R-3 Repository :: Browsing by Author "Hellums, Jesse D."

Browsing by Author "Hellums, Jesse D."

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A study of the effects of shearing stress on normal human blood platelets
(1973) Lemuth, Richard F.; Hellums, Jesse D.
As the disappearance of platelets in extracorporeal circulation has been established as a problem in surgical and experimental applications, an investigation was undertaken to determine the fragility of normal human blood platelets. A study of recognized procedures for platelet anticoagulation and storage dictated handling procedures and preparation of a platelet-rich plasma (PRP). An attempt was made to eliminate variables of platelet function—aggregation, adhesion, etc.—by careful choice of donors and adherence to recommended handling techniques. Viscometry equipment, originally used for shear of whole blood, was modified for application to PRP. Analyses were developed for determination of platelet destruction as a result of shear stress. Release of the platelet enzyme acid phosphatase and Coulter Counter platelet enumeration provided consistent, if not exactly comparable, destruction data. Phase microscopy platelet enumeration was not successful in determining platelet destruction. Results of applied shear to the PRP indicated a critical threshold shearing stress for platelet destruction in the range of 15-2 dynes/cm. This threshold is approximately one-tenth of that found for red blood cells through previous experimentation in this laboratory. Through varying of viscometer configuration, it was determined that the platelet shear destruction was not surface dependent. It was, however, increased by turbulent flow. A secondary set of results was obtained when a "storage lesion" developed prior to shear in ineffectively anticoagulated PRP. With appearance of this lesion, platelet destruction was drastically increased and inconsistent. The storage lesion correlated with increased acid phosphatase level in the platelet-poor plasma controls and with platelet aggregation observed after shear.
Effects of antiplatelet agents on polymorphonuclear leukocytes exposed to mechanical trauma
(1981) Rothberg, Jeanne Mays; McIntire, Larry V.; Hellums, Jesse D.; Martin, R. Russell
The effects of shear stress on polymorphonuclear leukocytes (PMNs) were investigated to ascertain whether mechanical trauma induces lysosomal granule release or membrane lysis. In addition, the effects of antiplatelet agents on PMN leukocyte response to shear stress were studied. The antiplatelet agents, known to elevate intracellular and cyclic adenosine monophosphate (cAMP), included the following: prostaglandin Ej (PGEi), prostaglandin I2 (PGI2), theophylline, RA-8 (dipyridamole), and the dipyridamole analog, RA-233. PMN leukocyte suspensions were exposed to shear stresses of 1 or 3 dynes/cm2 at room temperature (23°C) for 1 minutes. Leukocyte responses were monitored by the following measurements. The electronic particle count was measured to indicate shear-induced aggregation and lysis. 8-Glucuronidase in the extracellular fluid was assayed to indicate azurophilic granule release. Finally, lactic dehydrogenase release was measured to indicate cell lysis due to PMN exposure to shear stress. Results of the shear-induced effects on PMNs without drug treatment show that cell loss, granule release, and cell lysis increase with increasing shear stress. Furthermoré, PMNs exposed to a shear stress of 1 dynes/cm2 release approximately equal percentages of 3-glucuronidase and lactic dehydrogenase. Percent cell loss approximates enzyme release indicating cell disruption at this shear stress. At a shear stress of 3 dynes/cm2, 3-glucuronidase release is greater than LDH release indicating that the PMNs are expelling granules into the extracellular fluid at this higher shear stress without concomitant cytoplasmic enzyme release. Pretreatment of PMNs with PGEi plus RA-233 or PGEi plus theophylline produces an approximately 25% reduction of cell loss and release of 3-glucuronidase and LDH release after exposure to a shear stress of 3 dynes/cm2. No significant effects of PGEi plus RA-233 are observed at 1 dynes/cm2. Pretreatment of PMNs with PGI2 plus RA-233 produces no reduction of PMN response to mechanical stimuli. PMNs incubated with dipyridamole and exposed to a shear stress of 3 dynes/cm2 show a significant reduction in cell loss, granule release, and lysis. The suppressive effect of dipyridamole appears to be additive with the effect of PGEi. Leukocytes pretreated with PGEi plus dipyridamole and exposed to shear stresses of 3 dynes/cm2 show a greater decrease of cell loss and enzyme release than pretreatment with dipyridamole alone or PGEi plus a phosphodiesterase inhibitor.
