Effect of microRNA-106b on the invasion and proliferation of trophoblasts through targeting MMP-2 / 中华妇产科杂志

ABSTRACT

Objective To investigate the expression of microRNA-106b(miR-106b)in the placentas of patients with pre-eclampsia and its relationship with matrix metallopeptidase(MMP)-2,and its effect on the invasion and proliferation of trophoblasts. Methods (1) Placental tissues were collected from patients with mild pre-eclampsia (mPE, n=30) , severe pre-eclampsia (sPE, n=30) and normal pregnant women (n=40). Human choriocarcinoma cell lines JAR and JEG3 were assigned to the miR-106b mimics group, the mimics negative control group, the miR-106b inhibitor group and the inhibitor negative control group, respectively. (2) The target gene of miR-106b(such as MMP-2) was predicted by bioinformatics. Dual-luciferase reporting system was used to verify the regulation of miR-106b on the expression of MMP-2. (3) The expressions of miR-106b and MMP-2 were measured by quantitative real-time PCR (qRT-PCR) and western blot. (4) Cell proliferation was determined by MTT assay. (5) Invasive activities in each group were assessed by cell transwell invasion assays. Results (1) Predicting result of bioinformatics indicated that MMP-2 was one of the target genes of miR-106b. Dual-luciferase activity assay demonstrated that MMP-2 was the direct target of miR-106b(P＜0.01).(2) The results of qRT-PCR.①The expression of miR-106b in the placentas of mPE, sPE, normal pregnant women were 2.89±0.04, 1.96±0.03, 1.01±0.03, respectively (P＜0.05). And the expression of MMP-2 mRNA in the placentas of mPE, sPE, normal pregnant women were 1.87±0.05, 0.69±0.03, 2.78±0.03, respectively (P＜0.05).②The expression of miR-106b in the JAR cell line in the miR-106b mimics group, the mimics negative control group, the miR-106b inhibitor group and the inhibitor negative control group were 2.39 ± 0.03, 1.03 ± 0.04, 0.73 ± 0.03, 1.11 ± 0.04, respectively (P＜0.05). And its expression in the JEG3 cell line were 2.17±0.04, 1.18±0.04, 0.61±0.03 and 1.22±0.03, respectively (P＜0.05). ③The expression of MMP-2 mRNA in the JAR cell line in the miR-106b mimics group, the mimics negative control group, the miR-106b inhibitor group and the inhibitor negative control group were 0.45±0.15, 1.02±0.03, 2.28±0.03, 1.11±0.03, respectively (P＜0.05). And its expression in the JEG3 cell line were 0.58±0.03, 1.25±0.15, 2.25±0.03, 1.21±0.03, respectively (P＜0.05). (3) The results of western blot.①The expression of MMP-2 protein in the placentas of mPE, sPE, normal pregnant women were 1.63 ± 0.04, 0.55±0.03, 2.82±0.03, respectively (P＜0.05).②The expression of MMP-2 protein in the JAR cell line in the miR-106b mimics group, the mimics negative control group, the miR-106b inhibitor group and the inhibitor negative control group were 0.41 ± 0.03, 0.97 ± 0.03, 2.25 ± 0.03, 1.01 ± 0.03, respectively (P＜0.05). And its expression in the JEG3 cell line were 0.53±0.03, 1.20±0.03, 2.31±0.04, 1.19±0.03, respectively (P＜0.05). (4) miR-106b could inhibit the proliferation of JAR and JEG3 cells, cell proliferation rates in the miR-106b mimics group were lower than that in the mimics negative control group (P＜0.05). And cell proliferation rate in the miR-106b inhibitor group was higher than the inhibitor negative control group (P＜0.05) . (5) The numbers of JAR cell that passed the membrane in the miR-106b mimics group, the mimics negative control group. The miR-106b inhibitor group and the inhibitor negative control group were 61±15, 79±13, 134±13, 80±12, respectively( P＜0.05). And the numbers of JEG3 cell that passed were 57±12, 71±15, 128±15, 70± 14, respectively (P＜0.05). Conclusion The miR-106b could inhibit the invasion and proliferation of JAR and JEG3 cells through targeting MMP-2, and have a relationship with the pathogenesis of pre-eclampsia.