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Nitrite reduction to nitric oxide by deoxyhemoglobin vasodilates the human circulation

Abstract

Nitrite anions comprise the largest vascular storage pool of nitric oxide (NO), provided that physiological mechanisms exist to reduce nitrite to NO. We evaluated the vasodilator properties and mechanisms for bioactivation of nitrite in the human forearm. Nitrite infusions of 36 and 0.36 μmol/min into the forearm brachial artery resulted in supra- and near-physiologic intravascular nitrite concentrations, respectively, and increased forearm blood flow before and during exercise, with or without NO synthase inhibition. Nitrite infusions were associated with rapid formation of erythrocyte iron-nitrosylated hemoglobin and, to a lesser extent, S-nitroso-hemoglobin. NO-modified hemoglobin formation was inversely proportional to oxyhemoglobin saturation. Vasodilation of rat aortic rings and formation of both NO gas and NO-modified hemoglobin resulted from the nitrite reductase activity of deoxyhemoglobin and deoxygenated erythrocytes. This finding links tissue hypoxia, hemoglobin allostery and nitrite bioactivation. These results suggest that nitrite represents a major bioavailable pool of NO, and describe a new physiological function for hemoglobin as a nitrite reductase, potentially contributing to hypoxic vasodilation.

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Figure 1: Hemodynamic and metabolic measurements at baseline and during exercise (protocol, part I).
Figure 2: Effects of nitrite infusion.
Figure 3: Effects of low-dose nitrite infusion.
Figure 4: Formation of iron-nitrosylated hemoglobin and S-nitroso-hemoglobin after nitrite infusion in vivo and in vitro.
Figure 5: Production of NO gas and vasodilation are augmented by nitrite reaction with deoxyhemoglobin.

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References

  1. Ignarro, L.J. & Gruetter, C.A. Requirement of thiols for activation of coronary arterial guanylate cyclase by glyceryl trinitrate and sodium nitrite: possible involvement of S-nitrosothiols. Biochim. Biophys. Acta. 631, 221–231 (1980).

    Article  CAS  Google Scholar 

  2. Ignarro, L.J. et al. Mechanism of vascular smooth muscle relaxation by organic nitrates, nitrites, nitroprusside and nitric oxide: evidence for the involvement of S-nitrosothiols as active intermediates. J. Pharmacol. Exp. Ther. 218, 739–749 (1981).

    CAS  PubMed  Google Scholar 

  3. Moulds, R.F., Jauernig, R.A. & Shaw, J. A comparison of the effects of hydrallazine, diazoxide, sodium nitrite and sodium nitroprusside on human isolated arteries and veins. Br. J. Clin. Pharmacol. 11, 57–61 (1981).

    Article  CAS  Google Scholar 

  4. Gruetter, C.A., Gruetter, D.Y., Lyon, J.E., Kadowitz, P.J. & Ignarro, L.J. Relationship between cyclic guanosine 3':5′-monophosphate formation and relaxation of coronary arterial smooth muscle by glyceryl trinitrate, nitroprusside, nitrite and nitric oxide: effects of methylene blue and methemoglobin. J. Pharmacol. Exp. Ther. 219, 181–186 (1981).

    CAS  PubMed  Google Scholar 

  5. Matsunaga, K. & Furchgott, R.F. Interactions of light and sodium nitrite in producing relaxation of rabbit aorta. J. Pharmacol. Exp. Ther. 248, 687–695 (1989).

    CAS  PubMed  Google Scholar 

  6. Laustiola, K.E. et al. Exogenous GTP enhances the effects of sodium nitrite on cyclic GMP accumulation, vascular smooth muscle relaxation and platelet aggregation. Pharmacol. Toxicol. 68, 60–63 (1991).

    Article  CAS  Google Scholar 

  7. Rodriguez, J., Maloney, R.E., Rassaf, T., Bryan, N.S. & Feelisch, M. Chemical nature of nitric oxide storage forms in rat vascular tissue. Proc. Natl. Acad. Sci. USA 100, 336–341 (2003).

    Article  CAS  Google Scholar 

  8. Gladwin, M.T. et al. Role of circulating nitrite and S-nitrosohemoglobin in the regulation of regional blood flow in humans. Proc. Natl. Acad. Sci. USA 97, 11482–11487 (2000).

    Article  CAS  Google Scholar 

  9. Rassaf, T. et al. NO adducts in mammalian red blood cells: too much or too little? Nat. Med. 9, 481–483 (2003).

    Article  CAS  Google Scholar 

  10. Rassaf, T., Bryan, N.S., Kelm, M. & Feelisch, M. Concomitant presence of N-nitroso and S-nitroso proteins in human plasma. Free Radic. Biol. Med. 33, 1590–1596 (2002).

    Article  CAS  Google Scholar 

  11. Rassaf, T. et al. Evidence for in vivo transport of bioactive nitric oxide in human plasma. J. Clin. Invest. 109, 1241–1248 (2002).

