EM患者子宫内膜组织的ESCs中miR-539表达水平及其靶向MMP-9对ESCs侵袭和迁移的影响

The effects of expression level of miR-539 in ESCs of endometrium in patients with endometriosis and its targeting MMP-9 on the migration and invasion of endometrial stromal cells

目的探究子宫内膜异位症(EM)患者子宫内膜组织的子宫内膜基质细胞(ESCs)中miR-539表达水平,以及miR-539靶向基质金属蛋白酶9(MMP-9)对ESCs侵袭和迁移的影响。方法收集60例EM患者子宫内膜异位组织及47例正常子宫内膜组织,分离ESCs原代培养,实时荧光定量PCR检测miR-539和MMP-9 mRNA表达水平,并采用双荧光素酶报告基因检测其靶向关系。利用Lipofectamine 3000试剂转染ESCs,并将其分为miR-539 mimics组、mimics NC组、miR-539 inhibitors组和inhibitors NC组。CCK-8检测各组细胞增殖情况;划痕实验检测细胞迁移;Transwell检测细胞侵袭;Western blot法检测各组细胞中MMP-9蛋白表达水平。结果EM组miR-539 m RNA水平显著低于对照组,MMP-9 mRNA水平显著高于对照组(P<0.05)。双荧光素酶报告实验结果显示miR-539与MMP-9具有明显的靶向关系(P<0.05)。与mimics NC组相比,miR-539 mimics组ESCs细胞增殖、划痕愈合率、侵袭细胞数量及MMP-9蛋白表达水平均显著降低(P<0.05);与inhibitors NC组相比,miR-539 inhibitors组ESCs细胞增殖、划痕愈合率、侵袭细胞数量及MMP-9蛋白表达水平均显著升高(P<0.05)。结论miR-539可靶向调节MMP-9表达水平,影响ESCs的增殖、迁移和侵袭。

Objective To explore the expression level of miR-539 in endometrial stromal cells(ESCs)of endometrium in patients with endometriosis(EM),and the effects of miR-539 targeting matrix metalloproteinase 9(MMP-9)on the invasion and migration of ESCs.Methods The endometrial samples of 60 EM patients and 47 non-EM patients admitted to Hubei Province Maternal and Child Health Hospital from 2016 to 2020 were collected.The ESCs were isolated for primary culture.The expression levels of miR-539 and MMP-9 mRNA were detected with qPCR technology,and its targeting relationship was detected with dual luciferase reporter gene.ESCs were transfected with Lipofectamine 3000 reagent and divided into the miR-539 mimics group,the mimics NC group,the miR-539 inhibitor group and the inhibitor NC group.CCK-8 was used to detect cell proliferation in each group.Scratch test was used to detect cell migration.Transwell was used to detect cell invasion.Western blot method was used to detect the expression levels of MMP-9 protein in each group of cells.Results The level of miR-539 mRNA in endometrium was significantly lower in the EM group than that of the control group(P<0.05),and the level of MMP-9 mRNA was significantly higher than that of the control group(P<0.05).The results of double luciferase report showed that miR-539 had an obvious targeting relationship with MMP-9(P<0.05).Compared with the mimics NC group,the ESCs cell proliferation,scratch healing rate,number of invasive cells and MMP-9 protein level were significantly reduced in the miR-539 mimics group(P<0.05).Compared with the inhibitor NC group,the ESCs cell proliferation,scratch healing rate,number of invaded cells and MMP-9 protein level were significantly increased in the miR-539 inhibitor group(P<0.05).Conclusion MiR-539 can target and regulate the expression level of MMP-9 and affect the cell proliferation,migration and invasion abilities of ESCs.