Session Type: Abstract Submissions (ACR)
Background/Purpose: There is evidence of osteoclast activation in the inflamed joints of patients with rheumatoid arthritis (RA). Studies that focus on osteoblasts are not sufficient to illustrate the cell functioning in an RA joint. Osteoblast differentiation is regulated by complex environmental stimuli. Among them, Wnt family proteins are thought to promote osteoblast differentiation via stabilization of ß-catenin and activation of TCF transcription. It is now known that the Wnt pathway is blocked with several soluble proteins such as Dockcop-1 (DKK-1) or Sclerostin (SOST). Fibroblast-like synoviocyte (FLS) is a unique articular resident cell and is a therapeutic target for RA, because the cell is believed to play a role in the joint destruction observed in this disease. In this study, we aimed to determine if FLS has an inhibitory effect on Wnt signaling.
Methods: FLS derived from RA patients (Articular Engineering, USA) was cultured in Dulbecco’s modified Eagle’s medium with 10% fetal bovine serum. Supernatants were collected from the FLS culture for 24 hours, with or without TNF-α (100 ng/ml, Millipore). The supernatants were added to U2OS culture after the transfection of TCF reporter plasmid, which contains Wnt/ß-catenin responsive consensus sequence (Millipore), by using lipofectamine 2000 to monitor the activity of Wnt3a responsive promoter. Wnt3a (R&D) or lithium chloride (LiCl) was used to activate the Wnt signaling pathway. Anti DKK-1 antibodies were used to neutralize DKK-1 in uupernatants from the FLS. The DKK-1 productions from FLS were measured by ELISA after treatment with TNF-α (100 ng/mL), IL-6 (100 ng/mL) with soluble IL-6 receptor (100 ng/mL), or IL-17A (100 ng/mL) for 24 hours.
Results: Wnt3a (200 ng/mL) or LiCl (50 mM) clearly increased the luciferase activity, which was effectively suppressed by the addition of supernatants from the FLS culture. The supernatant from FLS treated with TNF-α exhibited a greater inhibitory effect on luciferase activity induced by Wnt3a or LiCl. The inhibitory effects were blocked by anti DKK-1 neutralizing antibodies. DKK-1 was detectable in the FLS culture medium by ELISA, and the concentrations of DKK-1 were higher in the FLS culture medium supplemented with TNF-α, but not with IL-6 or IL-17, compared to the control.
Conclusion: Our data suggests that FLS inhibits osteogenesis by inhibiting osteoblast differentiation, which is achieved by inhibition of the Wnt signaling pathway. DKK-1, an antagonist for Wnt signaling, is one of the soluble factors that might contribute to the inhibition of the Wnt signaling pathway by FLS. The inflammatory milieu may enhance the Wnt signal inhibition by FLS because TNF-α facilitates the secretion of DKK-1 from FLS. Detailed research is required for the clinical application of DKK-1 blockade in promoting bone regeneration in an inflamed joint of a patient with RA.
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ACR Meeting Abstracts - https://acrabstracts.org/abstract/fibroblast-like-synoviocytes-inhibit-wnt-signaling-pathway-by-secreting-dockcop-1/