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1 Department of Pharmacology, University of Tokushima School of Medicine, Tokushima, Japan; Department of Urology, University of Tokushima School of Medicine, Tokushima, Japan
2 Department of Clinical Pharmacology, Graduate School of Pharmaceutical Sciences, University of Tokushima, Tokushima, Japan
3 Department of Pharmacology, University of Tokushima School of Medicine, Tokushima, Japan
4 Department of Urology, University of Tokushima School of Medicine, Tokushima, Japan
* To whom correspondence should be addressed. E-mail: tamaki{at}basic.med.tokushima-u.ac.jp.
Nitric oxide (NO) is synthesized from L-arginine by nitric oxide synthase (NOS), and nitrite and nitrate are believed to be waste forms of NO. We previously reported an enzyme-independent pathway of NO generation from nitrite in acidic conditions. In this study, we show nitrite-derived NO formation in renal ischemia-reperfusion injury using electron paramagnetic resonance (EPR) spectroscopy. In this experiment, we utilized a stable isotope of 15N-nitrite as a source of nitrite in order to distinguish L-arginine-derived NO from 15N-nitrite-derived 15NO. Intravenous infusion of a stable isotope of 15N-nitrite (15NO2-) facilitated the formation of Hb15NO during renal ischemia, which demonstrated that the origin of NO was nitrite. The EPR signal of Hb15NO in kidney appeared after 40 min of renal ischemia, and renal reperfusion decreased the Hb15NO level in kidney and increased it in blood by contrast. In addition, the amount of HbNO was nitrite concentration-dependent, and this formation was NOS-independent. Our findings suggest that the nitrite can be an alternative source of NO in ischemic kidney and that it binds with hemoglobin and then is spread by the circulation after reperfusion.
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