Figure 2

αvβ5 integrin is involved in CCN4-mediated IL-6 production in synovial fibroblast.s (A-C) OASFs were pretreated for 30 minutes with α5β1, αvβ3, or αvβ5 integrin antibody (3 µg/ml) followed by stimulation with CCN4 for 24 hours, and IL-6 expression was examined by qPCR, ELISA, and Western blot. *, P <0.05 compared with IgG control. (D) OASFs were incubated with various concentrations of CCN4 for 24 hours, and the cell surface αvβ5 integrin expression was examined by flow cytometry. (E) OASFs were incubated with CCN4 (30 ng/ml) for 24 hours, and the mRNA expression of αv and β5 integrins was examined by qPCR. (F) OASFs were transfected for 24 hours with αv and β5 siRNA, and the integrins expression was examined by Western blot. (G) OASFs were cotransfected for 24 hours with αv and β5 siRNA followed by stimulation with CCN4 for 24 hours, and IL-6 expression was examined by qPCR. (H) OASFs were incubated with vitronectin (10 ng/ml) for 24 hours, and IL-6 expression was examined by qPCR. Results are expressed as the mean ± SEM (n = 3). *, P <0.05 compared with control; #, P <0.05 compared with the CCN4-treated group. OASFs, osteoarthritis synovial fibroblasts; IgG, immunoglobulin G; SEM, standard error of the mean.