Ectodomain shedding of VEGF183, a novel isoform of vascular endothelial growth factor, promotes its mitogenic activity in vitro.

Abstract

We have previously reported the discovery of VEGF183, a novel isoform of vascular endothelial growth factor (VEGF), whose nucleotide sequence revealed an 18-bp deletion in the exon 6A-encoded region of VEGF. The following study was done to characterize VEGF183 and to determine its biological activity in vitro and in vivo. Recombinant human VEGF183 was expressed in Escherichia coli (rhVEGF183) or in human 293 embryonic kidney cells (293-VEGF183) and tested for stimulation of permeability of dermal vessels in normal rats as well as for mitogenic activity and phosphorylation of mitogen-activated protein kinases (MAPK) in cultured human umbilical vein endothelial cells (HUVECs). While small amounts of VEGF183 were secreted into the conditioned media (CM) of 293 cells expressing VEGF183 (293-VEGF183 cells), most of the VEGF183 remained cell surface-bound and could be released into the CM following treatment with plasmin or heparin. CM from 293-VEGF183 cells treated with heparin or plasmin induced about a twofold increase in cell numbers and stimulated MAPK phosphorylation in HUVECs as compared with CM from untreated 293-VEGF183 cells or from heparin- or plasmin-treated 293 cells containing the vector alone. Intradermal injections of rhVEGF183 promoted increased permeability of dermal vessels to Evan's blue dye. Our study shows that VEGF183 is predominantly a cell-anchored protein that promotes increased vascular permeability in vivo but requires extracellular cleavage or release by heparin or plasmin to promote its mitogenic activity in vitro.