Controlled ovarian hyperstimulation reduces mouse embryo implantation capacity through uterine HIF-2α mediated pathway

Abstract

Controlled ovarian hyperstimulation (COH) is a cornerstone of assisted reproductive technologies, yet its effects on endometrial function and embryo implantation remain poorly understood, particularly regarding the role of hypoxia-inducible factor 2α (HIF-2α) signaling. Therefore, the objective of this study was to investigate whether COH-induced endometrial dysfunction impairs mouse embryo implantation through the HIF-2α pathway. A COH mouse model was established using gonadotropin-releasing hormone agonist (GnRH-a)/human menopausal gonadotropin (hMG)/human chorionic gonadotropin (hCG) administration. Embryo implantation status was evaluated on gestational days 5, 6, and 20; endometrial tissues were analyzed for HIF-2α pathway activity via immunohistochemistry (IHC), immunofluorescence (IF), Western blot (WB) and quantitative real-time PCR (qRT-PCR); histological changes were assessed by Jones silver staining and transmission electron microscopy (TEM); serum estradiol (E₂), progesterone (P₄), and prolactin (PRL) levels were measured by radioimmunoassay (RIA). COH mice exhibited reduced total embryo implantation rates (on day 5, 6 and 20), together with decreased serum E₂, P₄, and PRL levels. COH mice exhibited preserved luminal epithelium integrity with increased microvillus density and continuous basement membrane structure. qRT-PCR and WB demonstrated significantly downregulated HIF-2α expression at both mRNA and protein levels, accompanied by reduced downstream RAB27B (member of RAS oncogene family)/matrix metalloproteinase 9 (MMP9) and lysyl oxidase (LOX)/adrenomedullin (ADM) signaling, which associated with impaired luminal epithelium detachment and compromised trophoblast invasion. Together, these findings identify HIF-2α as a potential key mediator of COH-induced endometrial microenvironment alterations, revealing molecular mechanisms underlying implantation failure. Importantly, the HIF-2α-RAB27B/MMP9 and HIF-2α-LOX/ADM axes are highlighted as promising therapeutic targets to optimize assisted reproductive outcomes.