    Article  CAS  Google Scholar 

  12. Schechter, A.N., Gladwin, M.T. & Cannon, R.O., 3rd. NO solutions? J. Clin. Invest. 109, 1149–1151 (2002).

    Article  CAS  Google Scholar 

  13. Millar, T.M., Stevens, C.R. & Blake, D.R. Xanthine oxidase can generate nitric oxide from nitrate in ischaemia. Biochem. Soc. Trans. 25, 528S (1997).

    Article  CAS  Google Scholar 

  14. Millar, T.M. et al. Xanthine oxidoreductase catalyses the reduction of nitrates and nitrite to nitric oxide under hypoxic conditions. FEBS Lett. 427, 225–228 (1998).

    Article  CAS  Google Scholar 

  15. Godber, B.L. et al. Reduction of nitrite to nitric oxide catalyzed by xanthine oxidoreductase. J. Biol. Chem. 275, 7757–7763 (2000).

    Article  CAS  Google Scholar 

  16. Zhang, Z. et al. Generation of nitric oxide by a nitrite reductase activity of xanthine oxidase: a potential pathway for nitric oxide formation in the absence of nitric oxide synthase activity. Biochem. Biophys. Res. Commun. 249, 767–772 (1998).

    Article  CAS  Google Scholar 

  17. Li, H., Samouilov, A., Liu, X. & Zweier, J.L. Characterization of the magnitude and kinetics of xanthine oxidase-catalyzed nitrite reduction. Evaluation of its role in nitric oxide generation in anoxic tissues. J. Biol. Chem. 276, 24482–24489 (2001).

    Article  CAS  Google Scholar 

  18. Li, H., Samouilov, A., Liu, X. & Zweier, J.L. Characterization of the magnitude and kinetics of xanthine oxidase-catalyzed nitrate reduction: evaluation of its role in nitrite and nitric oxide generation in anoxic tissues. Biochemistry 42, 1150–1159 (2003).

    Article  CAS  Google Scholar 

  19. Zweier, J.L., Wang, P., Samouilov, A. & Kuppusamy, P. Enzyme-independent formation of nitric oxide in biological tissues. Nat. Med. 1, 804–809 (1995).

    Article  CAS  Google Scholar 

  20. Zweier, J.L., Samouilov, A. & Kuppusamy, P. Non-enzymatic nitric oxide synthesis in biological systems. Biochim. Biophys. Acta 1411, 250–262 (1999).

    Article  CAS  Google Scholar 

  21. Samouilov, A., Kuppusamy, P. & Zweier, J.L. Evaluation of the magnitude and rate of nitric oxide production from nitrite in biological systems. Arch. Biochem. Biophys. 357, 1–7 (1998).

    Article  CAS  Google Scholar 

  22. Modin, A. et al. Nitrite-derived nitric oxide: a possible mediator of 'acidic-metabolic' vasodilation. Acta Physiol. Scand. 171, 9–16 (2001).

    CAS  PubMed  Google Scholar 

  23. Demoncheaux, E.A. et al. Circulating nitrite anions are a directly acting vasodilator and are donors for nitric oxide. Clin. Sci. (Lond.) 102, 77–83 (2002).

    Article  CAS  Google Scholar 

  24. Agvald, P., Adding, L.C., Artlich, A., Persson, M.G. & Gustafsson, L.E. Mechanisms of nitric oxide generation from nitroglycerin and endogenous sources during hypoxia in vivo. Br. J. Pharmacol. 135, 373–382 (2002).

    Article  CAS  Google Scholar 

  25. Lauer, T. et al. Plasma nitrite rather than nitrate reflects regional endothelial nitric oxide synthase activity but lacks intrinsic vasodilator action. Proc. Natl. Acad. Sci. USA 98, 12814–12819 (2001).

    Article  CAS  Google Scholar 

  26. Cicinelli, E. et al. Different plasma levels of nitric oxide in arterial and venous blood. Clin. Physiol. 19, 440–442 (1999).

    Article  CAS  Google Scholar 

  27. Fox-Robichaud, A. et al. Inhaled NO as a viable antiadhesive therapy for ischemia/reperfusion injury of distal microvascular beds. J. Clin. Invest. 101, 2497–2505 (1998).

    Article  CAS  Google Scholar 

  28. McMahon, T.J. et al. Nitric oxide in the human respiratory cycle. Nat. Med. 3, 3 (2002).

    Article  Google Scholar 

  29. Cannon, R.O., 3rd et al. Effects of inhaled nitric oxide on regional blood flow are consistent with intravascular nitric oxide delivery. J. Clin. Invest. 108, 279–287 (2001).

    Article  CAS  Google Scholar 

  30. Gladwin, M.T. et al. S-nitrosohemoglobin is unstable in the reductive red cell environment and lacks O2/NO-linked allosteric function. J. Biol. Chem. 21, 21 (2002).

