Ovarian Stimulation Altered Uterine Fluid Extracellular Vesicles miRNA Affecting Implantation in Rats.

Abstract

Uterine fluid (UF) extracellular vesicle (EV) miRNA may affect implantation and could be the potential biomarker of endometrial receptivity (ER). Ovarian stimulation (OS) could damage the ER but its mechanism is still unclear. Here, we evaluate the affections of OS on UF EV miRNA expression and implantation. Female rats were divided into three groups: natural cycle or injection with GnRH-a following HP-HMG or u-FSH. UF was collected on the 5th day of gestation. Affinity membrane columns were utilized to isolate EVs from UF, obtained during implantation flushing. The EV miRNAs were sequenced, and five of them were validated by qRT-PCR. HTR-8/Svneo cells were transfected with miR-223-3p mimic and inhibitor, followed by conducting colony formation, invasion, migration, and adhesion assays to assess the cellular functions. In OS groups, the implantation rate decreased (p < 0.05), and the pinopode was damaged in the OS groups. The EVs were isolated from UF, and the differential expression key miRNAs were involved in several regulation pathways, such as cancer, endocrine, and cell cycles, which were correlated with ER and implantation. Among the miRNAs, miR-223-5p greatly differed and was most consistent with the sequencing results, followed by miR-223-3p and miR-98-5P. miR-223-3p promoted HTR-8/SVneo cells grow and ability of invasion, migration, and adhesion. OS altered UF EVs miRNAs affecting implantation in rats, and miR-223-3p might be the key molecule.