Figure 6

Role of Src kinase activity in VEGF-and IL-1β-induced endothelial cell permeability. (A) Effect of Src inhibitor on VEGF-and IL-1β-induced endothelial cell permeability. PRECs were grown to confluent monolayers on porous membranes (12-well transwell insert plate). The lower chamber was incubated with VEGF (25 ng/ml) or IL-1β (10 ng/ml) and in the presence or absence of PP2 (10 μM) for 6 h at 37°C; (B-C) PRECs were transiently transfected with DNA dominant negative Src (HA-Src KD K295 M) and constitutive active Src (HA-Src-CA Y527F). Transfected PRECs were treated with VEGF and IL-1β for 6 h at 37°C where the induction of permeability by growth factor in wild type cells was completely blocked in DN Src transfected cells (B) where the CA Src transfected cells resulted in increased permeability than the wild type cells(C). The flux of RITC-dextran from the upper to the lower chamber was measured 6 h after treatment. Values are expressed in relative fluorescence counts (RFCs) as mean ± SEM, with each condition performed at least in triplicate.