Summary
Ultrastructures of membrane skeletons in spherocytic and elliptocytic erythrocytes were investigated immunocytochemically. Erythrocytes obtained from patients with hereditary spherocytosis (HS) and hereditary elliptocytosis (HE) were split open mechanically to obtain exposed cytoplasmic sides of erythrocyte membranes and were immunostained with anti-spectrin antibody. Replica membranes were prepared by a quick-freezing and deep-etching method and were checked by electron microscopy. The in situ membrane skeletons of normal erythrocytes consisted mainly of reticular patterns of spectrin filaments, which formed networks on the cytoplasmic sides of the cell membrane. In contrast, the membrane skeletons of abnormally shaped erythrocytes (HS and HE) were much less filamentous and more granular than those of normal erythrocytes. This abnormal organization in erythrocyte membrane skeletons may be one of the factors that induce abnormally shaped erythrocytes in HS and HE patients.
Similar content being viewed by others
References
Agre P, Casella JF, Zinkham WH, McMillan C, Bennett V (1985) Partial deficiency of erythrocyte spectrin in hereditary spherocytosis. Nature 314:380–383
Agre P, Asimos A, Casella JF, McMillan C (1986) Inheritance pattern and clinical response to splenectomy as a reflection of erythrocyte spectrin deficiency in hereditary spherocytosis. N Engl J Med 315:1579–1583
Bennett V (1985) The membrane skeleton of human erythrocytes and its implications for more complex cells. Annu Rev Biochem 54:273–304
Bennett V (1989) The spectrin-actin junction of erythrocyte membrane skeletons. Biochem Biophys Acta 988:107–121
Burke BE, Shotton DM (1983) Erythrocyte membrane skeleton abnormalities in hereditary spherocytosis. Br J Haematol 54:173–187
Byers TJ, Branton D (1985) Visualization of the protein associations in the erythrocyte membrane skeleton. Proc Natl Acad Sci USA 82:6153–6157
Coetzer T, Zail S (1982) Spectrin tetramer-dimer equilibrium in hereditary elliptocytosis. Blood 59:900–905
Delaunay J, Alloisio N, Morlé L, Pothier B (1990) The red cell skeleton and its genetic disorders. Mol Aspects Med 11:161–241
Fowler VM (1986) New views of the red cell network. Nature 322:777–778
Fujikawa S (1983) Tannic acid improves the visualization of the human erythrocyte membrane skeleton by freeze-etching. J Ultrastruct Res 64:289–298
Goodman SR, Shiffer K (1983) The spectrin membrane skeleton of normal and abnormal human erythrocytes: a review. Am J Physiol 244:C121–141
Goodman SR, Shiffer KA, Casoria LA, Eyster E (1982) Identification of the molecular defect in the erythrocyte membrane skeleton of some kindreds with hereditary spherocytosis. Blood 60:772–784
Hainsfeld JF, Steck TL (1977) The sub-membrane reticulum of the human erythrocyte: a scanning electron microscope study. J Supramol Struct 6:301–317
Iolascon A, Miraglia del Giudice E, Camaschella C (1992) Molecular pathology of inherited erythrocyte membrane disorders: hereditary spherocytosis and elliptocytosis. Haematologica (Pavia) 77:60–72
Liu SC, Palek J, Prchal JT (1982) Defective spectrin dimer-dimer association in hereditary elliptocytosis. Proc Natl Acad Sci USA 79:2072–2076
Liu SC, Derick LH, Palek J (1987) Visualization of the hexagonal lattice in the erythrocyte membrane skeleton. J Cell Biol 104:527–536
Liu SC, Derick LH, Agre P, Palek J (1990) Alteration of the erythrocyte membrane skeletal ultrastructure in hereditary spherocytosis, hereditary elliptocytosis, and pyropoikilocytosis. Blood 76:198–205
Lupu F, Constantinescu E (1989) A new freeze-drying device for platinum replica studies of cell surface and cytoskeleton: an example using immunogold-labeled human erythrocytes. J Electron Microsc Tech 11:76–82
Lux SE, Tse WT, Menninger JC, John KM, Harris P, Shalev O, Chilcote RR, Marchesi SL, Watkins PC, Bennett V, McIntosh S, Collins FS, Francke U, Ward DC, Forget BG (1990) Hereditary spherocytosis associated with deletion of human erythrocyte ankyrin gene on chromosome 8. Nature 345:736–739
Marchesi VT (1983) The red cell membrane skeleton: recent progress. Blood 61:1–11
McGough Am, Josephs R (1990) On the structure of erythrocyte spectrin in partially expanded membrane skeletons. Proc Natl Acad Sci USA 87:5208–5212
McGuire M, Agre P (1988) Clinical disorders of the erythrocyte membrane skeleton. Haematol Pathol 2:1–14
McGuire M, Smith BL, Agre P (1988) Distinct variants of erythrocyte protein 4.1 inherited in linkage with elliptocytosis and Rh type in three white families. Blood 72:287–293
Nermut MV (1981) Visualization of the “membrane skeleton” in human erythrocytes by freeze-etching. Eur J Cell Biol 25:265–271
Ohno S (1985) Immunocytochemical study on the cytoplasmic side of cell membrane infected with vesicular stomatitis virus by quick-freezing and deep-etching replica method. Histochemistry 82:565–575
Ohno S (1992) An ultrastructural study of the cytoplasmic aspects of erythrocyte membranes by a quick-freezing and deep-etching method. J Anat 180:315–320
Ohno S, Fujii Y (1990) Three-dimensional and histochemical studies of peroxisomes in cultured hepatocytes by quick-freezing and deep-etching method. Histochem J 22:143–154
Ohno S, Fujii Y (1991) Three-dimensional studies of the cytoskeleton of cultured hepatocytes: a quick-freezing and deep-etching study. Virchows Arch [A] 418:61–70
Ohno S, Takasu N (1989) Three-dimensional studies of cytoskeletal organizations in cultured thyroid cells by quick-freezing and deep-etching method. J Electron Microsc 38:352–362
Shen BW, Josephs R, Steck TL (1984) Ultrastructure of unit fragments of the skeleton of the human erythrocyte membrane. J Cell Biol 99:810–821
Shen BW, Josephs R, Steck TL (1986) Ultrastructure of the intact skeleton of the human erythrocyte membrane. J Cell Biol 102:997–1006
Tchernia G, Mohandas N, Shohet SB (1981) Deficiency of skeletal membrane protein band 4.1 in homozygous hereditary elliptocytosis. J Clin Invest 68:454–460
Tsukita S, Tsukita S, Ishikawa H (1980) Cytoskeletal network underlying the human erythrocyte membrane. J Cell Biol 85:567–576
Ursitti JA, Pumplin DW, Wade JB, Bloch RJ (1991) Ultrastructure of the human erythrocyte cytoskeleton and its attachment to the membrane. Cell Motil Cytoskeleton 19:227–243
Author information
Authors and Affiliations
Rights and permissions
About this article
Cite this article
Ohno, S., Terada, N., Fujii, Y. et al. Immunocytochemical study of membrane skeletons in abnormally shaped erythrocytes as revealed by a quick-freezing and deep-etching method. Vichows Archiv A Pathol Anat 422, 73–80 (1993). https://doi.org/10.1007/BF01605136
Received:
Accepted:
Issue Date:
DOI: https://doi.org/10.1007/BF01605136