微小RNA-106通过调控PTEN/PI3K/AKT信号通路影响子宫内膜癌RL95-2细胞生物学行为的研究

MicroRNA-106 affects the biological behavior of endometrial cancer RL95-2 cells by regulating the PTEN/PI3K/AKT signaling pathway

目的探讨微小RNA-106(miR-106)通过调节PTEN/PI3K/AKT信号通路对子宫内膜癌细胞(RL95-2细胞)增殖、凋亡、侵袭和迁移的影响。方法采用实时荧光定量聚合酶链反应(qRT-PCR)检测子宫内膜癌组织标本及癌旁组织中miR-106表达。体外培养子宫内膜癌细胞RL95-2,分别转染抑制miR-106表达质粒(miR-106 inhibitor)及其阴性对照(NC inhibitor)、PTEN过表达质粒(PTEN)及其阴性对照(pcDNA)、PTEN+miR-106过表达质粒(miR-106 mimics)及其阴性对照(miR-NC),并依次作为miR-106 inhibitor组、NC inhibitor组、PTEN组、pcDNA组、PTEN+miR-106组及PTEN+miR-NC组,另设对照组(不做转染)。分别采用细胞计数试剂盒8(CCK-8)法、Transwell小室、流式细胞术检测RL95-2细胞增殖、侵袭和迁移、凋亡情况;采用蛋白免疫印迹(WB)法检测RL95-2细胞PTEN/PI3K/AKT通路相关蛋白表达。采用双荧光素酶报告基因实验验证miR-106与PTEN的靶向关系。结果miR-106在子宫内膜癌组织中表达水平高于癌旁组织,差异有统计学意义(P<0.05);双荧光素酶报告基因实验证实miR-106与PTEN存在靶向关系。抑制miR-106表达或过表达PTEN后,RL95-2细胞增殖能力、细胞迁移与侵袭数、p-PI3K、p-AKT蛋白表达水平降低,细胞凋亡率升高,差异均有统计学意义(P<0.05);过表达miR-106可显著逆转过表达PTEN对RL95-2细胞增殖、迁移与侵袭的抑制作用和凋亡的促进作用(P<0.05)。结论miR-106可通过靶向抑制PTEN表达并激活PI3K/AKT通路来促进人子宫内膜癌细胞的增殖、迁移及侵袭,抑制细胞凋亡。

Objective To investigate the effects of microRNA-106(miR-106)on the proliferation,apoptosis,invasion and migration of endometrial cancer cells(RL95-2 cells)by regulating PTEN/PI3K/AKT signaling pathway.Methods Real-time fluorescent quantitative polymerase chain reaction(qRT-PCR)was used to detect the expression of miR-106 in specimens of endometrial cancer tissues and adjacent tissues.Endometrial cancer RL95-2 cells were cultured in vitro,and were respectively transfected with plasmid inhibiting miR-106 expression(miR-106 inhibitor)and its negative control(NC inhibitor),overexpression plasmid of PTEN(PTEN)and its negative control(pcDNA),overexpression plasmid of PTEN and miR-106(miR-106 mimics)and its negative control(miR-NC),which were named as miR-106 inhibitor group,NC inhibitor group,PTEN group,pcDNA group,PTEN+miR-106 group and PTEN+miR-NC group in proper sequence,and the control group was set up without transfection.The cell counting kit 8(CCK-8)method,transwell chamber,and flow cytometry were used to detect the proliferation,invasion and migration,and apoptosis of RL95-2 cells;western blotting(WB)was used to detect the expression of PTEN/PI3K/AKT pathway related proteins in RL95-2 cells.Dual luciferase reporter gene experiment was used to verify the targeting relationship between miR-106 and PTEN.Results The expression of miR-106 in endometrial cancer tissues was significantly higher than that in adjacent tissues(P<0.05);the dual luciferase reporter gene experiment confirmed that miR-106 had a targeting relationship with PTEN.After inhibition of miR-106 expression or overexpression of PTEN,the proliferation ability,cell migration and invasion numbers of the RL95-2 cells as well as expression levels of p-PI3K and p-AKT proteins were significantly reduced,while the apoptosis rate was significantly increased(P<0.05);overexpression of miR-106 could significantly reverse the inhibitory effects of overexpression of PTEN on the proliferation,migration and invasion of RL95-2 cells and the promotion effect on apoptosis(P<0.05).Conclusion MiR-106 can target and inhibit the expression of PTEN and promote the proliferation,migration and invasion of human endometrial cancer cells by activating the PI3K/AKT pathway,and inhibit cell apoptosis.