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Biomedical subjects

G V Seaman

Publications and source records attributed to G V Seaman.

At least 19 recordsLinked to original sources
PubMed1996

Bioprocessing in microgravity: applications of continuous flow electrophoresis to rat anterior pituitary particles.

In this report we describe the results of a continuous flow electrophoresis (CFE) experiment done on STS-65 in which we tested the idea that intracellular growth hormone (GH) particles contained in a cell lysate prepared from cultured rat anterior pituitary cells in microgravity might have different electrophoretic mobilities from those in a synchronous ground control cell lysate. Collectively, the results suggested that CFE processing in microgravity was better than on earth; more sample could be processed/time (6 x) and more variant forms of GH molecules could be resolved as well. We had also hoped to carry out a pituitary cell CFE experiment, but failure of the hardware required that the actual cell electrophoresis trials be done on earth shortly after Shuttle landing. Data from these experiments showed that space-flown cells possessed a higher electrophoretic mobility than ground control cells, thereby offering evidence for the idea that exposure of cultured cells to microgravity can change their net surface charge-density especially when the cells are fed. Collectively, the results from this pituitary cell experiment document the advantage of using coupled cell culture and CFE techniques in the microgravity environment.

Animals↗
PubMed1991

Chronic blood hyperviscosity in subjects with acute stroke, transient ischemic attack, and risk factors for stroke.

The origin and significance of blood hyperviscosity in subjects with acute stroke has been controversial. It has been argued that viscous abnormalities simply reflect either elevated hematocrit or an acute-phase response to the stroke itself. To address these issues, we measured the factors that determine blood viscosity in a cross-sectional study of 430 subjects, including 135 with acute stroke, 89 with acute transient ischemic attacks of the brain, 115 with recognized risk factors for stroke, and 91 healthy controls. The at-risk group was balanced with the acute stroke group for types of risk factors and medication usage, and all four groups were balanced for age. The viscosity of whole blood at low rates of shear and the plasma viscosity were significantly elevated in both groups with cerebrovascular symptoms and in the at-risk group compared with the healthy controls. The severity of hyperviscosity was stroke group greater than transient ischemic attack group greater than at-risk group greater than healthy controls. Increased viscosity of whole blood was associated with an elevated plasma fibrinogen concentration and with a decreased albumin/globulin ratio. This study provides evidence that blood hyperviscosity is present not only in subjects with acute brain infarction, but also in those with risk factors for stroke, and that these abnormalities are, to a considerable degree, chronic.

Acute Disease↗
PubMed1990

Membrane active plasma factor in multiple sclerosis: characterization and isolation by recycling isoelectric focusing.

Recycling isoelectric focusing is a rapid, high resolution technique that has the capability of fractionating complex mixtures of proteins on a preparative scale on the basis of their isoelectric points (pIs). For this reason, it appeared to be an ideal tool to further characterize and isolate the surface active plasma component(s) which is abnormal in multiple sclerosis (MS). The normal control and the abnormal MS plasma components, or factors, proved to be stable under the conditions used in this technique, including deionization by electrodialysis, dialysis against distilled water, lyophilization and the presence of 3M urea and carrier distilled water, lyophilization and the presence of 3M urea and carrier ampholytes. The presence or absence of plasma factor activity was determined by incubating red blood cells in a test plasma, or plasma fraction, followed by the determination of the red blood cell electrophoretic mobility in the presence and absence of linoleic acid. Both the normal and MS factor had a pI of 4.0 +/- 0.1 under the conditions used. A high degree of purification was achieved and albumin was eliminated as a possible candidate for the factor(s).

Dialysis↗
PubMed1989

The importance of formaldehyde purity in studies on the electrokinetic charge of human red cells.

