Vascular endothelial growth factor stimulates the primary to secondary follicle transition in bovine follicles in vitro.

Little is known about the mechanisms regulating the growth of early preantral follicles, especially in nonrodent mammalian species. To test the hypothesis that vascular endothelial growth factor (VEGF) promotes the primary to secondary follicle transition, pieces of bovine fetal ovarian cortex were cultured with VEGF (0, 1, 10, or 100 ng/ml) for 0 or 10 days, followed by morphometric analysis. On day 0, cortical pieces contained mostly primordial follicles, but after 10 days in vitro most primordial follicles had activated, differentiating into primary follicles. VEGF had no effect on the numbers of primordial or primary follicles, compared with untreated controls, but all doses increased the number of secondary follicles. In the second experiment, a range of lower doses of VEGF (0.1-10 ng/ml) increased the number of secondary follicles in a dose-dependent manner. Analysis of VEGF transcripts by RT-PCR showed expression of mRNA for three isoforms of VEGF (VEGF121, 165, and 189) in fetal bovine ovarian cortex, with VEGF121 and 165 expressed predominantly and levels of mRNA for VEGF121 and 189 increasing after day 211 of gestation, when the first secondary follicles appear. Expression of mRNA for both VEGF receptors (flt-1 and flk-1) was also detected, but did not change with the development of fetal ovaries. Immunohistochemistry revealed positive staining for VEGF in blood vessels and in follicle cells of secondary follicles, consistent with Western blot analyses showing increases in VEGF protein as ovarian development progressed. Taken together, the results provide support for a role for VEGF in the primary to secondary follicle transition.