微小RNA-645靶向IFIT2对幽门螺杆菌感染胃癌细胞增殖和侵袭的影响

Effects of microRNA-645 targeting IFIT2 on proliferation and invasion of gastric cancer cells infected with Helicobacter pylori

目的探讨微小RNA-645(miR-645)在幽门螺杆菌(Hp)感染的胃癌细胞中的作用,重点分析miR-645在Hp感染胃癌细胞中的表达及其对细胞增殖、侵袭和迁移的影响。方法采用实时荧光定量PCR(qPCR)检测人正常胃上皮细胞(GES-1)和胃癌细胞(BGC-823和SGC-7901)的miR-645水平。在Hp感染的BGC-823细胞中转染miR-645抑制物(inhibitor)并将细胞分为3组:对照组、NC组和Inhibitor组。CCK-8法、划痕实验和Transwell小室实验分别检测Hp阳性BGC-823细胞的增殖、迁移和侵袭能力。荧光素酶报告基因实验验证miR-645与潜在靶基因干扰素诱导蛋白2(IFIT2)的关系。结果与Hp阴性细胞相比,miR-645在Hp阳性GES-1细胞(1.00±0.07 vs.1.81±0.17)、Hp阳性SGC-7901细胞(1.00±0.05 vs.3.47±0.15)和Hp阳性BGC-823细胞(1.00±0.06 vs.4.00±0.34)中的水平均升高(P<0.05)。Hp阳性SGC-7901细胞和BGC-823细胞的miR-645水平分别为1.92±0.08和2.22±0.19,均高于Hp阳性GES-1细胞的1.00±0.10(P<0.05)。与对照组和NC组相比,Inhibitor组Hp阳性BGC-823细胞的增殖活力在转染后48、72 h后均下降(P<0.05)。Inhibitor组Hp阳性BGC-823细胞的划痕愈合率和穿膜细胞数分别为(11.48±1.67)%和(139.20±8.27)个,低于对照组的(48.36±8.15)%和(288.91±11.92)个及NC组的(45.87±2.86)%和(272.50±10.15)个,差异有统计学意义(P<0.05)。生物信息学预测和荧光素酶报告分析确定了抑癌基因IFIT2是miR-645的下游靶基因,且Inhibitor组IFIT2水平高于对照组和NC组(P<0.05)。结论miR-645作为一种促癌miRNA,通过靶向抑癌基因IFIT2来促进Hp阳性胃癌细胞的增殖、迁移和侵袭过程。本研究有助于进一步阐明胃癌进展的机制,为胃癌的治疗提供一定理论基础。

Objective To investigate the role of microRNA-645(miR-645)in gastric cancer cells infected with Helicobacter pylori(Hp),and to analyze the expression of miR-645 in gastric cancer cells infected with Hp and its effects on cell proliferation,invasion and migration.Methods Real-time quantitative PCR(qPCR)was used to detect the miR-645 levels in human normal gastric epithelial cell(GES-1)and gastric cancer cells(BGC-823 and SGC-7901).BGC-823 cells infected with Hp were transfected with miR-645 inhibitor and divided into three groups:Control group,NC group and Inhibitor group.CCK-8 method,scratch test and Transwell chamber test were used to detect the proliferation,migration and invasion of Hp-positive BGC-823 cells.Luciferase reporter assay was used to verify the relationship between miR-645 and potential target gene interferon inducible protein 2(IFIT2).Results The miR-645 levels were increased in Hp-positive GES-1(1.00±0.07 vs.1.81±0.17),Hp-positive SGC-7901(1.00±0.05 vs.3.47±0.15)and Hp-positive BGC-823(1.00±0.06 vs.4.00±0.34)cells compared with the Hp-negative cells,respectively(P<0.05).The miR-645 levels in Hp-positive SGC-7901 and BGC-823 cells were 1.92±0.08 and 2.22±0.19,higher than 1.00±0.10 in HP-positive GES-1 cells(P<0.05).Compared with the Control group and NC group,the proliferation activity of Hp-positive BGC-823 cells in Inhibitor group decreased at 48 and 72 h after transfection(P<0.05).The scratch healing rate and number of penetrating cells of Hp-positive BGC-823 cells in the Inhibitor group were(11.48±1.67)%and 139.20±8.27,lower than(48.36±8.15)%and 288.91±11.92 in Control group and(45.87±2.86)%and 272.50±10.15 in NC group(P<0.05).Bioinformatics prediction and luciferase report analysis confirmed that tumor suppressor gene IFIT2 was the downstream target gene of miR-645,and the IFIT2 level in Inhibitor group was higher than those in Control group and NC group(P<0.05).Conclusion As a cancer promoting miRNA,miR-645 promotes the proliferation,migration and invasion of Hp-positive gastric cancer cells by directly and negatively targeting the tumor suppressor gene IFIT2.This study will help to further clarify the mechanism of gastric cancer progression and provide a theoretical basis for the treatment of gastric cancer.