Failure theory for red blood cells in couette flow
(1973) Jones, Jack A.; Hellums, Jesse D.
In this analysis, the red blood cell is idealized as a spherical liquid drop, encapsulated by a thin membrane. By transforming G. I. Taylor's equations for the surface loading on a liquid drop with no membrane in Couette flow, spherical loading components are calculated for a drop with a membrane. A solution developed by Fliigge for unsymmetrical loading of spherical shells is then used to derive the complete membrane stress pattern for spherical, membrane-encapsulated, liquid drops in couette flow. The maximum distortion energy is calculated for any given shear rate, and using the relation, a critical shear stress for short duration failure is found as a function of the critical distortion energy. Viscoelastic solutions based on the maximum normal strain failure theory and the maximum stress failure theory are then developed in order to predict the entire critical shear stress vs. time curve for hemolysis. Using data supplied by Rand's micropipette experiments on red cell membrane strength, the theories are then shown to roughly predict the entire couette flow hemolysis curve for times up to 1 seconds. Thus, the theory in this analysis provides a much greater understanding of the mechanism causing red blood cell damage in artificial heart valves, heart-lung machines, artificial kidneys, etc. In addition, due to the model cell's physical generality, the theory is applicable to other biological and non-biological systems.
Fixation of platelet aggregate size distribution
(1981) Goldblum, David K.; Hellums, Jesse D.; Solis, Robert Thomas; Rowley, Richard L.
An experimental study has been carried out on the use of aldehydes for fixation of human platelet aggregate size distributions. The objective of the work was to develop a methodology of stopping aggregation and disaggregation processes for subsequent analysis. The results are intended to facilitate study of rates of aggregation and disaggregation as influenced by various stimuli. Platelet aggregation was induced in citrate-anticoagulated platelet-rich plasma (PRP) by addition of adenine dinucleotide (ADP) in final concentration ranging from .5 to 2 yM. The aggregated PRP specimens were diluted (158.5 to 1) in a counting medium (isoton) for size distribution analysis. An electronic particle counter was used to study the aggregate size distributions in the range 13-11 urn in equivalent spherical diameter. Parameters used to monitor the size distributions were cumulative volume and cumulative population of the aggregates, mean aggregate size, and volume available for aggregation from free (unaggregated) platelets. In preliminary studies evidence was obtained that glutaraldehyde was a more promising fixative than formaldehyde. Glutaraldehyde in appropriate concentrations caused no important problems in resuspension or in aggregate size change for times of fixation of several minutes. Dilution of aggregated PRP specimens in isoton for counting induced rapid disaggregation. However, it was found that this disaggregation could be avoided by use of glutaraldehyde in the isoton counting diluent. Glutaraldehyde addition to both the aggregated PRP specimen and to the isoton counting diluent to final concentration of .48 wt% was selected as the recommended procedure. Detailed studies were made of aggregate size distributions fixed at various times in the aggregation process. The results indicate that the fixative stops the reactions and stabilizes the distribution for times of 3 to 5 minutes. Thus, the procedure should be useful in studies on rates of platelet aggregation.
Mantle convection at marginal stability
(1979) Warford, Andrew Craig; DeBremaecker, Jean-Claude; Hellums, Jesse D.; Lallemant, Hans G. Avé
The horizontal extent of convection cells in the earth's mantle can be estimated from the geometry of plate boundaries. The vertical dimensions can perhaps be estimated from the theory of marginal stability in variable viscosity fluids. For viscosity laws symmetrical about mid-depth the aspect ratio increases with increasing viscosity contrast, but the law of variation with depth has little effect. The value of the Rayleigh number is affected by both the viscosity law and the contrast. The aspect ratio for the asymmetric cases studied is much less affected except at very high contrasts (>3) and then only in the case of an exponentially varying viscosity. In all cases studied, the variation of the Rayleigh number with wavelength is smaller as the viscosity contrast increases, thus allowing for a fairly wide range of aspect ratios. The variation of velocity with depth indicates that motion takes place in the entire depth range except in the case of viscosity decreasing exponentially with depth and then only at high viscosity contrast (>2).
