Nichani P, Lazer T, Szaraz P, Power J, Gauthier-Fisher A, Kenigsberg S, Librach CL
UofT Department of Obstetrics and Gynaecology 33rd Annual Research Day, Toronto, Canada
Publication year: 2016

Abstract

Impact Statement: Leukaemia inhibitory factor receptor alpha (LIFRα) is present in human granulosa cells (GC) and its expression is increased in patients with high ovarian reserve (HOR), while decreased in low ovarian reserve (LOR).

Objective: GCs consist of subpopulations of differentiated and less differentiated cells that survive and divide in response to leukaemia inhibitory factor (LIF) in culture. The LIF pathway is one of the important factors involved in the 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 HOR, normal ovarian reserve (NOR), and LOR 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 (AFC) 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, cumulus cells (CMC) and follicular fluid (FF) were validated using Western blot, 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, CMCs). 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. These results suggest that different proportions of precursor GCs 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; cumulus cells; leukaemia inhibitory factor receptor alpha (LIFRα); pluripotent stem cell markers

Disclosures: This project was funded by the CReATe Fertility Centre and the D+H Sunnybrook Research Institute Studentship Award. There are no conflicts of interest to disclose.

Presentations

  1. Nichani P, Lazer T, Szaraz P, Power J, Gauthier-Fisher A, Kenigsberg S, Librach CL. (May 2016). Expression of leukaemia inhibitory factor receptor (LIFRα) in human luteinized granulosa cells is associated with ovarian reserve. UofT Department of OB-GYN 33rd Annual Research Day, Toronto, ON (poster). View poster here.