ECTS Abstracts (2015) 1 P222

Inflammatory cytokines affect the production of osteocyte-related signalling molecules by human bone cells cultured in close contact with their native matrix

Janak L Pathak1,2, Astrid D Bakker1, Frank P Luyten2, Patrick Verschueren2, Willem F Lems3, Jenneke Klein-Nulend1 & Nathalie Bravenboer4

1Department of Oral Cell Biology, Academic Centre for Dentistry Amsterdam (ACTA), University of Amsterdam and VU University Amsterdam, MOVE Research Institute Amsterdam, Amsterdam, The Netherlands; 2Skeletal Biology and Engineering Research Center, KU Leuven, Leuven, Belgium; 3Department of Rheumatology, VU University Medical Center, MOVE Research Institute Amsterdam, Amsterdam, The Netherlands; 4Department of Clinical Chemistry, VU University Medical Center, MOVE Research Institute Amsterdam, Amsterdam, The Netherlands.

Bone remodelling is disturbed in rheumatoid arthritis (RA) possibly because of elevated levels of circulating inflammatory cytokines. Osteocyte signalling plays a vital role in bone remodelling by affecting bone formation and/or bone resorption. Therefore, we aimed to investigate the effect of RA-serum containing inflammatory cytokines and exogenous recombinant inflammatory cytokines on human osteocyte signalling. Human trabecular bone chips were denuded by 2h collagenase treatment. Bone chips were cultured with or without 10% active RA-serum, and with or without recombinant IL-1β, IL-6, IL-17, TNFα (concentration: 10 ng/ml), or a cytokine cocktail (IL-1β, IL-6, TNFα) for 7 days. Live-dead staining was performed to assess cell viability. Gene expression of cytokines and osteocyte signalling proteins was analysed by qPCR. Only few cells were observed on the surface of bone chips at day 0, while approximately 80% of the surface was covered by cells after 7 days. Cells in or on the bone chips did express the osteocyte markers sclerostin, FGF23, DKK1, MEPE, IL-1β, and TNFα at day 0 and 7. Treatment with RA-serum, IL-1β, or TNFα enhanced gene expression of IL-1β (8-15–fold) and TNFα (2-3–fold). Treatment with IL-1β or TNFα, but not RA-serum, also enhanced gene expression of IL-6 (25-32–fold) and IL-8 (24-58–fold). The stimulatory effect of the cytokine cocktail on gene expression of IL-1β, IL-6, and IL-8 was significantly higher (80-120–fold) than the effect of the individual cytokines. IL-1β, TNFα, and the cytokine cocktail enhanced FGF23 expression (2-4–fold). Sclerostin expression was only enhanced by IL-1β (5-fold). RA-serum increased both sclerostin expression (2.5-fold), and DKK1 expression (2-fold). In conclusion, RA-serum and exogenous recombinant cytokines changed osteocyte signalling in cultured human denuded bone chips containing cells that express osteocyte markers, which suggests that osteocytes could provide a new target to prevent bone loss in inflammatory diseases.

Disclosure: The authors declared no competing interests.

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