Mathematical modelling of oxygen transport in the microcirculation
(1985) Yap, Eddy W. H.; Hellums, Jesse D.; McIntire, Larry V.; Olson, John S.
Many mathematical models of the transport of oxygen in the microcirculation have been proposed. It is the aim of the present study to examine the validity of some of the many simplifying assumptions that have been used. Due to the many experimental difficulties,mathematical modeling has proved most useful in elucidating the effect of the physical and chemical processes on. the oxygen trasport of the microcirculation. The validity of using the simpler variable rate coefficient kinetic model compared to the complicated but realistic Adair model is studied. It is found that the variable rate coefficient model is adequate for the normal physiological range of fluxes. The choice of which kinetic models to use only becomes important at high fluxes i.e. in the reaction kinetics rate-limiting regime. The neglect of oxyhemoglobin diffusion in aiding the oxygen transport is found to be valid only at high fluxes where again the rate-limiting step is the chemical kinetics. For physiological range of fluxes, the neglect of oxyhemoglobin diffusion introduces substantial errors in the estimation of the resistance to oxygen transport in the capillary. The simpler mathematical model incorporating the simplifying assumption of local chemical equilibrium and no hemoglobin diffusion predicted the capillary wall oxygen concentration within an error of 16% but gave large errors in predicting the resistance to oxygen transport in the capillary. The mixed-mean and space-averaged oxyhemoglobin concentrations for a hemoglobin solution flowing through the capillary are compared and found to be significantly different. Solutions for the simulation of oxygen transport from a hemoglobin solution flowing through the capillary in a parabolic velocity profile is presented in the form of the Nusselt number against the Graetz number. The Nusselt number from an in-vitro experimental capillary is compared to that calculated from a model simulating the experimental and is found to agree very closely.
Oxygen transport in hemoglobin solutions: applications in the microcirculation
(1979) Sheth, Bhupendrakumar V.; Hellums, Jesse D.; McIntire, Larry V.; Olson, John S.
Numerical solutions are obtained for the unsteady-state oxygen release process from hemoglobin layers of thicknesses and concentrations typical of the microcirculation. A modification of the CrankNicolson scheme is used to solve the coupled, non-linear, parabolic partial differential equations in layers of thicknesses 1.6 u, and 6. u. Two types of boundary conditions on oxygen are employed: 1) constant concentration, and 2) constant flux. Analytical solutions are presented for two cases: 1) oxyhemoglobin dissociating irreversibly to give oxygen and hemoglobin, and 2) oxygen and oxyhemoglobin assumed always in chemical equilibrium, the saturation curve being a straight line. The numerical results are presented as Nusseltt number versus time required for deoxygenation. Oxyhemoglobin diffusion accelerates the oxygen release process. The facilitation increases with decreasing oxygen concentration. It is shown that the assumption of oxyhemoglobin-oxygen equilibrium leads to significant error for conditions of the microcirculation. Thus the chemical reaction rate constants must be taken into account. Furthermore, it is shown that the reaction rate constants must be taken into account in a way compatible with the known equilibrium relationship. The analytical solutions are shown to differ significantly from the numerical results, but are useful for simple order of magnitude estimates.
Rheometric studies on the in vitro polymerization of microtubules
(1984) Nelson, Rickey L.; McIntire, Larry V.; Hellums, Jesse D.; Armeniades, C. D.
