Arthritis & Rheumatism, Volume 63,
November 2011 Abstract Supplement
Abstracts of the American College of
Rheumatology/Association of Rheumatology Health Professionals
Annual Scientific Meeting
Chicago, Illinois November 4-9, 2011.
IL-1beta Inhibits TNF-Mediated Differentiation of Human Osteoclast Precursors Grown in Presence of Fibroblast-Like Synoviocytes.
Harvey, Bohdan P., Syed, Farhan A., Kaymakcalan, Zehra
Inflammatory cytokines such as TNF-alpha (TNFa), IL-1beta (IL-1b) and IL-6 have been shown to contribute to osteoclastogenesis independently or in conjunction with M-CSF or RANKL, two key cytokines involved in osteoclast development. However, the role of TNFa as well as the other inflammatory cytokines in promoting the expression of these key osteoclastogenic factors in synovial tissue and the concomitant induction of osteoclast differentiation is poorly understood. In this study, we sought to determine whether TNFa, IL-1b or IL-6, separately or in combination, could induce human osteoclast differentiation directly in the presence of fibroblast-like synoviocytes (FLS).
Primary human osteoblasts or FLS from healthy or rheumatoid arthritis (RA) donors, were pre-treated with various combinations of TNFa, IL-1b, IL-6, soluble IL-6 receptor (sIL-6R), M-CSF and RANKL for 24 hr in the absence or presence of osteoprotegerin (OPG), a natural inhibitor of RANKL. Human osteoclast precursor cells were then added, and the co-cultures were continued for 7 days. Western blot analysis was performed on RA-FLS to evaluate RANKL protein expression. Osteoclast differentiation was determined by the appearance of large multinucleated cells staining positive for tartrate-resistant acid phosphatase (TRAP) and by bone-specific TRAP5b activity within culture supernatants.
RA-FLS were found to express RANKL protein in response to either TNFa or IL-1b. Pre-treatment of osteoblasts or RA-FLS with a combination of TNF and M-CSF led to the appearance of TRAP+ multinucleated cells and an increase in TRAP5b activity; whereas, the combination of TNF and RANKL was ineffective in inducing osteoclasts, suggesting that TNF could substitute for exogenous RANKL but not M-CSF. Osteoclast differentiation was observed with FLS from healthy and RA donors and was found to increase in a TNF dose dependent manner. Addition of the RANKL inhibitor OPG caused a significant reduction in TRAP+ multinucleated cells without altering the level of TRAP5b activity. A greater degree of inhibition was observed with the addition of IL-1b alone or in combination with IL-6 and sIL-6R, since the generation of both TRAP+ multinucleated cells and TRAP5b were significantly reduced even in the presence of exogenous RANKL.
These findings suggest that in a co-culture system with FLS, TNFa can induce an early stage of osteoclast differentiation independent of RANKL as indicated by the expression of TRAP5b in the presence of OPG. This finding is in agreement with the literature describing TRAP5b as a marker of an immature osteoclast. However, the generation of multinucleated cells requires TNF-induced RANKL expression from the FLS since the number of TRAP+ multinucleated cells was reduced with OPG treatment. Surprisingly, IL-1b prevented both the expression of TRAP5b and the generation of TRAP+ multinucleated cells that was induced by either TNF or RANKL, suggesting that this pro-inflammatory cytokine can inhibit osteoclast development at an early stage.
To cite this abstract, please use the following information:
Harvey, Bohdan P., Syed, Farhan A., Kaymakcalan, Zehra; IL-1beta Inhibits TNF-Mediated Differentiation of Human Osteoclast Precursors Grown in Presence of Fibroblast-Like Synoviocytes. [abstract]. Arthritis Rheum 2011;63 Suppl 10 :1774