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Biomedical subjects

S B Shohet

Publications and source records attributed to S B Shohet.

At least 91 records · Page 5Linked to original sources
PubMed1980

Deformability and spectrin properties in three types of elongated red cells.

Three types of unusual, elongated erythrocytes have been studied in our effort to understand factors that regulate the shape and deformability of red blood cells. Measurements of deformability, morphologic response to membrane-shape-altering drugs, spectrin phosphorylation, spectrin extractability, and casein kinase activity were measured in llama cells, irreversibly sickled cells, and nonhemolytic human elliptocytes. The results of these studies support the concept that the physical properties of spectrin are important in maintaining the shape and deformability of the erythrocyte. Furthermore, changes in spectrin extractability and phosphorylation appear to be associated with changes in the viscoelastic properties of the membrane. However, it seems that spectrin's regulatory role is permissive rather than direct and active.

Anemia, Sickle Cell↗
PubMed1980

Deformability of oxygenated irreversibly sickled cells.

The deformability characteristics of isolated subpopulations of irreversibly sickled cells (ISC) have been studied in an ektacytometer. Analysis of laser diffraction patterns of well-oxygenated cells subjected to shear stress in solutions of varying osmolality has demonstrated a profound influence of mean corpuscular hemoglobin concentration and intracellular viscosity on the deformability of ISC. Virtually undeformable at 290 mosM, ISC became almost totally deformable at 130 mosM. In addition, when ISC membranes were loaded with normal hemoglobin at low concentration, they deformed easily in isotonic medium, as did resealed normal cell membranes. The restoration of deformability of ISC upon reduction of their hemoglobin concentration and internal viscosity to normal levels suggests that altered membrane properties are not the primary determinant of decreased deformability in these cells. Rather, cellular dehydration induced by previous sickling would appear to contribute in a major way to their abnormal rheological behavior.

Anemia, Sickle Cell↗
PubMed1980

Analysis of factors regulating erythrocyte deformability.

Using a laser diffraction technique, we have studied factors that influence the deformability of erythrocytes. Variations in suspending medium osmolality and applied shear stress were employed to isolate the individual contributions to whole cell deformability of internal viscosity, surface area-to-volume ratio, and viscoelastic properties of the membrane. An experimental system was devised in which normal cells were modified in vitro to induce specific alterations in each factor. Measurements of deformability as a function of medium osmolality showed characteristic behavior of the modified cells. Reduced surface area-to-volume ratio was detected by an exaggeration of the normal decrease in deformability as medium osmolality was decreased. In contrast, increased internal viscosity was detected by an increase in deformability as osmolality was decreased. Finally, decreased membrane flexibility was detected by reduced deformation at low shear stress. These methods of analysis were applied to cells from patients with hereditary spherocytosis, hereditary pyropoikilocytosis, and hemoglobin CC disease to define the basis of reduced deformability. Hereditary spherocytes showed the combined effects of reduced surface area and increased internal viscosity. Hereditary pyropoikilocytes revealed the effects of severely reduced surface area-to-volume ratio. Hemoglobin CC cells showed only the effects of high internal viscosity. An increase in the membrane shear modulus (decreased membrane deformability) was not evident in these disorders.

Anemia, Hemolytic, Congenital↗
PubMed1980

Ektacytometric analysis of factors regulating red cell deformability.

Photometric analysis of laser diffraction patterns has been used to obtain quantitative measurements of deformability of specifically modified normal red cells. Variation of deformability with suspending medium osmolality and with applied shear stress was used to distinguish between changes in internal viscosity, surface area-to-volume ratio, and viscoelastic properties of the membrane in their influence on whole cell deformability.

Dextrans↗
PubMed1980

Influence of red cell water content on the morphology of sickling.

The response of sickle cells with varying water content to alterations in oxygen tension has been studied. Cells that were severely dehydrated while sickled retained the characteristic sickled morphology even after prolonged reoxygenation. When the cell water content was increased by reduction of the suspending medium osmolality, the cells unsickled. Cells that were dehydrated before deoxygenation were unable to assume the spiculated morphology typical of sicked cells. This was true both for high mean cell hemoglobin concentration (MCHC) discoid sickle cells and for irreversibly sickled cells. When such cells were resuspended in hypotonic medium before deoxygenation, they sickled with the characteristic morphology of sickle cells with normal MCHC. The morphological behavior of Ca-loaded sickled cells as well as irreversibly sickled cells showed a major influence of increased hemoglobin concentration and extremely high internal viscosity. Constraint on cell morphology by putative membrane rigidity was not observed.

Anemia, Sickle Cell↗
PubMed1980

Erythropoietic stress, macrocytosis, and hemoglobin switching in HbAA sheep.

