Background/Purpose: Synovial inflammation is a common manifestation of juvenile idiopathic arthritis (JIA), often accompanied by debilitating synovial hyperplasia. The current study aims to characterize the potential benefit of TGF-β activated kinase 1 (TAK1) inhibition on hyperproliferation and tissue destruction in IL-1β-activated JIA synovial fibroblasts (JIASFs).

Methods: To evaluate the effect of TAK1 inhibition, JIASFs were pretreated with a small-molecular inhibitor of TAK1, 5Z-7-Oxozeaenol (5Z) at 0.01–0.25 µM concentration followed by IL-1β (10 ng/ml) stimulation for 15 mins to study signaling events or 24 h to determine downstream inflammatory mediators using Western blotting and ELISA methods. To study the superiority of TAK1 inhibition over NF-κB or MAPKs pathway inhibition, JIASFs were pretreated for 2 h with chemical inhibitors of NF-κB (PDTC), ERK (PD98059), p38 (SB203580), or JNK (SP600125), followed by IL-1β stimulation. Trans-well migration and CyQUANT cell proliferation assays were used to determine the impact on JIASF’s function.

Results: Compared to normal foreskin fibroblasts (FSKs), JIASFs exhibited 50% higher expression of total TAK1 (n=4; p< 0.05) and ~50% lower expression of p53 (n=4; p< 0.05). IL-1β suppressed the expression of p53 in JIASFs by ~33% (n=4; p< 0.01) compared to untreated cells, while TAK1 inhibition restored the expression of p53 almost to its basal level in a dose-dependent manner (n=4; p< 0.05). The expression of anti-apoptotic Mdm2, Bcl-2, and Mcl-1 showed a modest decrease, with no change in cleaved PARP-1 or Bak expression, suggesting no significant impact of 5Z on apoptosis. Consistent with p53 increase, 5Z pretreatment increased the expression of p21 by 50% (n=4; p< 0.05) with a concomitant reduction in p-Rb (~55%), PCNA (~40%), and E2F1 (~50%) expression in IL-1β-activated JIASFs in vitro (n=4; p< 0.01). Comparison study with NF-κB and MAPK inhibitors showed only a modest effect on p53 and p21 restoration by ERK or p38 inhibition, suggesting that TAK1 inhibition is a more effective target to limit IL-1β-induced JIASF activation. Additionally, 5Z dose-dependently suppressed the IL-1β-induced expression of MMP-1 (n=4; p< 0.0001) and MMP-3 (n=4; p< 0.001) production in JIASFs. Consistent with the identified cellular changes, the trans-well migration and CyQUANT cell proliferation assay revealed that TAK1 inhibition significantly reduced IL-1β mediated JIASFs’ migration (p< 0.05) and proliferation (p< 0.05), respectively.

Conclusion: The current study provides evidence of the role of TAK1 in JIASFs aggressive and inflammatory phenotypes and underlines the prospect of therapeutically targeting TAK1 to limit cytokine-driven synovial inflammation in JIA.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/tgf-%ce%b2-activated-kinase-1-tak1-inhibition-suppresses-synovial-inflammation-and-tissue-destruction-mediated-by-activated-juvenile-idiopathic-arthritis-synovial-fibroblasts/