Developmental programming: impact of prenatal testosterone excess on ovarian cell proliferation and survival factors

PADMANABHAN V; SALVETTI NR; VEIGA-LOPEZ A; ORTEGA HH

Oregon

Congreso; 44th Annual Meeting, Society for the Study of Reproduction (SSR); 2011

SSR

Prenatal testosterone (T) excess leads to reproductive dysfunctions in sheep culminating in progressive loss of cyclicity.  At the follicular level, prenatal T excess induces 1) selective increase in androgen receptor in the stroma and granulosa cells of fetal days 90 and 140 ovaries as well as antral follicles of 10- and 21-month-old females, 2) an increase in estrogen receptor (ESR) 1 and decrease in ESR2 in granulosa cells of antral follicles of 10- and 21-month-old females, and 3) an increase in ovarian follicular recruitment and persistence resulting in multifollicular ovarian morphology.  The factors contributing to follicular persistence in prenatal T-treated sheep are unknown.  We hypothesized that prenatal T excess disrupts the developmental ontogeny of ovarian proliferation and apoptotic factors, thus contributing to the development of follicular persistence.  Pregnant Suffolk sheep were given T propionate, 100 mg i.m. twice weekly, in cottonseed oil from days 30-90 of gestation.  Changes in developmental expression of proliferating cell nuclear antigen (PCNA), antiapoptotic protein B-cell lymphoma 2 (Bcl-2), and proapoptotic protein Bcl-2–associated X protein (BAX) were determined in ovarian sections using streptavidin-biotin immunoperoxidase method at the following time points: fetal (days 90 and 140), prepubertal (22 weeks), end of first breeding (10 months), and second breeding (21 months) season (n=4-7/per age/treatment group).  Image analysis was performed using Image Pro-Plus 3.0.1 system.  Analyses of data (ANOVA, followed by Duncan’s multiple range tests) found age, follicle, and tissue specific changes in expression of PCNA, BAX, and Bcl-2 protein.  Prenatal T treatment induced a) an increase (p<0.05) in PCNA in theca and granulosa cells of antral follicles of 10 and 21 month old animals, b) a selective decrease (p<0.05) in BAX expression in granulosa cells of fetal day 90 primordial and primary follicles, and c) a selective decrease (p<0.05) in Bcl-2 in granulosa cells and theca cells of large preantral and antral follicles of 10 and 21-month old T females.  No changes were observed in stromal cells at any age.  Decreased expression of BAX may be the result of androgenic programming stemming from increased androgen receptor expression in T fetuses evident at this time.  The decrease in BAX is likely a contributing factor in enhancing follicular recruitment.  Increased expression of PCNA and decreased expression of Bcl-2 seen in antral follicles of 10 and 21 month old T females may stem from the altered equilibrium of androgen/estrogen receptor expression found in antral follicles of these animals at these ages.  The increase in PCNA is likely responsible for overcoming the effects of decreased Bcl-2 expression and arresting the follicles from undergoing atresia or differentiating further there by contributing to the follicular persistence seen in T females.  These findings are likely to be of translational relevance in addressing follicular persistence seen in women with PCOS, the reproductive and metabolic attributes of whom the T females recapitulates. Supported by NIH P01 HD44232 (VP).