Role of Hemoglobin Denaturation and Band 3 Clustering in Initiating Red Cell Removal

  • Philip S. Low
Part of the Advances in Experimental Medicine and Biology book series (AEMB, volume 307)


While multiple mechanisms likely exist to assure that a defective erythrocyte does not escape removal by macrophages, we believe that the more heavily used clearance pathways will have certain characteristics in common. First, the pathway should involve a change in components already present in the circulating erythrocytes, since de novo protein synthesis will have terminated before the erythrocyte reaches maturity. Second, the changes initiating the removal sequence must eventually be manifested on the exofacial surface of the cell, since a macrophage has little means of detecting an intracellular biochemical lesion. And finally, the exofacial changes recognized by the macrophage must be inducible by a change in the biochemistry of the cytoplasm, since cells that develop intracellular defects early in their lifespans are also removed early (e.g., sickle cells, (1) ß-thalassemic cells, (2) cells with enzyme deficiencies, (3) cells treated with oxidants, (4) etc). That is, a linkage of some sort must exist between the functional state of components in the cytoplasm and markers at the cell surface recognized by macrophages. The hypothesis outlined below describes how hemoglobin, the most abundant protein in the cytoplasm, and band 3, the most prominent protein in the membrane cooperate to establish this linkage, transducing information regarding the biochemical integrity of the cell to the reticuloendothelial system which is responsible for aged/abnormal cell clearance.


Sickle Cell Human Erythrocyte Complement Fixation Cluster Agent Globin Chain 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.


