Development of in vitro procedures which permit the measurement of the synergistic biological influences of polypeptide growth factors and steroid hormones on cellular processes represents an important objective in reaching an understanding of the molecular basis of many physiological events, including those associated with the proliferation of endothelial cells and the subsequent angiogenesis which occurs, for example, during the estrous cycle and wound repair. The present investigations were undertaken to examine the use of nonradioisotopic procedures for the bioassay of polypeptide growth factors, such as basic fibroblast growth factor (bFGF), known to exert effects on the proliferation and migration of endothelial cells, and to overcome some of the recognised sensitivity limitations with endothelial cell proliferation assays based on the [3H]thymidine uptake methods due to the cell passage number and culture conditions. The experimental results confirm these objectives with establishment of a simplified chromogenic procedure for the sensitive determination of the effects of the steroid hormones, estradiol and progesterone, on the proliferation of endothelial cells in culture, either directly or via mediation of the effects of bFGF at low concentration levels. This procedure has provided experimental data suggesting that a hitherto unrecognised synergy may exist between these steroids and bFGF, with both estradiol and progesterone significantly antagonising the bFGF-stimulated proliferative response with bovine aortic endothelial cells (BAEC) in culture. Progesterone alone was also capable of directly stimulating BAEC proliferation, albeit transiently. Neither estradiol or progesterone affected the proliferation of the control 3T3 fibroblast cell population, in the presence or absence of a stimulatory dose of bFGF.
- Basic fibroblast growth factor
- Chromogenic substrate
- Colorimetric measurement
- Endothelial cell proliferation
- In vitro bioassay
- Steroid hormone