COMPARATIVE-STUDIES BETWEEN FRESHLY ISOLATED AND SPONTANEOUSLY IMMORTALIZED BOVINE GRANULOSA-CELLS - PROTEIN SECRETION, STEROID-METABOLISM,AND RESPONSIVENESS TO GROWTH-FACTORS

A bovine granulosa cell line (BGC-1) has been obtained by spontaneous immortalization of primary cultures. BGC-1 cells have retained some ch aracteristics of primary cultures, such as the hormonal regulation of fibronectin biosynthesis. In the present study we have compared BGC-1 cells and primary cultures of bovine granulosa cells in terms of prote in secretion, steroid metabolism, and mitogenic responses to growth fa ctors. The pattern of protein secretion in BGC-1 cells was qualitative ly similar to that of primary cultures. The main differences were a hi gher proportion of fibronectin and the relative amounts of several oth er unidentified proteins. Progesterone levels in BGC-1 cultures were u ndetectable. When BGC-1 cells and primary cultures were incubated with [H-3]-pregnenolone, the former showed a lower conversion rate to prog esterone. In contrast, the conversion rate of [H-3]-progesterone to 5 alpha-reduced metabolites was markedly increased in BGC-1 cells. We al so examined the effects of epidermal growth factor (EGF), insulin like growth factor-I (IGF-I), and transforming growth factor-beta (TGF-bet a) on DNA synthesis under serum-free conditions. Both primary cultures and BGC-1 cells exhibited a stimulatory response to EGF and IGF-I on [H-3]-thymidine incorporation. Neither BGC-1 cells nor primary culture s showed a significant response to TGF-beta when added alone. However, in the presence of a combination of EGF and IGF-I, TGF-beta displayed an inhibitory effect on primary cultures while it stimulated DNA synt hesis in BGC-1 cells even further. The addition of conditioned medium from BGC-1 cells (BGC-1-CM) stimulated DNA synthesis on primary cultur es to a greater extent than the addition of conditioned medium from pr imary cultures. These results suggest that BGC-1 cells may be a useful model to study the regulation of granulosa cell function during the p eriod previous to the preovulatory stage of follicular development. Th e differential responses of the immortalized cells to growth regulator s may offer some clues on the mechanisms that control cell proliferati on in normal tissues. (C) 1995 Wiley-Liss, Inc.