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  1. 1.
    P. R. McCurdy and A. S. Sherman, Irreversibly sickled cells and red cell survival in sickle cell anemia, Amer. J. Med. 64:253 (1978).PubMedCrossRefGoogle Scholar
  2. 2.
    E. A. Rachmilewitz, E. Shinar, O. Shalev, U. Galili and S. L. Schrier, Erythrocyte membrane alterations in ß-thalassemia, Clinics in Haematology 14:163 (1985).PubMedGoogle Scholar
  3. 3.
    J. J. Kaneko, Comparative erythrocyte metabolism, Adv. Vet. Sci. Comp. Med. 18:117 (1974).PubMedGoogle Scholar
  4. 4.
    D. A. Bates and C. C. Winterbourn, Hemoglobin denaturation, lipid peroxidation and hemolysis in phenylhydrazine-induced anemia, Biochim. Biophys. Acta 798:84 (1984).PubMedCrossRefGoogle Scholar
  5. 5.
    S. Horn, J. Copas and N. Bashan, A lectin-like receptor on murine macrophage is involved in the recognition and phagocytosis of human red cells oxidized by Phenylhydrazine, Biochem. Pharm. 39:775 (1990).PubMedCrossRefGoogle Scholar
  6. 6.
    M. Beppu, M. Ochiai and K. Kikugawa, Macrophage recognition of periodate-treated erythrocytes: involvement of disulfide formation of the erythrocyte membrane proteins, Biochim. Biophys. Acta 979:35 (1989).PubMedCrossRefGoogle Scholar
  7. 7.
    P. S. Low, Interaction of native and denatured hemoglobins with band 3: consequences for erythrocytes structure and function, in “Red Blood Cell Membranes” P. Agre and J. C. Parker, ed., Marcel Dekker, Inc. NY, p. 237 (1989).Google Scholar
  8. 8.
    P. S. Low and R. Kannan, Effect of hemoglobin denaturation on membrane strucutre and IgG binding: role in red cell aging, in “The Red Cell: Seventh Ann Arbor Conference”, Alan R. Liss, Inc. p. 525 (1989).Google Scholar
  9. 9.
    S. M. Waugh, B. M. Willardson, R. Kannan, R. J. Labotka and P. S. Low, Heinz bodies induce clustering of band 3, glycophorin, and ankyrin in sickle cell erythrocytes, J. Clin. Invest. 78:1155 (1986).PubMedCrossRefGoogle Scholar
  10. 10.
    P. S. Low, S. M. Waugh, K. Zinke and D. Drenckhahn, The role of hemoglobin denaturation and band 3 clustering in red blood cell aging, Science 227:531 (1985).PubMedCrossRefGoogle Scholar
  11. 11.
    H. Muller and H. U. Lutz, Binding of autologous IgG to human red blood cells before and after ATP-depletion, Biochim. Biophys. Acta 729:249 (1983).PubMedCrossRefGoogle Scholar
  12. 12.
    P. Hochstein and S. K. Jain, Association of lipid peroxidation and polymerization of membrane proteins with erythrocyte aging, FASEB 40:183 (1981).Google Scholar
  13. 13.
    A. Elgsaeter, D. M. Shotton and D. Branton, Intermembrane particle aggregation in erythrocyte ghosts, Biochim. Biophys. Acta 426:101 (1976).PubMedCrossRefGoogle Scholar
  14. 14.
    F. Turrini, A. Naitana, L. Mannuzzu, G. P. Pescarmona and P. Arese, Increased red cell calcium, decreased calcium adenosine triphosphatase, and altered membrane proteins during Fava bean hemolysis in glucose-6-phosphate dehydrogenase-deficient (Mediterranean variant) individuals, Blood 66:302 (1985).PubMedGoogle Scholar
  15. 15.
    M. R. Clark, Senescence of red blood cells: progress and problems, Physiol. Rev. 68:503 (1988).PubMedGoogle Scholar
  16. 16.
    J. R. Barber and S. Clarke, Membrane protein carboxyl methylation increase with human erythrocyte age, J. Biol. Chem. 258:1189 (1983).PubMedGoogle Scholar
  17. 17.
    K. Yamamoto, M. Yamada and Y. Kato, Age-related and phenylhydrazine-induced activation of the membrane-associated cathepsin E in human erythrocytes, J. Biochem. 105:114 (1989).PubMedGoogle Scholar
  18. 18.
    M. M. B. Kay, K. Sorensen, P. Wong and P. Bolton, Antigenicity, storage and aging: physiologic autoantibodies to cell membrane and serum proteins and the senescent cell antigen, Molec. and Cell. Biochem. 49:65 (1982).Google Scholar
  19. 19.
    H. Vlassara, J. Valinsky, M. Brownlee, C. Cerami, S. Nishimoto and A. Cerami, Advanced glycolysation endproducts on erythrocyte cell surface induce receptor-mediated phagocytosis by macrophages. A model for turnover of aging cells, J. Exp. Med. 166:539 (1987).PubMedCrossRefGoogle Scholar
  20. 20.
