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  • 1
    Publication Date: 1984-08-01
    Description: ntracellular calcium (Ca) concentration in erythrocytes (RBCs) is controlled by a low passive influx through a relatively impermeable membrane and by active efflux catalyzed by Ca2+,Mg2+-ATPase. Since precipitation of alpha-globin chains in thalassemic RBCs may interfere with normal membrane function, we studied the RBC intracellular Ca content and the RBC membrane Ca2+,Mg2+-ATPase activity in two groups of patients with nonsplenectomized (n = 9) and splenectomized (n = 9) beta- thalassemia intermedia and in two groups of matched controls. The mean +/- SD Ca concentration in the nonsplenectomized (n = 12) and splenectomized (n = 6) controls were 6.1 +/- 6.0 and 5.8 +/- 3.4 mumol Ca per liter of RBCs, respectively, compared with 26.0 +/- 7.6 (P less than .001) and 85 +/- 24.4 (P less than .001) in the nonsplenectomized and splenectomized thalassemia patients, respectively. The mean +/- SD Ca2+,Mg2+-ATPase activity in the eight nonsplenectomized patients was 0.77 +/- 0.58 mumol inorganic phosphate (Pi) per milligram of protein per hour compared with 0.66 +/- 0.41 in the controls (P = NS). Similar values were obtained for the splenectomized patients and their controls. No correlation was found between either the intracellular Ca content or the Ca2+,Mg2+-ATPase activity with the peripheral nucleated RBC count. These findings suggest that there is a major defect in the membrane of the thalassemic RBC leading to an increased Ca content that is more pronounced in splenectomized patients.
    Print ISSN: 0006-4971
    Electronic ISSN: 1528-0020
    Topics: Biology , Medicine
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  • 2
    Publication Date: 1986-11-01
    Description: When amphipathic cationic drugs are added to intact human RBCs, the RBCs first undergo a stomatocytic shape change and then, if relatively large amounts of drug are added and if the metabolic state of the RBC is appropriate, endocytic vacuoles form. Vanadate has a structural similarity to the transition state of phosphate, which presumably accounts for its ability to inhibit phosphohydrolases, although other actions of vanadate have been described. Vanadate inhibited three forms of drug-induced endocytosis in intact RBCs despite the fact that the three drugs chosen (primaquine, chlorpromazine, and vinblastine) are known to have differing requirements for RBC ATP. Vanadate also inhibited the stomatocytic shape change produced by primaquine, chlorpromazine, and vinblastine, but not the stomatocytosis produced by low pH. Vanadate had no effect on RBC echinocytosis produced by lysophosphatidylcholine. In studying endocytosis in hypotonic, leaky, “white” ghosts, we discovered that vanadate inhibited only the endocytosis produced by Mg-ATP and not the endocytosis produced by manipulations that directly attack the cytoskeletal proteins. These findings suggest that ATP hydrolysis has a role in some forms of amphipathic cation-induced stomatocytosis and endocytosis in intact RBCs. In addition, studies in ghosts support the idea that Mg-ATP does indeed produce “energized” endocytosis dependent on utilization or hydrolysis of ATP.
    Print ISSN: 0006-4971
    Electronic ISSN: 1528-0020
    Topics: Biology , Medicine
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  • 3
    Publication Date: 1996-03-01
    Description: The life threatening anemia in beta-thalassemia major (Cooley's anemia) is characterized by profound intramedullary lysis, the cause of which is incompletely understood. Using marrow obtained from beta thalassemia major patients undergoing allogeneic bone marrow transplantation in Pesaro Italy, it became possible to directly study the mechanism of the intramedullary hemolysis. Based on our previous studies, we hypothesized that the unmatched alpha globin chains would interfere with normal assembly of erythroid precursor membrane proteins. Patient and control erythroid precursors were reacted with monospecific polyclonal rabbit antibodies directed against spectrin, band 3, and band 4.1 and with a monoclonal anti-alpha globin chain antibody. Using laser confocal fluorescence microscopy, normal erythroid precursors show no alpha globin chain accumulation and exhibited uniformly smooth rim fluorescence of the three membrane proteins. In some thalassemic precursors, spectrin appeared to interact with large alpha globin accumulations, and in many of these cells the spectin appeared clumped and discontinuous. Band 4.1 interacted strongly with accumulations of alpha globin in thalassemic precursors to produce bizarrely clumped zones of abnormal band 4.1 distribution. Band 3 was incorporated smoothly into thalassemic erythroblast membranes. However, the proerythroblasts and basophilic erythroblasts were significantly deficient in band 3. Thus, accumulations of alpha globin in beta- thalassemia major colocalized with and disrupt band 4.1 and spectrin assembly into the membrane. The cause of deficient band 3 incorporation into thalassemic proerythroblast membranes remains unknown. These profound membrane alterations would likely contribute to the intramedullary lysis seen in Cooley's anemia.
