The overproduction of
nitric oxide (NO) by
inducible nitric oxide synthase (iNOS) may contribute to the pathophysiology of intestinal injury induced by
ischemia-reperfusion. The aim of the present study was to examine the effect of selective iNOS inhibition by a cyclic amidine analogue,
ONO-1714, on reperfusion-induced small intestinal injury and
inflammation in rats. Intestinal damage was induced in male Sprague-Dawley rats by clamping both the superior mesenteric artery and the celiac trunk for 30 min, followed by reperfusion. The
luminal nitrite concentration in the small intestine was measured by Griess reaction and the iNOS
mRNA expression by RT-PCR. The severity of the intestinal mucosal injury and
inflammation were evaluated by several
biochemical markers and by the histological findings. The rats which were killed after
ischemia-reperfusion had increased
luminal concentrations of
nitrite and iNOS
mRNA expression, in addition to severe intestinal
inflammation characterized by significant increases in
myeloperoxidase activity, a marker of neutrophil infiltration, and by the mucosal content of CINC-1
cytokine, a neutrophil
chemotactic cytokine. Administration with
ONO-1714 significantly inhibited the
luminal NO production. Reperfusion after 30-min
ischemia resulted in an increase in
luminal protein and
hemoglobin concentrations, with levels reaching a maximum after 60 min of reperfusion. In contrast, pre-treatment with
ONO-1714 2h before the
ischemia inhibited the increases in
luminal protein and
hemoglobin concentration in a dose-dependent manner (0.001-0.1mg/kg). The contents of the
thiobarbituric acid-reactive substances (a marker of oxidative lipid peroxidation) were significantly increased by
ischemia-reperfusion, and this increase was reduced by
ONO-1714. After reperfusion, the increase in tissue-associated
myeloperoxidase activity, an index of neutrophil infiltration, was significantly inhibited by pre-treatment with
ONO-1714.
ONO-1714 also inhibited increases in intestinal CINC-1
protein and
mRNA expression, as determined by ELISA and RT-PCR, respectively. In conclusion, the improvement of reperfusion-induced intestinal injury by
ONO-1714 suggested that an excess of NO, produced by iNOS, may have contributed to the initiation/amplification of intestinal inflammatory injury by various mechanisms, including nitrosative and oxidative damage as well as the enhancement of inflammatory
cytokine release.