Microtubules form weakly associated networks with many properties similar to networks of rigid rods. To study the in vitro polymerization of bovine brain tubulin, a Fluids Rheometer was used in the oscillatory mode. The maximum elastic modulus for microtubule networks in the standard PIPES buffer was observed to be proportional to the tubulin concentration over much of the investigated range. A maximum elastic modulus of 2 dynes/cm2 occurred at a tubulin concentration of 8 mg/ml. At even higher concentrations, a decrease in the maximum elastic modulus was observed. Other additions to the standard buffer primarily affected the interactions between microtubules as opposed to shifting the overall polymer-heterodimer equilibrium. 9% deuterium oxide increased the maximum elastic modulus by a factor of 4, while 25% glycerol led to approximately a doubling in the maximum elastic modulus when compared to microtubule networks in the standard buffer. Microtubule associated proteins (MAPs) were shown to inititate the formation of microtubules when added back to purified tubulin solutions. Even though more microtubules were formed as the MAPs were added, the elasticity decreased with increasing MAPs additions because shorter, more numerous microtubules were present. The interactions between bovine brain tubulin and rabbit muscle actin were also investigated. The resulting networks were found to be extremely strain sensitive but had elastic moduli considerably greater than the sum of the individual components. The MAPs fraction was found to cross-link actln filaments with the maximum interaction occurring at .3 mg MAPs/mg actln, corresponding to 1 mole of MAPs per 23 moles of actin. The extent of phosphorylation state of the MAPs also affected the interaction. At an actin concentration of 1 mg/ml, MAPs with a lower phosphate content would form a network with a maximum elastic modulus of 4 dynes/cm. However, at approximately the same actin and MAPs concentrations the maximum elastic modulus from MAPs with a higher phosphate content was 25% lower at 3 dynes/cm.
Shear induced tissue factor activity in human leukocytes
(1983) Pankowsky, Dan A.; McIntire, Larry V.; Hellums, Jesse D.; Armeniades, C. D.
There is a growing amount of evidence a significant link exists between the body's immune system and blood coagulation pathways. Specifically, the monocyte, a peripheral blood leukocyte, has been found to be the source of leukocyte tissue factor activity (TFA). This TFA activates the extrinsic blood clotting pathway. It is becoming apparent that there are two pathways for monocyte TFA production a lymphocyte dependent pathway and a lymphocyte independent pathway. Problems of blood clotting and thrombus formation in clinical situations include the use of extra-corporeal circulation and artificial heart valves. In addition disease states such as atherosclerosis and rheumatic heart disease are often associated with (or in some instances possibly caused by) coagulation abnormalities. These disease states also have in common non-physiologic levels of shear stress applied to the blood. The possibility exists that there is a significant link between the applied shear stress in the blood and the development of pathological coagulation. Since monocytes are known to be a source of TFA in response to various stimuli, this study was undertaken to measure monocyte TFA in response to shear stress. It was found that mechanical trauma could induce tissue factor production and that shear-induced TFA required the presence of lymphocytes. That the functional capability to produce TFA was there after shear stress in cells not producing TFA was demonstrated by subsequent treatment with endotoxin. Therefore, in clinical situations where mechanical trauma to leukocytes is a problem, the possibility of leukocyte TFA needs to be considered.
Shear stress effects on epithelium cell cultures in vitro
(1983) Stathopoulos, Nikos A.; Hellums, Jesse D.; Armeniades, C. D.; McIntire, Larry V.
This study aims to determine the influence of shear stress on cell cultures under carefully controlled conditions. Human embryonic kidney cells grow^as an attached, confluent monolayer on a flat substrate were subjected to steady, uniform laminar flow in a specially designed flow chamber, in which shear stress was precisely defined and controlled. Experiments performed for shear stresses ranging from .2 to 6. N/m, with a varied shear stress exposure time from 2 to 24 hours. The influence of the shear field was slight at low shear stress (.26 N/m) but it had a dramatic effect on cell morphology at higher shear stresses (above 2.6 N/m ) and long exposure times (24 hours). It was also found that shear stress and time stimulated the release of urokinase enzyme. The significance of such flow studies is presented in relation to their possible use for biosynthesis of urokinase enzyme by kidney epithelial cells/
The effect of platelet anti-aggregating Agent RA-233 and prostagland in E1 upon PMN leukocyte response to mechanical trauma administered in vitro
(1980) Stockwell, Douglas James; McIntire, Larry V.; Martin, R. Russell; Hellums, Jesse D.
