P-716 exosomes from human follicular fluid present a different miRNA and proteins composition in case of PCOS and impact granulosa cell activities

Abstract Study question how human exosomes from PCOS ovarian cells could change the activity of granulosa cells ? Summary answer miRNA and proteins presented in exosomes from human follicular fluid change cell proliferation and steroid secretion of granulosa cells by activating the STAT3 pathway. What is known already Main follicular fluid (FF) factors include proteins, metabolites, as well as extracellular vesicles (exosomes). All these secretory products participate to intertalk between different cell types in the ovary that contribute to follicular development and oocyte maturation. Recent studies have demonstrated that FF-derived exosomes contents, especially miRNAs, play a role in Polycystic Ovary Syndrome (PCOS) etiology and can be used as diagnostic and therapeutic biomarkers. However, no study until now has been analysed on same patients proteomic and miRNA sequencing of exosomes purified from blood and follicular fluid from control and PCOS patients. Study design, size, duration A cohort study involving 22 PCOS patients and 98 control patients was performed from 2021-2023 for the analysis of exosomes purified from follicular fluid and in 10 patients from blood. EVs were analysed for proteomic and miRNA sequencing. The biological effect of exosomes was performed on human granulosa cells lineage KGN from 2023 and still ongoing. Participants/materials, setting, methods 22 PCOS patients and 98 control patients was used from 2021-2023 for the exosomes purification from follicular fluid and blood. The study was carried out in accordance with the Declaration of Helsinki principles and free informed consent was obtained from all participants. exosomes were assessed by mass spectrophotometry and sequencing with a NextSeq500. steroid concentrations measured in medium was performed by ELISA. Cell viability and proliferation was determined by using CCK8 and BrDU assays, respectively. Main results and the role of chance To better understand the importance of exosome in ovary, the objective of our project was to obtain a “global” view of the content and activity of exosomes from ovarian fluid (follicular fluid) surrounded the germ cells purified from PCOS patients versus control patients by using transcriptomic and proteomic approaches. First results have observed the presence of approximately 1195 proteins in exosomes purified from human follicular fluid and 183 miRNA. Furthermore, in all patients and whatever the status, exosomes present similar morphology and diameter. Our study found that the protein STAT3 is overexpressed in exosomes from PCOS. STAT3 interacts with many proteins associated with the PCOS phenotype observed in granulosa cells. When granulosa cells were exposed to PCOS-derived exosomes along with STAT3 inhibitors, cell proliferation, migration, and steroid production returned to normal levels. In addition, the miR30c-5p is highly expressed and inhibition with antagomir induced a return to a control phenotype. In summary, the study showed that the content of follicular fluid-derived exosomes are specific and can directly regulate granulosa cell activity. The miRNA and proteins content of these exosomes differs in PCOS including inflammation, steroidogenesis, and cytoskeleton architecture. Limitations, reasons for caution The number of PCOS and normal patients is limited. Wider implications of the findings Exosomes allow cells to communicate with each other by transporting molecules from one cell to another. This transfer can alter the behavior of cells. Exosomes could present a different activity in function of pathology or could be modified to present a positive effect during IVF or cumulus maturation. Trial registration number No