Mechanical properties of oxygenated red blood cells in sickle cell (HbSS) disease

Abstract

Little data exist for the mechanical properties of individual irreversible or reversible sickle cells (ISC and RSC, respectively), nor is the process of ISC formation well understood. For oxygenated ISC and density-fractionated RSC, we have used micropipette techniques to measure cell surface area (SA) and volume (V), membrane shear elastic modulus (mu), time constant for viscoelastic shape recovery (tc), and hence to calculate membrane surface viscosity (eta = mu X tc). Volume loss associated with increasing cell density was accompanied by a proportionately smaller surface area decrease; SA/V ratio thus increased for denser cells, with ISC having the highest values. Membrane area loss by fragmentation must thus be accompanied by an accelerated decrease in cell volume. ISC had relatively rigid membranes (mu 130% above normal controls) and tc close to normal values, so that their effective membrane viscosity was more than double control. RSC had viscoelastic properties close to control, but showed wider variation between sickle cell donors and within samples. Measurements on density-separated RSC showed that, on average, mu was nearly constant, but that tc was longer for the densest cells, with their eta approaching ISC levels. A small subpopulation of RSC were found that had mu close to ISC values. Hypotonically swollen ISC (with internal hemoglobin concentration decreased to normal levels) retained their increased membrane stiffness but had markedly decreased tc, so that their eta approached normal values. The results show that elevated hemoglobin concentration influences the viscoelastic behavior of ISC and RSC, but that an irreversible change in membrane elasticity also occurs for ISC. These data suggest that ISC formation occurs via a two- stage process: (1) accelerated volume loss leading to increased cytoplasmic and effective membrane viscosity; (2) a sharp rise in membrane rigidity, presumably linked to membrane structural alteration.

Sickle Red Cell Microrheology and Sickle Blood Rheology

“…Decreased cellular deformability of oxygenated sickle cell anemia RBC has been documented using a variety of techniques, including viscometry, filtration, ektacytometry, and micropipet aspiration of individual cells (17,32,35,49,61,102,127,141). In these measurements, reduction in RBC deformability is manifested, respectively, by increased viscosity of blood (32), decreased filtration rate of dilute cell suspension through narrow pores (127,141), decreased ability of cells to undergo deformation in shear fields (17,35), and increased aspiration pressures needed to induce entry of cells into micropipets (49,61,102).…”

“…Decreased cellular deformability of oxygenated sickle cell anemia RBC has been documented using a variety of techniques, including viscometry, filtration, ektacytometry, and micropipet aspiration of individual cells (17,32,35,49,61,102,127,141). In these measurements, reduction in RBC deformability is manifested, respectively, by increased viscosity of blood (32), decreased filtration rate of dilute cell suspension through narrow pores (127,141), decreased ability of cells to undergo deformation in shear fields (17,35), and increased aspiration pressures needed to induce entry of cells into micropipets (49,61,102). These studies documented the abnormal rheologic properties of oxygenated sickle cell anemia RBC and, in addition, showed that there is marked heterogene-ity in the extent of rheologic abnormalities of RBC in individual blood samples and among blood samples from different individuals (3,7,49,79,102,132).…”

Erythrocyte heterogeneity in sickle cell disease: effect of deoxygenation on intracellular polymer formation and rheology of sub‐populations

“…With increasing density, there was a progressive increase in MCHC and ISC%, a right shift in the OSC, and impaired deformability of oxygenated erythrocytes. This rheological impairment of dense SS cells with a relatively high MCHC has also been demonstrated by viscometric (Kaul et al, 1983), micropipette aspiration (Evans et al, 1984;Nash et al, 1984) and ektacytometric techniques (Clark et al, 1980;Sorette et al, 1987). The increase in viscosity on deoxygenation of solutions of sickle haemoglobin is also dependent on haemoglobin concentration (Chien et al, 1982).…”

Influence of sickle hemoglobin polymerization and membrane properties on deformability of sickle erythrocytes in the microcirculation

“…The methods most commonly used to study erythrocyte suspensions are viscometry (Dintenfass, 1964;Charache and Conley, 1964;Chien et al, 1970Chien et al, , 1982Drasler et al, 1989), filtration (Weed et al, 1969;Messer and Harris, 1970;Chien et al, 1971;Lessin et al, 1977;Reinhart et al, 1984;Arai et al, 1990), and ektacytometry Mohandas, 1975, 1977;Groner et al, 1980;Sorette et al, 1987). The micropipette technique is useful to study single cell deformability, including cell membrane properties (Evans and Hochmuth, 1976;LaCelle et al, 1977;Nash et al, 1984). In addition to cell suspension studies, viscometry is also useful for determining the rheological properties of Hb S solutions (Briehl, 1981;Chien et al, 1982;Danish et al, 1987;Drasler et al, 1989).…”

“…The deformability properties of erythrocytes are critical determinants of flow in the microcirculation (Skalak, 1969;Evans and Hochmuth, 1976;Chien et al, 1982Chien et al, , 1987Linderkamp and Meiselman, 1982;Nash and Meiselman, 1982;Nash et al, 1984Nash et al, , 1986Pfafferott et al, 1985;Shohet and Mohandas, 1988). Abnormalities of these properties are thought to be the principal cause of the pathophysiology of sickle cell disease and other hemoglobinopathies (Schechter et al, 1987).…”