Free radical production requires both inducible nitric oxide synthase and xanthine oxidase in LPS-treated skin

Proceedings of the National Academy of Sciences of the United States of America
Kozo NakaiRonald P Mason

Abstract

Free radical formation has been investigated in diverse experimental models of LPS-induced inflammation. Here, using electron spin resonance (ESR) and the spin trap alpha-(4-pyridyl-1-oxide)-N-tert-butylnitrone, we have detected an ESR spectrum of alpha-(4-pyridyl-1-oxide)-N-tert-butylnitrone radical adducts in the lipid extract of mouse skin treated with LPS for 6 h. The ESR spectrum was consistent with the trapping of lipid-derived radical adducts. In addition, a secondary radical-trapping technique using dimethyl sulfoxide (DMSO) demonstrated methyl radical formation, revealing the production of hydroxyl radical. Radical adduct formation was suppressed by aminoguanidine, N-(3-aminomethyl)benzylacetamidine (1400W), or allopurinol, suggesting a role for both inducible nitric oxide synthase (iNOS) and xanthine oxidase (XO) in free radical formation. The radical formation was also suppressed in iNOS knockout (iNOS(-/-)) mice, demonstrating the involvement of iNOS. NADPH oxidase was not required in the formation of these radical adducts because the ESR signal intensity was increased by LPS treatment in NADPH oxidase knockout (gp91(phox-/-)) mice as much as it was in the wild-type mouse. Nitric oxide (*NO) end products were increa...Continue Reading

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