Figure 5

NF-κB is involved in the potentiation of IL-6 production by CCN4. (A) OASFs were transfected with IL-6 luciferase plasmids before incubation with CCN4 for 24 hours. Luciferase activity was then assayed. (B-D) OASFs were pretreated for 30 minutes with PDTC (10 µM) and TPCK (10 µM) or transfected for 24 hours with IKKα and IKKβ mutant followed by stimulation with CCN4 for 24 hours. Media, total RNA, and total protein were collected to measure IL-6 by ELISA, qPCR, and Western blot. (E) OASFs were incubated with CCN4 for the indicated time intervals, and p-IKKα/β, p-IκBα, and p-p65 expression was determined by Western blot analysis. Results are expressed as the mean ± SEM (n = 3). *, P <0.05 compared with control; #, P <0.05 compared with CCN4-treated group. OASFs, osteoarthritis synovial fibroblasts; PDTC, pyrrolidine-dithiocarbamate; SEM, standard error of the mean, TPCK, L-1-tosylamido-2-phenylenylethyl chloromethyl ketone.