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Inactivation of nitric oxide synthase after prolonged incubation of mouse macrophages with IFN-gamma and bacterial lipopolysaccharide.

Y Vodovotz, N S Kwon, M Pospischil, J Manning, J Paik and C Nathan
J Immunol April 15, 1994, 152 (8) 4110-4118;
Y Vodovotz
Beatrice and Samuel A. Seaver Laboratory, Cornell University Medical College, New York, NY 10021.
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N S Kwon
Beatrice and Samuel A. Seaver Laboratory, Cornell University Medical College, New York, NY 10021.
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M Pospischil
Beatrice and Samuel A. Seaver Laboratory, Cornell University Medical College, New York, NY 10021.
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J Manning
Beatrice and Samuel A. Seaver Laboratory, Cornell University Medical College, New York, NY 10021.
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J Paik
Beatrice and Samuel A. Seaver Laboratory, Cornell University Medical College, New York, NY 10021.
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C Nathan
Beatrice and Samuel A. Seaver Laboratory, Cornell University Medical College, New York, NY 10021.
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Abstract

Large amounts of nitric oxide (NO) are produced by the inducible isoform of NO synthase (iNOS) in many cell types once the iNOS gene is transcriptionally activated. In primary mouse peritoneal macrophages elicited by thioglycolate broth, expression of iNOS follows treatment with IFN-gamma and is synergistically increased by the addition of bacterial LPS. Expression of iNOS is suppressible at transcriptional and translational levels by certain cytokines and microbial products. The present study describes a novel form of inactivation of iNOS that is post-translational and nondegradative. Mouse peritoneal macrophages cultured in the presence of IFN-gamma alone or IFN-gamma plus LPS rapidly depleted the medium of L-arginine, a substrate for iNOS, and stopped producing NO. Repletion of L-arginine permitted cells treated with IFN-gamma alone to resume NO production for at least 5 days, leading to the release of more NO than macrophages were previously believed capable of generating. L-Arginine repletion also boosted NO production by macrophages cultured for up to 2 to 3 days in the presence of IFN-gamma plus LPS, but thereafter, iNOS was inactive in these cells whether or not L-arginine was repleted. Activity of iNOS could be restored by adding both L-arginine and fresh IFN-gamma with or without LPS, likely reflecting the synthesis of new enzyme. However, the inactivation of iNOS seen late in culture with a single application of IFN-gamma plus LPS could be attributed neither to loss of iNOS protein nor to its autoinactivation by NO. Thus, LPS, a co-inducer of iNOS, causes macrophages to inactivate iNOS about 3 days after the onset of its induction. The mechanism, which remains to be identified, is novel for iNOS, in that it decreases neither its amount nor its apparent molecular mass.

  • Copyright © 1994 by American Association of Immunologists

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The Journal of Immunology
Vol. 152, Issue 8
15 Apr 1994
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Inactivation of nitric oxide synthase after prolonged incubation of mouse macrophages with IFN-gamma and bacterial lipopolysaccharide.
Y Vodovotz, N S Kwon, M Pospischil, J Manning, J Paik, C Nathan
The Journal of Immunology April 15, 1994, 152 (8) 4110-4118;

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Inactivation of nitric oxide synthase after prolonged incubation of mouse macrophages with IFN-gamma and bacterial lipopolysaccharide.
Y Vodovotz, N S Kwon, M Pospischil, J Manning, J Paik, C Nathan
The Journal of Immunology April 15, 1994, 152 (8) 4110-4118;
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Print ISSN 0022-1767        Online ISSN 1550-6606