Electrokinetic measurements and rheological studies conducted in parallel have previously shown red cell surface charge to play a role in governing aggregative behavior and bulk flow properties of red cell suspensions. For these and other types of model investigations, aldehyde stabilized cells have been widely used. In this communication, the influence of the purity of formaldehyde was investigated. It was found that (a) the direct dissolution of commercially available paraformaldehyde in water or suitably buffered saline results in impure solutions which, if utilized in the fixation of human erythrocytes, produces cells which have significantly different electrophoretic properties from native cells; (b) the basis for the differences is the presence of metallic impurities in some commercially available paraformaldehyde preparations; (c) the impurities and thus the anomalous electrokinetic properties of the fixed cells may be eliminated by generating formaldehyde gas from paraformaldehyde by heating the latter to 203-210 degrees C; (d) alternatively, the impurities may be eliminated by addition of disodium ethylenediamine tetraacetate dihydrate to fixative solutions prepared directly from paraformaldehyde.

Adult↗
PubMed1989

Blood modeling using polystyrene microspheres.

The steady flow viscosity at shear rates 0 to 120 sec-1 and dynamic viscoelasticity at frequencies 0.02 to 0.8 Hz were determined for aqueous suspensions of uniform polystyrene microspheres of 1.0 micron diameter. Rheological properties of the microsphere suspensions were Newtonian for particle concentrations up to 32%. By introducing dextran and calcium chloride into the particle suspensions, non-Newtonian behavior was produced similar to that observed for human blood. The cooperative effects of dextran and calcium ions promoted aggregation of particles at a concentration as low as 12%. Thus, a suspension of uniform sized spherical polystyrene particles in aqueous solution of dextran may be made to mimic blood by controlling the surface charge on the polystyrene spheres using addition of calcium ions to the medium.

Blood↗
PubMed1988

Increased vascular resistance due to a reduction in red cell deformability in the isolated hind limb of swine.

This study investigated whether red cells with reduced deformability impeded flow through the microcirculation. Red cells were made less deformable in their normal biconcave disc shape by incubation with 2% formaldehyde (fRBCs). The blood supply to the right hind limb was isolated in 26 swine and the femoral artery was instrumented with two fine catheters, a flow probe, and an inflatable occluder. Flow was measured over a range of different perfusion pressures during adenosine-induced vasodilation under control conditions (C) and during an infusion of fRBCs at 1 ml/kg per minute (not to exceed 20 ml) into the femoral artery. At the same perfusion pressure (P), flow was significantly reduced 5 min after the fRBC infusion: Flow at P = 20 mm Hg, C = 41 ml/min vs fRBC = 10 ml/min; at P = 40, C = 160 ml/min vs fRBC = 79 ml/min; and at P = 60, C = 278 ml/min vs fRBC = 147 ml/min, with P less than 0.02 for all comparisons. Flow was still significantly reduced 15 min after the fRBCs, but by 30 min, it had returned to the control value. Chromium-51-labeling of red cells revealed that about one-third of fRBCs was trapped in the microcirculation compared to less than 3% of normal cells. This reduction in flow with fRBC infusion was not altered by alpha blockade, indicating that adrenergically mediated spasm was not responsible for the reduced flow. Aspirin 35 mg/kg iv completely prevented the reduction in flow despite an absence of change in the percentage of fRBCs trapped. Thus, red cells with reduced deformability infused into the circulation caused a significant, but transient, reduction in flow. The reduction in flow was not primarily related to entrapment of the abnormal RBCs, but may be mediated through platelet aggregation or release of potent vasoconstrictor substances from platelets or endothelial cells.

Adenosine↗
PubMed1987

Fractionation of K-562 cells on the basis of their surface properties by partitioning in two-polymer aqueous-phase systems.

The K-562 cell line is a culture of human leukemia stem cells originally derived from a patient with chronic myelogenous leukemia in blast crisis. We have subjected such cells, in the log phase of growth, to countercurrent distribution in a charge-sensitive dextran-polyethylene glycol aqueous-phase system, a method that fractionates cells on the basis of subtle differences in their surface properties, and found that: (1) The cell population is heterogeneous since it is composed of cells with different partition ratios. (2) There is a correlation between increasing cell partition ratios and increasing cell electrophoretic mobilities. (3) Cells under different parts of the distribution curve have dissimilar ratios of cells in different parts of the cell cycle, a phenomenon that may, at least partially, be the basis for the subfractionation of these cells. There is a clear tendency for cells in G0 + G1 + early S to decrease and for those in late S + G2 + M to increase with increasing partition ratios. (4) Sialic acid is a major surface charge component of the cells as evidenced by a dramatic drop in their partition ratios after treatment with neuraminidase.

