Analysis of the mechanism for the anti-inflammatory effect of the anti-rheumatic drug auranofin

Abstract

Effects of auranofin, an orally active chrysotherapeutic agent, were examined on the production of prostaglandin E2 (PGE2) and nitric oxide (NO) in rat peritoneal macrophages and in RAW 264.7 cells, a murine macrophage-like cell line. Auranofin (1-10 microM) inhibited PGE2 production in rat peritoneal macrophages stimulated with 12-O-tetra-decanoylphorbol 13-acetate (TPA, 16.2 nM) at 8-20 h, but did not affect PGE2 production at 4 h. However, in non-stimulated rat peritoneal macrophages, auranofin increased PGE2 production at 4 h and had no effect on PGE2 production at 8-20 h. It was proved that auranofin (1-10 microM) increased COX (cyclooxygenase)-1-dependent PGE2 production and inhibited COX-2-dependent PGE2 production in rat peritoneal macrophages. Auranofin showed no effect on the enzyme activities of the purified COX-1 and COX-2 proteins. Furthermore, auranofin did not affect the COX-1 protein level, but inhibited the TPA-induced expression of COX-2 protein. Therefore, it was suggested that auranofin inhibited PGE2 production by inhibiting the COX-2 protein induction in TPA-stimulated macrophages. In RAW 264.7 cells, auranofin (0.3-3 microM) inhibited lipopolysaccharide-induced NO synthesis by inhibiting the induction of NO synthase (NOS) protein expression. Auranofin did not affect the enzyme activity of iNOS (inducible NOS). Finally, using rat peritoneal macrophages, the effects of auranofin on PGE2 production and NO production were determined. Auranofin (10 microM) strongly inhibited the production of PGE2 and NO, and the induction of COX-2 protein and NOS protein by TPA. Indomethacin, a COX inhibitor, partially inhibited NO production at the concentration at which PGE2 production was completely inhibited. On the other hand, L-NG-monomethyl-L-arginine acetate (L-NMMA), a NOS inhibitor, partially inhibited PGE2 production. NO production was completely inhibited at the same concentration as shown above. These findings suggest that PGE2 production and NO production partially affect each other. Therefore, the inhibition of PGE2 production by auranofin might be partly due to the inhibition of NO production, and the inhibition of NO production by auranofin be partly due to the inhibition of PGE2 production. In conclusion, auranofin inhibits both PGE2 production and NO production by inhibiting the upregulation of mRNA levels of COX-2 and NOS.