    K. Miyahara and M. J. Spiro, Nonuniform loss of membrane glycoconjugates during in vivo aging of human erythrocytes: studies of normal and diabetic red cell saccharides, Arch. Biochem. Biophys. 232:310 (1984).PubMedCrossRefGoogle Scholar
  21. 21.
    M. A. Zago, D. T. Covas, M. S. Figueiredo, C. Bottura, Red cell pits appear preferentially in old cells after splenectomy, Acta Haemat. 76:54 (1986).PubMedCrossRefGoogle Scholar
  22. 22.
    A. Fazi, E. Piatti, A. Accorsi and M. Magnani, Cell age dependent decay of human erythrocytes glucose-6-phosphate isomerase, Biochim. Biophys. Acta 998:286 (1989).PubMedCrossRefGoogle Scholar
  23. 23.
    C. Seaman, S. Wyss and S. Piomelli, The decline in energetic metabolism with aging of the erythrocyte and its relationship to cell death, Amer. J. Hemat. 8:31 (1980).CrossRefGoogle Scholar
  24. 24.
    H. Q. Campwala and J. F. Desforges, Membrane-bound hemichrome in density-separated cohorts of normal (AA) and sickled (SS) cells, J. Lab. Clin. Med. 99:25 (1982).PubMedGoogle Scholar
  25. 25.
    D. A. Sears, J. M. Friedman and D. R. White, Binding of intracellular protein to the erythrocyte membrane during incubation: the production of Heinz bodies, J. Lab. Clin. Med. 86:722 (1975).PubMedGoogle Scholar
  26. 26.
    J. G. Selwyn, Heinz bodies in red cells after splenectomy and phenacetin administration, Brit. J. Haematol. 4:173 (1955).CrossRefGoogle Scholar
  27. 27.
    T. J. Mueller, C. W. Jackson, M. E. Dockter and M. Morrison, Membrane skeletal alterations during in vivo mouse red cell aging: increase in the band 4.1a:4.1b ratio, J. Clin. Invest. 79:492 (1987).PubMedCrossRefGoogle Scholar
  28. 28.
    T. Suzuki and G. Dale, membrane proteins in senescent erythrocytes, Biochem. J. 257:37 (1989).PubMedGoogle Scholar
  29. 29.
    S. K. Jain, Evidence for membrane lipid peroxidation during the in vivo aging of human erythrocytes, Biochim. Biophys. Acta 937:205 (1988).PubMedCrossRefGoogle Scholar
  30. 30.
    A. Brovelli, C. Seppi and C. Balduini, Modification of membrane protein organization during in vitro aging of human erythrocytes, Int. J. Biochem. 16:1115 (1984).PubMedCrossRefGoogle Scholar
  31. 31.
    S. P. Sutera, R. A. Gardner, C. W. Boylan, G. L. Carroll, K. C. Chang, J. S. Marvel, C. Kilo, B. Gonen and J. R. Williamson, Age-related changes in deformability of human erythrocytes, Blood 65:275 (1985).PubMedGoogle Scholar
  32. 32.
    J. M. Rifkind, K. Araki and E. C. Hadley, The relationship between the osmotic fragility of human erythrocytes and cell age, Arch. Biochem. Biophys. 222:582 (1983).PubMedCrossRefGoogle Scholar
  33. 33.
    D. Aminoff, M. A. Ghalambor and C.J. Henrich, GOST, galactose oxidase and sialyl transferase, substrate and receptor sites in erythrocyte senescence, in “Erythrocyte Membranes 2. Recent Clinical and Experimental Advances”, W. C. Kruckerberg, J. W. Eaton and G. J. Brewer, ed., Liss: New York, p. 269 (1981).Google Scholar
  34. 34.
    U. Galili, I. Flechner, A. Knyszynski, D. Danon and E.A. Rachmilewitz, The natural anti-±-galactosyl IgG on human normal senescent red blood cells, Br. J. Haematol. 62:317 (1986).PubMedCrossRefGoogle Scholar
  35. 35.
    T. Shiga, M. Sekiya, N. Maeda, K. Kon and M. Okazaki, Cell age-dependent changes in deformability and calcium accumulation of human erythrocytes, Biochim. Biophys. Acta 814:289 (1985).PubMedCrossRefGoogle Scholar
  36. 36.
    S. M. Waugh and P. S. Low, Hemichrome binding to band 3: nucleation of Heinz bodies on the erythrocyte membrane, Biochemistry 24:34 (1985).PubMedCrossRefGoogle Scholar
  37. 37.
    S. M. Waugh, J. A. Walder and P. S. Low, Partial characterization of the copolymerization reaction of erythrocyte membrane band 3 with hemichromes, Biochemistry 26:1777 (1987).PubMedCrossRefGoogle Scholar
  38. 38.
    P. P. DaSilva, Translational mobility of the membrane intercalated particles of human erythrocyte ghosts, J. Cell. Biol. 53:777 (1972).CrossRefGoogle Scholar
  39. 39.
    K. Schlüter and D. Drenckhahn, Co-clustering of denatured hemoglobin with band 3: its role in binding of autoantibodies against band 3 to abnormal and aged erythrocytes, Proc. Natl. Acad. Sci.USA 83:6137 (1986).PubMedCrossRefGoogle Scholar
  40. 40.
