Abnormal vascular growth and remodeling are components of multiple systemic diseases, including diabetes, atherosclerosis, hypertension, and cancer. Diverse pathological changes involving the vasculature occur in these diseases. A clear understanding of the mechanisms leading to abnormal vascular growth and remodeling should address, at least in part, the interaction between local growth factors in the vasculature and their modulators. A leading candidate in this process is the angiogenic peptide, basic fibroblast growth factor (FGF-2) which is widely distributed in the extracellular matrix of vascular endothelial cells. The applicant has recently identified, purified, and sequenced a new group of modulators of the FGF axis, the soluble FGF receptors. These proteins are present in blood and in the extracellular matrix of vascular endothelial cells. Recently, the applicant has obtained exciting preliminary data showing that the soluble FGF receptors completely inhibit the mitogenic action of FGF-2 on the proliferation of vascular endothelial cells in vitro. Based on these findings is the hypothesis that the truncated FGF receptors control the angiogenic activity of the endogenous stores of FGF-2 in the extracellular matrix.
Five Specific Aims are proposed to test this hypothesis.
Specific Aim 1 is directed at identifying the specific isoform(s) of the truncated FGF receptors in the extracellular matrix of microvascular endothelial cells and measure the binding affinity for FGF-2 for the ECIVI-associated, truncated FGF receptors. Included within this aim is the characterization of truncated receptors using isoform specific antibodies, expression of the truncated receptor in a mammalian expression system, measurement of the affinity of the receptor for FGF-2, and the examination of heparin requirements for growth factor binding.
Specific Aim 2 is directed at clarifying how the truncated FGF receptors in the ECM modulate the angiogenic activity of the extracellular matrix stores of FGF-2.
Specific Aim 3 is designed to characterize the conditions which regulate the deposition of the truncated FGF receptor(s) into the ECM. More specifically, the applicant plans to test the activities of matrix metalloproteinases, substances which modulate the biological activity of FGF, substances which modulate endothelial cell growth, and activators of protein kinase C on the appearance of the truncated receptors in ECIVI.
Specific Aim 4 is designed to characterize the conditions which regulated the release of truncated FGF receptors from FGF-2 in the ECIVI. The effects of pH, plasmin and other proteases, and enzymes which degrade the extracellular matrix will be examined.
Specific Aim 5 is designed to determine how conditions which alter the levels of truncated FGFRs in the extracellular matrix modulate the biological activity of the extracellular matrix stores of FGF- 2.
| Hanneken, A (2001) Structural characterization of the circulating soluble FGF receptors reveals multiple isoforms generated by secretion and ectodomain shedding. FEBS Lett 489:176-81 |
