Abstract
Introduction
Androgens play a critical role in female reproduction. Two major receptors, the classical androgen receptor (AR) and solute carrier family 39 member 9 (SLC39A9), have been shown to regulate androgen function in animal studies. Specifically, mouse AR knockout studies showed impaired follicular development, fewer ovulated oocytes, and diminished fertilization rates, whereas SLC39A9 knockout studies in zebrafish showed impaired oocyte germinal vesicle breakdown, meiotic resumption, and early embryogenesis. However, the presence of these receptors in human oocytes has not been studied. The purpose of this study is to investigate the presence and localization of AR and SLC39A9 in human oocytes.
Methods
Discarded deidentified oocytes were obtained with consent from participants undergoing in vitro fertilization, as approved by the Old Dominion University Institutional Review Board. Oocytes were fixed in 4% paraformaldehyde, incubated with primary and labeled secondary antibodies, and mounted on slides using mounting medium with DAPI. Oocytes were imaged using the BioTek Cytation C10 Confocal Imaging Reader. Human granulosa cells and sperm that were bound to the zona pellucida were used as positive controls since previous studies have demonstrated the presence of AR and SLC39A9 on these cells. Oocytes exposed only to secondary antibodies were negative controls.
Results
A total of 60 oocytes were labeled from 19 different participants with a mean age of 34.1 years (range 22 – 42). Of the 33 oocytes labeled for SLC39A9, 100% displayed SLC39A9 in the cytoplasm. Of the 17 MII oocytes labeled for SLC39A9, 6 (35.3%) displayed SLC39A9 in the polar body. AR labeling was performed on 24 oocytes, with 100% displaying AR in the cytoplasm. Of the 9 MII oocytes labeled for AR, only 1 (11.1%) displayed AR in the polar body.
Conclusion
SLC39A9 and AR are both present in the cytoplasm of human oocytes at the GV, M1, and M2 stages of maturity, without polarity to the polar body or nucleus. This is expected for AR, which is typically located in the cytoplasm when not bound to androgens. However, SLC39A9 is a transmembrane protein and is typically found on the plasma membrane, not commonly within the cytoplasm. Ongoing studies focus on the subcellular localization of each protein, with the goal of providing insight into the function of oocyte AR and SLC39A9 in human fertility.