Objective: To examine the regulatory role of interleukin‐22 (IL‐22) in the expression of RANKL and induction of osteoclastogenesis in rheumatoid arthritis (RA).Methods: Concentrations of IL‐22 and RANKL in the serum and synovial fluid of RA patients were measured using enzyme‐linked immunosorbent assay. RA synovial fibroblasts were treated with recombinant human IL‐22 (rhIL‐22), and the expression of RANKL messenger RNA (mRNA) and protein was measured using real‐time polymerase chain reaction, Western blotting, and intracellular immunostaining. Human monocytes were cocultured with IL‐22–prestimulated RA synovial fibroblasts and macrophage colony‐stimulating factor, and osteoclastogenesis was assessed by counting the multinucleated cells (those staining positive for tartrate‐resistant acid phosphatase).Results: The IL‐22 concentration in the synovial fluid was higher in RA patients than in patients with osteoarthritis (OA). The serum IL‐22 concentration was also higher in RA patients than in OA patients and healthy volunteers, and this correlated with serum titers of rheumatoid factor and anti–cyclic citrullinated peptide antibodies. In RA synovial fibroblasts treated with rhIL‐22, the expression of RANKL mRNA and protein was increased in a dose‐dependent manner. IL‐22–induced RANKL expression was down‐regulated significantly by the inhibition of p38 MAPK/NF‐κB or JAK‐2/STAT‐3 signaling. In human monocytes cocultured with IL‐22–prestimulated RA synovial fibroblasts in the absence of exogenous RANKL, the monocytes differentiated into osteoclasts, but this osteoclastogenesis decreased after p38 MAPK/NF‐κB or JAK‐2/STAT‐3 signaling was inhibited.Conclusion: These results show that IL‐22 up‐regulates RANKL expression in RA synovial fibroblasts and induces osteoclastogenesis. These effects are mediated by the p38 MAPK/NF‐κB and JAK‐2/STAT‐3 signaling pathways.

Objective: Stromal cell–derived factor 1 (SDF‐1) is a potent chemoattractant for memory T cells in inflamed rheumatoid arthritis (RA) synovium. This study was undertaken to investigate the effect of interleukin‐17 (IL‐17) and CD40–CD40L interaction on SDF‐1 production in RA fibroblast‐like synoviocytes (FLS).Methods: Synovial fluid (SF) and serum levels of SDF‐1 in RA patients were measured by enzyme‐linked immunosorbent assay (ELISA). The SDF‐1 produced by cultured RA FLS was evaluated by real‐time polymerase chain reaction and ELISA after FLS were treated with IL‐17 and inhibitors of intracellular signal molecules. The SDF‐1 level was also determined after FLS were cocultured with T cells in the presence and absence of IL‐17.Results: Concentrations of SDF‐1 in the sera and SF were higher in RA patients than in osteoarthritis patients, although the increase in the serum levels did not reach statistical significance. The production of SDF‐1 in RA FLS was enhanced by IL‐17 stimulation. This effect of IL‐17 was blocked by inhibitors of phosphatidylinositol 3‐kinase (PI 3‐kinase), NF‐κB, and activator protein 1 (AP‐1). When FLS were cocultured with T cells, SDF‐1 production was up‐regulated, especially in the presence of IL‐17, but FLS were inhibited by neutralizing anti–IL‐17 and anti‐CD40L antibodies. Addition of RA SF to cultured RA FLS significantly up‐regulated SDF‐1 messenger RNA expression, which was hampered by pretreatment with anti–IL‐17 antibody.Conclusion: SDF‐1 is overproduced in RA FLS, and IL‐17 could up‐regulate the expression of SDF‐1 in RA FLS via pathways mediated by PI 3‐kinase, NF‐κB, and AP‐1. Our findings suggest that inhibition of the interaction between IL‐17 from T cells and SDF‐1 in FLS may provide a new therapeutic approach in RA.

Introduction: C-reactive protein (CRP) is one of the biomarkers for the diagnosis and assessment of disease activity in rheumatoid arthritis (RA). CRP is not only the by-product of inflammatory response, but also plays proinflammatory and prothrombotic roles. The aim of this study was to determine the role of CRP on bone destruction in RA. Methods: CRP levels in RA synovial fluid (SF) and serum were measured using the immunoturbidimetric method. The expression of CRP in RA synovium was assessed using immunohistochemical staining. CD14+ monocytes from peripheral blood were cultured with CRP, and receptor activator of nuclear factor-κB ligand (RANKL) expression and osteoclast differentiation were evaluated using real-time PCR, counting tartrate resistant acid phosphatase (TRAP)-positive multinucleated cells and assessing bone resorbing function. CRP-induced osteoclast differentiation was also examined after inhibition of Fcγ receptors. Results: There was a significant correlation between CRP levels in serum and SF in RA patients. The SF CRP level was correlated with interleukin (IL)-6 levels, but not with RANKL levels. Immunohistochemical staining revealed that compared with the osteoarthritis synovium, CRP was more abundantly expressed in the lining and sublining areas of the RA synovium. CRP stimulated RANKL production in monocytes and it induced osteoclast differentiation from monocytes and bone resorption in the absence of RANKL. Conclusions: CRP could play an important role in the bony destructive process in RA through the induction of RANKL expression and direct differentiation of osteoclast precursors into mature osteoclasts. In the treatment of RA, lowering CRP levels is a significant parameter not only for improving disease activity but also for preventing bone destruction.

Objective. To assess bone mineral density (BMD), serum levels of soluble receptor activator of nuclear factors-κB ligand (sRANKL) and osteoprotegerin (OPG) in patients with ankylosing spondylitis (AS), and to determine their relationships with disease activities.

Vascular endothelial growth factor (VEGF) has angiogenic, inflammatory, and bone-destructive roles in rheumatoid arthritis (RA). We aimed to determine the unique role of VEGF in osteoclastogenesis in RA. VEGF-induced receptor activator of nuclear factor ҡB ligand (RANKL) expression was determined in RA synovial fibroblasts by real-time PCR, luciferase assays, and ELISA. Osteoclastogenesis in peripheral blood monocytes cultured with VEGF was assessed by determining the numbers of tartrate-resistant acid phosphatase (TRAP)-positive multinucleated cells. Synovial fluid RANKL was correlated with VEGF concentration in the RA patients. VEGF stimulated the expression of RANKL in RA synovial fibroblasts. The RANKL promoter activity was upregulated by VEGF in the synovial fibroblasts transfected with RANKL-reporter plasmids. The VEGF-induced RANKL expression was decreased by the inhibition of both VEGF receptors (VEGFR) 1 and 2, Src, protein kinase C (PKC) and p38 MAPK. VEGF induced osteoclast differentiation from monocytes in the absence of RANKL and this was decreased by the inhibition of VEGFR1 and 2, Src, PKC and p38 MAPK. On coculturing with VEGF-prestimulated RA synovial fibroblasts, the monocytes differentiated into osteoclasts, and the osteoclastogenesis decreased by inhibition of Src and PKC pathways. VEGF plays dual roles on osteoclastogenesis in RA: direct induction of osteoclastogenesis from the precursors and stimulation of RANKL production in synovial fibroblasts, which is mediated by Src and PKC pathways. The axis of VEGF and RANKL could be a potential therapeutic target for RA-associated bone destruction.