Hemoglobin switching and macrocytosis were studied in homozygous hemoglobinAA sheep. An abrupt initiation of erythropoietic stress, accompanied by a pulsed elevation of circulating erythropoietin levels, was induced by phlebotomy. Sequential blood samples were separated according to density on Stractan gradients to isolate cells newly entering the circulation from the marrow each day. Analysis of hemoglobin phenotype and cell volume distribution in these young reticulocytes revealed a distinct temporal separation in the appearance of hemoglobin C and increased cell volume. The appearance of macrocytes within 24 hr of erythropoietin elevation suggests that macrocytosis could be the result of the action of erythropoietin during the late stages of erythroid maturation. The 72-hr delay in the appearance of hemoglobin C indicates that commitment to a particular hemoglobin phenotype occurs at an early stage of differentiation and involves immature erythroid stem cells. The results of this study show that these consequences of erythropoietic stress are initiated at two different developmental stages, resulting in the production of macrocytosis and hemoglobin switching.

Animals↗
PubMed1980

Inaccuracies associated with the automated measurement of mean cell hemoglobin concentration in dehydrated cells.

Because of discrepancies between electronically and manually measured values of mean cell hemoglobin concentration (MCHC) encountered in studies of pathologic red cells, we studied the effect of cell water content on MCHC measurements by both methods. A series of red cell samples with varying water contents (54%-164% normal) were prepared from normal cells using the antibiotic nystatin. MCHC was then measured, using the microhematocrit centrifuge and three different electronic cell counters in common laboratory use. For MCHC values above 36 g/dl as measured by the spun hematocrit method, all three electronic counters under estimmated the MCHC, with increasing error as the true MCHC increased. For MCHC values below 30 g/dl, the values from two conductivity based instruments agreed with those from the spun hematocrit method, whereas one instrument based on light scattering overestimated the MCHC. These results indicate that inaccuracies in the measured mean cell volume (MCV) of dehydrated or otherwise undeformable cells may lead to spurious values for MCHC when electronic cell counters are used.

Erythrocyte Indices↗
PubMed1979

Hereditary acanthocytosis associated with the McLeod phenotype of the Kell blood group system.

Some boys with X-linked chronic granulomatous disease (CGD) have red cells of the rare McLeod phenotype in the Kell blood group system. Only one example of this phenotype has previously been described in a non-CGD subject. We have studied a 10-year-old boy and a maternal brother who do not have CGD and whose red cells are of the McLeod type . The boy presented as a haematological problem with red-cell abnormalities. These were acanthocytosis, anisocytosis and 'tailing' in the osmotic fragility curve, changes now known to occur with the McLeod phenotype. Subsequent studies revealed his rare blood group. A family study has established that an uncle also has acanthocytic red cells and the McLeod phenotype. In addition the boy's sister, mother and maternal grandmother all show red-cell mosaicism with double populations of McLeod acanthocytes and normal red cells of common Kell type. The gene that determines inheritance of the McLeod phenotype is X-linked and the mosaicism present in female carriers is believed to result from X chromosome inactivation by the Lyon effect. The study provides further evidence that the McLeod phenotype arises by inheritance of a variant X-linked modifying gene and not through inheritance of a variant gene at the Kell autosomal locus. It also represents the first occasion that a person of rare blood group has been recognized because of an associated anomaly in red cell morphology.

Acanthocytes↗
PubMed1979

Erythrocyte membrane lipid reorganization during the sickling process.

In order to study possible alterations in membrane lipids during sickling, we have measured the difference in susceptibility to lipid peroxidation, binding of trinitrobenzenesulfonic acid (TNBS) to aminophospholipids, and fatty acid uptake in cells containing sickle haemoglobin under aerobic and anaerobic conditions. We have also examined TNBS binding in irreversibly sickled cells in an attempt to evaluate the permanent effects of any such alterations. We found that when erythrocytes were sickled by deoxygenation, the susceptibility to lipid peroxidation and binding of TNBS to aminophospholipids was markedly increased, while normal control cells showed no change. These effects appeared to be specific for the sickled state rather than a nonspecific consequence of cell age or the concentration of sickle haemoglobin within the cell. In contrast, fatty acid incorporation into membrane phospholipids, representing potential lipid renewal, was decreased in the sickled state. Cell fractions enriched in irreversibly sickled cells showed increased TNBS labelling in air and only modest rises with anoxia. Taken together, these data imply a rearrangement of membrane lipids during the sickling process and suggest a permanent reorganization of membrane lipids in the irreversibly sickled cell.

Aerobiosis↗
PubMed1979

Variability in erythrocyte deformability among various mammals.

