miR-21靶向PTEN调控AKT/FoxO1信号通路对胃癌细胞凋亡的机制研究  

Mechanism of miR-21 Targeting PTEN to Regulate AKT/FoxO1 Signaling Pathway on Apoptosis in Gastric Cancer Cells

作　　者：王艳花[1] 聂亚楠 齐俊娟 季艳霞[1] WANG Yanhua(Handan Central Hospital,Hebei Handan 056000,China)

机构地区：[1]河北省邯郸市中心医院,河北邯郸056000 [2]河北省邯郸市邯钢医院,河北邯郸056000

出　　处：《河北医学》2023年第12期1985-1992,共8页Hebei Medicine

基　　金：2020年度河北医学科研课题计划(编号:20200195)。

摘　　要：目的:探究miR-21靶向PTEN调控AKT/FoxO1信号通路对胃癌细胞凋亡的影响。方法:将胃癌SGC-7901细胞分为miR-NC inhibitors组(SGC-7901细胞中转染miR-NC inhibitors质粒)、miR-21 inhibitors组(SGC-7901细胞中转染miR-21 inhibitors质粒)、miR-21 inhibitors+sh-PTEN组(SGC-7901细胞中转染miR-21 inhibitors和sh-PTEN质粒);Control组(不转染任何质粒的SGC-7901细胞)。qRT-PCR检测细胞中miR-21 mRNA表达;采用CCK-8、Transwell小室法和流式细胞仪检测细胞增殖、侵袭能力和凋亡率;蛋白质印迹检测细胞中PTEN、AKT、p-AKT、PI3K、p-PI3K、FoxO1蛋白表达;双荧光素酶报告检测miR-21和PTEN的靶向关系。结果:人正常胃黏膜上皮细胞GES-1(1.00±0.10)相比,胃癌细胞SGC-7901中miR-21(1.89±0.17)表达明显升高(P<0.05)。Control组相比,miR-21 inhibitors组细胞中miR-21表达(0.83±0.10)、增殖率(45.31±4.92)%、侵袭数目(62.34±5.83)个和p-PI3K/PI3K(0.42±0.05)、p-AKT/AKT(0.51±0.05)比值均明显降低,细胞凋亡率(23.48±3.51)%、PTEN(0.98±0.10)和FoxO1(0.76±0.08)蛋白表达明显升高(P<0.0001)。pcDNA-NC组相比,pcDNA-PTEN组细胞增殖率(48.26±5.01)、侵袭细胞数(65.37±6.02)个和细胞中p-PI3K/PI3K(0.46±0.05)、p-AKT/AKT(0.55±0.06)比值均明显降低,细胞凋亡率(25.61±3.27)%及细胞中PTEN(0.91±0.09)和FoxO1(0.70±0.08)蛋白表达升高(P<0.05)。预测发现PTEN的3'UTR端与miR-21有碱基互补结合点位。miR-NC组相比,转染野生型PTEN(PTEN-WT)时miR-21组(0.32±0.03)荧光素酶活性明显降低(P<0.05)。miR-21 inhibitors组相比,miR-21 inhibitors+sh-PTEN组细胞增殖率(90.25±9.14)%和侵袭细胞数(125.69±10.31)个和细胞中p-PI3K/PI3K(0.80±0.08)、p-AKT/AKT(0.76±0.08)比值明显增加,细胞凋亡率(6.24±1.32)和细胞中PTEN(0.30±0.04)、FoxO1(0.38±0.05)蛋白表达降低(P<0.0001)。结论:敲减miR-21可靶向负调控PTEN表达,调控AKT/FoxO1信号通路,抑制胃癌细胞的增殖和侵袭,促进凋亡。Objective:To investigate the effect of miR-21 targeting PTEN to regulate AKT/FoxO1 signaling pathway on apoptosis in gastric cancer cells.Methods:Gastric cancer SGC-7901 cells were divided into miR-NC inhibitors group(SGC-7901 cells transfected with miR-NC inhibitors plasmid),miR-21 inhib-itors group(SGC-7901 cells transfected with miR-21 inhibitors plasmid),miR-21 inhibitors+sh-PTEN group(SGC-7901 cells transfected with miR-21 inhibitors and sh-PTEN plasmids);and control group(SGC-7901 cells not transfected with any plasmids).qRT-PCR was performed to detect miR-21 mRNA ex-pression in cells;CCK-8 was used to detect cell proliferation ability;Transwell assay for cell invasion shift a-bility;flow cytometry for apoptosis rate;protein blotting for PTEN,AKT,p-AKT,PI3K,p-PI3K,FoxO1 protein expression in cells;dual luciferase reporter for targeting relationship between miR-21 and PTEN.Re-sults:Compared with normal human gastric mucosal epithelial cell GES-1(1.00±0.10),the expression of miR-21(1.89±0.17)in gastric cancer cell SGC-7901 was significantly increased(P<0.05).Compared with the control group,the expression of miR-21(0.83±0.10),proliferation rate(45.31±4.92)%,number of invasions(62.34±5.83),and p-PI3K/PI3K(0.42±0.05),p-AKT/AKT(0.51±0.05)ratios were significantly reduced in the miR-21 inhibitors group,while the apoptosis rate(23.48±3.51)%,PTEN(0.98±0.10),and FoxO1(0.76±0.08)protein expression were significantly increased(P<0.001).Compared with the pcDNA-NC group,the cell proliferation rate(48.26±5.01),number of invasive cells(65.37±6.02),and p-PI3K/PI3K(0.46±0.05),p-AKT/AKT(0.55±0.06)ratios in the pcD-NA-PTEN group were significantly reduced,while the cell apoptosis rate(25.61±3.27)%and the expres-sion of PTEN(0.91±0.09)and FoxO1(0.70±0.08)proteins in the cells were increased(P<0.05).It was predicted that the 3'UTR end of PTEN had complementary binding sites with miR-21.Compared with the miR-NC group,the luciferase activity of the miR-21 group(0.32±0.03)was significantly reduced(P<0.05)when tran

关 键 词：胃癌 MIR-21 PTEN AKT/FoxO1信号通路 凋亡 

分 类 号：R735.2[医药卫生—肿瘤]