HEPATIC NITRIC-OXIDE PRODUCTION FOLLOWING ACUTE ENDOTOXEMIA IN RATS IS MEDIATED BY INCREASED INDUCIBLE NITRIC-OXIDE SYNTHASE GENE-EXPRESSION

In the present studies, we analyzed the effects of acute endotoxemia o n hepatocyte nitric oxide production and functional activity, Treatmen t of rats with 5 mg/kg of lipopolysaccharide (LPS), which induces acut e endotoxemia, caused an increase in nitric oxide production in the li ver, as measured by electron paramagnetic spin trapping, which was evi dent within 6 hours. This was associated with expression of inducible nitric oxide synthase (iNOS) messenger (m) RNA in hepatocytes and in s inusoidal cells throughout the liver lobule. Acute endotoxemia also ca used alterations in hepatic structure, including hypertrophy, vacuoliz ation, and chromosomal emargination, however these changes were not ap parent for 24 to 48 hours. Hepatocytes isolated from endotoxemic rats released increased amounts of nitric oxide, measured by nitrite produc tion, in response to interferon gamma (gamma-IFN) alone or in combinat ion with LPS, tumor necrosis factor alpha, macrophage-colony stimulati ng factor, granulocyte/macrophage-colony stimulating factor, or hepato cyte growth factor. These results show that hepatocytes are sensitized by acute endotoxemia to respond to inflammatory mediators and growth factors. Increased nitrite production by hepatocytes was due to increa sed expression of iNOS mRNA and protein and was correlated with the ti me following induction of acute endotoxemia. Thus, cells isolated 48 h ours after induction of acute endotoxemia released significantly more nitrite than cells recovered after 6 hours, a response that was not du e to alterations in hepatocyte viability, Hepatocytes isolated hom end otoxemic rats also exhibited a marked increase in proliferative capaci ty when compared with cells from control rats. Nitric oxide production by hepatocytes in vitro was associated with inhibition of cell growth and protein synthesis, which was reversed by the nitric oxide synthas e inhibitor, N-G-monomethyl-l-arginine (L-NMMA), Agarose gel electroph oresis showed extensive cytoplasmic DNA fragmentation in hepatocytes t reated with LPS and gamma-IFN, a characteristic of apoptosis, which wa s also reversed by L-NMMA. These results, together with our findings t hat treatment of rats with an inhibitor of nitric oxide synthase parti ally reversed the structural alterations in the liver associated with acute endotoxemia suggest that nitric oxide may contribute to the path ophysiologic response to this bacterially derived toxin.