Mechanisms of Human Erythrocytic Bioactivation of Nitrite

Nitrite signaling likely occurs through its reduction to nitric oxide (NO). Several reports support a role of erythrocytes and hemoglobin in nitrite reduction, but this remains controversial, and alternative reductive pathways have been proposed. In this work we determined whether the primary human...

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Bibliographic Details
Published in:	The Journal of biological chemistry Vol. 290; no. 2; pp. 1281 - 1294
Main Authors:	Liu, Chen, Wajih, Nadeem, Liu, Xiaohua, Basu, Swati, Janes, John, Marvel, Madison, Keggi, Christian, Helms, Christine C., Lee, Amber N., Belanger, Andrea M., Diz, Debra I., Laurienti, Paul J., Caudell, David L., Wang, Jun, Gladwin, Mark T., Kim-Shapiro, Daniel B.
Format:	Journal Article
Language:	English
Published:	United States Elsevier Inc 09-01-2015 American Society for Biochemistry and Molecular Biology
Subjects:	Blood Platelets - metabolism Carbonic Anhydrases - drug effects Electron Spin Resonance Spectroscopy Erythrocyte Erythrocytes - metabolism Hemoglobin Hemoglobins - isolation & purification Hemoglobins - metabolism Humans Nitric Oxide Nitric Oxide - biosynthesis Nitric Oxide - metabolism Nitrite Nitrite Reductases - metabolism Nitrites - metabolism Oxidation-Reduction Platelet Redox Signaling Signal Transduction Spectroscopy Sulfonamides - administration & dosage Thiophenes - administration & dosage Erythrocyte Spectroscopy Platelet Hemoglobin Nitric Oxide Nitrite Redox Signaling
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Summary:	Nitrite signaling likely occurs through its reduction to nitric oxide (NO). Several reports support a role of erythrocytes and hemoglobin in nitrite reduction, but this remains controversial, and alternative reductive pathways have been proposed. In this work we determined whether the primary human erythrocytic nitrite reductase is hemoglobin as opposed to other erythrocytic proteins that have been suggested to be the major source of nitrite reduction. We employed several different assays to determine NO production from nitrite in erythrocytes including electron paramagnetic resonance detection of nitrosyl hemoglobin, chemiluminescent detection of NO, and inhibition of platelet activation and aggregation. Our studies show that NO is formed by red blood cells and inhibits platelet activation. Nitric oxide formation and signaling can be recapitulated with isolated deoxyhemoglobin. Importantly, there is limited NO production from erythrocytic xanthine oxidoreductase and nitric-oxide synthase. Under certain conditions we find dorzolamide (an inhibitor of carbonic anhydrase) results in diminished nitrite bioactivation, but the role of carbonic anhydrase is abrogated when physiological concentrations of CO2 are present. Importantly, carbon monoxide, which inhibits hemoglobin function as a nitrite reductase, abolishes nitrite bioactivation. Overall our data suggest that deoxyhemoglobin is the primary erythrocytic nitrite reductase operating under physiological conditions and accounts for nitrite-mediated NO signaling in blood.Erythrocytes contribute to nitrite-mediated NO signaling, but the mechanism is unclear. Deoxyhemoglobin accounts for virtually all NO made from nitrite by erythrocytes with no contributions from other proposed pathways. Deoxyhemoglobin is the primary erythrocytic nitrite reductase operating under physiological conditions. Reduction by deoxyhemoglobin accounts for nitrite-mediated NO signaling in blood mediating vessel tone and platelet function.
Bibliography:	ObjectType-Article-1 SourceType-Scholarly Journals-1 ObjectType-Feature-2 content type line 23 Supported by National Institutes of Health Grant T32 GM095440. Filers of a patent application on the use of nitrite salts in cardiovascular disease. Both authors share first authorship of this paper and contributed to it equally.
ISSN:	0021-9258 1083-351X
DOI:	10.1074/jbc.M114.609222