Follicle-stimulating hormone stimulated differentiation and progesterone production of bovine granulosa cells in culture

Abstract

Abstract Progesterone production of granulosa cells cultured in vitro is stimulated and cell differentiation increased, by follicle-stimulating hormone (FSH). This study examined whether the increased progesterone production observed when bovine granulosa cells are cultured occurs because (1) progesterone production by undifferentiated and/or differentiated cells is increased or (2) the differentiation of granulosa cells is stimulated. Viable bovine granulosa cells (2−3×105) from follicles 5–8 mm in diameter were cultured in the presence of 0, 1, 10 and 100 μu FSH (1 μu ≡ 1 μg NIH-FSH-S1) for 6 days at 37°C in a humidified atmosphere of 5% CO2 in air in 1 ml of a 1:1 mixture of Dulbecco's modified Eagle medium: Ham's F10 medium supplemented with 365 μg ml−1 l -glutamine, 100 U ml−1 penicillin and 100 μg ml−1 streptomycin. Progesterone production, total DNA and protein, and cell diameter were determined sequentially over the culture period. The increases in progesterone production (ng μg−1 DNA per 24 h), cytoplasmic:nuclear ratio (μg protein μg−1 DNA) and cell diameter (μm) over 6 days culture indicated that granulosa cells underwent differentiation in the presence of FSH. Progesterone production of undifferentiated granulosa cells (diameter 14 μm or less) was stimulated by FSH (P 0.05) during a 6 day culture period. FSH stimulated (P 0.05) progesterone production by differentiating granulosa cells (8.7±0.5 ng μg−1 DNA per 24 h). In conclusion, the increase in progesterone production of FSH-stimulated granulosa cells cultured in vitro appears to be mainly due to an increase in the number of differentiating cells with a constant rather than an increasing progesterone production per cell.