Skip to main content
NCBI home page
The Journal of Clinical Investigation logoLink to The Journal of Clinical Investigation
. 1976 Oct;58(4):797–802. doi: 10.1172/JCI108531

Mechanisms of methylene blue stimulation of the hexose monophosphate shunt in erythrocytes.

E N Metz, P Balcerzak, A L Sagone Jr
PMCID: PMC333241  PMID: 965487

Abstract

The response of the hexose monophosphate shunt in erythrocytes was studied with the ionization chamber-electrometer apparatus to measure continuously 14CO2 derived from 14C-labeled substrates. The effect of methylene blue at high (0.1 mM) and low (1 muM) concentrations was evaluated under different gas mixtures; air, carbon monoxide, and 6% carbon monoxide in air. The latter gas mixture results in nearly 100% carboxyhemoglobin but provides a physiologic partial pressure of oxygen. The extent to which pentose is recycled through the shunt in response to methylene blue stimulation was examined with radioactive glucose substrates labeled on the first, second, and third carbon positions. Generation of hydrogen peroxide after stimulation of erythrocytes with methylene blue was evaluated by the catalase-aminotriazole trapping technique, [14C]formate oxidation, and oxidation of reduced glutatione. Stimulation of the shunt with 1 muM methylene blue was markedly impaired in the absence of oxyhemoglobin, but stimulation with 0.1 mM methylene blue was only slightly impaired under the carbon monoxide-air mixture. The higher concentration of methylene blue produced evidence of hydrogen peroxide generation of all three techniques. Despite the evidence for the involvement of oxygen, oxyhemoglobin, and hydrogen peroxide in the response to methylene blue, cells containing methemoglobin induced by sodium nitrite or from a patient with congenital methemoglobinemia responded normally to methylene blue in the absence of oxygen. These experiments indicate that the reactions induced by methylene blue in erythrocytes are more complex than generally thought and that high concentrations are associated with production of peroxide.

Full text

PDF
797

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. BEUTLER E., DURON O., KELLY B. M. Improved method for the determination of blood glutathione. J Lab Clin Med. 1963 May;61:882–888. [PubMed] [Google Scholar]
  2. BRIN M., YONEMOTO R. H. Stimulation of the glucose oxidative pathway in human erythrocytes by methylene blue. J Biol Chem. 1958 Jan;230(1):307–317. [PubMed] [Google Scholar]
  3. COHEN G., HOCHSTEIN P. GENERATION OF HYDROGEN PEROXIDE IN ERYTHROCYTES BY HEMOLYTIC AGENTS. Biochemistry. 1964 Jul;3:895–900. doi: 10.1021/bi00895a006. [DOI] [PubMed] [Google Scholar]
  4. COHEN G., HOCHSTEIN P. Glucose-6-phosphate dehydrogenase and detoxification of hydrogen peroxide in human erythrocytes. Science. 1961 Dec 1;134(3492):1756–1757. doi: 10.1126/science.134.3492.1756. [DOI] [PubMed] [Google Scholar]
  5. Chaudhry A. A., Sagone A. L., Jr, Metz E. N., Balcerzak S. P. Relationship of glucose oxidation to aggregation of human platelets. Blood. 1973 Feb;41(2):249–258. [PubMed] [Google Scholar]
  6. Davidson W. D., Tanaka K. R. Continuous measurement of pentose phosphate pathway activity in erythrocytes. An ionization chamber method. J Lab Clin Med. 1969 Jan;73(1):173–180. [PubMed] [Google Scholar]
  7. Davidson W. D., Tanaka K. R. Factors affecting pentose phosphate pathway activity in human red cells. Br J Haematol. 1972 Sep;23(3):371–385. doi: 10.1111/j.1365-2141.1972.tb08884.x. [DOI] [PubMed] [Google Scholar]
  8. GOLUBOFF N., WHEATON R. Methylene blue induced cyanosis and acute hemolytic anemia complicating the treatment of methemoglobinemia. J Pediatr. 1961 Jan;58:86–89. doi: 10.1016/s0022-3476(61)80064-4. [DOI] [PubMed] [Google Scholar]
  9. Jacob H. S., Jandl J. H. Effects of sulfhydryl inhibition on red blood cells. 3. Glutathione in the regulation of the hexose monophosphate pathway. J Biol Chem. 1966 Sep 25;241(18):4243–4250. [PubMed] [Google Scholar]
  10. Klebanoff S. J., Pincus S. H. Hydrogen peroxide utilization in myeloperoxidase-deficient leukocytes: a possible microbicidal control mechanism. J Clin Invest. 1971 Oct;50(10):2226–2229. doi: 10.1172/JCI106718. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. MURPHY J. R. Erythrocyte metabolism. II. Glucose metabolism and pathways. J Lab Clin Med. 1960 Feb;55:286–302. [PubMed] [Google Scholar]
  12. Roth E. F., Jr, Nagel R. L., Neuman G., Vanderhoff G., Kaplan B. H., Jaffé E. R. Metabolic effects of antisickling amounts of nitrogen and nor-nitrogen mustard on rabbit and human erythrocytes. Blood. 1975 Jun;45(6):779–788. [PubMed] [Google Scholar]
  13. SZEINBERG A., MARKS P. A. Substances stimulating glucose catabolism by the oxidative reactions of the pentose phosphate pathway in human erythrocytes. J Clin Invest. 1961 Jun;40:914–924. doi: 10.1172/JCI104330. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Sagone A. L., Jr, Balcerzak S. P., Metz E. N. The response of red cell hexose monophosphate shunt after sulfhydryl inhibition. Blood. 1975 Jan;45(1):49–54. [PubMed] [Google Scholar]
  15. Sagone A. L., Jr, Metz E. N., Balcerzak S. P. Effect of inorganic phosphate on erythrocyte pentose phosphate pathway activity. Biochim Biophys Acta. 1972 Jan 28;261(1):1–8. doi: 10.1016/0304-4165(72)90306-6. [DOI] [PubMed] [Google Scholar]
  16. Smith R. P., Thron C. D. Hemoglobin, methylene blue and oxygen interactions in human red cells. J Pharmacol Exp Ther. 1972 Dec;183(3):549–558. [PubMed] [Google Scholar]
  17. Spiro T. G., Strekas T. C. Resonance Raman spectra of heme proteins. Effects of oxidation and spin state. J Am Chem Soc. 1974 Jan 23;96(2):338–345. doi: 10.1021/ja00809a004. [DOI] [PubMed] [Google Scholar]
  18. TARLOV A. R., KELLERMEYER R. W. The hemolytic effect of primaquine. XI. Decreased catalase activity in primaquine-sensitive erythrocytes. J Lab Clin Med. 1961 Aug;58:204–216. [PubMed] [Google Scholar]
  19. TEPHLY T. R., ATKINS M., MANNERING G. J., PARKS R. E., Jr ACTIVATION OF A CATALASE PEROXIDATIVE PATHWAY FOR THE OXIDATION OF ALCOHOLS IN MAMMALIAN ERYTHROCYTES. Biochem Pharmacol. 1965 Apr;14:435–444. doi: 10.1016/0006-2952(65)90216-9. [DOI] [PubMed] [Google Scholar]

Articles from Journal of Clinical Investigation are provided here courtesy of American Society for Clinical Investigation

RESOURCES