The aim of this study was to characterize the effects of auranofin (2,3,4,6-tetra-O-acetyl-l-thio-β-d-gluco-pyranosato-S) on cyclooxygenase expression and prostaglandin E₂ synthesis on cultured human synovial fibroblast-like cells (synoviocytes). Synoviocytes were treated with auranofin in the presence or absence of interleukin-1β. Cultured supernatants were harvested for prostaglandin E₂ synthesis. Cyclooxygenase-1 and -2 expression was analyzed with Western and Northern blotting. Translocation of nuclear factor-κ B p65 was determined by immunostaining. Cytotoxicity was measured with ⁵¹Cr release assay. Auranofin attenuated interleukin-1β-induced prostaglandin E₂ production of the cells in a dose-dependent fashion. Auranofin selectively suppressed interleukin-1β-induced cyclooxygenase-2 mRNA and protein expression of the cells without alteration of cyclooxygenase-1 expression. Also, auranofin interfered with interleukin-1β-induced translocation of nuclear factor-κ B. These inhibitory effects did not originate in the cytotoxicity of the agent. Our data indicate that auranofin inhibits interleukin-1β-induced prostaglandin E₂ synthesis and cyclooxygenase-2 expression via suppression of nuclear factor-κ B activation on synoviocytes.