    Google Scholar 

  31. Gladwin, M.T., Lancaster, J.R., Freeman, B.A. & Schechter, A.N. Nitric oxide's reactions with hemoglobin: a view through the SNO-storm. Nat. Med. 9, 496–500 (2003).

    Article  CAS  Google Scholar 

  32. Schechter, A.N. & Gladwin, M.T. Hemoglobin and the paracrine and endocrine functions of nitric oxide. N. Engl. J. Med. 348, 1483–1485 (2003).

    Article  CAS  Google Scholar 

  33. Doyle, M.P., Pickering, R.A., DeWeert, T.M., Hoekstra, J.W. & Pater, D. Kinetics and mechanism of the oxidation of human deoxyhemoglobin by nitrites. J. Biol. Chem. 256, 12393–12398 (1981).

    CAS  PubMed  Google Scholar 

  34. Luchsinger, B.P. et al. Routes to S-nitroso-hemoglobin formation with heme redox and preferential reactivity in the beta subunits. Proc. Natl. Acad. Sci. USA 100, 461–466 (2003).

    Article  CAS  Google Scholar 

  35. Xu, X. et al. Measurements of nitric oxide on the heme iron and β-93 thiol of human hemoglobin during cycles of oxygenation and deoxygenation. Proc. Natl. Acad. Sci. USA 100, 11303–11308 (2003).

    Article  CAS  Google Scholar 

  36. Fernandez, B.O., Lorkovic, I.M. & Ford, P.C. Nitrite catalyzes reductive nitrosylation of the water-soluble ferri-heme model Fe(III)(TPPS) to Fe(II)(TPPS)(NO). Inorg. Chem. 42, 2–4 (2003).

    Article  CAS  Google Scholar 

  37. Watanabe, S. & Ogata, M. Generation of superoxide and hydrogen peroxide during interaction of nitrite with human hemoglobin. Acta Med. Okayama 35, 173–178 (1981).

    CAS  PubMed  Google Scholar 

  38. Kosaka, H., Imaizumi, K. & Tyuma, I. Mechanism of autocatalytic oxidation of oxyhemoglobin by nitrite. An intermediate detected by electron spin resonance. Biochim. Biophys. Acta 702, 237–241 (1982).

    Article  CAS  Google Scholar 

  39. Kosaka, H. & Tyuma, I. Mechanism of autocatalytic oxidation of oxyhemoglobin by nitrite. Environ. Health Perspect. 73, 147–151 (1987).

    Article  CAS  Google Scholar 

  40. Jia, L., Bonaventura, C., Bonaventura, J. & Stamler, J.S. S-nitrosohaemoglobin: a dynamic activity of blood involved in vascular control. Nature 380, 221–226 (1996).

    Article  CAS  Google Scholar 

  41. Stamler, J.S. et al. Blood flow regulation by S-nitrosohemoglobin in the physiological oxygen gradient. Science 276, 2034–2037 (1997).

    Article  CAS  Google Scholar 

  42. Huang, K.T. et al. Modulation of nitric oxide bioavailability by erythrocytes. Proc. Natl. Acad. Sci. USA 98, 11771–11776 (2001).

    Article  CAS  Google Scholar 

  43. Reiter, C.D. et al. Cell-free hemoglobin limits nitric oxide bioavailability in sickle-cell disease. Nat. Med. 8, 1383–1389 (2002).

    Article  CAS  Google Scholar 

  44. Panza, J.A., Casino, P.R., Kilcoyne, C.M. & Quyyumi, A.A. Role of endothelium-derived nitric oxide in the abnormal endothelium- dependent vascular relaxation of patients with essential hypertension. Circulation 87, 1468–1474 (1993).

    Article  CAS  Google Scholar 

  45. Yang, B.K., Vivas, E.X., Reiter, C.D. & Gladwin, M.T. Methodologies for the sensitive and specific measurement of S-nitrosothiols, iron-nitrosyls, and nitrite in biological samples. Free Radic. Res. 37, 1–10 (2003).

    Article  CAS  Google Scholar 

  46. Crawford, J.H., White, C.R. & Patel, R.P. Vasoactivity of S-nitrosohemoglobin: role of oxygen, heme, and NO oxidation states. Blood 101, 4408–4415 (2003).

    Article  CAS  Google Scholar 

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Acknowledgements

This work was supported by Clinical Center and National Heart, Lung and Blood Institute intramural funds (R.O.C. and M.T.G.), National Institutes of Health grant HL58091 (D.B.K.-S.), RO1HL70146 (R.P.P.) and Medical Scientist Training Program T32GM08361. We thank V. Annavajjhala for helpful laboratory assistance.

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Correspondence to Richard O Cannon III or Mark T Gladwin.

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Cosby, K., Partovi, K., Crawford, J. et al. Nitrite reduction to nitric oxide by deoxyhemoglobin vasodilates the human circulation. Nat Med 9, 1498–1505 (2003). https://doi.org/10.1038/nm954

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