microRNA-375在肝纤维化中的作用及其调控信号通路的研究

目的通过提高肝星状细胞(HSC)中微小核糖核酸(microRNA-375,miR-375)的表达水平,探讨miR-375在肝纤维化进程中的作用及其靶向信号通路。方法通过瞬时转染将miR-375mimic或者miR-375 mimic negative control(NC)转染到大鼠肝星状细胞(HSC-T6),将细胞分为mimic组、NC组和空白对照组。通过实时荧光定量聚合酶链反应(qRT-PCR)、免疫印迹实验(Western blot)检测细胞中蛋白激酶B(Akt)基因、3-磷酸肌醇依赖性蛋白激酶1(PDK1)基因表达水平的变化。分别用细胞流式技术、细胞计数实验(CCK-8)试剂盒和细胞迁移实验检测过表达的miR-375对HSC-T6的细胞周期、增殖和迁移能力的影响。结果与其他两组相比,mimic组细胞中Akt、PDK1含量明显下降(P0.05),细胞停滞在G0/G1期的比例升高(P0.05)、增殖受到抑制(P0.05),细胞迁移的差异无统计学意义(P0.05)。结论miR-375可抑制HSC的增殖,负性调控肝纤维化进程。

Function and signaling pathway regulated by micro RNA-375 in hepatic fibrosis

Objective To explore the function and signaling pathway regulated by microRNA-375 (miR-375 )in hepatic fibrosis,the expression levels of miR-375 in hepatic stellate cells (HSC)were increased. Methods The miRNA-375 mimic or miRNA-375 mimic negative control (NC)was transduced into the HSC-T6 cell lines. All cells were divided into three groups and designated as the mimic group,the NC group,and the blank control group. The qRT-PCR and western blot were used to test the change of expression levels of Akt and PDK1 gene. The change of HSC-T6 cell cycle,cell proliferation and cell migration were assessed by cell cycle assay,cell proliferation assay and cell migration assay. Results The expression levels of Akt and PDK1 gene in the mimic group were found lower than those of other groups (P<0.05 ),while the G0/G1 phase fractions and the cell growth inhibition rate of the mimic group were higher than those of other groups (P<0.05). However, there was no significant difference in cell migration of the three groups (P>0.05). Conclusions miRNA-375 suppresses the process of liver fibrosis by inhibiting the proliferation of HSC and arresting the cell cycle of them at the G0/G1 phaseinvitro.