    Print ISSN: 0006-4971
    Electronic ISSN: 1528-0020
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  • 4
    Publication Date: 1980-10-01
    Description: In order to study 45Ca distribution within erythrocytes, a method was devised that had minimal deleterious effects on the treated erythrocytes. It was observed that newly introduced 45Ca was predominantly recoverable from the cytosol and exchanged relatively slowly with membrane-associated Ca. Younger erythrocytes appeared to have relatively more 45Ca in membrane-associated sites. Erythrocytes from patients with sickle cell anemia had significantly more 45Ca in membrane-associated sites than did normal controls or patients with reticuloctosis due to a variety of disorders. There are theoretical reasons for considering the possibility that the distribution of 45Ca between cytosol and membrane-associated sites could modulate some of the properties of the erythrocyte membrane.
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    Electronic ISSN: 1528-0020
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  • 5
    Publication Date: 1986-06-01
    Description: The heavy chain of myosin can be detected in human red cell membranes by immunoblot analysis with antiplatelet myosin antibodies. Neonatal red cell membranes have more immunoreactive myosin than adult membranes. Membranes from young adult red cells contain more immunoreactive myosin than membranes from old adult red cells. In contrast, young and old neonatal red cells have equivalent mounts of myosin. Erythrocyte myosin is present in a membrane fraction enriched in integral membrane proteins but is not found in cytoskeletal preparations.
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    Electronic ISSN: 1528-0020
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  • 6
    Publication Date: 1987-07-01
    Description: The protein composition of ghosts, inside-out vesicles (IOV), and membrane skeletons (MS) of erythrocytes (RBC) from splenectomized (spx) and nonsplenectomized (non-spx) patients with beta-thalassemia major and beta-thalassemia intermedia was determined. Ghosts from spx thalassemia intermedia patients had a significant increase in their globin content (which was mostly heme reactive) and contained extra polypeptides in the protein 4.2 to 5 and 6-globin areas. The Triton- extracted MS from all of the thalassemic patients showed two major abnormalities: they retained up to twice the amount of protein 3 when compared with controls; they had a significant increase in their globin content, the concentration of which was independent of their protein 3 content. Analysis of the IOV revealed no differences between those prepared from normal controls and those of the patients. MS from spx thalassemia intermedia patients were grossly abnormal when examined by scanning electron microscopy and they exhibited aggregates of material that on transmission electron microscopy suggested the presence of globin precipitates. We propose that, although the integral protein composition, as reflected in the IOV, from severely affected beta- thalassemics is intact, their MS assembly is deranged. The altered skeletal structure of thalassemic RBC could result from attachment of denatured globin to the skeleton components. These abnormalities may contribute to the premature cell death seen in severe beta-thalassemia.
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    Electronic ISSN: 1528-0020
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  • 7
    Publication Date: 1977-08-01
    Print ISSN: 0006-4971
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  • 8
    Publication Date: 1984-08-01
    Description: ntracellular calcium (Ca) concentration in erythrocytes (RBCs) is controlled by a low passive influx through a relatively impermeable membrane and by active efflux catalyzed by Ca2+,Mg2+-ATPase. Since precipitation of alpha-globin chains in thalassemic RBCs may interfere with normal membrane function, we studied the RBC intracellular Ca content and the RBC membrane Ca2+,Mg2+-ATPase activity in two groups of patients with nonsplenectomized (n = 9) and splenectomized (n = 9) beta- thalassemia intermedia and in two groups of matched controls. The mean +/- SD Ca concentration in the nonsplenectomized (n = 12) and splenectomized (n = 6) controls were 6.1 +/- 6.0 and 5.8 +/- 3.4 mumol Ca per liter of RBCs, respectively, compared with 26.0 +/- 7.6 (P less than .001) and 85 +/- 24.4 (P less than .001) in the nonsplenectomized and splenectomized thalassemia patients, respectively. The mean +/- SD Ca2+,Mg2+-ATPase activity in the eight nonsplenectomized patients was 0.77 +/- 0.58 mumol inorganic phosphate (Pi) per milligram of protein per hour compared with 0.66 +/- 0.41 in the controls (P = NS). Similar values were obtained for the splenectomized patients and their controls. No correlation was found between either the intracellular Ca content or the Ca2+,Mg2+-ATPase activity with the peripheral nucleated RBC count. These findings suggest that there is a major defect in the membrane of the thalassemic RBC leading to an increased Ca content that is more pronounced in splenectomized patients.
    Print ISSN: 0006-4971
    Electronic ISSN: 1528-0020
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  • 9
    Publication Date: 1981-11-01
    Print ISSN: 0006-4971
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  • 10
    Publication Date: 1980-10-01
    Description: In order to study 45Ca distribution within erythrocytes, a method was devised that had minimal deleterious effects on the treated erythrocytes. It was observed that newly introduced 45Ca was predominantly recoverable from the cytosol and exchanged relatively slowly with membrane-associated Ca. Younger erythrocytes appeared to have relatively more 45Ca in membrane-associated sites. Erythrocytes from patients with sickle cell anemia had significantly more 45Ca in membrane-associated sites than did normal controls or patients with reticuloctosis due to a variety of disorders. There are theoretical reasons for considering the possibility that the distribution of 45Ca between cytosol and membrane-associated sites could modulate some of the properties of the erythrocyte membrane.
    Print ISSN: 0006-4971
    Electronic ISSN: 1528-0020
    Topics: Biology , Medicine
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