Intraphagosomal Peroxynitrite as a Macrophage-derived Cytotoxin against Internalized Trypanosoma cruzi CONSEQUENCES FOR OXIDATIVE KILLING AND ROLE OF MICROBIAL PEROXIREDOXINS IN INFECTIVITY

Macrophage-derived radicals generated by the NADPH oxidase complex and inducible nitric-oxide synthase (iNOS) participate in cytotoxic mechanisms against microorganisms. Nitric oxide ((NO)-N-center dot) plays a central role in the control of acute infection by Trypanosoma cruzi, the causative agent of Chagas disease, and we have proposed that much of its action relies on macrophage-derived peroxynitrite (ONOO- + ONOOH) formation, a strong oxidant arising from the reaction of (NO)-N-center dot with superoxide radical (O-2(radical anion)). Herein, we have shown that internalization of T. cruzi trypomastigotes by macrophages triggers the assembly of the NADPH oxidase complex to yield O-2(radical anion) during a 60-90-min period. This does not interfere with IFN-gamma-dependent iNOS induction and a sustained (NO)-N-center dot production (similar to 24 h). The major mechanism for infection control via reactive species formation occurred when (NO)-N-center dot and O-2(radical anion) were produced simultaneously, generating intraphagosomal peroxynitrite levels compatible with microbial killing. Moreover, biochemical and ultrastructural analysis confirmed cellular oxidative damage and morphological disruption in internalized parasites. Overexpression of cytosolic tryparedoxin peroxidase in T. cruzi neutralized macrophage-derived peroxynitrite-dependent cytotoxicity to parasites and favored the infection in an animal model. Collectively, the data provide, for the first time, direct support for the action of peroxynitrite as an intraphagosomal cytotoxin against pathogens and the premise that microbial peroxiredoxins facilitate infectivity via decomposition of macrophage-derived peroxynitrite.