Our long term goals are to understand the relation between hydration and polymerization in the sickle red cell and to predict its behavior as it moves through the hypertonic environment of the renal medulla. (Severe sickling commonly occurs in the kidney, probably because of the strong concentration dependence of HbS polymerization.) We will investigate and characterize the sources of membrane lesions that are responsible for perturbations of membrane transport that lead to different hydration and polymerization states of these cells. We will investigate differences between osmotic equilibrium properties of normal and sickle cells and the extent that these differences account for volume regulation problems encountered by sickle cells in various states. This will include measurements of how the osmotic coefficient of sickle cell Hb and fixed charge changes with concentration, pH and 02 tension, and how these results are altered as the Hb polymerizes. We will also examine the extent that hydration, polymerization, and transport properties of sickle cells are perturbed by the hypertonic and high urea environments characteristic of the renal medullary circulation. This will include measurements of how the sickle cell permeabilities of Na+, K+, Ca++, urea and glutathione depend on the hypertonic salt and urea concentrations in their environment and how these factors are modified by pH and hypoxia. Further, the kinetics of Hb polymerization in response to concentration jumps under simulated renal medulla environments will be studied, and particular attention will be paid to inhibition of polymerization by intracellular urea. Data obtained in the above projects will be assembled in a model designed to predict the fate of a sickle red cell as it enters the renal medulla. Finally we will investigate biochemical mechanisms responsible for these membrane perturbations. We will test the hypothesis that the membrane defects are a result of oxidative damage that accrues when the membrane is deformed by osmotic stress.

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
Research Project (R01)
Project #
5R01HL037593-03
Application #
3353404
Study Section
(SRC)
Project Start
1986-09-30
Project End
1989-09-29
Budget Start
1988-09-30
Budget End
1989-09-29
Support Year
3
Fiscal Year
1988
Total Cost
Indirect Cost
Name
University of California Berkeley
Department
Type
Schools of Arts and Sciences
DUNS #
094878337
City
Berkeley
State
CA
Country
United States
Zip Code
94704
Mannuzzu, L M; Moronne, M M; Macey, R I (1993) Estimate of the number of urea transport sites in erythrocyte ghosts using a hydrophobic mercurial. J Membr Biol 133:85-97
Gross, C T; Salamon, H; Hunt, A J et al. (1991) Hemoglobin polymerization in sickle cells studied by circular polarized light scattering. Biochim Biophys Acta 1079:152-60
Moronne, M M; Mehlhorn, R J; Miller, M P et al. (1990) ESR measurement of time-dependent and equilibrium volumes in red cells. J Membr Biol 115:31-40
van den Berg, J J; Kuypers, F A; Roelofsen, B et al. (1990) The cooperative action of vitamins E and C in the protection against peroxidation of parinaric acid in human erythrocyte membranes. Chem Phys Lipids 53:309-20
Karan, D M; Macey, R I (1990) Temperature- and concentration-dependence of urea permeability of human erythrocytes determined by NMR. Biochim Biophys Acta 1024:271-7
Yousef, L W; Macey, R I (1989) A method to distinguish between pore and carrier kinetics applied to urea transport across the erythrocyte membrane. Biochim Biophys Acta 984:281-8
Todd, A P; Mehlhorn, R J; Macey, R I (1989) Amine and carboxylate spin probe permeability in red cells. J Membr Biol 109:41-52
Todd, A P; Mehlhorn, R J; Macey, R I (1989) Amine spin probe permeability in sonicated liposomes. J Membr Biol 109:53-64
Macey, R I; Yousef, L W (1988) Osmotic stability of red cells in renal circulation requires rapid urea transport. Am J Physiol 254:C669-74
Orme, F W; Moronne, M M; Macey, R I (1988) Modification of the erythrocyte membrane dielectric constant by alcohols. J Membr Biol 104:57-68

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