VEGF enhances phosphorylation of MAPK family members of signaling proteins in HIMEC. Activation of p44/42 MAPK, JNK and p38 MAPK in HIMEC by VEGF was assessed using Western blot analysis. Total cell lysates from cultured HIMEC stimulated with VEGF (10–50 ng/ml), were subjected to SDS-PAGE and immunoblotted with phospho-specific anti-ERK 1/2, JNK and p38 MAPK antibodies. VEGF enhances phosphorylation of all three MAPK members, as compared with total ERK. The time of induction ranged from 1 to 120 min as detailed in the materials and methods section. Maximum detection of p42/44 MAPK was seen at 30 min (shown); Maximum activation of p38 MAPK was detected at 10 min in response to VEGF; JNK maximum activation was seen at 5 min following VEGF activation. Coomassie staining of the polyacrylamide gel confirms equal amounts of total protein loaded. The representative figure shown is from one of three independent experiments.