POTENT ANTI-INFLAMMATORY EFFECTS OF DENBINOBIN MEDIATED BY DUAL INHIBITION OF EXPRESSION OF INDUCIBLE NO SYNTHASE AND CYCLOOXYGENASE 2

Inducible NO synthase (iNOS) and cyclooxygenase 2 (COX-2) have been suggested to play important roles in various inflammatory diseases. We explored the anti-inflammatory potential of a natural compound, denbinobin (5-hydroxy-3,7-dimethoxy-1,4-phenanthraquinone), by examining its effects on the expression and activity of iNOS and COX-2 in LPSactivated macrophages. Denbinobin markedly decreased the LPS (1 mu g/mL)-induced increase in iNOS and COX-2 gene and protein expression, as well as levels of the downstream products NO and prostaglandin E-2, in a concentration-dependent manner (0.3-3 mu M). In clarifying the mechanisms involved, denbinobin was found not only to inhibit LPS-induced nuclear factor kappa B (NF-kappa B) activation, an effect highly correlated with its inhibitory effect on LPS-induced inhibitory. B kinase activation, inhibitory. B degradation, NF-kappa B phosphorylation, and binding of NF-kappa B to the kappa B motif of the iNOS and COX-2 promoters, but also suppressed phosphorylation of mitogen-activated protein kinases. Reporter gene assays and Western blotting revealed that denbinobin significantly suppressed NF-kappa B activation. Furthermore, denbinobin also downregulated the LPS-mediated CD14/toll-like receptor 4 complex level and TNF-alpha, IL-1 beta, and IL-10 mRNA expression. Our results demonstrate that denbinobin exerts potent anti-inflammatory activity, suggesting that it might provide a new therapeutic approach to inflammatory diseases.