Hsa-miR518c-5p Regulates Trophoblast Invasion And Migration Through P120/WISP2 And Participates In The Pathogenesis Of Preeclampsia

Objective.Preeclampsia(PE)is a disease with high morbidity and mortality in pregnancy.Poor placental trophoblast invasion contributes to the pathogenesis of PE.hsa-miR518c-5p regulates carcinoma cells migration and invasion,however,its regulatory role in trophoblasts is unrevealed.The expression pattern of hsa-miR518c-5p in PE has not been reported.In the present study,the effect of hsa-miR518c-5p on the invasion and migration of trophoblast cells is investigated.Methods.Placental tissues of PE patients were collected,the expression of hsa-miR518c-5p in placentae was detected by quantitative real-time PCR(qRT-PCR)and in situ hybridization of DIG-labeled probe with hsa-miR518c-5p in paraffin sections of human placentas.The correlation between hsa-miR518c-5p and maternal blood pressure was analyzed.To explore the biofunction of hsa-miR518c-5p in trophoblast,hsa-miR518c-5p was overexpressed or silenced in the trophoblast cell line HTR-8/SVneo.Transwell analysis,wound healing assay,villous explant in vitro culture were performed to investigate the migration and invasion of HTR8 cells.Edu staining and PI staining followed by flow cytometry detection were performed to determine proliferation of HTR8 cells.In the mechanistic study,mRNA-sequencing was conducted and CTNND1,a gene coding p120 catenin(p120),was found downregulated in miR-518c-5p mimics transfected HTR8 cells.Luciferase assays and western blotting were also performed to verify the regulatory relationship of hsa-miR518c-5p to p120.In addition,the relationship between p120 and WISP2 was verified by immunofluorescence staining and co-immunoprecipitations.The regulation of p120/WISP2 signaling on trophoblast invasion and migration was verified by a series of rescue experiments.Results.The expression of hsa-miR518c-5p was decreased in placental tissue of PE patients,and its expression was negatively correlated with systolic and diastolic blood pressure of pregnant women.Knockdown of hsa-miR518c-5p in HTR-8/SVneo resulted in activation of Snail/E-cadherin signalling pathway and decreased cell invasive ability.Transcriptome sequencing and WB experiments confirmed that the expression of p120 was inhibited by hsa-miR518c-5p,and luciferase experiments showed that p120 was the target gene of hsa-miR518c-5p.Using forward and reverse co-immunoprecipitation experiments,it was clarified that p120 regulates the expression of WSP2 protein by binding to WISP2 protein molecule and then affects the downstream Snail/E-cadherin pathway.Finally,the rescue experiments showed that forced overexpression of p120 attenuated hsa-miR518c-5p mediated regulation of WISP2/Snail/E-cadherin pathway and trophoblast invasion and migration.Conclusion.The expression level of hsa-miR518c-5p is decreased in the placenta of PE patients.hsa-miR518c-5p promotes trophoblast invasion and migration by targeting p120 mRNA.The p120 regulates WISP2 protein accumulation by directly interaction with WISP2 protein,and therefore regulates the downstream Snail/E-cadherin signaling.The study provides a new insight in understanding the trophoblast shallow invasion caused pathology of PE.