The effect of platelet anti-aggregate agent RA-233 and prostaglandin upon PMN response to mechanical trauma was studied using the Rice University ROM-8 viscometer. Suspensions of PMN leukocytes in phosphatebuffered saline (PBS) were sheared at 1 and 3 dynes per square centimeter for ten minutes in a 37°C environment. The leukocyte suspensions were tested for alterations in total leukocyte count, changes in morphology, and variances in the PMN differential count. PMN leukocyte function was tested using several adapted assays including adhesion to glass slides, phagocytosis, and release or loss of enzymes marking the various granule populations present in PMN cell cytoplasm. Leukocytes which received 1 x 1^-6M RA-233 and 3 x 1^-6M PGE1 ten minutes prior to shearing, showed significant preservation of enzyme activity marking specific granules and reduced release of enzyme activities marking both specific and azurophilic granules, while control suspensions showed losses of enzyme activity agreeing with the results of earlier work by Dewitz (4). PMN phagocytosis, as measured by a chemiluminescence assay was significantly decreased by shear stresses of 1 and 3 dynes/cm, and was not preserved by incubation with the anti-platelet agents. PMN leukocyte adhesion to serum-coated glass slides after shear stress exposure was increased while PMN leukocytes incubated with RA-233 and PGE1 showed reduced overall adhesion in the controls, but similar increases with shear stress exposure. PMN leukocyte suspensions showed a sharp drop in electronic particle count, (EPC), following shear stress exposure, due to the combined effects of aggregation and lysis. The latter effect was apparently reduced at the higher shear stress levels by incubation with the anti-platelet agents. These results and the observations of several other groups, reviewed in the first section, lead us to propose a possible leukocyte preservational effect for the two platelet antiaggregating agents tested. The significance of shear stress trauma is reconfirmed once again by observations made in this series of experiments. The possibility that mechanical trauma may be a mediator of leukocyte dysfunction during extracorporeal circulation is discussed in the light of this new experimentation.
The effects of oxygen tension, ATP concentration, and incubation in various serum solutions on shear stress-induced hemolysis
(1976) Tadano, Ken; Hellums, Jesse D.
The red blood cell is susceptible to damage by foreign surface interaction and hydrodynamic shear stressing; both of these hemolytic sources are present in blood processing machines which are used to treat chronic and acute disorders. The present investigation attempts to elucidate several facets of the erythrocyte degradation process produced by hydrodynamic shear stress. In this study, a constant shear stress viscometer was used to subject normal human erythrocytes suspended in altered serum environments to known shearing forces. The response of the cells was determined by hemolysis measurements. This study examined 1) the effect of oxygenation on hemolysis, 2) the effect of ATP concentration on hemolysis, and 3) the effects of incubation in various suspending mediums upon erythrocyte characteristics and the relationship between these altered characteristics and cell trauma.
The effects of shear stress on the clottability of fibrinogen in plasma
(1975) Peters, Bruce Craig; Hellums, Jesse D.
One problem encountered in the use of circulatoryassist devices has been alterations of the coagulation mechanism. This study was made to determine if one source of this coagulation problem could be due to increased shear stress in the extracorporeal systems. A plasma pool was collected from thirteen donors and lyophilized to provide identical plasma samples for all experimental results. Tests were made with various shear stress and shear exposure times to determine the effects of these variables. A constant shear stress viscometer was used in this examination, and two systems with different surface to volume ratios were available. The surfaces were siliconized before the plasma was introduced to the viscometer. To determine the magnitude of any surface losses, tests were made with radioactive Iodine (I125) labeled fibrinogen. The tests with the xa.l>eXed fibrinogen showed that no loss of fibrinogen occurred due to adhesion to the surface. Studies made with constant time exposures and varying shear stress indicated that clottability does decrease with increased shear stress up to 1 dynes/cm, at which point an apparent minimum clottability occurred. This loss was attributed to conformational changes causing steric interferences with polymerization sites. The minimum was considered to be caused by a maximum in the steric hindrance of the polymerization sites due to conformational changes. Studies done at constant shear stress, varying time indicate that loss of clottability is a function of both exposure time and shear stress. Variation of the surface to volume ratio suggested that the clottability loss is probably surface dependent.
The response of human blood platelets to shear stress in rotational and capillary viscometers
(1985) Roper, Vaughan Phillip; Hellums, Jesse D.; Philpott, Charles W.; McIntire, Larry V.
