The Functional Mechanism Of MiR-181a-5p In Invasion And Migration Of Placental Trophoblast Cells

BackgroundAbnormalities of trophoblast cell differentiation can lead to abnormal invasion and migration of trophoblast cells,which can lead to pregnancy-related diseases such as pre-eclampsia(PE).Preeclampsia is an idiopathic hypertension syndrome that is unique to multiple organs and multiple systems during pregnancy.It occurs more than 20 weeks after pregnancy.It is mainly manifested in hypertension,proteinuria and other symptoms.The incidence rate is about 3% to 5%,which is one of the important causes of maternal and infants deaths.Micro RNAs(miRNAs)are short-chain and non-coding small molecules that regulate post-transcriptional gene expression.During pregnancy,miRNAs participate in a series of reactions,including endometrium preparation,genetic recombination associated with immune responses,placental development,and angiogenesis,and have long been considered to be associated with placental dysplasia and pregnancy complications.It has confirmed that a large number of miRNAs are expressed in the placenta,and abnormal expression of miRNAs in the placenta may be one of the causes of preeclampsia.However,the effects of miRNA on placental development and its functional mechanism need to be further explored.In our previous work,it has shown that miR-181a-5p is up-regulated in plasma and placental expression in patients with severe pre-eclampsia.In order to understand the pathogenesis of severe preeclampsia,this project focuses on the role and molecular mechanism of miR-181a-5p in placental trophoblast invasion and migration.ObjectivesThis study elucidated the role and potential molecular mechanisms of miRNA-181a-5p in placental trophoblast invasion and migration,in order to gain a deeper understanding of the roles of miRNA in placental development and pre-eclampsia.Materials and Method1.According to the inclusion and exclusion criteria to collect placental tissue samples.2.By using four free databases(including TargetScan 7.0,PITA,PicTar,and miRanda)to predict miR-181a-5p directly target genes,IGF2BP2 was selected as the miR-181a-5p target gene and subsequent experiments were designed.3.In sPE placenta,RT-q PCR was used to detect the expression level of miRNA-181a-5p and Western Blot was used to detect the IGF2BP2.4.Transwell assay was used to study the effect of miR-181a-5p on invasion and migration ability of HTR-8/SVneo cells.5.After constructing the luciferase vector and co-transfecting it with the miR-181a-5p mimic or inhibitor,the activity of the luciferase reporter gene was measured by Dual-luciferase assay,and RT-qPCR and Western Blot were performed to exam the effect of miR-181a-5p expression on IGF2BP2 mRNA and protein levels in HTR-8/SVneo cells to verify whether miR-181a-5p directly inhibits IGF2BP2.6.Transwell assay was used to investigate the effect of overexpression or inhibition of IGF2BP2 on the invasion and migration of HTR-8/SVneo cells.7.After IGF2BP2 and miR-181a-5p were co-overexpressed in HTR-8/SVneo cells,the expression of IGF2BP2 was detected by Transwell assay and Western Blot,so as to explore whether the effect of miR-181a-5p inhibit invasion and migration is mediated through direct inhibition of IGF2BP2.Results1.In sPE placenta,miRNA-181a-5p expression was significantly up-regulated,but IGF2BP2 protein expression was significantly down-regulated.2.miR-181a-5p inhibited invasion and migration of HTR-8/SVneo cells,while IGF2BP2 promoted invasion and migration of HTR-8/SVneo cells.3.miR-181a-5p directly inhibits IGF2BP2.4.miR-181a-5p inhibits IGF2BP2 through a conserved binding site on IGF2BP2 mRNA 3’-UTR.5.miR-181a-5p inhibits invasion and migration of trophoblast cells by directly inhibiting IGF2BP2.ConclusionThis study demonstrated that miR-181a-5p inhibits invasion and migration of placental trophoblast cells,and its role is mediated through direct inhibition of IGF2BP2,thus revealing the potential role of miR-181a-5p in the pathogenesis of severe preeclampsia.