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目的观察5-氮杂-2’-脱氧胞苷(5-Aza)对TGF-β1诱导的HSC-T6细胞株活化状态及miR-29b基因甲基化的影响。方法采用MTT、qRT-PCR及Western blot技术检测5-Aza治疗组细胞活化状态及miR-29b含量;BSP及TA克隆检测miR-29b启动子区CpG岛甲基化的改变。结果相较TGF-β1组,5-Aza组细胞增殖率下调为58.62%(P<0.05)且Ⅰ型胶原及α-SMA的mRNA及蛋白均显著下降(P<0.05)。miR-29b在5-Aza治疗后呈时间依赖性上升(P<0.05)。BSP及TA克隆显示5-Aza组miR-29b甲基化率为7.27%,呈现下调(P<0.05)。结论5-Aza可负性调控肝纤维化进程,这与miR-29b启动子区去甲基化进程密切相关。
Objective To investigate the effect of 5-Aza-5-Aza on the activation of HSC-T6 cells induced by TGF-β1 and the methylation of miR-29b gene. Methods MTT, qRT-PCR and Western blot were used to detect the activation of cells and the content of miR-29b in 5-Aza treatment group. The methylation of CpG island in miR-29b promoter region was detected by BSP and TA cloning. Results Compared with TGF-β1 group, the proliferation rate of 5-Aza group was decreased by 58.62% (P <0.05) and the mRNA and protein of type Ⅰ collagen and α-SMA were significantly decreased (P <0.05). miR-29b increased in a time-dependent manner after 5-Aza treatment (P <0.05). BSP and TA cloning showed that the methylation rate of miR-29b in 5-Aza group was 7.27%, which was down-regulated (P <0.05). Conclusions 5-Aza can negatively regulate the progress of hepatic fibrosis, which is closely related to the process of demethylation of miR-29b promoter region.