MURINE MACROPHAGES USE OXYGEN-DEPENDENT AND NITRIC OXIDE-DEPENDENT MECHANISMS TO SYNTHESIZE S-NITROSO-ALBUMIN AND TO KILL EXTRACELLULAR TRYPANOSOMES

Reactive nitrogen intermediates were synthesized spontaneously in cult ures of macrophages from Trypanosoma brucei brucei-infected mice by an inducible nitric oxide (NO) synthase. This was inhibited by the addit ion of nitro-L-arginine, In this paper, we report the kinetics of the fixation of macrophage-derived NO on bovine serum albumin by using an enzyme-linked immunosorbent assay. S nitrosylation was confirmed by th e Saville reaction, using mercuric chloride, It is known that reactive oxygen intermediates (ROI) are also synthesized by stimulated macroph ages. The fact that NO is able to bind cysteine only under aerobic con ditions led us to investigate the role of macrophage-derived ROI in th e formation of S-nitrosylated proteins by activated macrophages. The i mmunoenzymatic signal decreased by 66 and 30% when superoxide dismutas e and catalase, respectively, were added to the culture medium of macr ophages from infected mice. In addition, the decrease in S-nitrosylate d albumin formation correlated with the protection of extracellular tr ypanosomes from the cytostatic and cytotoxic activity of NO. Melatonin , a hydroxyl radical scavenger resulting from the decomposition of per oxynitrous acid, had no effect. All these data support the concept tha t an interaction between NO and ROI promoted the production of S-nitro so-albumin by activated macrophages from infected mice.