Expression of CXCL16 /CXCR6 in fibroblast-like synoviocytes in rheumatoid arthritis and its role in synoviocyte proliferation / 北京大学学报(医学版)

ABSTRACT

Objective:It has been found that serum CXCL16 concentration in rheumatoid arthritis (RA) patients are significantly higher than those in osteoarthritis (OA) and normal subjects, and are positively correlated with disease activity and bone erosion.However, how is CXCL16 involved in the pathogenesis of RA is unclear.To evaluate the expression of CXCL16 and its receptor CXCR6 in fibroblast-like synoviocytes (FLS) of rheumatoid arthritis (RA) patients, and to explore the role of CXCL16 in the proliferation of RA-FLS.Methods: FLS were isolated from knee synovial tissues obtained from 8 patients of RA, 7 osteoarthritis (OA) and 3 normal controls.The diagnosis of RA was in line with the 1987 American Rheumatology Association (ACR) RA classification criteria, osteoarthritis met the 1996 ACR revised knee osteoarthritis classification criteria.Control synovium were obtained from trauma caused knee joint injury in healthy individuals who required surgery.Human knee FLS were cultured by tissue explants adherent method.FLS between passages 3 and 5 were used in the experiment.Expression of CXCL16 and its receptor CXCR6 were performed in Western blot analysis.FLS proliferation follo-wing stimulation with TNF-α and different concentrations of CXCL16 was examined by cell counting kit-8 (CCK-8).Expression of phosphorylated AKT (pAKT) in RA-FLS stimulated by CXCL16 was quantified by Western blot.Different concentrations of recombinant human CXCL16 were added to the culture medium of RA-FLS.After 48 h culture, supernantants were collected, and TNF-α, IL-6, RANKL and MMP3 in culture supernatants of RA-FLS were determined by enzyme-linked immunosorbent assays (ELISA) operated following the kit instructions.Results: Expression of CXCL16 and CXCR6 in RA-FLS was significantly higher than that of OA and controls (P<0.05), but no significant difference was found between OA-FLS and control FLS.Proliferation of RA-FLS was markedly up-regulated after stimulation of CXCL16 (P <0.05).In the case of the CXCL16 stimulated OA-FLS and control FLS, the FLS proliferation remained basically unchanged.Expression of phosphorylated AKT in RA-FLS increased remarkably in condition of CXCL16 (50,100, 200 μg/L) stimulation.The levels of IL-6 and RANKL in culture supernatants of RA-FLS were obviously increased under CXCL16 (200 μg/L) stimulation, while TNF-α and MMP-3 levels in the culture supernatants remained unchanged after CXCL16 (200 μg/L) stimulation.Conclusion: This study shows that the expression of CXCL16 and its receptor was highly elevated in RA-FLS.Recombinant CXCL16 promoted RA-FLS proliferation and activation in vitro.All these indicate that CXCL16 play an important role in the pathogenesis of RA, anti-CXCL16 treatment may help to relieve inflammation and bone damage of RA patients.However, due to the limitations of this study, the role of CXCL16 and its receptors in RA-FLS remains to be elucidated by further research.