Human monocytic U937 cells transfected with human hepatic inducible nitricoxide synthase exhibit leishmanicidal activity

In mice, the high inducible synthesis of nitric oxide (NO) resulting from i nducible NO synthase (iNOS, NOS2) expression by macrophages (M phi) is cons idered an essential component of the protective immune response against inf ection by intracellular pathogens. Conversely, in humans, the question of a role for NO as an antimicrobial defense mechanism has been the subject of much debate. Recently, however, iNOS expression by human M phi and formatio n of NO or its derivatives have been reported both in vivo and in vitro, st rongly suggesting that human M phi are indeed capable of inducible NO synth esis, However, the: conditions allowing NO production by human M phi in cul ture remain poorly defined, rendering more difficult the study of the effec tor functions of NO in these cells. To alleviate this problem, cells of the U937 monocytoid line were engineered to express iNOS by transfection with human hepatic iNOS (DFGiNOS), leading to production of NO on supplementatio n with the cofactor tetrahydrobiopterin. We report that U937 cells, when di fferentiated with 1,25-dihydroxyvitamin D-3 and retinoic acid, acquire a ph enotype allowing infection by Leishmania parasites and maintain viable intr acellular microorganisms up to 72 h past-infection, Leishmania survival in DFGiNOS cells is strongly decreased when the cells are treated with tetrahy drobiopterin. Intracellular killing is evident by 24 h and increases up to 72 h post-infection, and is inhibited by L-N-5-(1-iminoethyl)ornithine, an inhibitor of NO synthesis. In contrast, superoxide anion does not appear to play a role in the killing of Leishmania by DGFiNOS U937 cells. The releva nce of this model to the study of the mechanisms of intracellular killing b y human macrophages is discussed.