Research Article

HMGB1 Acts in Synergy with Lipopolysaccharide in Activating Rheumatoid Synovial Fibroblasts via p38 MAPK and NF-κB Signaling Pathways

Figure 5

HMGB1 and LPS synergistically activated NF-κB and p38 MAPK. MAPK: mitogen-activated protein kinase; p-p38: phosphorylated p38; p-IκB: phosphorylated IκB. (a) Levels of intracellular molecules in whole cell lysates were analyzed by Western blotting after the SF were stimulated with 10 ng/mL of LPS and/or 100 ng/mL of HMGB1 for 30 min. To test whether RAGE mediate the effect of HMGB1 plus LPS, SF were pretreated with RAGE-Fc (10 μg/mL) for 30 min prior to the stimulation with HMGB1 plus LPS. (b) Densitometry analysis of the Western blot bands which was adjusted by the density of corresponding band of β-actin. (c) Cultured SF were pretreated with NF-κB inhibitor Bay11-7085 (20 μM) or p38 inhibitor SB203580 (10 μM) for 1 h and then incubated with 10 ng/mL of LPS and/or 100 ng/mL of HMGB1 for 48 h. The concentrations of IL-6, MMP-3, and MMP-13 proteins in culture supernatants were measured by ELISA. Data are expressed as mean ± SD ( ). * and ** compared with SF stimulated with HMGB1 plus LPS.
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