Inhibition of nitric oxide synthase attenuates peroxynitrite generation, but augments neutrophil accumulation in hepatic ischemia-reperfusion in rats.

The role of nitric oxide (NO) and peroxynitrite in the process of neutrophil adhesion and infiltration was investigated in a model of hepatic ischemia-reperfusion. Male Fischer rats were subjected to 30 min of hepatic no-flow ischemia followed by 4 h of reperfusion (I/R). I/R induced liver injury as evidenced by a 13.7-fold increase in plasma alanine aminotransferase activity. Induction of liver injury was associated with an increase in neutrophil accumulation in ischemic lobes of livers [215 +/- 27 polymorphonuclear neutrophil leukocytes/50 high-power field (HPF), P < .05 compared with sham control] and 8-fold augmentation of inducible NO synthase (NOS) activity. However, NO levels in the liver decreased; this decrease may be caused by peroxynitrite formation by the reaction of NO with superoxide. Sections of ischemic lobes of the liver tissue of I/R animals exhibited marked immunoreactivity with anti-nitrotyrosine antibody, which indicates the presence of nitrotyrosine. Administration of Nw-nitro-L-arginine methyl ester (10 mg/kg i.v. before reperfusion) attenuated total and inducible NOS activity in both ischemic and nonischemic lobes of liver, and reduced NO levels in plasma and liver. However, NOS inhibition aggravated liver injury as alanine aminotransferase increased by 61% compared with rats subjected to reperfusion injury. Neutrophil accumulation was enhanced in ischemic (436 +/- 48/50 HPF, P < .05 compared with I/R animal) and nonischemic lobes of livers (34 +/- 3.2/50 HPF, P < .05 compared with sham control). NOS inhibition also attenuated immunohistochemically detected nitrotyrosine formation, but increased superoxide production in the liver. The NO-dependent regulation of neutrophil accumulation in the liver may be linked closely to P-selectin and intracellular adhesion molecule-1 expression because inhibition of NOS resulted in significant increases in gene expression of these two adhesion molecules (determined by reverse transcription-polymerase chain reaction analysis). These results suggest that NO is important in attenuating neutrophil accumulation and liver damage in ischemia-reperfusion injury. Inhibition of NOS activity reduces peroxynitrite formation but aggravates liver injury and increases neutrophil accumulation, which suggests that the anti-inflammatory function of NO is more important than the cytotoxic potential of peroxynitrite in acute inflammation.