Cell Cycle↗
PubMed1986

Effect of surface modification of rat erythrocytes of different ages on their partitioning behavior in charge-sensitive two-polymer aqueous phases.

Partitioning differences between cells in two-polymer aqueous phase systems originate from subtle differences between the surface properties of cells. Because of the exponential relation between the parameters affecting the partition ratio (P) and the P itself, differences in membrane components suspected of effecting the differential partitioning of closely related cell populations cannot be directly established by conventional chemical assay techniques. In order to study the chemical nature of the components responsible for the age-related changes in surface properties of rat red cells we have devised an approach which uses a combination of isotopic labeling of erythrocyte subpopulations of distinct cell age with different enzyme and/or chemical treatments followed by countercurrent distribution in charge-sensitive two-polymer aqueous phase systems. These studies show that: neuraminidase-susceptible sialic acid is not responsible for the cell age-related surface differences detected by partitioning; the component(s) responsible for the cell age-related surface differences can be extracted (from aldehyde-fixed red cells) with ethanol or cleaved with dilute sulfuric acid. Our data are consistent with the hypothesis that ganglioside-linked sialic acid is the chemical moiety responsible for the cell charge-associated surface differences among rat red blood cells of different ages.

Age Factors↗
PubMed1984

Polystyrene latex particles as indicators of abnormal plasma properties in multiple sclerosis.

Polystyrene latex particles were incubated in plasma from multiple sclerosis (MS) patients or normal subjects. Compared with plasma from healthy controls, MS plasma produced a significantly lower mean electrophoretic mobility of the latex particles after the addition of linoleic acid. The effect was similar to that found when erythrocytes were used as the indicator particle. However, the result with latex particles demonstrates that the phenomenon originates from a plasma component of MS patients rather than from an intrinsic difference in membranes of the red blood cell.

Culture Media↗
PubMed1983

The effect of neuraminidase on the relative surface charge-associated properties of rat red blood cells of different ages.

Approx. 70% of the sialic acid on the rat erythrocyte surface is susceptible to cleavage by neuraminidase (Vibrio cholerae). Neuraminidase treatment results in a reduction in the partition coefficient (K) of the red cells in a charged dextran-poly(ethylene glycol) aqueous phase system and in the electrophoretic mobility of the cells. Countercurrent distribution of rat neuraminidase-treated erythrocytes, containing 59Fe-labeled mature red cells of distinct age, indicates that (a) the electrophoretic mobilities of red cells in different cavities along the extraction train increase with increasing K, as is the case with untreated erythrocytes, and (b) the cell age-related differences in surface charge-associated properties are neither eliminated nor altered by the enzyme action.

Animals↗
PubMed1983

Action of hydroxyethyl starch on the flow properties of human erythrocyte suspensions.

Hydroxyethyl starch (HES) has often been used as a plasma expander, but questions still remain concerning the mechanisms by which it produces changes in the rheological properties of blood and erythrocyte (RBC) suspensions under various flow conditions. The present investigation has shown that the dynamic viscosity of HES (232,000 and 565,000 daltons) solutions rises in a nonlinear fashion with increasing HES concentration, and for a given concentration of HES exhibits Newtonian behavior at shear rates between 0.15 to 124 sec-1. At low (less than 0.9 sec-1) shear rates the apparent viscosity of a 40% RBC suspension increases with lower concentrations of HES because of RBC aggregation. At higher concentrations of HES, increases in suspension viscosity are due to an increase in the viscosity of the continuous phase since the RBC are largely disaggregated. At high (greater than 36 sec-1) shear rates the relative viscosity (eta/eta O) of RBC suspensions slowly decreases with increasing HES concentration. At low shear rates eta/eta O increases and then decreases with increasing HES concentration. Evidence of the concentration-dependent effects of HES on RBC aggregation is provided not only by the viscometric analysis but also from measurements of erythrocyte sedimentation rate (ESR) and the zeta sedimentation ratio (ZSR). HES is a more potent aggregating agent in phosphate buffered saline (PBS) than it is in plasma. Polymer size has only a slight effect on the extent of RBC aggregation produced, but does have a significant effect on the concentration of polymer at which maximum aggregation occurs. The viscosity-corrected electrophoretic mobility of RBC in HES rises monotonically with the concentration of HES in the suspending medium. Decreases in the extent of RBC aggregation with increasing polymer concentrations probably result from an increase in the electrostatic repulsive forces between the cells.

