Fig. 4. Effect of U0126 on ERK, JNK and p38 phosphorylations. (A) Immunoblots of total lysates from untreated cells (C), cells treated with TPA (TPA) and 10 µM U0126 in the absence (U) or presence of TPA (U + TPA) for 30 minutes, 2 and 5 days, using anti-phospho-active ERK and JNK antibodies. For normalization, filters were re-probed with antibodies that recognize total proteins. (B) Immunoblots of total lysate from cells incubated for 30 minutes with control medium (C30 min, C5d) or with U0126-containing conditioned medium derived from cells treated with U0126 for 30 minutes (U 30 min) and 5 days (U 5d), using anti-phospho-active ERK antibody. The data shown are representative of four independent experiments.