Employing both microscopic and photometric methods the rheology of pathological red cell aggregation was studied in model experiments. Suspensions of washed human red blood cells in dextran solutions containing rising concentrations of dextrans (M.W. 40000, 70000, 110000, 250000, 500000) were used. At low concentrations ( 70000)red cell suspensions formed aggregates similar to the ones found in normal human blood. At higher concentrations, the aggregates were similar to those observed in pathological human blood. The aggregates were studied under the condition of stasis, slow flow and at shear rate of their hydrodynamic dispersion. Besides, the flow behavior of the dispersed cells at high shear rates was studied. We found:

1.

In all samples the rate of spontaneous aggregate re-formation in stasis (following hydrodynamic desaggregation) rose with rising dextran concentration up to 5.0 g-%.




2.

The shear resistance of the aggregates, as measured by the shear stress necessary to keep them dispersed, rose up to concentrations of 2.5 g-%, but fell at higher concentrations.




3.

Only with dextran of a molecular weight above 110000 coarse agglomerates could be produced at high concentrations. Loose elastic meshes were rapidly produced at high concentrations of Dx 70.




4.

When subjected to steady state low shear (7 sec−1) only the agglomerates, but not the meshes rapidly grew in size.

Microrheology and light transmission of blood. III. The velocity of red cell aggregate formation.

The rheological behavior of normal and pathological red cell aggregates in viscometric flow (artificial flow in cone plate chamber) is studied by direct microscopy, (rheoscopy) viscometry and photometry. Marked differences between normal and pathological blood are measured in the microrheological properties of red cell aggregates; only discreet differences are measured by blood viscometry (macrorheology). Both in normal and abnormal blood, red cell aggregation is a reversible process in the presence of adequate shear forces; their respective influences on apparent blood viscosity at low rates of shear are complex functions of shear rate, shear time, hematocrit and plasma viscosities. Pathological red cell aggregation (RCA) forms more rapidly and extensively than normal RCA. The pathological aggregates frequently have a tendency to grow at low rates of shear and they are highly shear resistant.

Red cell aggregation in blood flow. I. New methods of quantification.

Rheological properties were studied of young and old human erythrocytes from healthy adults. Viscosity measurements of packed erythrocyte suspensions as well as filtration of cells through polycarbonate sieves show that young cells are more flexible than aged ones. Since deformability of erythrocytes is the product of cell shape, flexibility of the membrane and fluidity of the intracellular hemoglobin, we studied the manner in which these factors are relevant to the diminished flexibility of aged erythrocytes. The biconcave cell shape is maintained during the process of aging. The viscosity of packed ghost suspensions from aged erythrocytes is increased versus that of young ones. The diminished flexibility of old ghosts correlates well with their smaller cell volume. The fluidity of the hemoglobin in the interior of the cells is decreased as indicated by an increased hemoglobin content of the isolated ghosts. We conclude that aged erythrocytes loose their deformability as a result of both a decreased fluidity of the intracellular hemoglobin and a diminished flexibility of the membrane.

Rheological properties of young and aged human erythrocytes

The rheological properties of erythrocytes were studied in 12 heterozygous and nine homozygous beta thalassaemia patients, using filtration of erythrocytes through polycarbonate sieves with a pore diameter of 5 micron and viscosity measurements of erythrocyte and ghost suspensions. Erythrocyte deformability of all patients was decreased. In the heterozygous patients moderately diminished flexibility of cells was due to microcytosis of erythrocytes. Rigidity of erythrocytes in homozygous patients was due to altered cell shape, diminished fluidity of haemoglobin, and in splenectomized patients to the presence of inclusion bodies. Membrane flexibility of erythrocytes remained unaltered. We suggest that the decreased fluidity of haemoglobin as well as the presence of inclusion bodies result from the excess of alpha chains in erythrocytes of homozygous patients. The haemoglobin fluidity of erythrocytes of one patient with thalassaemia intermedia was unchanged. The flexibility of these cells was only moderately altered. In four patients who needed transfusions every 1--4 weeks deformability of transfused erythrocytes was likewise decreased when compared to controls. Erythrocyte suspensions obtained from the splenic artery and vein during splenectomy showed decreased flexibility of transfused cells after their passage through the spleen.

Rheological properties of erythrocytes in heterozygous and homozygous beta thalassaemia.

The rheological properties of erythrocytes of 14 patients with iron deficiency anemia were studied by filtration of cells through polycarbonate filters with a nominal pore diameter of 5 Μm and by viscosity measurements of erythrocyte suspensions with a hematocrit of 80%. Erythrocytes of the patients passed through the filter pores more slowly than the cells from controls. The diminished deformability of the erythrocytes of the patients was solely due to an unfavorable ratio of cell surface area to microcytic cell volume. The viscosity of the ghost suspensions of the patients showed a normal flexibility. The hemoglobin content of the isolated ghosts was diminished, indicating an in-increased hemoglobin fluidity in the interior of the intact cells. The viscosity of erythrocytes of the patients was slightly increased at low shear rates but was normal at intermediate and high shear rates. We suggest that the decreased erythrocyte flexibility of microcytosis at low shear rates is no longer present at higher shear rates because of an increased fluidity of the intracellular hemoglobin. We discuss whether or not this mechanism also operates in vivo. The in vitro diminished deformability of erythrocytes explains the shortened survival of the patients' erythrocytes in vivo.

Deformability of erythrocytes in iron deficiency anemia.

A viscometer chamber (combination of cone plate and Couette-type) adaptable to the standard Wells-Brookfield-Microviscometer is described. The instrument allows measurements at shear rates between 0.23 and 160 sec−1; it incorporates a guard ring against surface artefacts. Measurements with normal human blood, standardized for hematocrit value, give excellent agreement with data obtained previously on the GDM-viscometers.

H. Schmid-Schönbein

Papers

A simple device allowing blood viscometry at low rates of shear with the Wells-Brookfield-viscometer.