Detection and comparison of nitric oxide in clinically normal horses and those with naturally acquired small intestinal strangulation obstruction

The purpose of this study was to determine whether nitric oxide (NO) is pre sent in clinically normal horses under basal conditions and if it increases secondary to naturally acquired small intestinal strangulation obstruction . Thirty-one horses were used; 20 horses with naturally acquired small inte stinal strangulation obstruction and 11 clinically normal horses with no si gns of gastrointestinal tract disease. Jugular venous blood, abdominal flui d, and urine were collected for NO quantification. Plasma, abdominal fluid, and urine were stored at -70 degrees C until analyzed for NO using a chemi luminescent method. Biopsy specimens collected from the affected jejunal se gment, during anesthesia or after immediately after euthanasia, or from the mid-jejunum of control horses, were divided into subsections for fixation in zinc formalin and cryopreservation in OCT gel. Nicotinamide adenine dinu cleotide phosphate (reduced) (NADPH) diaphorase histochemical stains were p erformed on cryopreserved tissues and inducible nitric oxide synthase (iNOS ) and nitrotyrosine immunohistochemical stains were performed on formalin-f ixed, paraffin-embedded tissues. There were significantly greater plasma an d abdominal fluid NO concentrations in affected horses as compared with con trols, but there were no significant differences between horses for urine N O concentrations. There was a significant decrease in NADPH diaphorase stai n in mucosal epithelium, vasculature, and leukocytes, and in submucosal ple xi in affected horses compared with control horses. There was a significant increase in iNOS staining in mucosal and submucosal leukocytes and in muco sal leukocyte nitrotyrosine staining of the affected compared with control horses. Endothelial NOS and neuronal NOS are present under basal conditions in the jejunum of horses and probably mediate physiologic or cytoprotectiv e effects. Plasma and abdominal fluid, but not urine, NO concentrations inc rease subsequent to small intestinal strangulation obstruction; this may be associated with increased mucosal and submucosal iNOS staining in leukocyt es, which was likely due to increased expression subsequent to stimuli asso ciated with ischemia. The increased nitrotyrosine staining in mucosal leuko cytes of affected horses likely reflects the presence of peroxynitrite subs equent to increased NO and superoxide production and may reflect a cytotoxi c role of NO in small intestinal strangulation obstruction in horses.