抑制miR-18a的水平增加HTR8人正常滋养细胞雌激素受体1的表达并促进细胞凋亡

Inhibition of miR-18a increases expression of estrogen receptor 1 and promotes apoptosis in human HTR8 trophoblasts

目的研究子痫前期相关微小RNA-18a(miR-18a)对人正常滋养细胞(HTR8)中雌激素受体1(ER1)的调控作用及对HTR8细胞周期及凋亡的影响。方法使用miR-18a前体分子(pre-miR-18a)分组转染HTR8细胞,实验分为pre-miRNA阴性对照组、miR-18a过表达组、miR-18a抑制组。反转录PCR(RT-PCR)检测转染后各组细胞中miR-18a mRNA的表达水平;RT-PCR及Western blot法分别检测各组细胞中miR-18a的靶基因ER1在mRNA和蛋白水平的表达;流式细胞术检测转染后各组细胞周期及凋亡的变化。结果与pre-miRNA阴性对照组相比,miR-18a过表达和miR-18a抑制均获得明显效果;抑制miR-18a表达后,ER1 mRNA和蛋白水平增加;过表达和抑制miR-18a对HTR8细胞周期无明显影响;抑制miR-18a可增加HTR8细胞凋亡。结论抑制miR-18a的水平增加HTR8人正常滋养细胞雌激素受体1的表达并促进细胞凋亡。

Objective To investigate the role of pre-eclampsia related miR-18 a in regulating estrogen receptor 1（ ER1）expression,cell cycle and apoptosis in HTR8 human trophoblasts. Methods HTR8 cells were transfected with synthesized negative control（ NC） miRNA,miR-18 a and miR-18 a inhibitor,respectively. The mRNA expressions of miR-18 a and ER1 were determined by reverse transcription PCR（ RT-PCR）. The protein expression level of ER1 was examined by Western blotting. The cell cycle and apoptosis were evaluated by flow cytometry. Results The mRNA level of miR-18 a was significantly up-regulated in miR-18a-transfected group,while down-regulated in miR-18 a inhibitor group as compared with the NC group in HTR8 cells. Compared with the NC group,both mRNA and protein expression levels of ER1 were significantly higher in the miR-18 a inhibitor group. Cell cycle analysis showed that no significant difference was observed among the three groups. The apoptosis rate was significantly higher in the miR-18 a inhibitor group as compared with the other two groups.Conclusion Inhibition of miR-18 a increases the expression of ER1 and promotes apoptosis in HTR8 human trophoblasts.