VEGF stimulates tyrosine phosphorylation of beta-catenin and small-pore endothelial barrier dysfunction

The purpose of this study was to test the hypothesis that tyrosine phosphor ylation signaling events and protein kinase C (PKC) activation mediate vasc ular endothelial growth factor-A(165) (VEGF)-induced endothelial cell (EC) proliferation and barrier dysfunction in bovine pulmonary artery EC monolay ers. A size-selective permeability assay showed that VEGF stimulated a dela yed, prolonged (6-45 h), concentration-dependent (50-200 ng/ml, similar to 1-4 nM) increase in the number of predominantly small-"pore" transport path ways (<60 Angstrom) across EC monolayers. The tyrosine kinase inhibitor her bimycin A (KA) and the selective PKC inhibitor bisindolyl-maleimide (BIM) p revented this phenomenon. After 6-24 h, VEGF-treated monolayers displayed a n HA- and BIM-sensitive reorganization of beta-catenin adherens junctions w ith fingerlike projections and the loss of beta-catenin at sites of small p aracellular hole formation. HA and BIM prevented the VEGF-induced increase in EC growth. HA blocked the VEGF-induced rapid and prolonged (10 min-45 h) increases in the phosphotyrosine (PY) contents of VEGF receptor 2, phospho lipase C-gamma 1, paxillin, and beta-catenin as well as similar to 140- and 128-to 117-kDa proteins, whereas BIM inhibited only the tyrosine phosphory lation of beta-catenin. These data suggest that VEGF initiates increased EC growth and chronic, small-pore endothelial barrier dysfunction by PY signa ling through beta-catenin that depends on PKC.