In this study, we investigated the mechanisms underlying the anti-inflammatory effects of honokiol in tumor necrosis factor (TNF)-α-stimulated rheumatoid arthritis synovial fibroblasts (RASFs). RASFs pre-treated with honokiol (0-20 μM) were stimulated with TNF-α (20 ng/ml). The levels of prostaglandin E2 (PGE2), nitric oxide (NO), soluble intercellular adhesion molecule-1 (sICAM-1), transforming growth factor-β1 (TGF-β1), monocyte chemotactic protein-1 (MCP-1), and macrophage inflammatory protein-1α (MIP-1α) in supernatants were determined by enzyme-linked immunosorbent assay (ELISA) and Griess assay. In addition, protein expression levels of cyclooxygenase-2 (COX-2), inducible nitric oxide synthase (iNOS), and phosphorylated Akt, nuclear factor kappa B (NFκB), and extracellular signal-regulated kinase (ERK)1/2 were determined by western blot. The expression of NFκB-p65 was assessed by immunocytochemical analysis. TNF-α treatment significantly up-regulated the levels of PGE2, NO, sICAM-1, TGF-β1, MCP-1, and MIP-1α in the supernatants of RASFs, increased the protein expression of COX-2, iNOS, and induced phosphorylation of Akt, IκB-α, NFκB, and ERK1/2 in RASFs. TNF-α-induced expression of these molecules was inhibited in a dose-dependent manner by pre-treatment with honokiol. The inhibitory effect of honokiol on NFκB-p65 activity was also confirmed by immunocytochemical analysis. In conclusion, honokiol is a potential inhibitor of TNF-α-induced expression of inflammatory factors in RASFs, which holds promise as a potential anti-inflammatory drug.