沉默PRMT5基因对胃癌细胞株SGC-7901细胞周期的影响及机制研究

Effect and mechanism of PRMT5 gene silencing on cell cycle and apoptosis of gastric cancer cell line SGC-7901

目的探究沉默蛋白质精氨酸甲基转移酶5(PRMT5)基因对胃癌细胞株SGC-7901的细胞周期、凋亡的影响及相关机制。方法选取对数生长期的正常胃黏膜上皮细胞系GES-1和人胃癌细胞株SGC-7901,采用蛋白质印迹法检测两种细胞中PRMT5蛋白的相对表达量。将SGC-7901细胞分为control组(不做处理)、NC组(转染阴性对照载体)、PRMT5小干扰RNA(siRNA)组(转染PRMT5 siRNA)、PRMT5 siRNA+胰岛素样生长因子-Ⅰ(IGF-Ⅰ)组(转染PRMT5 siRNA,加入IGF-Ⅰ并使其终浓度为10 ng/mL)。转染48 h后,采用蛋白质印迹法检测各组PRMT5、蛋白激酶B(AKT)、磷酸化AKT(p-AKT)、哺乳动物雷帕霉素靶蛋白(mTOR)、p-mTOR的相对表达量。采用流式细胞术检测各组的细胞周期和细胞凋亡率。结果与GES-1细胞比较,SGC-7901细胞中PRMT5蛋白的相对表达量较高,两组差异有统计学意义(P<0.05)。与control组、NC组比较,PRMT5 siRNA组PRMT5蛋白的相对表达量较低,G0/G1期细胞的占比、细胞凋亡率较高,而S期、G2/M期细胞的占比较低,差异均有统计学意义(P均<0.05)。与PRMT5 siRNA组比较,PRMT5 siRNA+IGF-Ⅰ组PRMT5蛋白的相对表达量较高,G0/G1期细胞的占比、细胞凋亡率较低,而S期、G2/M期细胞的占比较高,差异均有统计学意义(P均<0.05)。与control组、NC组比较,PRMT5 siRNA组p-AKT/AKT、p-mTOR/mTOR均较低,差异均有统计学意义(P均<0.05);与PRMT5 siRNA组比较,PRMT5 siRNA+IGF-Ⅰ组p-AKT/AKT、p-mTOR/mTOR较高,差异均有统计学意义(P均<0.05)。结论沉默PRMT5基因可阻滞胃癌细胞株SGC-7901的细胞周期,并促使其凋亡,其作用机制可能与抑制PI3K/AKT信号通路有关。

Objective This paper intends to study the effect of protein arginine methyltransferase 5(PRMT5)gene silencing on the cell cycle and apoptosis of gastric cancer cell line SGC-7901,and to explore related mechanisms.Methods The normal gastric mucosal cell line GES-1 and the gastric cancer cell line SGC-7901 in the logarithmic phase were collected.The expression of PRMT5 protein in cells were detected by Western blotting.The SGC-7901 cells were divided into the control group(no treatment),the NC group(transfected with negative control vector),the PRMT5 siRNA group(transfected with PRMT5 siRNA),and the PRMT5 siRNA+insulin-like growth factor-Ⅰ(IGF-Ⅰ)group(PRMT5 siRNA was transfected and IGF-Ⅰ was added to a final concentration of 10 ng/mL).After 48 hours of transfection,the expression of PRMT5 protein,protein kinase B(AKT),p-AKT,mammalian target of rapamycin(mTOR),and p-mTOR were detected by Western blotting.The cell cycle and apoptosis rate were detected by flow cytometry.Results Compared with GES-1 cells,the expression of PRMT5 protein in the SGC-7901 cells is higher,with a statistically significant difference(P<0.05).Compared with the control group and the NC group,the expression of PRMT5 protein in the PRMT5 siRNA group is lower,the proportion of cells in the G0/G1 phase and the apoptosis rate are higher,while the proportion of cells in the S phase and the G2/M phase is lower,with a statistically significant difference(P<0.05).Compared with the PRMT5 siRNA group,the expression of PRMT5 protein in the PRMT5 siRNA+IGF-Ⅰ group is higher,the proportion of cells in the G0/G1 phase and the apoptosis rate are lower,while the proportion of cells in the S phase and the G2/M phase is higher,with a statistically significant difference(P<0.05).Compared with the control group and the NC group,the p-AKT/AKT and p-mTOR/mTOR in the PRMT5 siRNA group are lower,with a statistically significant difference(P<0.05).Compared with the PRMT5 siRNA group,the p-AKT/AKT and p-mTOR/mTOR in the PRMT5 siRNA+IGF-Ⅰgroup are higher,with a statistically significant differences(P<0.05).Conclusions PRMT5 gene silencing can block the cell cycle of gastric cancer cell line SGC-7901 and promote its apoptosis.It is speculated that its mechanism is related to the inhibition of PI3K/AKT signaling pathway.