PROLIFERATION AND PROGESTERONE PRODUCTION OF OVINE LUTEAL CELLS FROM SEVERAL STAGES OF THE ESTROUS-CYCLE - EFFECTS OF FIBROBLAST GROWTH-FACTORS AND LUTEINIZING-HORMONE

This study was conducted to evaluate the effects of fibroblast growth factor 1 (FGF-1), fibroblast growth factor 2 (FGF-2), or luteinizing hormone (LH) on proliferation and progesterone secretion of ovine luteal cells from days 5, 10, or 15 after estrus (estrus = day 0; n = 4 or 5 ewes/day). After enzymatic dispersion, luteal cells were incubated in the presence or absence of various doses of FGF-1, FGF-2, LH, or fetal bovine serum (FBS) (positive control) in serum-free media for 7 days in 24-well plates. Cells were counted on day 7 of culture and media analyzed for progesterone concentration. For all treatments, maximal effects (E(max)) and dissociation constants (K-D) were calculated. In addition luteal cells were cultured in eight-chamber slides and treated as above, but on day 7 of culture cells were fixed and stained for the presence of 3 beta-hydroxy-Delta(5)-steroid dehydrogenase (3 beta HSD). The number of steroidogenic (3 beta HSD positive) cells per unit area was counted for control cultures (no treatment) and cultures treated with the most effective doses of FGF-1, FGF-2, LH, or FBS in proliferation and (or) progesterone assays. FGF-1, FGF-2, and FBS stimulated (p < 0.05) proliferation of luteal cells from all stages of luteal development in a dose-dependent manner. In addition, LH increased (p < 0.01) the number of 3 beta HSD-positive cells across all stages of luteal development. Moreover, LH and FBS increased (p < 0.05) progesterone secretion by luteal cells from all stages in a dose-responsive manner, but the effects of FGF-1 and FGF-2 were variable. For proliferation, the E(max) of all factors was greatest (p < 0.01) on day 5, whereas the K-D values were similar across days of the estrous cycle. For progesterone production, the E(max) and K-D of LH and FBS were similar and did not differ across the estrous cycle. These data demonstrate that luteal cells from the early luteal phase of the estrous cycle exhibit the greatest ability to proliferate and (or) increase their progesterone secretion in response to FGF-1, FGF-2, LH, or FBS. In addition, although LH does not affect the total number of luteal cells in culture, it does increase the number of steroidogenic cells. These data indicate that in addition to LH, fibroblast growth factors may be involved in regulation of luteal growth and differentiation in ewes.