STEROIDOGENIC PROPERTIES OF A SPONTANEOUSLY ESTABLISHED PORCINE GRANULOSA-CELL LINE (PGC-2)

A spontaneously established porcine granulosa cell line (PGC-2) was cl oned through the continuous culturing of primary granulosa cells colle cted from equine chorionic gonadotropin (eCG)-treated prepubertal gilt s. This established cell line has undergone similar to 100 passages an d shows contact-inhibition of growth. PGC-2 stained with a monoclonal antibody (mAb) directed against cytokeratin, indicating its epithelial nature, but not with a mAb directed against vimentin, suggesting that it is not fibroblast-derived. Immunoblotting revealed that PGC-2 expr esses cadherin, an epithelial Ca++-dependent cell adhesion molecule. T he cells were dependent on serum for growth and had a doubling time of similar to 20 hr when cultured with 10% fetal bovine serum. The cell line was examined for the presence of FSH receptors, cAMP responses, a nd steroidogenic capabilities. The cell line lacks FSH receptors as as sessed by radiolabelled-ligand binding, and no transcripts for FSH rec eptor were detected by Northern blotting of total cellular RNA. Neithe r FSH nor cholera toxin (0.5 ng/mL) stimulated increases in cAMP level s in these cells, whereas forskolin (10 mu M) induced a fivefold incre ase in cAMP production. When a higher concentration of cholera toxin ( 300 ng/mL) was used, however, cAMP levels doubled by 2 hr. Despite a l ack of responsiveness to purified oFSH or oLH, the cells were capable of progesterone and estradiol production when provided with the approp riate substrates. We conclude that PGC-2 display properties that are s imilar to immature granulosa cells and may provide a suitable in vitro model for the study of granulosa cell function. (C) 1996 Wiley-Liss, Inc.