Non-invasive Analysis of Reactive Oxygen Species Generated in NH⁴OH-induced Gastric Lesions of Rats using a 300 MHz In Vivo ESR Technique

Abstract

Free radicals are reportedly involved in mucosal injury, including NH⁴OH-induced gastric lesions, but the kind, location and origin of radical generation have yet to be clarified. We developed the non-invasive measurement of reactive oxygen species (ROS) in stomach, and applied to mucosal injury. NH⁴OH-induced gastric lesions were prepared in rats, which were then given a nitroxyl probe intragastrically or intravenously, and the spectra of the gastric region were obtained by in vivo 300 MHz electron spin resonance (ESR) spectroscopy. The spectral change of the nitroxyl probe administered intragastrically was significantly enhanced 30 min after NH⁴OH administration, but no change occurred when the probe was given by intravenous injection. The enhanced change was confirmed to be due to •OH generation, because it was completely suppressed by mannitol, catalase and desferrioxamine (DFO), and was not observed in neutropenic rats. NH⁴OH-induced neutrophil infiltration of the gastric mucosa was suppressed by intravenous injection of superoxide dismutase (SOD) or catalase, or by administration of allopurinol. The present study provided the direct evidence in NH⁴OH-treated living rats that •OH produced from O₂^•- derived from neutrophils caused gastric lesion formation, while O₂^•- or H²O² derived from the xanthine oxidase system in endothelial cells was involved in neutrophil infiltration.

• Hydroxyl radical
• Free radicals
• Oxidative injury
• ESR