Expression of the genes encoding basic fibroblast growth factor and its receptor in human granulosa cells

The role of basic fibroblast growth factor (bFGF) in granulosa cell ontogeny has been previously demonstrated. In this study we evaluated the possible intraovarian origin of bFGF. Human granulosa cells were maintained in primary culture and their cytoplasmic extract was purified by affinity chromatography on a heparin-sepharose column. The column was then eluted with 10 mM Tris-HCl containing increasing concentrations of NaCl. The chromatographic fractions were tested in a bioassay using bovine adrenal capillary endothelial cells (ACE) as targets. A peak of mitogenic activity was detected in the fraction eluted with the highest salt concentration. This chromatographic profile is similar to that of bFGF. The in situ synthesis of this bFGF-like protein was then demonstrated by reverse transcription-polymerase chain reaction. Using oligonucleotide primers specific for the bFGF gene, a single major band of DNA, corresponding to the expected size, was amplified. The identity of this fragment with the bFGF corresponding sequence was further demonstrated by restriction enzyme analysis. Moreover, RT-PCR was also employed to amplify a DNA band specific for the bFGF receptor gene. These data indicate that human granulosa cells are able to synthesize both bFGF and its receptor and, thus, bFGF might participate to the autocrine mechanisms regulating their growth and differentiated functions.