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. 2007 Feb 5;27(8):2997–3007. doi: 10.1128/MCB.01485-06

FIG. 6.

FIG. 6.

Furin is required for TNF-α-induced MMP/nSMase/SK-1 activation. (A) Time course of furin activation in wt SMC, furin-silenced SMC (pretreated with specific siRNA for 24 h), SMC/PDX (SMC transfected with pCR/CMV-PDX plasmid), and SMC treated with the furin inhibitor decanoyl-Arg-Val-Lys-Arg-chloromethyl-ketone (FI) at 25 μM for 16 h and then treated with TNF-α (2 ng/ml) and time course of furin in MMP2+/+ or MMP2−/− fibroblasts treated with TNF-α (2 ng/ml). (B) Western blot of MT1-MMP activation induced by TNF-α in wt SMC or SMC/PDX. Membranes were blotted with anti-MT1-MMP and anti-β-actin antibodies (the arrow indicates the active form of MT1-MMP). (C to F) Determination of MT1-MMP, MMP2, nSMase, and SK-1 activities in wt SMC, SMC/PDX, FI-treated SMC, and furin-silenced SMC stimulated with TNF-α (2 ng/ml). (G) TNF-α-induced ERK1/2 phosphorylation in wt SMC and SMC/PDX. Western blots were labeled with anti-activated-ERK1/2 and ERK2 antibodies. Results are representative of three separate experiments. (H) DNA synthesis induced by TNF-α in wt SMC, furin-silenced SMC, SMC/PDX, and FI-treated SMC. Results are means ± SEM from three or four separate experiments. *, P < 0.05 (comparison between cells treated or not with TNF-α).