Intrafollicular level of steroid hormones and the expression of androgen receptor in the equine ovary at puberty

Steroidogenic activity in the equine ovary from birth to puberty has been poorly investigated. This study aimed to examine the capability of the ovarian follicles of prepubertal and pubertal fillies to produce steroid hormones and to evaluate the expression and cellular localization of androgen receptor (AR) in their ovaries. The ovaries of 6-18 month-old fillies were divided into two groups: prepubertal (PrP) - without preovulatory follicle (pF) and corpus luteum (CL), and ovulating/postpubertal (Ov/pB) - with pF and/or CL in at least one of the gonads. Adult mares (Me) were used as a control. The concentration of progesterone (P4), testosterone (T) and estradiol (E2) in follicular fluid (FF) was measured by radioimmunoassay. AR distribution was assessed by immunohistochemistry, while AR protein expression was examined by Western blot analysis. In the female groups, E2 concentration in FF of small follicles (<10 mm) was low and increased with the diameter of the follicle reaching the greatest value in pF (Ov/pB and Me group). In follicles (11-30 mm) of PrP fillies, the concentration of E2 was similar to that from Ov/pB fillies, but less than half (P < 0.05) than in Me follicles. In FF from all classes of follicles of Ov/pB fillies, the concentration of all steroids was similar to that in Me. AR immunolocalization, predominantly nuclear, was observed in all types of follicular cells (granulosa and theca cells) as well as in stroma and luteal cells. The pattern of staining was dependent on the follicle size and the group of females. In smaller antral follicles and in pF, the nuclear AR staining in granulosa cells was stronger than that found in follicles of 21-25 mm. In theca interna cells of pF, both nuclear and faint cytoplasmic reactions were seen. In luteal cells, AR labeling was noted in the nuclei and the cytoplasm: the strongest one in the early CL and almost negative in the late CL. AR protein expression in filly and mare ovarian tissues was confirmed by Western blot analysis and detected as a single band at approximately 110 kDa. In summary, the ovaries of fillies aged at least 6 months are capable of active steroidogenesis. ARs are present either in the cell nuclei or cytoplasm of all compartments of the equine ovary. AR expression in follicular and stroma cells may indicate the sensitivity of the filly ovarian tissue to androgens, the impact of androgens on folliculogenesis and the development of the equine ovary via a receptor-mediated pathway.