Studies of the vascular endothelial growth factors, VEGFs, and their receptors, focusing on VEGF-B

Abstract: The inner lining of all blood- and lymphatic vessels are composed of endothelial cells. Their growth and differentiation are in part regulated by polypeptide growth factors and their receptors. Five endothelial cell-specific receptor tyrosine kinases are known, including three receptors for vascular endothelial growth factors (VEGFs) and a receptor for angiopoietins, Tie-2, as well as the related orphan receptor Tie-1. Vascular endothelial growth factor (VEGF) is a prime regulator of endothelial cell proliferation, angiogenesis, vasculogenesis and vascular permeability. The family of VEGF-related molecules has recently been expanded and is presently composed of five members, VEGF, placenta growth factor (PIGF), VEGF-B, VEGF-C, and VEGF-D. In addition, there are three viral homologues, collectively called VEGF-E, encoded by different strains of the Orf virus. This work describes the cloning and characterisation of VEGF-B. VEGF-B is a secreted disulphide linked dimer, capable of forming heterodimers with VEGF. VEGF-B was found to be predominantly expressed in embryonic and adult muscle tissues, including the heart. The structure for both the mouse and the human gene for VEGF-B was solved and found to contain 7 exons and span about 4 kb of DNA. Two isoforms arise from alternative splicing which involves the use of overlapping reading frames. The two forms have a common N-terminal domain of 115 amino acids, but differ in their C-terminal regions affecting their affinity for heparin and thus release and bio-availability. While VEGF-B167 is non-glycosylated, highly basic and retained at the cell surface upon secretion, VEGF-B186 is O-linked glycosylated and readily secreted from cells. Both isoforms of VEGF-B were found to specifically bind VEGFR-1. Proteolytic cleavage of VEGF-B186 appears to result in an increased VEGFR-1 affinity. Mutational analysis of VEGF-B provided evidence for the structural conservation with VEGF and PDGF. Disruption of dimer formation leads to severely reduced VEGFR-1 binding, suggesting that covalent dimerisation is important for VEGF-B function. This work also covers a comparison at the cellular level of the in vivo receptor binding sites for VEGF and VEGF-C. VEGF is a key modulator of angiogenesis during development and in adult tissues, while the related VEGF-C has been shown to induce both lymphangiogenesis and angiogenesis. In adult tissues, where VEGFR-3 is largely confined to the lymphatic vasculature, the binding of VEGF-C occurs preferentially to this receptor, while in the developing embryos the binding sites include almost all vessel structures. The elucidation of the properties of the VEGF-related growth factors, VEGF-B and VEGF-C, increase our understanding of the complexity of the regulatory signals for endothelial cells, this could have important medical implications.