Effect of Src and MMP inhibitors on EGFR phosphorylation. (A) SKOV-3 cells were treated with 100 nM PAF for 0, 5, 10, 15, 20, or 25 min. Following PAF treatment, cells were lysed and lysates were evaluated by Western blotting. Data were normalized to total Src protein expression and are expressed as the fold change (average ± S.E.M.) in phospho-Src compared to vehicle-treated cells. Representative blots for phosphor/total-EGFR are shown. (B) SKOV-3 cells were treated with the Src inhibitor PP2 (20 μM) for 1 hour before stimulation with PAF (100 nM) or EGF (5 ng/ml) for 5 min. Cells were harvested and subjected to Western blot. (C) SKOV-3 cells were pretreated for 1 h with the metalloproteinase inhibitor GM6001 (20 μM). Cells were then stimulated with either PAF (100 nM) or EGF (5 ng/ml) for 5 min before they were harvested and immunoblotting. (D) SKOV-3 cells were pretreated for 1 h with increasing concentrations of the metalloproteinase inhibitor GM6001 before they were harvested and subjected to Western blot. The data shown are representative of at least three independent experiments. Data were analyzed by Student’st-test. * p < 0.05.