Blood Sedimentation↗
PubMed1983

Plasma protein interactions at biological interfaces.

The adsorption of plasma proteins, especially fibrinogen at the endoendothelial interface and to the surfaces of the blood cellular elements in contact with blood is relevant to both hemostasis and thrombogenesis. The intact endothelium is the most thrombo-resistant surface known and the exact nature of this structure holds the answer to developing adequate biocompatibility for implants and prosthetic devices. Under physiological conditions the endoendothelial surface, blood cellular elements and plasma proteins all possess net negative charges which will play a role in their interactions. The density and charge sign of ionizable surface groups will determine the local value of pH at that surface. Distributions of surface charge are possible which could lead to local pH values which vary from one region to another by several units of pH. Erythrocytes and platelets possess adsorbed fibrinogen and the endothelial surface is likely also to attract fibrinogen. The presence of one or more layers of fibrinogen on the endothelial lining is, therefore, a well founded physicochemical expectation. Aggregation of the blood cellular elements is mediated by calcium ions, fibrinogen and other agents depending upon the cell type. The presence of fibrinogen is essential for platelet aggregation induced by adenosine diphosphate and for the observed changes in platelet surface charge. There is evidence that in general electrostatic interactions are important in the contact relationships between the blood cellular elements and other surfaces either natural or foreign.

Adenosine Diphosphate↗
PubMed1982

Early electrochemical events in lectin-lymphocyte interaction. I. Mechanism of lectin-mediated changes in the electrophoretic mobility of lymphocytes.

Concanavalin A, at extremely low concentrations, will produce significant increases in the electrophoretic mobility of murine splenic T lymphocytes. It has been established that the alteration in cellular surface charge is mediated by a factor produced by those lymphocytes that have reacted directly with con A. We originally conjectured that the mobility change might be the consequence of an alteration in the distribution of the charged moieties of membrane glycoproteins. The results of experiments conducted at low temperature raise some questions about this mechanism. Further experiments have been performed to establish the nature of the physicochemical alterations in the peripheral zone of the factor-stimulated lymphocytes that are manifest as changes in cellular surface charge. The results of these studies indicate that, subsequent to the interaction of T lymphocytes with con A, there is a reduction in the number of positively charged amino groups effective at the electrophoretic surface of the cells.

Animals↗
PubMed1982

Evaluation of density gradient separation methods.

A new method for evaluating the effectiveness of isopycnic separation processes is developed and explored. This technique is designed to measure the ability of a separation technique to attain the goals of a sorting process. The performance of a separation process is expressed in terms of an inconsistency number, I, whose value ranges from zero to unity. Processes which function perfectly are characterized by an I value of zero. Separation processes which merely subdivide a sample are characterized by an I value of unity. The validity of using I to measure the performance of separation processes was established by demonstrating that alterations in the performance of a well characterized separation technique were fully reflected by changes in the I value for the process. The capacity of four different techniques for ordering red cells with respect to their density was then shown to lie in a definite hierarchy. This hierarchy can be expressed as: Murphy technique (I = 0.21) greater than albumin density gradient (I = 0.43) greater Stractan density gradient (I = 0.53) greater than Percoll density gradient (I = 0.73). The performance was found to be dependent on the exact operating conditions employed and the specific fraction of red cells isolated.

Cell Separation↗
PubMed1981

The automated analytical electrophoresis microscope.

The components of an automated computer-controlled analytical electrophoresis microscope (AEMS) are described. Computer tracking of migrating cells projected under phase contrast onto a vidicon permits the rapid taking of multiple velocity measurements per cell so that reliable determinations of electrophoretic mobilities thereby result. The computer-controlled cell search and tracking algorithms allow high speed operation so that statistically valid profiles and data bases can be collected rapidly. Initial electrophoretic mobility evaluations have been carried out on populations of lymphocytes, erythrocytes, and platelets.

Animals↗