Figure 3

HMGB1 in complex with IL-1β stimulates SFs to TNF, IL6, IL-8 and MMP-3 production. Synovial fibroblasts were stimulated for nine hours with A) HMGB1, IL-1β or HMGB1-IL-1β complexes. Addition of HMGB1-IL-1β complexes stimulates RASFs and OASFs to a 1 to 2 log-fold increased number of TNF producing cells compared to HMGB1 simulation alone. Squares (RA) and dots (OA) represent results from each donor; the horizontal line indicates the median values. B) HMGB1-IL-1β complexes at indicted concentrations induced a significantly enhanced production of IL-6 and IL-8 after 24 hours of stimulation compared to HMGB1 simulation alone whereas no production of IL-10 or IL-1β could be detected. Pooled data from RAFSs and OASFs where the horizontal line indicates the median values. C) Enhanced MMP-3 secretion was evident with HMGB1-IL-1β stimulation after 24 hours of stimulation as recorded by ELISA. Irrespective of the level of spontaneous MMP-3 production, stimulation with HMGB1-IL-1β complex enhanced the production in 8/9 cell lines compared to HMGB1 simulation alone. Squares (RA) and dots (OA) represent results from each donor, horizontal line indicates the median values. RASF n = 4, OASF n = 5. P-values were calculated using Kruskal-Wallis nonparametric ANOVA test. * (P < 0.05) ** (P < 0.01) *** (P < 0.001).