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Toll-like receptor-4 signalling is specifically tak1-independent in synovial fibroblasts
  1. Ben T van den Brand1,
  2. Shahla Abdollahi-Roodsaz1,
  3. Onno J Arntz1,
  4. Michael Kracht2,
  5. Jeroen Geurts1,
  6. Wim B van den Berg1,
  7. Fons A J van de Loo1
  1. 1Department of Rheumatology, Radboud University Nijmegen Medical Centre, Nijmegen, The Netherlands
  2. 2University Giessen, Rudolf-Buchheim-Institute of Pharmacology, Frankfurter Strasse 107, 35392 Giessen, Germany


Background and objective Activated synovial fibroblasts are key players in the pathogenesis of rheumatoid arthritis (RA) by driving inflammation and joint destruction. Numerous molecules including cytokines and toll-like receptor (TLR) ligands induce pro-inflammatory signalling and gene expression through a hierarchical network of kinases. Upstream mitogen-activated protein kinase kinase kinases (MAP3Ks) represent an attractive target for RA treatment. In this study the authors sought to determine the role of the MAP3K transforming growth factor β activated kinase1 (TAK1) in cytokine and TLR-mediated signalling.

Materials and methods TAK1 activity was inhibited using either a small molecule inhibitor 5Z-7-oxozeaenol or lentivirally overexpressed kinase-inactive TAK1-K63W mutant in murine embryonic and human dermal and synovial fibroblasts. Fibroblasts were stimulated with interleukin 1 (IL-1), tumour necrosis factor (TNF), TLR2 or TLR4 agonists and responses were evaluated using transcriptional reporters and analysis of gene expression of collagenases (matrix metalloproteinases (MMP3, MMP13)), cytokines (IL-1β, IL-6) and chemokines (IL-8, MCP-1).

Results TAK1 inhibition abrogated cytokine- and TLR-induced activation of nuclear factor κ-light-chain-enhancer of activated B cells (NF-κB) and Saa3-promoter reporters in mouse and human dermal fibroblasts. In synovial fibroblasts, TAK1 crucially regulated IL-1 and TNF mediated NF-κB, but not Saa3-promoter activation. Furthermore, TAK1 was required for inducible mRNA expression of IL-1β, IL-6, IL-8, MMP3 and MMP13, but not MCP-1, in response to IL-1, TNF and TLR2 agonist. Unexpectedly, TLR4-induced NF-κB activation and gene expression was fully TAK1-independent as found using the TAK1 inhibitor or kinase inactive mutant. Western blots demonstrated IL-1-inducible p38 and p65 phosphorylation and inhibition by the TAK1 inhibitor 5Z-7-oxozeaenol, whereas no effect of TAK1 inhibitor on lipopolysaccharides signalling in RA-SF.

Conclusions In general, TAK1 plays a prominent role in regulation of IL-1- and TNF mediated signalling in fibroblasts. Interestingly, TLR4 signalling is specifically TAK1-independent in synovial fibroblasts. Consequently, therapeutic TAK1 inhibition in arthropaties may not dampen the damage-associated molecular pattern-mediated TLR4 activation of synovial fibroblasts.

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