Mechanisms of human erythrocytic bioactivation of nitrite

J Biol Chem. 2015 Jan 9;290(2):1281-94. doi: 10.1074/jbc.M114.609222. Epub 2014 Dec 3.


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.

Keywords: Erythrocyte; Hemoglobin; Nitric Oxide; Nitrite; Platelet; Redox Signaling; Spectroscopy.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Blood Platelets / metabolism
  • Carbonic Anhydrases / drug effects
  • Electron Spin Resonance Spectroscopy
  • Erythrocytes / metabolism*
  • Hemoglobins / isolation & purification
  • Hemoglobins / metabolism*
  • Humans
  • Nitric Oxide / biosynthesis*
  • Nitric Oxide / metabolism
  • Nitrite Reductases / metabolism
  • Nitrites / metabolism
  • Oxidation-Reduction*
  • Signal Transduction
  • Sulfonamides / administration & dosage
  • Thiophenes / administration & dosage


  • Hemoglobins
  • Nitrites
  • Sulfonamides
  • Thiophenes
  • nitrosyl hemoglobin
  • Nitric Oxide
  • dorzolamide
  • Nitrite Reductases
  • Carbonic Anhydrases