Rheumatology Advance Access published online on September 11, 2008
Rheumatology, doi:10.1093/rheumatology/ken363
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IL-6 trans-signalling directly induces RANKL on fibroblast-like synovial cells and is involved in RANKL induction by TNF-
and IL-17
1Product Research Department, Chugai Pharmaceutical Co. Ltd, Gotemba, Shizuoka, Japan.
Correspondence to:
M. Mihara, Product Research Department, Chugai Pharmaceutical Co. Ltd, 1-135, Komakado, Gotemba, Shizuoka 412-8513, Japan. E-mail: miharamsh{at}chugai-pharm.co.jp
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Abstract |
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Objectives. We investigated the influence of cytokines on the expression of RANK ligand (RANKL) in fibroblast-like synoviocytes from RA patients (RA-FLS).
Methods. RA-FLS were stimulated by IL-6, TNF-
, IL-17 and IL-1β with or without soluble IL-6 receptor (sIL-6R) for 24 h. The expression of RANKL was measured by real-time PCR, western blotting and immunostaining. In proliferation assay, RA-FLS were cultured with cytokines for 3 days. RA-FLS were co-cultured with RAW cell in the presence of IL-6/sIL-6R for 3 days and then NFATc1 mRNA expression in RAW cells was examined. RA-FLS was cultured with parthenolide [PAR, signal transducer and activator of transcription (STAT) inhibitor] or PD98059 (PD, mitogen-activated protein kinase inhibitor) in the presence of IL-6/sIL-6R and then the influence of these drugs on phosphorylation of STAT3 and ERK1/2, and RANKL expression was examined.
Results. RANKL expression was induced by IL-6/sIL-6R (but not IL-6 alone) and by IL-1β. On the other hand, TNF- and IL-17 did not induce RANKL expression, although TNF-, IL-17 or IL-1β stimulated cell growth and IL-6 production. However, in the presence of sIL-6R, TNF- or IL-17 induced RANKL expression. By the co-culture of RA-FLS, NFATc1 mRNA expression was induced in RAW cells. Finally, IL-6/sIL-6R induced phosphorylation of STAT3 and ERK1/2 in RA-FLS, and was completely inhibited by PAR and PD, respectively. PAR completely inhibited IL-6/sIL-6R-induced RANKL expression, but PD did not.
Conclusions. IL-6/sIL-6R directly induced RANKL expression in RA-FLS and it is essential for RANKL induction by TNF- and IL-17. Moreover, RANKL induction by IL-6/sIL-6R is mediated by the janus kinase/STAT signalling pathway.
KEY WORDS: Fibroblast-like synoviocytes, Receptor activator of nuclear factor-
B ligand, Tocilizumab, Soluble IL-6 receptor, Interleukin-6, Tumour necrosis factor-, Interleukin-17, Interleukin-1, Janus kinase/signal transducer and activator of transcription, Mitogen-activated protein kinase
Submitted 18 April 2008; Accepted 6 August 2008
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