Part 1 of this study concerns the dependence of shear-induced platelet aggregation on platelet concentration. Uniform shear stesses of 5, 75, 1, 15, 2, and 25 dynes per square centimeter were applied to platelet suspensions by Couette flow viscometry for 3 seconds. Relative-Platelet-Aggregation (RPA) is defined as the number of platelets aggregated during shear divided by the number present before shear. Whole blood from a group of normal, healthy, non-smoking human subjects was centrifuged to prepare platelet plasma with various platelet concentrations. Aggregation was determined by electronic particle counts both before and after the specimens were exposured to shear in the viscometer. RPA was found to be proportional to initial platelet concentration at all shear stresses studied. RPA increased with shear stress up to 15 dynes per square centimeter, where RPA was a maximum (82 percent for a platelet concentration of 31, per microliter). Shear stresses higher that 15 resulted in lower levels of aggregation (57 percent at a stress of 25). Part IX outlines the procedures and equipment needed to investigate shear stress-induced platelet reactions at very small times of specimen exposure to shear. A review of the role of ionic calcium as a platelet activator is presented. Internal platelet calcium can be localized and quantitated by electron microscopy. Measurement of calcium redistribution can be used as an aggregation stimulation indicator. A capillary viscometer-nitrogen freezer apparatus and freeze drying system were designed and constructed to prepare specimen for these purposes.
The response of normal human blood platelets to exposure to 2.45 GHz continuous wave radiation
(1980) Piana, Martha Louise; Hellums, Jesse D.; Wilson, William L.; McIntire, Larry V.
Human blood platelets in platelet-rich plasma were irradiated at room temperature with 2.45 GHz continuous wave microwave radiation in a rectangular waveguide chamber for an exposure duration of two hours. Measurements of the change in platelet count, release of lC-radiolabeled serotonin, increase in the level of LDH activitiy in the supernatant platelet-free plasma, alteration in the aggregation response to exogenous ADP, and decrease in the intracellular potassium ion concentration were assayed to determine any damage to the platelets. In all experiments, microwave heated platelets were compared with platelets warmed to the same temperature by use of an adjustable block-heater and with platelets maintained at room temperature. Preliminary studies indicated that the incident power flux was not a crucial factor for the power fluxes considered, 3.35 to 2.12 mW/cm^2. The subsequent experiments were performed at the maximum power flux of 2.12 mW/cm^2, which corresponds to an absorbed power of 112.62 mW/cm^2. At this power flux, the final temperature was 41°C. There were no significant differences between the block-heated and microwaved samples in the release of serotonin, the increase in LDH activity, and the decrease in intracellular potassium ion concentration . Significant differences were observed between the block-heated and microwaved samples in the platelet count and in the aggregation response to exogenous ADP. However, these observed differences were small, and there was more alteration of platelet response from block-heating than from irradiation. Therefore, it may be concluded that the observed response of human blood platelets exposed to 2.45 GHz continuous wave radiation at a power level of 112.62 mW/cm^2 is entirely due to temperature elevation caused by energy absorption.
Variations in glucose diffusion coefficients in biological flocs
(1974) Pipes, David Meggs; Characklis, William G.; Hellums, Jesse D.; Kessick, Michael A.
The diffusion coefficient of glucose was measured in biological flocs under various growth conditions. Significant differences were observed with different carbon sources and carbon to nitrogen ratios. The diffusion coefficients were calculated from direct measurements of mass flux through a series of membrane filters separated by a metal template which contained the floe. The template assembly was prepared by packing the floe material into the template and placing a .45|j, membrane filter on either side of the template. The assembly was then placed in a modified plastic Millipore filter apparatus having the lower portion filled with a glucose solution (1 mg/1) and the top section containing deionized water. Increasing glucose concentration in the top was related to mass flux through the floe by measuring the flux through the membrane filters separately. Measurements were replicable within 8%. A 2^3 factorial analysis, using carbon source (glucose or methanol), carbon to nitrogen ratio (5:1 or 5:1), and temperature (2°C or 3°C) as independent variables, indicated significant effects at the 99% level for all three variables.

Browsing by Author "Hellums, Jesse D."

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