胃癌中miRNA-522-3p的表达及其靶向抑制RSU1表达对胃癌细胞生物学功能的影响

miRNA-522-3p expression in gastric cancer and the effect of its targeted inhibition of RSU1 expression on the biological function of gastric cancer cells

目的探讨miRNA-522-3p(miR-522-3p)在胃癌组织和细胞中的表达及其对RSU1表达的调控, 以及体外对胃癌细胞生物学功能的影响。方法采用实时荧光定量聚合酶链反应(qRT-PCR)检测山西白求恩医院2019年5月至2020年6月确诊的50例胃癌患者肿瘤组织和癌旁组织中miR-522-3p的相对表达水平。将胃癌MGC-803细胞和BGC-823细胞分为miR-522-3p抑制剂组(转染miR-522-3p抑制剂)和空载体组(转染空载体), 采用CCK-8法检测细胞增殖能力, 细胞划痕实验检测细胞的划痕愈合能力, 流式细胞术检测细胞凋亡能力;通过双荧光素酶报告基因实验检测miR-522-3p和RSU1的靶向相关性, 蛋白质印迹法检测RSU1蛋白的变化。结果 qRT-PCR结果显示, 与癌旁组织比较, 胃癌组织中miR-522-3p的相对表达水平上调, 差异具有统计学意义(0.84±0.31比0.48±0.22, t=2.93, P<0.05)。不同肿瘤长径和病理分级的胃癌组织中miR-522-3p的相对表达水平差异有统计学意义(均P<0.05)。体外实验表明, 48、72 h时miR-522-3p抑制剂组MGC-803细胞、BGC-823细胞的细胞增殖率均下降(均P<0.05), MGC-803、BGC-823细胞转染miR-522-3p抑制剂后, 能抑制MGC-803、BGC-823细胞的划痕愈合能力。双荧光素酶报告基因实验验证miR-522-3p能靶向结合RSU1的3’’UTR, 并影响其荧光活性;蛋白质印迹法结果显示, miR-522-3p抑制剂可促进RSU1蛋白的表达。结论 miR-522-3p可能通过调控RSU1表达参与胃癌的进展, 其可能是潜在的治疗靶标。

Objective To investigate the expression of miRNA-522-3p(miR-522-3p)in gastric cancer tissues/cells and its regulation of RSU1 expression and to analyze the effect of miR-522-3p on biological function of gastric cancer cells in vitro.Methods miR-522-3p relative expression levels in tumor tissues and adjacent tissues of 50 patients clinically diagnosed as gastric cancer in Shanxi Bethune Hospital from May 2019 to June 2019 were detected by using real-time quantitative polymerase chain reaction(qRT-PCR).MGC-803 cells and BGC-823 cells in gastric cancer were divided into miR-522-3p inhibitor group(transfected with miR-522-3p inhibitor)and empty vector group(transfected with empty vector).The cell proliferation was detected by using CCK-8 assay,cell scratch assay was used to detect the scratch healing ability of cells,and flow cytometry was used to detect the apoptosis.The target correlation between miR-522-3p and RSU1 was detected by using double luciferase reporter gene assay,the change of RSU1 protein was detected by using Western blot.Results qRT-PCR showed that compared with adjacent cancer tissues,the relative expression level of miR-522-3p was up-regulated,and the difference was statistically significant(0.84±0.31 vs.0.48±0.22,t=2.93,P<0.05).There were no statistically significant differences in the relative expression level of miR-522-3p in gastric cancer tissues with different tumor diameter and pathological grade(all P<0.05).In vitro experimental assay showed that the cell proliferation rates of MGC-803 and BGC-823 cells in miR-522-3p inhibitor group at 48 h and 72 h were decreased(all P<0.05).Furthermore,MGC-803 and BGC-823 tranfected with miR-522-3p inhibitor could effectively inhibit the scratch healing ability of MGC-803 and BGC-823 cells.Dual-luciferase reporter gene assay verified that miR-522-3p targeted to RSU13'UTR and affected its fluorescence activity.Western blot results showed that miR-522-3p could promote RSU1 protein expression.Conclusions miR-522-3p is involved in the progression of gastric cancer probably via the regulation of RSU1 expression,and it may be a potential therapeutic target.