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Hoffmann, R. A wiki for the life sciences where authorship matters. Nature Genetics (2008)

Bacterial lipopolysaccharide-stimulated nitric oxide generation is unrelated to concurrent cytotoxicity of bovine alveolar macrophages.

Nitric Oxide (NO) is a reactive metabolite produced by stimulated macrophages, and it has been demonstrated to exert cytotoxic actions on a number of microbes, parasites, and tumor cells. In addition, NO has been reported to have an autotoxic effect on murine macrophages, its site of synthesis. We have investigated the relationship of NO generation to cytotoxicity of bovine alveolar macrophages (AM) in vitro, and have also assessed the effects of several modulators of cellular function on this relationship. NO was generated in cultures of AM using sodium nitroprusside (SNP) and measured as [nitrite]. Cellular viability of AM reflected a strong, negative correlation with the concentration of NO/nitrite in supernatants (r = -0.987). Supernatants with nitrite concentrations in excess of 30 microMs were correlated with cytotoxicity. AM stimulated with the potent combination of endotoxin (Lipopolysaccharide, LPS; 10 ng/ml) and recombinant bovine IFN gamma (100 U/ml) also exhibited cytotoxicity over a 48-hour incubation period, and cells deteriorated to an average viability of 72.3% as compared to unstimulated control macrophages. In some cases the viability of macrophages was much lower. Even though LPS-mediated cytotoxicity occurred, the [nitrite] produced in supernatants during the 48-hour period (12.23 microMs) was well below the minimum concentration of SNP-generated NO required to induce cytotoxicity to macrophages. N(G)-monomethyl-L-arginine (N(G)MMA, 2 mM) is a competitive inhibitor of NO synthesis and was found to reduce nitrite concentrations from 12.23 microMs to 1.56 microMs in supernatants of LPS-stimulated AM, but this reduction did not promote increased viability of AM. Other modulators of cellular function including phenylbutazone (PBZ, 100 microMs), flunixin meglumine (FM, 100 microMs) and interleukin-4 (IL-4, 100 ng/ml) modestly inhibited synthesis of NO, but did not improve cellular viability. These results suggest that relatively high concentrations of exogenously-generated NO are toxic to AM in vitro, but the quantity of endogenously-generated NO synthesized by LPS-stimulated bovine AM is usually below the threshold for toxicity. Cytotoxicity occurs independently of NO synthesis, and factors other than NO are apparently responsible for LPS-related cytotoxicity to bovine macrophages.[1]


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