MiR-107对柯萨奇病毒B3感染细胞糖原合成酶激酶-3β蛋白及β-连环蛋白表达的影响

Effect of miR-107 on expression of glycogen synthase kinase-3β and β-catenin in CVB3-infected cells

摘要:

【背景】MiR-107异常表达可引起肿瘤细胞中Wnt/β-catenin信号通路主要蛋白表达发生改变，但其能否在柯萨奇病毒B3（coxsackievirus B3，CVB3）感染的人宫颈癌细胞（HeLa cells）中发挥同样作用却未见报道。【目的】探讨miR-107能否影响CVB3感染HeLa细胞中的糖原合成酶激酶-3β（GSK-3β）蛋白、P-GSK-3β蛋白和β连环蛋白（β-catenin）的表达水平。【方法】体外培养HeLa细胞，感染CVB3不同时间，通过显微镜观察HeLa细胞的形态学变化、实时荧光定量PCR实验检测HeLa细胞中miR-107表达量、免疫印迹实验检测HeLa细胞中的GSK-3β、P-GSK-3β、β-catenin蛋白及病毒衣壳蛋白（VP1）的表达水平。【结果】CVB3感染HeLa细胞6 h后，细胞病变效应明显，miR-107表达量及GSK-3β、P-GSK-3β和VP1蛋白的表达水平随CVB3感染时间（0—8 h）的延长逐渐增加，而β-catenin蛋白的表达水平逐渐减少。过表达miR-107的CVB3感染6 h的HeLa细胞死亡细胞增多，GSK-3β、P-GSK-3β和VP1蛋白表达水平增加（P<0.05），β-catenin蛋白表达水平减少（P<0.001）；抑制miR-107的CVB3感染6 h的HeLa细胞GSK-3β、P-GSK-3β及VP1蛋白表达水平明显减少（P<0.05），β-catenin蛋白表达水平明显增加（P<0.05）。【结论】MiR-107异常表达可影响CVB3感染HeLa细胞中Wnt/β-catenin信号通路蛋白和病毒衣壳蛋白的表达水平。

关键词:柯萨奇病毒B3;病毒性心肌炎;miR-107;Wnt/β-catenin

Abstract:

[Background] Abnormal expression of miR-107 can change the expression of the main proteins of Wnt/β-catenin signaling pathway in tumor cells, while miR-107 plays the same role in human cervical cancer cells (HeLa cells) infected with coxsackievirus B3 (CVB3) has not been reported. [Objective] To investigate whether miR-107 can affect the expression levels of glycogen synthase kinase-3β (GSK-3β), P-GSK-3β, and β-catenin in CVB3-infected HeLa cells. [Methods] The HeLa cells were cultured in vitro and infected with CVB3 for different time periods. The morphological changes of HeLa cells were observed by microscopy. The expression of miR-107 in HeLa cells was measured by real-time fluorescent quantitative PCR. The protein levels of GSK-3β, P-GSK-3β, β-catenin and viral capsid protein (VP1) in HeLa cells were determined by western blotting. [Results] After CVB3 infection for 6 h, the cytopathic effect was obvious. The expression level of miR-107 and the protein levels of GSK-3β, P-GSK-3β, and VP1 increased, while the protein level of β-catenin decreased, with the prolongation of CVB3 infection time (0−8 h). MiR-107 overexpression in the HeLa cells infected with CVB3 for 6 h increased the cell deaths, up-regulated the protein levels of VP1, GSK-3β, and P-GSK-3β (P<0.05), and down-regulated the protein level of β-catenin (P<0.001). The inhibition of miR-107 expression in the HeLa cells infected with CVB3 for 6 h reduced the cell deaths, down-regulated the protein levels of VP1, GSK-3β, and P-GSK-3β (P<0.05), and up-regulated the protein level of β-catenin (P<0.05). [Conclusion] Abnormal expression of miR-107 can affect the expression levels of proteins in the Wnt/β-catenin signaling pathway and VP1 in CVB3-infected HeLa cells.

Key words:coxsackievirus B3;viral myocarditis;miR-107;Wnt/β-catenin