Impact Statement: LIFRα is present in human GCs and its expression is increased in patients with high ovarian reserve, while decreased in low ovarian reserve
Objective: GCs consist of subpopulations of differentiated and less differentiated cells which survive and divide in response to leukaemia inhibitory factor (LIF) in culture. The LIF pathway is one of the important factors involved in growth initiation of human primordial to primary follicles through its receptor, LIFR. The objective of this study was to determine whether GCs from patients with high, normal and low ovarian reserve vary in their expression of the LIFRα.
Methods: This study was approved by the University of Toronto REB and informed consent was obtained from all participants (age: 24 – 42, BMI: 19.5 – 33). Aspirated follicular cells (AFCs) were collected after IVF-related transvaginal oocyte aspiration (LOR: n = 3, NOR: n = 6, HOR: n = 6). GCs were obtained via differential centrifugation and Ficoll gradient was used to eliminate red and white blood cell contamination. LIFRα (+) cells were detected using flow cytometry. CD45 was used as a negative control. LIFRα protein and gene expression in GCs, CM cells and follicular fluid (FF) were validated using Western blotting, immunocytochemistry and qPCR.
Results: Preliminary results from flow cytometry comparing HOR, NOR and LOR GCs (pooled, n=3) show that LIFRα (+) CD45 (-) cells were predominantly present in HOR samples comparing to NOR, while LOR cells were mostly LIFRα (-) CD45 (-). CD45 (+) cells represented ~3% of the cell population. Immunocytochemistry (IHC) in GCs and CMs showed co-expression of LIFRα in FSHR+ve cells. Western blot analysis confirmed IHC results. Quantitative PCR (qPCR) results revealed a 6.5-fold increase in LIFRα expression in HOR as compared to NOR, with low expression values for LOR.
Conclusion: LIFRα was detected in ovarian somatic cells (GCs, CMs). Significantly higher expression was found in GCs from HOR patients, compared to NOR and LOR patients, suggesting a higher proportion of precursor cells. Moreover, low LIFR expression was found in NOR. This results suggest that different proportion of precursor granulosa cells exist in the follicles of HOR and LOR when compared with NOR patients. We are currently studying the significance of this interesting finding in order to better understand how folliculogenesis is altered in patients with HOR, and whether LIFRα expression could be a surrogate marker for oocyte quality and maturity.
Keywords: granulosa cells (GCs), cumulus cells (CMs), leukaemia inhibitory factor receptor alpha (LIFRα), pluripotent stem cell markers
Funding & Disclosures: This project was jointly-funded by CReATe Program Inc. and by a D+H Sunnybrook Research Institute Studentship Award. There are no conflicts of interest to disclose.