    F. Turrini, P. Arese, J. Yuan and P. S. Low, Clustering of integral membrane proteins of the human erythrocyte membrane stimulates autologous IgG binding, complement deposition and phagocytosis, submitted for publication (1991).Google Scholar
  41. 41.
    R. Kannan, R. Labotka and P. S. Low, Isolation and characterization of the hemichrome-stabilized membrane protein aggregates from sickle erythrocytes, J. biol. Chem. 263:13766 (1988).PubMedGoogle Scholar
  42. 42.
    R. Kannan, Mechanism of aging of human red cells, Ph. D. Dissertation, Purdue University, 71 (1990).Google Scholar
  43. 43.
    R. Kannan, J. Yuan and P. S. Low, Isolation and characterization of antibody-enriched complexes from membranes of density fractionated human erythrocytes, manuscript submitted (1991).Google Scholar
  44. 44.
    M. Morrison, C. W. Jackson, T. J. Mueller, T. Huang, M. E. Dockter, W. S. Walker, J. A. Singer and H. H. Edwards, Does cell density correlate with red cell age?, Biomed. Biochim. Acta 42:S107 (1983).PubMedGoogle Scholar
  45. 45.
    M. J. Clague and R. J. Cherry, A comparative study of band 3 aggregation in erythrocyte membranes by melittin and other cationic agents, Biochim. Biophys. Acta 980:93 (1989).PubMedCrossRefGoogle Scholar
  46. 46.
    S. W. Hui, C. M. Stewart and R.J. Cherry, Electron microscopic observation of the aggregation of membrane proteins in human erythrocyte by mellitin, Biochim. Biophys. Acta 1023:335 (1990).PubMedCrossRefGoogle Scholar
  47. 47.
    G. Lelkes, G. Lelkes, K. S. Merse and S. R. Hollan, Intense, reversible aggregation of intramembrane particles in non-haemolyzed human erythrocytes, Biochim. Biophys. Acta 732:48 (1983).PubMedCrossRefGoogle Scholar
  48. 48.
    M. TenBrinke and J. DeReget, Cr-half time of heavy and light human erythrocytes, Scand. J. Haematol. 7:336 (1970).CrossRefGoogle Scholar
  49. 49.
    H. U. Lutz, A naturally occurring autoantibody to band 3 protein of human red blood cells and its possible role in removal of senescent red cells, in “Red Cell Membrane Glycoconjugates and Related Genetic Markers”, J-P Cartron, P. Rouger and C. Salmon, eds., p. 273 (1983).Google Scholar
  50. 50.
    M. Beppu, A. Mizukami, M. Nagoya and K. Kikugawa, Binding of anti-band 3 autoantibody to oxidatively damaged erythrocytes, J. Biol. Chem. 265:3226 (1990).PubMedGoogle Scholar
  51. 51.
    M. M. B. Kay, Localization of senescent cell antigen on band 3, Proc. Natl. Acad. Sci. USA 81:5753 (1984).PubMedCrossRefGoogle Scholar
  52. 52.
    A. G. Ehlenberger and V. Nussenzweig, The role of membrane receptors for C3b and C3d in phagocytosis, J. Exp. Med. 145:357 (1977).PubMedCrossRefGoogle Scholar
  53. 53.
    H. U. Lutz, F. Bussolino, R. Flepp, S. Fasler, P. Stammler, M. D. Kazatchkine and P. Arese, Naturally occurring anti-band 3 antibodies and complement together mediate phagocytosis of oxidatively stressed human erythrocytes, Proc. Natl. Acad. Sci. USA 84:7368 (1987).PubMedCrossRefGoogle Scholar
  54. 54.
    N. Yousaf, J. C. Howard and B. D. Williams, Studies in the rat of antibody-sensitized and N-ethylmaleimide-treated erythrocyte clearance by the liver: effects of immune complex infusion and complement activation, Immunolgy 64:193 (1988).Google Scholar
  55. 55.
    A. Hermanowski-Vosatka, P. A. Detmers, O. Götze, S. C. Silverstein and S. D. Wright, Clustering of ligand on the surface of a particle enhances adhesion to receptor-bearing cells, J. Biol. Chem. 263:17822 (1988).PubMedGoogle Scholar
  56. 56.
    M. M. B. Kay, Role of physiologic autoantibody in the removal of senescent human red cells, J. Supramol. Struct. 9:555 (1978).PubMedCrossRefGoogle Scholar
  57. 57.
    G. M. Shaw, D. Aminoff, S. P. Balcerzak and A. F. LoBuglio, Clustered IgG on human red blood cell membranes may promote human lymphocyte antibody-dependent cell-mediated cytotoxicity, J. Immunol. 125:501 (1980).PubMedGoogle Scholar
  58. 58.
    R. A. Rifkind, Heinz body anemia: an ultrastructural study. II. Red cell sequestration and destruction, Blood 26:433 (1965).PubMedGoogle Scholar

Copyright information

© Plenum Press, New York 1991

Authors and Affiliations

  • Philip S. Low
    • 1
  1. 1.Department of ChemistryPurdue UniversityWest LafayetteUSA

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