microRNA-145在宫颈癌中的表达及其对Wnt/β-catenin信号通路的抑制作用

The expression of microRNA-145 in cervical cancer and its inhibitory effects on Wnt/β-catenin signaling pathway

目的:探讨microRNA-145在宫颈癌中表达的临床意义及其对Wnt/β-catenin信号通路的调控作用。方法:收集2002年1月至2004年12月解放军464医院62例宫颈癌患者的组织标本,采用实时荧光定量PCR方法检测microRNA-145表达并将其分为高表达组和低表达组,分析其与临床病理参数的关系。将microRNA-145表达质粒和无义对照质粒分别转染入宫颈癌HeLa中,分为过表达microRNA-145组和对照组,采用细胞免疫荧光染色法分析细胞中β-catenin表达定位的变化,双荧光素酶报告系统检测细胞中TCF/LEF转录活性及与Cateninδ-1的直接作用,采用Western blot法检测microRNA-145对Cateninδ-1、C-MYC和CyclinD1表达的影响。结果:microRNA-145低表达组患者的总生存期为(41.28±2.00)个月,高表达组为(46.06±0.95)个月,两者比较差异具有统计学意义(P<0.05),低表达组患者预后较差。过表达microRNA-145组HeLa细胞中的β-catenin主要分布于胞浆,对照组主要位于细胞核和胞浆;过表达microRNA-145组细胞中TCF/LEF转录活性受到抑制,伴随着Cateninδ-1、C-MYC和CyclinD1表达下调,microRNA-145与Cateninδ-1可直接结合发挥作用。结论:microRNA-145可通过与Cateninδ-1直接作用,抑制Wnt/β-catenin信号通路转导,从而发挥抑制宫颈癌演进的生物学功能。

Objective:To explore the clinic-pathological significance of microRNA-145 expression in human cervical cancer and its effects on Wnt/β-catenin signaling pathway.Methods:Real-time PCR was used to detect the expression of microRNA-145 in 62 cervical cancer samples.The correlation between microRNA-145 expression and the clinic-pathological parameters and its prognostic significance was analyzed.MicroRNA-145-expressing plasmid and non-sense plasmid were transfected into human cervical cancer HeLa cells,assigned into overexpressed microRNA-145 group and control group.Immunofluorescence staining was performed to detect the expression location ofβ-catenin.Top Flash luciferase reporter assay was performed to investigate the effects of microRNA-145 on the transcriptional activity of TCF/LEF and direct interactions with Cateninδ-1.Western blot was used to detect the effects of microRNA-145 on the expression of Cateninδ-1,C-MYC,and CyclinD1.Results:The patients with low microRNA-145 expression showed poorer prognosis((41.28±2.00)months vs.(46.06±0.95)months,P<0.05)β-catenin immunofluorescence was distributed within the cytoplasm in the microRNA-145-overexpressed HeLa cells,but mainly within the nucleus and cytoplasm in the control cells.The luciferase reporter system indicated that the transcriptional activity of TCF/LEF was inhibited in the microRNA-145-overexpressed HeLa cells,and validated Cateninδ-1 was a target of miR-145.The expression of Cateninδ-1,C-MYC,and CyclinD1 was decreased in the microRNA-145-overexpressed HeLa cells.Conclusions:microRNA-145 may inhibit cervical cancer progression via Cateninδ-1 and inhibit the Wnt/β-catenin signaling pathway.