长链非编码RNA MAFG-AS1通过靶向miR-143-3p/AKT2轴促进胃癌的进展

目的:探讨长链非编码RNA(long non-coding RNA,lncRNA)MAFG-AS1在胃癌(gastric cancer,GC)的发生和发展中的生物学功能及可能的作用机制。方法:分别用GEPIA(Gene Expression Profiling Interactive Analysis)和Kaplan-Meier Plotter数据库分析MAFG-AS1在GC组织中的表达水平,及与GC患者预后的关系。采用实时荧光定量PCR法检测胃黏膜正常上皮GES-1细胞和人GC细胞系BGC-823、SGC-7901和MGC-803中MAFG-AS1的表达水平。采用脂质体转染法将MAFG-AS1过表达重组质粒转入MGC-803细胞,分别采用CCK-8法和EdU染色法检测MAFG-AS1过表达对MGC-803细胞增殖的影响,Transwell小室实验检测对MGC-803细胞迁移的影响。利用StarBase数据库预测MAFG-AS1和微RNA(microRNA,miRNA,miR)-143-3p之间,miR-143-3p和AKT2(AK mouse plus Transforming or Thymoma 2)基因之间的靶向关系,并分别采用双荧光素酶报告基因实验进行验证。采用实时荧光定量PCR法检测MAFG-AS1、miR-143-3p和AKT2 mRNA在癌及癌旁组织中的表达水平,分别应用Pearson对MAFG-AS1和miR-143-3p之间,miR-143-3p和AKT2 mRNA之间,MAFG-AS1和AKT2 mRNA之间表达水平的相关性进行分析。采用脂质体转染法将miR-143-3p-模拟物(miR-143-3p-mimics)或特异性针对AKT2基因的siRNA(siAKT2)转入MAFG-AS1过表达的MGC-803细胞中,采用蛋白印迹法检测对AKT2蛋白表达水平的影响,再分别采用CCK-8法和EdU染色法检测对MGC-803细胞增殖的影响,Transwell小室实验检测对MGC-803细胞迁移的影响。结果:MAFG-AS1在GC组织和细胞中高表达(P值均<0.001),其高表达水平与患者总生存期、首次进展生存期以及后进展生存期呈负相关(P值均<0.05)。MAFG-AS1与miR-143-3p表达呈负相关(P<0.05),与AKT2 mRNA表达呈正相关(P<0.05)。过表达MAFG-AS1可促进细胞增殖和迁移(P值均<0.001),而上调miR-143-3p表达可削弱此作用(P值均<0.05)。结论:MAFG-AS1通过调控miR-143-3p/AKT2轴在GC的进展中发挥促进作用。

LncRNA MAFG-AS1 promotes proliferation and migration of gastric cancer cells by regulating the miR-143-3p/AKT2 axis

Objective:To investigate the role and mechanism of long non-coding RNA(lncRNA)MAFG-AS1 in the initiation and development of gastric cancer(GC).Methods:Gene Expression Profiling Interactive Analysis(GEPIA)and Kaplan-Meier Plotter database were used to analyze the expression level of MAFG-AS1 in GC tissues and its relationship with the prognosis of GC patients.Real-time fluorescent quantitative PCR was used to detect the expression level of MAFG-AS1 in normal gastric epithelial GES-1 cells and human GC BGC-823,SGC-7901 and MGC-803 cells.The recombinant plasmid expressing MAFG-AS1 was transfected into MGC-803 cells by liposome,and the effect of MAFG-AS1 overexpression on the proliferation of MGC-803 cells was detected by CCK-8 assay and EdU staining.Transwell assay was used to detect the effect of MAFG-AS1 overexpression on the migration of MGC-803 cells.The StarBase database was used to predict the relationship between MAFG-AS1 and miR-143-3p,miR-143-3p and AK mouse plus Transforming or Thymoma 2(AKT2)gene,and the relationship was verified by dual-luciferase reporter assay.Real-time fluorescent quantitative PCR was used to detect the expression levels of MAFG-AS1,miR-143-3p and AKT2 mRNA in tumor and para-tumor tissues.Pearson correlation analysis was conducted to evaluate the relevance between the expressions of MAFG-AS1,miR-143-3p and AKT2 mRNA.The miR-143-3p-mimics or siRNA targeting AKT2 gene(siAKT2)were transfected into MGC-803 cells overexpressing MAFG-AS1 by liposome,then Western blotting was used to detect the protein expression of AKT2,CCK-8 assay and EdU staining were used to detect the MGC-803 cells proliferation,and the Transwell assay was used to detect the MGC-803 cells migration.Results:MAFG-AS1 was highly expressed in GC tissues and cells(all P<0.001),and its high expression level was negatively correlated with the survival(all P<0.05).MAFG-AS1 was negatively correlated with miR-143-3p expression(P<0.05),and positively correlated with AKT2 mRNA expression(P<0.05).Overexpression of MAFG-AS1 could promote cell proliferation and migration(all P<0.001),while the up-regulation of miR-143-3p expression could weaken this effect(all P<0.05).Further studies showed that MAFG-AS1 up-regulated the expression of AKT2 by sponging miR-143-3p(P<0.05).Conclusion:LncRNA MAFG-AS1 promotes GC progression through regulating miR-143-3p/AKT2 axis.