Exosomal miR-199a-3p derived from follicular fluid modulates the immune microenvironment in polycystic ovary syndrome by regulating macrophage polarization

Polycystic ovary syndrome (PCOS) is characterized by chronic low-grade inflammation of the ovaries. This disorder of the immune microenvironment is closely related to abnormal follicular development and ovulation disorders. This study aims to investigate whether the exosomes derived from the follicular fluid of PCOS patients regulate macrophage polarization by delivering miR-199a-3p, and further participate in the molecular mechanism of ovarian immune microenvironment imbalance. Exosomes were extracted from the follicular fluid of both PCOS patients and control subjects. The expression levels of miR-199a-3p in exosomes were quantified through Quantitative reverse transcription PCR, while the concentrations of inflammatory cytokines were measured through ELISA. Bioinformatics analysis predicted mTOR as a potential target of miR-199a-3p. To investigate functional interactions, PCOS-derived exosomes were co-cultured with THP-1-derived macrophages, either in the presence or absence of miR-199a-3p mimics. After 48 h of incubation, the expression profiles of inflammatory factors(TNF-α, IL-6, IL-10 and TGF-β1), miR-199a-3p, M1/M2 macrophage polarization markers (CD86 and CD206), and mTOR were systematically analyzed. PCOS patients exhibited significantly upregulated miR-199a-3p in follicular fluid exosomes, along with elevated pro-inflammatory factors (TNF-α, IL-6) and decreased levels of anti-inflammatory cytokines (IL-10, TGF-β1). In an in vitro exosome-macrophage co-culture experiment, miR-199a-3p overexpression enhanced the pro-inflammatory phenotype, further increasing TNF-α and IL-6 while decreasing IL-10 and TGF-β1. It also promoted M1 polarization (upregulated expression of CD86) and suppressed M2 polarization (downregulation of CD206 expression) through mTOR downregulation. These findings demonstrate that exosomes derived from the follicular fluid of PCOS patients transfer miR-199a-3p to macrophages. This transfer disrupts the delicate balance of M1/M2 polarization, thereby fostering the establishment of a pro-inflammatory microenvironment within the ovarian tissue. Follicular fluid exosomes from PCOS patients may target and inhibit the expression of mTOR in macrophages by delivering highly expressed miR-199a-3p, thereby hindering M2 polarization and promoting M1 polarization, ultimately leading to a shift of the local ovarian immune microenvironment towards a pro-inflammatory state. These findings offer novel molecular insights into exosome-mediated immune regulation within the pathogenesis of PCOS, highlighting a promising direction for the development of cell-free therapeutic strategies. This study has been registered with the International Traditional Medicine Clinical Trial Registry under the registration number ITMCTR2025001857, with the registration date recorded as September 26, 2025.