Figure 1.

Effect of growth factors on the phosphorylation of the p44^mapk/p42^mapk at Tyr 204 in EGCG pretreated VSMCs. (A) VSMCs were cultured in dishes (diameter: 3 cm) and cultivated until confluence. Then the medium was replaced by serum-free medium containing 50 μM EGCG. After 24 h the medium was replaced by serum-free medium without EGCG, and VSMCs were stimulated with PDGF-BB (A) and EGF (B) for different time periods. VSMCs were pretreated with different concentrations of EGCG and then stimulated for 5 min with 50 ng/ml PDGF-BB (C), 100 nM Ang II (E), 50 ng/ml EGF (D), and 5% FCS (F). EGCG-treated VSMCs were stimulated with 50 ng/ml PDGF-BB, 100 nM Ang II, and 50 ng/ml EGF for 5 min. (G) Cells were treated with 50 μM EGCG for 24 h and then stimulated with 50 ng/ml PDGF-BB, 50 ng/ml EGF, 100 nM Ang II, and 5% FCS for 5 min. Cells were lysed, and 20 μg of protein were analyzed with SDS-PAGE. MAP kinase was detected after blotting on polyvinylidene difluoride membranes by a specific MAP kinase antibody that recognizes the catalytically activated p44^mapk/p42^mapk. Cells were stimulated with 50 ng/ml PDGF-BB, 100 nM Ang II, and 50 ng/ml EGF for 5 min.