Figure 3

Effect of 6-ME on VEGF-induced phosphorylation of MEK1/2 and ERK1/2 and transcription of DUSP1 and DUSP5. HUVE cells were serum starved for 2 h in M199 and then stimulated with VEGF (50 ng/ml) (A & B) or FGF (2.5 ng/ml) (C), in the absence or presence of 6-ME, for 15 min. Then cell lysates were collected with 1% SDS lysis buffer supplemented with PMSF and immunoblotting followed using antibodies against endogenous phospho-MEK1/2, MEK1/2, phospho-ERK1/2, ERK1/2 and actin. Graphs show normalized intensity values ± s.d. derived from three independent experiments. (D) HUVE cells were stimulated by VEGF (50 ng/ml) in the absence or presence of 6-ME (20, 10μM) for 30 min. Then, total RNA was isolated and qRT-PCR experiments followed using primers for DUSP1 and DUSP5.