Inhibition of Nitric Oxide Synthase Expression and Activity in Macrophages by 3-Hydroxyanthranilic Acid,a Tryptophan Metabolite

Indoleamine 2,3-dioxygenase (IDO) and nitric oxide synthase (NOS) type II are induced in macrophages by interferon (IFN)-γ and lipopolysaccharide (LPS). Nitric oxide has been previously shown to inhibit IDO activity. We studied whether metabolites of tryptophan via the IDO pathway could alter NOS II activity. In RAW 264.7 cells, the phenolic antioxidant 3-hydroxyanthranilic acid (OH-AA), but not anthranilic acid, inhibited citrulline synthesis by NOS II at sub-millimolar concentrations, when added 1 h before IFN-γ and LPS. OH-AA inhibited NOS II activity in cytosolic extracts, suggesting a direct action of OH-AA on NOS II protein. Moreover, expression of NOS II mRNA and activation of the nuclear factor κB (NF-κB) in RAW 264.7 cells were decreased by a pretreatment with OH-AA, but not anthranilic acid, before addition of IFN-γ and LPS. This pretreatment also inhibited activation of NF-κB in response to TNF-α in lymphoblastoid J.Jhan5-1 cells. Finally, expression of a long terminal repeat of the human immunodeficiency virus (HIV-LTR)-driven luciferase reporter gene, controlled by NF-κB activation, was severely decreased by OH-AA or 3-hydroxykynurenine in J.Jhan5-1 cells. Other tryptophan derivatives were inactive. These data identify OH-AA as an aminophenolic tryptophan derivative inhibiting NF-κB activation and impairing both NOS II expression and activity in a millimolar concentration range.