miRNA-193a-3p靶向TGF-β2基因诱导肝癌细胞凋亡的分子机制研究

Molecular mechanism of miRNA-193a-3p induced apoptosis of hepatocellular carcinoma cells through targeting TGF-β2

目的探讨微小RNA(micro RNA,miRNA)-193a-3p在肝癌组织中的表达及其对肝癌细胞迁移、凋亡的影响和潜在的分子机制。方法利用实时荧光定量PCR(real-time quantitative PCR,qRT-PCR)检测miR-193a-3p在肝癌及其癌旁正常组织中的表达;将miR-193a-3p mimic转染肝癌HepG2细胞系(NC为对照组),利用CCK-8(cell counting kit-8)实验、Transwell实验及流式细胞仪评估HepG2细胞的活性、迁移及凋亡情况;利用生物信息学分析、qRT-PCR及双荧光素酶报告实验确证miR-193a-3p对下游靶标转化生长因子(transforming growth factor,TGF)-β2的影响;在miR-193a-3p过表达的HepG2细胞中转染TGF-β2质粒,检测过表达TGF-β2对miR-193a-3p诱导HepG2细胞活性、迁移及凋亡的影响。结果miR-193a-3p在肝癌组织中的表达显著低于正常肝脏组织,差异有统计学意义(P<0.01):与NC组比较,miR-193a-3p组细胞的增殖活性及迁移数量减少,细胞凋亡率显著增加,差异有统计学意义(P<0.05),且以时间依赖性方式诱导HepG2细胞凋亡:miR-193a-3p与靶基因TGF-β2的互补结合序列在进化上高度保守,TGF-β2是miR-193a-3p的下游靶标;与miR-193a-3p+空质粒组比较,miR-193a-3p+TGF-β2组肝癌细胞的增殖活性及迁移数量增多,细胞凋亡率显著减少,差异有统计学意义(P<0.05)。结论miR-193a-3p在肝癌组织中表达减少,miR-193a-3p通过靶向调控TGF-β2基因抑制HepG2细胞的活性和迁移,促进HepG2细胞的凋亡。

Objective To investigate the expression of microRNA(miRNA)-193a-3p in hepatic carcinoma tissue and its effect on migration and apoptosis of hepatocellular carcinoma cells.Methods The expression of miR-193a-3p in hepatic carcinoma tissue and its adjacent normal tissues was detected by real-time quantitative PCR(qRT-PCR);miR-193a-3p mimic was transfected into HepG2 cell line(NC as the control group).The viability,migration and apoptosis of HepG2 cells were evaluated by cell counting kit-8(CCK-8)assay,transwell assay and flow cytometry analysis.Bioinformatics analysis,qRT-PCR and double Luciferase report experiment were used to confirm the effect of miR-193a-3p on downstream target gene transforming growth factor(TGF)-β2.TGF-β2 plasmid was transfected to HepG2 cells with miR-193a-3p overexpression to detect the effect of TGF-β2 on the viability,migration and apoptosis of miR-193a-3p-induced HepG2 cells.Results The expression of miR-193a-3p in hepatic carcinoma tis-sue was significantly lower than that in normal liver tissue(P<0.01);Compared with NC group,the proliferation via-bility and migration of cells in miR-193a-3p group decreased,and the apoptosis rate increased significantly(P<0.05),and HepG2 cell apoptosis was induced by miR-193a-3p in a time-dependent manner;The complementary binding sequence between miR-193a-3p and the target gene TGF-β2 was highly conserved.TGF-β2 was the target gene of miR-193a-3p.Compared with miR-193a-3p+empty plasmid group,the viability and migration number of HepG2 cells in the miR-193a-3p+TGF-β2 groups increased,and the apoptosis rate decreased significantly(P<0.05).Conclusion The expression of miR-193a-3p is decreased in hepatic carcinoma tissue,and miR-193a-3p inhibits the viability and migration of HepG2 cells and promotes the apoptosis of HepG2 cells by targeting TGF-β2 gene.