HORMONAL-REGULATION OF THECAL CELL-FUNCTION DURING ANTRAL FOLLICLE DEVELOPMENT IN BOVINE OVARIES

The hormonal regulation of thecal cell function was investigated with cells isolated at various stages of antral follicle development. Bovine thecal cells were isolated from small antral, medium antral, and large Graffian follicles (small, medium, and large ovarian follicles). Serum-free cultures of thecal cells were established and viable for a minimum of 6-8 days of culture. The purity of the thecal cell population was characterized cytochemically and was found to contain less than 5% endothelial cell and/or granulosa cell contamination. The steroidogenic capacity of this purified population of thecal cells inserum-free culture was examined through an analysis of androgen and progesterone production. Androgen production was high during the first 3 days of culture, then declined to undetectable levels. Production of androstenedione was approximately 10-fold higher than production of testosterone. Progesterone production remained relatively constant throughout the 8-day culture period. hCG was found to stimulate androgen production during days 1-3 of culture, but had a negligible effect on progesterone production. In contrast, hCG stimulated progesterone production during days 3-6 of culture, but had a negligible effect on androgen production. Insulin stimulated progesterone production during days 3-6 of culture, but had no effect on androgen or progesterone production during days 1-3 of culture. The minimum effective concentrations of hCG and insulin required to stimulate steroidogenesis of the thecal cells ranged from approximately 1-10 ng/ml. Addition of serum to the cultures decreased androgen production and suppressed the hormone responsiveness of the cells. Thecal cells in culture appear to alter their steroidogenic capacity from an androgen-producing cell to a progesterone-producing cell. Analysis of the developmental regulation of thecal cell function revealed that androgen production and hormone responsiveness were relatively constant in small, medium, and large follicles. In contrast, progesterone production and hormone responsiveness were highest in small follicles, intermediate in medium follicles, and lowest in large follicles. A more general analysis of the developmental regulationof thecal cell function examined the secretion of radiolabeled proteins. A large number of radiolabeled proteins were secreted by thecal cells, ranging in molecular mass from 5-500 kDa. Interestingly, insulin and hCG had no major effect on secretion of proteins by cells isolated from any of the stages of development examined. The major developmental change in radiolabeled secreted proteins was a decline in the abundant secretion of proteins greater than 100 kDa by thecal cells from small follicles as follicle development progressed. Combined results suggest the following. 1) Bovine thecal cells in culture appear to alter steroidogenic capacity, reflected in a switch in predominant secretion of androgen to progesterone. 2) hCG/LH has a major role in the stimulation of thecal cell steroidogenesis, and insulin at nearly physiological concentrations may regulate progesterone production. 3) As a small antral follicle develops into a large Graffian follicle the ability of the thecal cells to produce androgen remains relatively constant, while the ability to produce progesterone and several secreted proteins declines. The serumfree culture of bovine thecal cells is anticipated to be useful to elucidate the importance of thecal cells in the hormonal and developmental regulation of ovarian physiology. © 1990 by The Endocrine Society.