This proposal is designed to study the complex interplay of the components of the human red blood cell (RBC) membrane that result in its unique functional capacities. Abnormalities of these capabilities are very well exemplified by the pathophysiology of the human thalassemias, disorders of world wide importance, that cause anemia sufficient to produce morbidity and even mortality. While these inherited disorders may eventually be managed by the tools of molecular biology, we propose that they can be ameliorated by applying what we plan to learn in this project. The pathophysiology of alpha and beta thalassemia is different, and it is hypothesized that each is caused by excess beta or alpha globin chains binding to the RBC membrane and its skeleton, producing misshapen RBC with altered cellular hydration and deranged membrane material properties, leading to intramedullary hemolysis (ineffective erythropoiesis) and peripheral red cell destruction. Experiments are proposed to determine the mechanism underlying the apoptosis contributes to intramedullary lysis in beta thalassemia major and to contrast alpha vs. beta thalassemia and discover: how much of the excess unmatched globin chains bind; what specific derangements of membrane function are produced and how; which proteins show evidence of oxidant attack, whether the RBC abnormalities are preconditioned by events occurring in the marrow during erythroid maturation (beta thalassemia) perhaps altering the orderly assembly of membrane proteins, or whether senescence is particularly damaging (alpha thalassemia). Methods to be used include: analysis of membrane proteins by gel electrophoresis and Western blotting; isolation and purification of populations of thalassemic reticulocytes and beta thalassemic marrow erythroblasts for analysis; Northern blot studies to detect mRNA's mediating apoptotic programs; immunofluorescent confocal laser microscopy to identify protein distribution in marrow erythroblasts and circulating reticulocytes and RBC; ektacytometry and micropipette analysis to provide correlative data on cellular and membrane material properties; methods to identify protein thiol oxidation and tyrosine oxidation. Because we will compare marrow erythroblasts with reticulocytes and mature RBC, we can detect the site and stage of the alterations. The results should allow us to pinpoint key elements in the pathophysiology and perhaps clarify which of the current therapeutic clinical protocols is likely to produce cellular and thus clinical benefit.

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Research Project (R01)
Project #
5R01DK013682-34
Application #
2136804
Study Section
Hematology Subcommittee 2 (HEM)
Program Officer
Badman, David G
Project Start
1979-06-01
Project End
1998-06-30
Budget Start
1995-07-01
Budget End
1996-06-30
Support Year
34
Fiscal Year
1995
Total Cost
Indirect Cost
Name
Stanford University
Department
Internal Medicine/Medicine
Type
Schools of Medicine
DUNS #
800771545
City
Stanford
State
CA
Country
United States
Zip Code
94305
Schrier, Stanley L; Centis, Filippo; Verneris, Michael et al. (2003) The role of oxidant injury in the pathophysiology of human thalassemias. Redox Rep 8:241-5
Angelucci, Emanuele; Bai, Hua; Centis, Filippo et al. (2002) Enhanced macrophagic attack on beta-thalassemia major erythroid precursors. Haematologica 87:578-83
Schrier, Stanley L (2002) Pathophysiology of thalassemia. Curr Opin Hematol 9:123-6
Centis, F; Tabellini, L; Lucarelli, G et al. (2000) The importance of erythroid expansion in determining the extent of apoptosis in erythroid precursors in patients with beta-thalassemia major. Blood 96:3624-9
Pootrakul, P; Sirankapracha, P; Hemsorach, S et al. (2000) A correlation of erythrokinetics, ineffective erythropoiesis, and erythroid precursor apoptosis in thai patients with thalassemia. Blood 96:2606-12
Schrier, S L; Bunyaratvej, A; Khuhapinant, A et al. (1997) The unusual pathobiology of hemoglobin constant spring red blood cells. Blood 89:1762-9
Aljurf, M; Ma, L; Angelucci, E et al. (1996) Abnormal assembly of membrane proteins in erythroid progenitors of patients with beta-thalassemia major. Blood 87:2049-56
Yuan, J; Bunyaratvej, A; Fucharoen, S et al. (1995) The instability of the membrane skeleton in thalassemic red blood cells. Blood 86:3945-50
Yuan, J; Angelucci, E; Lucarelli, G et al. (1993) Accelerated programmed cell death (apoptosis) in erythroid precursors of patients with severe beta-thalassemia (Cooley's anemia) Blood 82:374-7
Advani, R; Rubin, E; Mohandas, N et al. (1992) Oxidative red blood cell membrane injury in the pathophysiology of severe mouse beta-thalassemia. Blood 79:1064-7

Showing the most recent 10 out of 15 publications