Fig. 5.

Effects of IL-33 on RANKL-induced osteoclast differentiation in peripheral blood (PB) and synovial fluid (SF) monocytes. a PB and b SF CD14+ monocytes were cultured with various doses of IL-33 in the presence of 25 ng/ml M-CSF and 10 ng/ml RANKL. After 21 days of culturing, TRAP-positive multinucleated cells, mature osteoclasts, were counted. Figures represent data from one of three independent experiments, and the bars represent mean ± SEM. Gene expression levels of osteoclast markers such as TRAP, NFATc1, DC-STAMP, OC-STAMP, and ATP6v0d2 were measured using real-time PCR. Data were normalized to the expression level of beta-actin and reported in relative expression units. *p < 0.05, **p < 0.01, and ***p < 0.001 indicate significant difference from the value for nil condition and ^##p < 0.01 indicates significant difference between two conditions. c Immunoblotting was performed for TRAF6, phospho-Src, Src, phospho-JNK, JNK, phospho-ERK, ERK, phospho-p38, p38, phospho-Akt, Akt, phospho-IκBα, IκBα, phospho-c-Jun, c-Jun, and beta-actin in PB CD14+ monocytes treated with IL-33 and untreated controls in the presence of 10 ng/ml RANKL for 1 h. Data were normalized to the expression level of beta-actin and reported in relative expression units. Bars show mean ± SEM from 3 independent experiments. *p < 0.05, **p < 0.01, and ***p < 0.001