Deformability is an important aspect of erythrocyte physiology and has been extensively studied using human red cells. We have studied erythrocytes from 25 different animals using a viscometric technique. Erythrocyte diameters ranged from 3.3 microns in the goat to 11.4 microns for the elephant seal. Erythrocytes from most species deformed readily when a fluid shear stress was applied. A deformability index of the stressed cell defined as (length - width)/(length + width) correlated with cell size. The erythrocytes of four animals (pygmy goat, goat, Batanga horse, and miniature horse) deformed less than most species. Camel and llama erythrocytes, which were ellipsoidal, did not deform but oriented in the stress field.

Animals↗
PubMed1979

Reconstitution of spectrin-deficient, spherocytic mouse erythrocyte membranes.

To study directly the role of spectrin in erythrocyte membrane function, we have designed a reconstituted membrane system using erythrocyte membranes from spectrin-deficient mice and purified spectrin from normal mice. The normal spectrin is inserted into the spectrin-deficient spherocytes by exchange hemolysis. Thereafter, raising the ionic strength and temperature reseals the cells and, with time, facilitates binding of the spectrin to the spectrin-deficient membranes. The binding is apparently specific as shown by its dependence upon the concentration of undenatured spectrin and the concentration of salt used, as well as by the immunofluorescent appearance of the reconstituted cells after treatment with specific antispectrin antibody. In terms of in vitro cellular behavior, the reconstituted preparations show marked changes in comparison to the untreated spherocytes. In particular, membrane stability, as measured by the reduction of myelin figure formation and lipid loss, is considerably enhanced. In addition, membrane fusion, which occurs readily with the untreated spherocytes, is virtually eliminated. Finally, the osmotic behavior of the native spherocytes is appreciably altered, such that the early phase of osmotically induced swelling, as measured in a high-speed stop-flow apparatus, is delayed and modified. Taken together, these findings indicate specific roles for spectrin in the stabilization of the erythrocyte membrane, in the limitation of membrane fusion, and in the modulation of the membrane's response to osmotic stress.

Animals↗
PubMed1978

Effects of abnormal cation transport on deformability of desiccytes.

We have studied the deformability of subpopulations of red cells from a patient with "desiccytosis," a disorder characterized by increased membrane permeability to potassium and associated with a probable increase in sodium-sodium exchange. Cells become increasingly dehydrated after maturation because of continued potassium loss without compensatory sodium gain, and they exhibit a progressive increase in mean cell hemoglobin concentration (MCHC). This increase in MCHC causes the cells to become underformable at shear stress values which result in extensive deformation of normal cells. Reduction of MCHC to approximately normal levels by suspending the cells in hypotonic medium restores normal deformability to all but 0.1--0.2% of the cells. These results suggest that the major factor leading to premature destruction in this disorder is whole cell rigidity conferred by increased intracellular hemoglobin concentrations, rather than any associated membrane rigidity.

Anemia, Hemolytic↗
PubMed1978

Bilayer balance and regulation of red cell shape changes.

Discocytic human red cells undergo discocyte-echinocyte and discocyte-stomatocyte transformations under the action of a wide variety of lipid-soluble anionic and cationic agents respectively. These shape transformations are explained by the bilayer couple hypothesis of Sheetz and Singer to be the result of preferential distribution of the anionic agents in the outer half of the bilayer and the cationic agents in the inner half of the bilayer. We demonstrate that echinocytogenic effects indeed occur when the naturally occurring phospholipid lysophosphatidylcholine (LPC) is localized in the outer half of the bilayer, and stomatocytogenic effects occur when LPC is in the inner half. However, in contrast to the bilayer couple hypothesis, our results show that simple equivalent membrane surface area expansion on each layer is insufficient to maintain the discocytic shape and there exists a differential concentration effect of LPC on the two halves of the bilayer.

Erythrocyte Membrane↗
PubMed1978

Monovalent cation transport in irreversibly sickled cells.

Using discontinuous density gradients of Stractan II, we have separated sickle cell blood into discrete subpopulations of reticulocytes, mature discoid cells, and irreversibly sickled cells (ISCs). We have measured active and passive fluxes of monovalent cations in mature discoid cells, ISCs, and normal control cells, also separated upon density gradients. These measurements revealed a decreased active cation transport in ISC-rich populations. However, parallel measurements of Na, K-ATPase activity showed normal ouabain-sensitive ATPase activity in ISCs. Passive permeability to external Rb was also normal in ISCs. The observation of depressed pump activity in intact ISCs, contrasted with normal ATPase activity in ISC membranes, suggests the presence of factors in the intact cell which inhibit the active transport of Na and K in ISCs.

Adenosine Triphosphatases↗