title: Mechanisms of human erythrocytic bioactivation of nitrite.
abstract: 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.
authors.affiliation: From the Department of Physics and.
authors.forename: Chen
authors.initials: C
authors.lastname: Liu
authors.affiliation: From the Department of Physics and.
authors.forename: Nadeem
authors.initials: N
authors.lastname: Wajih
authors.affiliation: From the Department of Physics and.
authors.forename: Xiaohua
authors.initials: X
authors.lastname: Liu
authors.affiliation: From the Department of Physics and the Translational Science Center Wake Forest University, Winston-Salem, North Carolina 27109, the Departments of Radiology and.
authors.forename: Swati
authors.initials: S
authors.lastname: Basu
authors.affiliation: From the Department of Physics and.
authors.forename: John
authors.initials: J
authors.lastname: Janes
authors.affiliation: From the Department of Physics and.
authors.forename: Madison
authors.initials: M
authors.lastname: Marvel
authors.affiliation: From the Department of Physics and.
authors.forename: Christian
authors.initials: C
authors.lastname: Keggi
authors.affiliation: Physics Department, University of Richmond, Richmond, Virgina 23173.
authors.forename: Christine C
authors.initials: CC
authors.lastname: Helms
authors.affiliation: From the Department of Physics and.
authors.forename: Amber N
authors.initials: AN
authors.lastname: Lee
authors.affiliation: From the Department of Physics and.
authors.forename: Andrea M
authors.initials: AM
authors.lastname: Belanger
authors.affiliation: the Translational Science Center Wake Forest University, Winston-Salem, North Carolina 27109, Hypertension and Vascular Research Center and.
authors.forename: Debra I
authors.initials: DI
authors.lastname: Diz
authors.affiliation: the Translational Science Center Wake Forest University, Winston-Salem, North Carolina 27109, the Departments of Radiology and Biomedical Engineering and.
authors.forename: Paul J
authors.initials: PJ
authors.lastname: Laurienti
authors.affiliation: Pathology-Comparative Medicine, Wake Forest University School of Medicine, Winston-Salem, North Carolina 27157, and.
authors.forename: David L
authors.initials: DL
authors.lastname: Caudell
authors.affiliation: Heart, Lung, Blood, and Vascular Medicine Institute and.
authors.forename: Jun
authors.initials: J
authors.lastname: Wang
authors.affiliation: Heart, Lung, Blood, and Vascular Medicine Institute and the Department of Medicine, Division of Pulmonary, Allergy, and Critical Care Medicine, University of Pittsburgh, Pittsburgh, Pennsylvania 15213.
authors.forename: Mark T
authors.initials: MT
authors.lastname: Gladwin
authors.affiliation: From the Department of Physics and the Translational Science Center Wake Forest University, Winston-Salem, North Carolina 27109, Hypertension and Vascular Research Center and Biomedical Engineering and shapiro@wfu.edu.
authors.forename: Daniel B
authors.initials: DB
authors.lastname: Kim-Shapiro
mesh_terms: D001792:Blood Platelets / Q000378:metabolism; D002256:Carbonic Anhydrases / Q000187:drug effects; D004578:Electron Spin Resonance Spectroscopy; D004912:Erythrocytes / Q000378:metabolism*; D006454:Hemoglobins / Q000302:isolation & purification / Q000378:metabolism*; D006801:Humans; D009569:Nitric Oxide / Q000096:biosynthesis* / Q000378:metabolism; D009572:Nitrite Reductases / Q000378:metabolism; D009573:Nitrites / Q000378:metabolism; D010084:Oxidation-Reduction; D015398:Signal Transduction; D013449:Sulfonamides / Q000008:administration & dosage; D013876:Thiophenes / Q000008:administration & dosage
chemical_list: D006454:Hemoglobins; D009573:Nitrites; D013449:Sulfonamides; D013876:Thiophenes; C008657:nitrosyl hemoglobin; D009569:Nitric Oxide; C062765:dorzolamide; D009572:Nitrite Reductases; D002256:Carbonic Anhydrases
keywords: Erythrocyte; Hemoglobin; Nitric Oxide; Nitrite; Platelet; Redox Signaling; Spectroscopy