Figure 3

TNF-α induces MMP-9 expression via ERK1/2 phosphorylation. (A) Cells were pretreated with U0126 for 1 h and then incubated with TNF-α (15 ng/ml) for 24 h. (B) Cells were pretreated with U0126 (3 μM) for 1 h and then incubated with TNF-α (15 ng/ml) for 6 h. The isolated RNA samples were analyzed for the levels of MMP-9 mRNA by real-time PCR. (C) Cells were pretreated with or without U0126 (3 μM) for 1 h and then stimulated with TNF-α for the indicated time intervals. Phosphorylation of ERK1/2 was determined by Western blot using an anti-phospho-ERK1/2 antibody. Data are expressed as mean±SEM of three independent experiments. ^#P < 0.01, as compare with the cells treated with TNF-α alone. (D) Cells were transfected with ERK2 siRNA for 24 h and then incubated with TNF-α (15 ng/ml) for 24 h. (A,D) MMP-9 expression was determined as described in Figure 1. The cell lysates were determined by Western blot using an anti-ERK2 or anti-GAPDH antibody.