Abstract: 417

FAS-MEDIATED APOPTOSIS OF BOVINE GRANULOSA CELLS (GC) IN DEFINED MEDIA IS PREVENTED BY INSULIN-LIKE GROWTH FACTOR-1 (IGF).

Susan M. Quirk^{1 *}, Robert G. Cowan¹ , Rebecca M. Harman.¹
Department of Animal Science, Cornell University, Ithaca, NY. ¹

We previously showed that bovine GC cultured in DMEM-F12 plus 5% FBS express Fas antigen (FA) mRNA but are resistant to apoptosis induced by soluble human Fas ligand (FL) unless pretreated with interferon gamma (IFN). We examined FL-induced killing of GC cultured in media with FBS vs defined media containing DMEM-F12 plus insulin, transferrin, selenium, and BSA (ITS). GC isolated from abattoir ovaries were plated in media with 10% FBS. Media was changed at 48h to either 5% FBS or ITS. Cells were treated at 72h with the same media ± 100 ng/ml FL and ± 200 U/ml IFN. Cells were counted at 96h and FA mRNA measured by quantitative RT-PCR. GC cultured in FBS and treated with FL had little (7±3%) killing in the absence of IFN and 31±2% killing with IFN (P<0.05). GC treated with FL in ITS had 27±8% killing without IFN and 27±9% killing with IFN. Expression of FA mRNA was not different in cells cultured in FBS vs ITS and increased 7-fold with IFN in both media (P<0.01). Because FBS contains IGF activity, we tested the effect of human IGF on FL-induced killing. GC pretreated with IGF in ITS media at 48h and treated with FL had 2±4% killing without IFN and 29±7% killing with IFN (P<0.05), while GC not pretreated with IGF had 29±8% killing without IFN and 31±8% with IFN. In conclusion, killing of GC by FL does not require IFN treatment except in the presence of FBS. Basal levels of FA mRNA in cells cultured in ITS without IFN are sufficient to allow Fas-mediated killing, suggesting that elevated expression of FA is not essential for killing. Inhibition of Fas-mediated killing by serum may be due to IGF activity, since addition of IGF to ITS media mimics results observed in media with FBS. Thus, growth factors such as IGF may be critical in regulating the sensitivity of GC to FL.

    This abstract is being presented on Tuesday, August 3 at 8:00 AM to 10:15 AM at CUB 2nd Floor Ballroom.