Suppression of PU.1-linked TLR4 expression by cilostazol with decrease of cytokine production in macrophages from patients with rheumatoid arthritis.

Background and Purpose: The present study assessed the effects of cilostazol on LPS-stimulated TLR4 signal pathways in synovial macrophages from patients with rheumatoid arthritis (RA). These effects were confirmed in collagen-induced arthritis (CIA) in mice.
Experimental Approach: Expression of TLR4, PU.1, NF-κB p65 and IκBα on synovial fluid macrophages from RA patients was determined by Western blotting, and cytokines were measured by ELISA. Anti-arthritic effects were evaluated in CIA mice.
Key Results: Intracellular cAMP was concentration-dependently raised by cilostazol (1-100 μM). Cilostazol significantly suppressed LPS-stimulated increase of TLR4 expression by blocking PU.1 transcriptional activity in RA macrophages. In addition, cilostazol decreased LPS-induced myeloid differentiation factor 88 (MyD88) expression, but not that of TNF receptor-associated factor 6 (TRAF6). Cilostazol also suppressed IkBα degradation and NF-κB p65 nuclear translocation. Moreover, LPS-induced increase of cytokine production (TNF-α, IL-1β) was inhibited by cilostazol, an effect which was accompanied by suppression of IκBα degradation, and NF-κB p65 nuclear translocation. However, expression of anti-inflammatory IL-10 was elevated by cilostazol and forskolin/IBMX. In mice with CIA, post-treatment with cilostazol (30 mg kg⁻¹ day⁻¹) decreased expression of TLR4 in knee joints in association with decreased recruitment of macrophages. Consequently, synovial inflammation, proteoglycan depletion and bone erosion were significantly inhibited by cilostazol treatment.
Conclusions and Implications: Cilostazol down-regulated LPS-stimulated PU.1-linked TLR4 expression and TLR4/MyD88/NF-κB signal pathways, and then suppressed inflammatory cytokine production in synovial macrophages from RA patients. Also cilostazol markedly inhibited the severity of CIA in mice.
(© 2012 The Authors. British Journal of Pharmacology © 2012 The British Pharmacological Society.)