Search

Red blood cell (RBC) aggregation induced by fibrinogen is a major determinant of the non-Newtonian flow behavior of human blood and has been suggested as a possible contributing factor for thrombogenesis. Given the elevated fibrinogen levels and the high incidence of thrombotic accidents in nephrotic syndrome (NS), a study was designed to assess RBC aggregation in this disease. Three different aggregation assays were used: (1) changes in light transmission through static cell suspensions, aggregation half time (AHT); (2) low gravity centrifugation of blood in nearly vertical tubes, zeta sedimentation ratio (ZSR); (3) direct observation of dilute RBC suspensions, microscopic aggregation index (MAI). The results indicate that RBC aggregation, as measured by AHT and ZSR, is increased significantly in NS patients versus healthy controls (P less than 0.001 for both methods); this increase was not a function of renal insufficiency. The AHT and ZSR data were well correlated with plasma fibrinogen levels (r = 0.75 and 0.84, respectively). No differences were observed with the MAI method, since with this technique the aggregation process reaches an asymptotic plateau for the range of fibrinogen found in NS. Implications of increased RBC aggregation and thus increased blood viscosity and flow disturbances on thrombogenesis are discussed.

The membrane shear elastic modulus (mu) and the time constant for extensional shape recovery (tc) were measured for normal, control human red blood cells (RBC) and for RBC heat treated (HT) at 48 degrees C. Three separate methods for the measurement of mu were compared (two used a micropipette and one employed a flow channel), and the membrane viscosity (n) was calculated from the relation n = mu. tc. The deformability of HT and control cells was evaluated using micropipette techniques, and the bulk viscosity of RBC suspensions at 40% hematocrit was measured. The shear elastic modulus, or "membrane rigidity", was more than doubled by heat treatment, although both the absolute value for mu and the estimate of the increase induced by heat treatment varied depending on the method of measurement. Heat treatment caused smaller increases in membrane viscosity and in membrane bending resistance, and only minimal changes in cell geometry. The deformability of HT cells was reduced: 1) the pressure required for cell entry (Pe) into 3 micrometers pipettes was increased, on average, by 170%; 2) at an aspiration pressure (Pa) exceeding Pe, longer times were required for cell entry into the same pipettes. However, when Pa was scaled relative to the mean entry pressure for a given sample (i.e, Pa/Pe), entry times were similar for control and HT cells. Bulk viscosity of HT RBC suspensions was elevated by approximately 12% on average (shear rates 75 to 1500 inverse seconds). These findings suggest that alteration of RBC membrane mechanical properties, similar to those induced by heat treatment, would most affect the in vivo circulation in regions where vessel dimensions are smaller than cellular diameters.

Red blood cell aggregation induced by fibrinogen is a major determinant of the non-Newtonian flow behavior of blood and has been suggested as a possible contributing factor for thrombogenesis. Given the elevated fibrinogen levels and the incidence of thrombotic accidents in pregnancy, a study was designed to assess red blood cell aggregation at selected gestational periods. Three separate in vitro aggregation assays were used: (1) aggregation half time, (2) zeta sedimentation ratio, and (3) microscopic aggregation index. Our results indicate that red blood cell aggregation is increased throughout normal pregnancy (i.e., at 10, 25, and 36 weeks and during labor) in comparison to that in nonpregnant women. Significant correlations between plasma fibrinogen concentration and zeta sedimentation ratio (p less than 0.001) or aggregation half time (p less than 0.02) were demonstrated, but the correlation coefficient for the microscopic aggregation index technique did not reach significance (p greater than 0.20). Type O-positive red blood cells suspended in plasma from pregnant women also exhibited increased aggregation (p less then 0.001), thus suggesting plasma rather than cellular factors for the enhanced aggregation. Possible influences of abnormal fibrinogen structure during pregnancy are considered and the implications of increased red blood cell aggregation vis-à-vis altered blood viscosity are discussed.

A new method for altering red cell morphology by high-speed centrifugation of cells through a physiological medium is described. Cell shape is preserved for microscopic analysis by allowing the sedimenting cells to pass from the physiological medium into a glutaraldehyde fixative solution. Examination of the deformed, fixed cells indicates that the vast majority resemble spheres with a flat, triangular tail. Measurements of the overall length of deformed cells show a nearly linear relationship between cell length and centrifugal force; average cell length increased from 8 to 11 micrometer as the centrifugal field was increased from 2,000 to 15,000 g. These data suggest that this centrifugal technique may be useful for evaluating cellular deformability and, potentially, the material properties of red cells.

To determine whether increased numbers of circulating endothelial cells, a possible indicator of endothelial injury, are present in subjects with sickle cell disease, we measured circulating endothelial cells in 30 normal subjects and in 23 subjects with sickle cell anemia. Mean circulating endothelial cells were significantly higher (P less than 0.025) in the sickle cell subjects than in the normal subjects. Circulating endothelial cells were significantly higher than normal in 10 sickle cell subjects studied during painful crisis (P less than 0.01) but not in 13 sickle cell subjects studied while in the steady state. To control for the known stimulatory effect of cigarette smoking on circulating endothelial cells, we analyzed the results for smokers and nonsmokers separately. Mean circulating endothelial cells were not significantly higher in sickle cell subjects who smoked (n = 10) than in normal subjects who smoked (n = 8), but were significantly higher (P less than 0.05) in sickle cell nonsmokers (n = 13) than in normal nonsmokers (n = 22). Among nonsmoking sickle cell subjects, mean circulating endothelial cells were significantly higher than normal (P less than 0.01) during painful crisis (n = 7), but not in the steady state (n = 6). We conclude that circulating endothelial cells are significantly increased in sickle cell crisis, and may indicate the occurrence of acute endothelial injury during episodes of microvascular occlusion.

The effects of low levels of glutaraldehyde uptake (less than 120 mumol/10(10) cells) on the physicochemical properties of human red blood cells (RBC) were investigated. Salient effects include: by different measures of cell deformability, the extent of glutaraldehyde uptake required to decrease cellular deformability was shown to range from approximately 8 to 30 mumol/10(10) cells; osmotically stressed red cells exhibit complete hemolysis when the level of glutaraldehyde uptake is less than 28 mumol/10(10) cells and no hemolysis when uptake is less than 70 mumol/10(10) cells with the extent of hemolysis decreasing in an approximately linear manner with glutaraldehyde uptake between these limits; glutaraldehyde uptake of up to 58 mumol/10(10) cells does not change the cells' density, mean cell volume or ability to retain potassium.

Hemoglobin solutions prepared from hemolyzed human erythrocyte packs have Newtonian flow properties. Diluted solutions are also Newtonian. All solutions have a viscosity lower than the apparent viscosity of erythrocyte suspensions of equal oxygen-carrying capacity. The presence of cell debris in hemoglobin solutions causes non-Newtonian (pseudoplastic or rheopectic) flow behavior.