miR-889-3p调控vimentin乙酰化对妊娠糖尿病胎盘滋养层细胞侵袭功能的影响

miR-889-3p attenuates invasive function of placental trophoblast cells in gestational diabetes mellitus by promoting vimentin acetylation

目的:探讨微小RNA-889-3p(miR-889-3p)调节妊娠糖尿病(GDM)胎盘滋养层细胞侵袭功能的作用机制。方法:将2020年1~12月的30名GDM产妇和30名正常葡萄糖耐量产妇选入本研究。通过qRT-PCR检测miR-889-3p在产妇胎盘、人肿瘤细胞系、人滋养层HTR-8/SVneo细胞和人脐静脉内皮细胞(HUVEC)中的表达水平。进行细胞培养、细胞转染、MTT测定、EdU检测、伤口划痕实验和Transwell实验,以确定miR-889-3p的沉默和过表达对HTR-8/SVneo细胞生物功能的影响。通过生物信息学分析确定miR-889-3p与波形蛋白(vimentin)的结合关系。分别使用双萤光素酶报告基因检测系统和蛋白质印迹分析miR-889-3p过表达对双萤光素酶活性和vimentin乙酰化的影响。结果:HTR-8/SVneo细胞的相对miR-889-3p表达水平高于其他肿瘤细胞和HUVEC,GDM产妇胎盘的相对miR-889-3p表达水平低于正常葡萄糖耐量产妇胎盘(P<0.05)。miR-889-3p过表达抑制HTR-8/SVneo细胞生长、迁移和侵袭(P<0.05),而miR-889-3p敲减促进细胞生长、迁移和侵袭(P<0.05)。通过生物信息学分析确定vimentin是miR-889-3p的直接靶点。在转染vimentin-WT的HTR-8/SVneo细胞中,miR-889-3p模拟物的施用抑制了萤光素酶活性(P<0.05),并显著提高vimentin乙酰化水平(P<0.05)。上调vimentin部分拮抗miR-889-3p过表达对细胞生长、迁移和侵袭的抑制作用(P<0.05),而下调vimentin减弱了由miR-889-3p敲减诱导的细胞生长、迁移和侵袭增强(P<0.05)。结论:胎盘组织中miR-889-3p的下调参与GDM病理过程。miR-889-3p通过促进vimentin乙酰化来减弱滋养层细胞的侵袭功能。

AIM:To explore the mechanism of microRNA-889-3p(miR-889-3p)regulating the invasion function of placental trophoblastic cells in gestational diabetes mellitus(GDM).METHODS:From January to December2020,30 parturient women with GDM and 30 parturient women with normal glucose tolerance were enrolled in this study.The expression levels of miR-889-3p in maternal placenta,human cancer cell lines,human trophoblast HTR-8/SVneo cells and human umlibical vein endothelial cells(HUVEC)were detected by qRT-PCR. Cell culture,cell transfection,MTT assay,EdU assay,wound scratch assay,and Transwell analysis were carried out to determine the effects of silencing and overexpression of miR-889-3p on the HTR-8/SVneo cells. The binding relationship between miR-889-3p and vimentin was determined by bioinformatics analysis. The effects of miR-889-3p overexpression on dual-luciferase activity and vimentin acetylation were analyzed by dual-luciferase reporter assay and Western blotting,respectively.RESULTS:Relative miR-889-3p expression level in HTR-8/SVneo cells was higher than that in cancer cells and HUVEC,and was lower in the GDM placentas than that in the normal placentas(P<0. 05). miR-889-3p overexpression inhibited the growth,migration and invasion of HTR-8/SVneo cells(P<0. 05),while miR-889-3p knockdown promoted cell growth,migration and invasion(P<0. 05). Vimentin was identified as a direct target of miR-889-3p by bioinformatics analysis. In the HTR-8/SVneo cells transfected with vimentin-WT,administration of miR-889-3p mimic significantly inhibited luciferase activity and increased vimentin acetylation level(P<0. 05). Up-regulation of vimentin partially antagonized the inhibitory effect of miR-889-3p overexpression on cell growth,migration and invasion(P<0. 05),while enhancement of cell growth,migration and invasion induced by miR-889-3p knockdown was attenuated by down-regulation of vimentin(P<0. 05).CONCLUSION:Down-regulation of miR-889-3p in placental tissue is involved in the pathological process of GDM. miR-889-3p attenuates the invasive function of trophoblast cells by up-regulating vimentin acetylation.