC group,and GES-1 cells without treatment were chosen as control group.The cell proliferation was tested by CCK-8,the apoptosis rate was evaluated by flow cytometry,the amount of miR-7 of cells was detected by real-time quantitative PCR,and RELA protein was measured by Western blot.Results Compared with control group,the expression of miR-7 was significantly upregulated in 1∶50 group,1∶100 group after infected with Hp for 24 h(P<0.05).Compared with control group,the expression of miR-7 was significantly upregulated in 24 h group and 48 h group at the ratio of cell and bacteria of 1∶100(P<0.05),and the amount of RELA protein was reduced significantly in 48 h group(P<0.05).Compared with control group and NC group,the cell proliferation was significantly decreased in mimics group,the cell apoptosis was significantly increased,and the level of RELA protein was decreased significantly(all P<0.05).Conclusion Hp downregulates the expression of miR-7 target gene RELA contributing to cell apoptosis through upregulating miR-7 in infected cells,which is one of the mechanisms of gastric epithelial cell damage induced by Hp.