PDGF receptor phosphorylation and MAP kinase activation.

(A) PDGF receptor phosphorylation was investigated at 5-minute PDGF-BB exposure (10 ng/mL), with or without bFGF. Increased phosphorylation of PDGF-Rα and PDGF-Rβ was found in the presence of bFGF/PDGF-BB compared with PDGF-BB alone (average increase, 52% ± 18%). Anti–PDGF-BB neutralizing antibody (Ab-PDGF-BB) markedly inhibited PDGF receptor phosphorylation. This experiment was carried out 3 times with similar results. One representative experiment is shown. (B) Phosphorylation of PDGF-Rα was investigated in BAECs exposed to PDGF-AA, which is a PDGF-Rα–specific agonist, in the absence and in the presence of bFGF. Under these conditions, bFGF/PDGF-AA–treated cells showed increased phosphorylation compared with PDGF-AA alone. Receptor phosphorylation was markedly reduced in DN-PDGF-Rα–transfected cells. This experiment was carried out 3 times, with similar results. One representative experiment is shown. (C) ERK1/2 phosphorylation induced by bFGF alone, PDGF-BB alone, and bFGF/PDGF-BB was detected with an antibody specifically recognizing the activated forms at 44 kd and 42 kd, respectively. A marked phosphorylation decrease was found in the presence of bFGF/ PDGF-BB compared with bFGF alone. This experiment was carried out 5 times with similar results. One representative experiment is shown.