Involvement of c-Src in diperoxovanadate-induced endothelial cell barrier dysfunction

Reactive oxygen species (ROS) generated by activated leukocytes play an imp ortant role in the disruption of endothelial cell (EC) integrity, leading t o barrier dysfunction and pulmonary edema. Although ROS modulate cell signa ling, information remains limited regarding the mechanism(s) of ROS-induced EC barrier dysfunction. We utilized diperoxovanadate (DPV) as a model agen t to explore the role of tyrosine phosphorylation in the regulation of EC b arrier function. DPV disrupted EC barrier function in a dose-dependent mann er. Tyrosine kinase inhibitors, genistein, and PP-2, a specific inhibitor o f Src, reduced the DPV-mediated barrier dysfunction. Consistent with these results, DPV-induced Src activation was attenuated by PP-2. Furthermore, DP V increased the association of Src with cortactin and myosin light chain ki nase, indicating their potential role as cytoskeletal targets for Src. Tran sient overexpression of either wild-type Src or a constitutively active Src mutant potentiated the DPV-mediated decline in barrier dysfunction, wherea s a dominant negative Src mutant attenuated the response. These studies pro vide the first direct evidence for Src involvement in DPV-induced EC barrie r dysfunction.