CagA作用于胃癌细胞调控miR-142-3p表达及机制

Effect and mechanism of CagA on miR-142-3p expression regulation in gastric cancer cells

目的幽门螺杆菌(Helicobacter pylori,Hp)主要毒力因子细胞毒素相关蛋白A(cytotoxin-associated gene A,CagA)在胃癌的发生发展中具有重要作用。微小RNA(miRNA)可通过影响下游mRNA分子的表达进而发挥促癌或抑癌作用,本研究探讨CagA相关的miRNA及其参与胃癌进展的作用机制,以期为CagA感染胃癌患者的诊断与治疗提供新思路。方法在GEO数据库和TCGA数据库中筛选与Hp和胃癌均相关的miRNA,并对其进行基于TCGA数据的临床病理资料分析。构建CagA原核表达系统,以不同浓度CagA转染胃癌AGS细胞,采用实时荧光定量聚合酶链式反应(qRT-PCR)法检测目标miRNA的表达。最后借助DIANA mirPath 3.0在线网站对miRNA进行GO和KEGG分析。结果通过GEO、TCGA数据库及细胞实验确认miR-142-3p与感染CagA的胃癌相关。人群水平上发现mir-142与男性胃癌Ⅰ期患者、女性45~54和55~64岁胃癌患者预后均明显相关(P<0.05)。细胞水平上发现miR-142-3p在胃癌AGS细胞中表达上调(P<0.01),且表达水平受CagA蛋白感染的影响(P<0.01)。预测发现PIK3R2、PIK3R5和PIK3CD可能作为miR-142-3p下游靶标参与miR-142-3p在胃癌进展中的作用。结论miR-142-3p可能是CagA感染胃癌患者的潜在的治疗靶标,并可能通过PIK3R2/PIK3R5/PIK3CD发挥对胃癌进展的促进作用。

Objective Cytotoxin associated protein A(CagA),the main virulence factor of Helicobacter pylori(Hp),plays an important role in the occurrence and development of gastric cancer.Micro RNA(miRNA)can promote or inhibit cancer by affecting the expression of downstream mRNA molecules.The aim of the study is to explore CagA-related miRNAs and effects of the miRNAs on gastric cancer for providing evidences to the diagnosis and treatment of gastric cancer correlated with Hp-CagA infection.Methods MiRNAs related to both Hp infection and gastric cancer were screened in the Gene Expression Omnibus(GEO)database and relevant clinicopathological data were analyzed based on the Cancer Genome Atlas(TCGA)dataset.The prokaryotic expression system of CagA was constructed and AGS cells were transfected with different concentrations of CagA.Quantitative real-time polymerase chain reaction(qRT-PCR)was used to detect the expression of the targeted miRNAs after transfection.Finally,Gene Ontology(GO)and Kyoto Encyclopedia of Genes and Genomes(KEGG)analysis of the miRNA were predicted and analyzed with DIANAmirpath3.0 online websites.Results MiR-142-3p was confirmed being related Hp-CagA infection based on GEO database,TCGA dataset and cell experiment.In clinical studies,mir-142 expression was associated with the prognosis of male patients with stage I gastric cancer and female patients aged 45-54 and 55-64 years(all P<0.05).In cytological studies,the expression of miR-142-3p was up-regulated in gastric cancer cells(P<0.01)and the level of miR-142-3p expression was affected by Hp-CagA infection(P<0.01).Other study results suggested that as downstream targets,phosphoinositide-3-kinase regulatory subunit 2(PIK3R2),PIK3R5 and phosphoinositide-3 kinase,catalytic subunit delta(PIK3CD)may be closely related to the effect of miR-142-3p in progressing course of gastric cancer.Conclusion MiR-142-3p may promote gastric cancer progressing through pathways of PIK3R2/PIK3R5/PIK3CD and may be a potential therapeutic target.