Expression of extracellular matrix components and cytokine receptors in human fibrocytes during rheumatoid arthritis.

Purpose: Fibroblast-like synoviocytes (FLS) represent one of the principal effectors of joint damage in rheumatoid arthritis (RA). Recent discovery of the circulating fibrocyte, a potential precursor of FLS, has raised issues regarding the characterization of fibrocytes with respect to their morphology and their biological role. In this study, we evaluated the morphology of fibrocytes in vitro and their ability to produce different extracellular matrix (ECM) components in comparison with two populations of RA FLS: synovial fluid FLS (fd-FLS) and intimal lining FLS (td-FLS). We also studied the expression of ECM regulators and a set of cytokine receptors involved in the pathogenesis of RA. Materials and Methods: Fibrocytes were cultured from peripheral blood of patients with RA. FLS were cultured from synovial fluids and tissues. ECM proteins (collagen I (col I) and fibronectin), Matrix metalloproteinases (MMP) (MMP3, and MMP9), ECM regulators (ß catenin, TCF4, and c-fos), and cytokine receptors (CXCR1, CXCR2, CXCR3, IL1RI, IL1RII, and IL6Rα) were analyzed using qRT-PCR and/or western blot. Results: Our results demonstrated that fibronectin and MMP3 levels were higher in FLS compared to fibrocytes. Although MMP9 was expressed in the three cell types, its level was greater in fibrocytes than in td/fd FLS. The three cell types expressed CXCR3, IL1RI, IL1RII, and IL6Rα, while the expression of CXCR1 and CXCR2 was restricted to fibrocytes. Conclusion: Our results demonstrated that fibrocytes express ECM molecules and cytokines receptors. The observed differences between fibrocytes and FLS may be due to their distinct functions or differentiation state during RA.Abbreviations: RA: Rheumatoid ArthritisFLS: fibroblast-like synoviocytestd: tissue derivedfd: fluid derivedSF: Synovial FluidWnt: WinglessMMP: Matrix MetalloproteinaseCIA: murine collagen induced arthritisECM: Extracellular matrixcol I: Collagen ITCF/LEF: T-cell factor/lymphoid enhancer-binding factorAP1: Activator Protein 1.

sFRP3 and DKK1 Regulate Fibroblast-Like Synoviocytes Markers and Wnt Elements Expression Depending on Cellular Context.

CONTEXT: Fibroblast-like synoviocytes (FLS) from rheumatoid arthritis (RA) display pathogenic behavior. Various members of the Wnt pathway, especially the canonical Wnt/ß-catenin cascade, may contribute to autonomous RA FLS activation. It has been shown that the two Wnt inhibitors: sFRP3 and DKK1 contribute to several critical aspects of joint biology. However, their effects on RA FLS are poorly characterized. The aim of our study was to investigate the effects of sFRP3 and DKK1 on FLS markers, Wnt components, and target oncogenes expression by RA FLS and compare the findings to osteoarthritic (OA) FLS. MATERIALS AND METHODS: RA and OA FLS were treated with sFRP3 and DKK1 for 6 days. Wnt signaling components (Wnt5a, LRP5 and ß-catenin), Wnt target oncogenes (cyclin E1 and WISP1), and FLS markers (fibronectin and MMP3) were analyzed using western blotting and/or qRT-PCR. RESULTS: Our data indicated that sFRP3 down-regulated the key gene ß-catenin in RA FLS. sFRP3 decreased fibronectin, a well-known downstream effectors gene of Wnt/ß-catenin pathway, and LRP5 expression in both RA and OA FLS. In OA FLS, sFRP3 induced increased expression of Wnt5a and MMP3 but did not affect their levels in RA FLS. On the other hand, DKK1 increased fibronectin expression in RA FLS and decreased its expression in OA FLS. CONCLUSION: Our results confirm the involvement of Wnt signaling in FLS transformation and show that two inhibitors of the same cascade can regulate differently the same elements and that a single inhibitor can initiate signaling depending on cellular context. ABBREVIATIONS: FLS: fibroblast-like synoviocytes; RA: rheumatoid arthritis; Wnt: Wingless; Fz: frizzled; LRP: Fz/low-density lipoprotein receptor protein; WISP1: Wnt1 inducible signaling pathway protein 1; sFRP: secreted Fz-related proteins; DKK: Dickkopf; OA: osteoarthritis; DMEM: Dulbecco's modified Eagle's medium; FBS: fetal bovine serum; PBS: phosphate buffered saline; SDS-PAGE: sodium dodecyl sulfate-polyacrylamide gel electrophoresis; ECL: enhanced chemiluminescence detection solution; MMP3: metaloproteinase 3; qRT-PCR: quantitative real-time polymerase chain reaction; S.D: standard deviation; CRD: cysteine-rich domain; MeCP2: methyl-CpG-binding protein; RANKL: nuclear factor-kappa B ligand.