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J Biol Chem, Vol. 275, Issue 15, 10826-10830, April 14, 2000

Reactive Nitrogen and Oxygen Species Attenuate Interleukin- 8-induced Neutrophil Chemotactic Activity in Vitro*

Etsuro SatoDagger , Keith L. SimpsonDagger , Matthew B. Grisham§, Sekiya Koyama, and Richard A. RobbinsDagger ||

From the Dagger  Research Service, Southern Arizona Veterans Health Care System, and the Department of Medicine, University of Arizona, Tucson, Arizona 85723, the § Department of Molecular and Cellular Physiology, LSU Medical Center, Shreveport, Louisiana 71130, and the  First Department of Internal Medicine, Shinshu University School of Medicine, 3-1-1 Asahi, Matsumoto 390-8621, Japan

Peroxynitrite, formed by the reaction between nitric oxide and superoxide, has been shown to induce protein nitration, which compromises protein function. We hypothesized that peroxynitrite may regulate cytokine function during inflammation. To test this hypothesis, the neutrophil chemotactic activity (NCA) of interleukin-8 (IL-8) incubated with peroxynitrite was evaluated. Peroxynitrite attenuated IL-8 NCA in a dose-dependent manner (p < 0.01) but did not significantly reduce NCA induced by leukotriene B4 or complement-activated serum. The reducing agents, dithionite, deferoxamine, and dithiothreitol, reversed and exogenous L-tyrosine abrogated the peroxynitrite-induced NCA inhibition. Papa-NONOate [N-(3-ammoniopropyl)-N-(n-propyl)amino]diazen-1-ium-1,2-dialase or sodium nitroprusside, NO donors, or a combination of xanthine and xanthine oxidase to generate superoxide did not show an inhibitory effect on NCA induced by IL-8. In contrast, small amounts of SIN-1, a peroxynitrite generator, caused a concentration-dependent inhibition of NCA by IL-8. Consistent with its capacity to reduce NCA, peroxynitrite treatment reduced IL-8 binding to neutrophils. Nitrotyrosine was detected in the IL-8 incubated with peroxynitrite by enzyme-linked immunosorbent assay. These findings are consistent with nitration of tyrosine by peroxynitrite with subsequent inhibition of IL-8 binding to neutrophils and a reduction in NCA and suggest that oxidants may play an important role in regulation of IL-8-induced neutrophil chemotaxis.

* This work was supported by a Merit Review grant from the Department of Veterans Affairs and a grant from Rotary International.The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

|| To whom correspondence and reprint requests should be addressed: Research Health Care Group, Southern Arizona Health Care System, 3601 S. 6th Ave., Tucson, AZ 85723. Tel.: 520-629-1824; Fax: 520-629-1801; E-mail: Richard.Robbins2@med.va.gov.

Copyright © 2000 by The American Society for Biochemistry and Molecular Biology, Inc.
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