title: Macrophage cytotoxicity against Entamoeba histolytica trophozoites is mediated by nitric oxide from L-arginine.
abstract: The killing of Entamoeba histolytica trophozoites by phagocytes involves oxidative and nonoxidative mediators. In this study, we determine whether L-arginine-derived nitric oxide (NO) is involved in the killing of E. histolytica trophozoites by activated murine macrophages in vitro. Elicited peritoneal and bone marrow-derived macrophages activated with IFN-gamma alone or with IFN-gamma and LPS killed 62 to 73% of amebae, concomitant with increased levels of nitrate (NO2). Depletion of L-arginine by addition of arginase to culture medium abrogated macrophage amebicidal activity. NG-monomethyl L-arginine, an L-arginine analog, competitively inhibited NO2 release and amebicidal activity in a dose-dependent fashion, without affecting H2O2 production; however, the addition of excess L-arginine competitively restored macrophage amebicidal effects. In culture, sodium nitrite and sodium nitroprusside were cytotoxic to E. histolytica and this was reversed by the addition of myoglobin. Exogenously added FeSO4 prevented macrophage cytotoxicity. Addition of superoxide dismutase, a scavenger of O2-, partially inhibited amebicidal activity, without influencing NO2 production. Untreated and LPS-exposed macrophages produced high levels of H2O2 independent from NO2 production and amebicidal effects. However, the addition of catalase, a scavenger of H2O2, inhibited both amebicidal activity and NO2 production by activated macrophages. Our results demonstrate that NO is the major cytotoxic molecule released by activated macrophages for the in vitro cytotoxicity of E. histolytica and that O2- and H2O2 may be cofactors for the NO effector molecule.
authors.affiliation: Institute of Parasitology, McGill University, Ste-Anne de Bellevue, Québec, Canada.
authors.forename: J Y
authors.initials: JY
authors.lastname: Lin
authors.forename: K
authors.initials: K
authors.lastname: Chadee
mesh_terms: D000818:Animals; D001119:Arginase / Q000494:pharmacology; D001120:Arginine / Q000031:analogs & derivatives / Q000494:pharmacology / Q000502:physiology*; D003602:Cytotoxicity, Immunologic; D004748:Entamoeba histolytica / Q000276:immunology*; D006861:Hydrogen Peroxide / Q000378:metabolism; D007501:Iron / Q000378:metabolism; D008262:Macrophage Activation; D008264:Macrophages / Q000276:immunology*; D051379:Mice; D008807:Mice, Inbred BALB C; D009569:Nitric Oxide / Q000378:metabolism*; D013481:Superoxides / Q000378:metabolism; D019323:omega-N-Methylarginine
chemical_list: D013481:Superoxides; D019323:omega-N-Methylarginine; D009569:Nitric Oxide; D001120:Arginine; D006861:Hydrogen Peroxide; D007501:Iron; D001119:Arginase