NICOTINAMIDE INHIBITS NITRIC-OXIDE SYNTHASE MESSENGER-RNA INDUCTION IN ACTIVATED MACROPHAGES

Nitric oxide (NO) is a potent mediator involved in many biological fun ctions including inflammation and non-specific immunity. Murine macrop hages possess the prototype of high-output NO synthase which is not co nstitutively expressed but induced within a few hours by immunological stimuli. In this study, we explored the possibility of controlling th e activity of the inducible NO synthase by interfering with the transd uction signal which triggers its induction, in the RAW 264.7 macrophag e cell line. We found that nicotinamide, an inhibitor of ADP-ribosylat ion, prevented NO synthase induction in RAW 264.7 cells after stimulat ion with interferon gamma (IFN-gamma) and lipopolysaccharide (LPS). Fu rthermore, the level of NO synthase mRNA was measured by Northern-blot analysis and we found that nicotinamide prevents expression of NO syn thase mRNA in IFN-gamma- and LPS-stimulated cells. Nicotinamide was al so found to inhibit other macrophage functions expressed in response t o IFN-gamma, i.e. tumour necrosis factor secretion and the expression of the Ia antigen of the major histocompatibility complex. Analysis of the pattern of ADP-ribosylated proteins revealed that nicotinamide as well as cholera toxin prevented the ADP-ribosylation of a 107-117 kDa protein found constitutively ADP-ribosylated in stimulated and non-st imulated macrophage extracts. Together, our results indicate ADP-ribos ylation as a crucial point of the signalling pathway which leads to NO synthase mRNA induction.