Inhibition of LPS-stimulated NO production in mouse macrophage-like cells by azulenes

We investigated the effect of twenty-seven azulenes on nitric oxide (NO) production by mouse macrophage-like Raw 264.7 cells. No azulene derivative alone induced NO production by the Raw 264.7 cells, but inhibited lipopolysaccharide (LPS)-stimulated NO production to various extents. The ability of azulenes to inhibit NO generation by activated macrophages was generally increased when their cytotoxic activity declined. Western blot and RT-PCR analyses demonstrated that the most potent compound, 1,3-difluoroazulene [11], slightly inhibited the expression of inducible NO synthase (iNOS), but only at extremely high concentrations. ESR spectroscopy showed that [11] did not produce radical under alkaline condition, nor scavenged O2- (generated by HX-XOD reaction) or NO (generated by NOC-7). These data suggest that the inhibitory effect of [11] may be produced via a mechanism other than iNOS induction and a radical-mediated mechanism.