title: Relationship of tumor necrosis factor alpha, the nitric oxide synthase pathway, and lipopolysaccharide to the killing of gamma interferon-treated macrophage-like RAW264.7 cells by Rickettsia prowazekii.
abstract: Macrophage-like RAW264.7 cells are killed by the combination of gamma interferon (IFN-gamma) treatment and infection with Rickettsia prowazekii. The roles of tumor necrosis factor alpha (TNF-alpha), the nitric oxide synthase pathway, and lipopolysaccharide (LPS) in this killing were investigated. R. prowazekii, both the Breinl and Madrid E strains, induced RAW264.7 cells to produce TNF-alpha. However, dead rickettsiae (which cannot kill the IFN-gamma-treated RAW264.7 cells) induced the production of as much TNF-alpha as viable rickettsiae. Inhibition of the production of TNF-alpha (by the addition of actinomycin D or emetine during the rickettsial infection) or neutralization of TNF-alpha (by the addition of polyclonal rabbit anti-mouse TNF-alpha serum both during the IFN-gamma treatment and during the rickettsial infection) did not inhibit the killing of the RAW264.7 cells. Addition of polymyxin B (which inhibits many effects of LPS) during the IFN-gamma treatment did not inhibit the ability of IFN-gamma to prepare the RAW264.7 cells to be killed by R. prowazekii. Suppression of nitrite production by addition of the nitric oxide synthase inhibitor aminoguanidine both during the IFN-gamma treatment and during the rickettsial infection also did not inhibit the killing of the RAW264.7 cells. R. prowazekii-mediated killing of the RAW264.7 cells was dramatically suppressed in cultures treated with IFN-gamma plus LPS compared with that in cultures treated with IFN-gamma alone, and inhibition of nitric oxide synthase restored the rickettsia-induced killing of the RAW264.7 cells in cultures treated with IFN-gamma plus LPS. These data indicate that (i) TNF-alpha, LPS, and the nitric oxide synthase pathway are not required in order for IFN-gamma to prepare RAW264.7 cells to be killed by R. prowazekii; (ii) neither TNF-alpha nor the nitric oxide synthase pathway is responsible for the killing of the IFN-gamma-treated RAW264.7 cells by R. prowazekii; and (iii) in cultures treated with IFN-gamma plus LPS and then incubated with rickettsiae, a nitric oxide synthase pathway-dependent mechanism inhibits the killing of the RAW264.7 cells.
authors.affiliation: Department of Microbiology and Immunology, University of South Alabama College of Medicine, Mobile 36688.
authors.forename: J
authors.initials: J
authors.lastname: Turco
authors.forename: H H
authors.initials: HH
authors.lastname: Winkler
mesh_terms: D000594:Amino Acid Oxidoreductases / Q000502:physiology*; D000818:Animals; D016923:Cell Death; D002478:Cells, Cultured; D007371:Interferon-gamma / Q000494:pharmacology*; D008070:Lipopolysaccharides / Q000633:toxicity*; D008264:Macrophages / Q000187:drug effects / Q000502:physiology*; D051379:Mice; D019001:Nitric Oxide Synthase; D009573:Nitrites / Q000378:metabolism; D012283:Rickettsia prowazekii / Q000472:pathogenicity*; D014409:Tumor Necrosis Factor-alpha / Q000502:physiology*
chemical_list: D008070:Lipopolysaccharides; D009573:Nitrites; D014409:Tumor Necrosis Factor-alpha; D007371:Interferon-gamma; D019001:Nitric Oxide Synthase; D000594:Amino Acid Oxidoreductases