Recent evidence indicates that the increase in follicle numbers seen in PCOS occurs early in folliculogenesis, and that androgens are implicated in this. In primates and sheep, androgen excess in-utero results in ovarian changes similar to those in PCOS. We have recently shown using a novel in ovo model, that testosterone added to implanted human tissue increased primary follicles compared to untreated tissue (Qureshi et al. 2005). This is similar to the published results of follicle counts in polycystic ovaries (Webber et al. 2003, Maciel 2004). It is important therefore to determine the follicle stage at which androgen receptors (AR) are first expressed. The problem has been the sensitivity of the techniques available for use with the limited amount of material.
The aim of this study therefore was to determine the origin of AR mRNA expression in individual, well characterised pre-antral follicles isolated from human, ovarian cortex using nested RT-PCR and correlate this to the appearance of mRNA for follicle stimulating hormone receptor (FSHr) in the same follicles. Ovarian tissue was obtained with informed consent from women undergoing TAH/BSO (n=3) and cortical biopsies from women undergoing elective Caesarean sections (n=4). Follicles were manually isolated from tissue following brief enzymatic digestion, staged under light microscope and photographed. Individual follicles were lysed, snap-frozen and stored at −80C, prior to reverse transcription. To confirm the presence of cDNA, nested PCR for beta-actin was run on each follicle. 61/67 follicles examined were positive for beta-actin, and of these 15 were positive for AR and 4 for FSHr. The AR positive follicles were: 0 primordials, 3 transitionals (primordial-primary), 6 primaries, 3 transitionals (primary-secondary), 2 secondaries and 1 pre-antral. The FSHr positive follicles were: 3 primaries and 1 pre-antral. We have shown that early follicles acquire AR prior to FSHr, demonstrating that androgens could be exerting their effects from initiation of follicle growth onwards. This adds further weight to the hypothesis that androgens cause disordered early folliculogenesis in PCO.
01 - 05 Apr 2006
European Society of Endocrinology