端粒酶抑制因子X1过表达对人食管癌EC1细胞增殖及凋亡的影响

Effect of overexpression of telomerase inhibitor X1 on proliferation and apoptosis of human esophageal carcinoma EC1 cells

目的探究端粒酶抑制因子X1(PinX1)对人食管癌EC1细胞增殖及凋亡的影响。方法体外培养人食管癌细胞系EC1细胞株,并将其分为空白对照(NG)组(空白对照细胞)、pcDNA组(转染阴性对照质粒)、pcDNA-PinX1组(转染PinX1过表达质粒)、N-乙酰-L-半胱氨酸(NAC)组(加入信号通路抑制剂)。采用实时荧光定量PCR法检测各组PinX1 mRNA的表达水平。分别采用CCK-8法和流式细胞术检测各组细胞的增殖、凋亡情况。采用荧光探针法检测各组活性氧簇(ROS)水平。采用蛋白质印迹法检测各组腺苷酸激活蛋白激酶/信号转导与转录激活因子3(AMPK/STAT3)信号通路相关蛋白的表达水平。结果经成功转染细胞后,与NG组、pcDNA组比较,pcDNA-PinX1组细胞的增殖抑制率、凋亡率、ROS水平、p-AMPK/AMPK均升高,而p-STAT3/STAT3均降低,差异均有统计学意义(P均<0.05)。与pcDNA-PinX1组比较,NAC组细胞的增殖抑制率、凋亡率、ROS水平、p-AMPK/AMPK均降低,p-STAT3/STAT3升高,差异均有统计学意义(P均<0.05)。结论PinX1过表达可抑制人食管癌EC1细胞增殖并诱导其凋亡,可能通过促进ROS/AMPK/STAT3信号通路活化来实现。

Objective This paper intends to investigate the effect of telomerase inhibitor X1(PinX1)on the proliferation and apoptosis of human esophageal carcinoma EC1 cells.Methods Human esophageal cancer cell line EC1 was cultured in vitro and assigned to the blank control(NG)group,the pc DNA group(transfected negative control plasmid),the pc DNA-Pinx1 group(transfected PinX1 overexpressed plasmid),and the N-acetyl-L-cysteine(NAC)group(added signaling pathway inhibitors).Real-time fluorescence quantitative PCR was used to detect the expression of PinX1 mRNA in each group.Cell proliferation and apoptosis were detected by using the CCK-8 method and flow cytometry,respectively.The reactive oxygen species(ROS)in each group were detected by fluorescence probe,and the expression of adenylate activated protein kinase/signal transduction and transcriptional activator 3(AMPK/STAT3)signaling pathways was detected by Western blotting.Results After successful transfection,compared with the NG group and the pc DNA group,the proliferation inhibition rate,the apoptosis rate,the ROS level,and the p-AMPK/AMPK in the pc DNA-Pinx1 group are increased,while p-STAT3/STAT3 are decreased,with statistically significant differences(P<0.05).Compared with the pc DNA-PinX1 group,the proliferation inhibition rate,the apoptosis rate,the ROS level,and p-AMPK/AMPK in the NAC group are decreased,while p-STAT3/STAT3 is increased,with statistically significant differences(P<0.05).Conclusion The overexpression of PinX1 can inhibit the proliferation and induce the apoptosis of human esophageal carcinoma EC1 cells,possibly by promoting the activation of ROS/AMPK/STAT3 signaling pathway.