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Biomedical subjects

E F Leonard

Publications and source records attributed to E F Leonard.

At least 19 recordsLinked to original sources
PubMed2004

Adjustment of dry weight in hemodialysis patients using intradialytic continuous multifrequency bioimpedance of the calf.

BACKGROUND: Current concepts of dry weight (DW) prescription are largely based on clinical symptoms because of the difficulty in assessing extracellular fluid volume (ECV) during dialysis. Intradialytic changes in ECV can be recorded as changes in extracellular resistance [Re] by continuous regional calf multifrequency bioimpedance spectroscopy (BIS). We hypothesized that relative changes in calf Re (Re at time '0' over Re at time 't' [R(e-0)/R(e-t)]) will become very small when ECV is reduced towards normal and individual dry weight is reached. METHOD: Intradialytic continuous calf BIS was recorded repeatedly in 15 hemodialysis (HD) patients. The first measurement was performed at the prevailing clinical dry weight (CDW). Next measurements were made after post-HD body weight was gradually decreased by 0.2-0.3 kg per treatment. This procedure was iterated over several subsequent treatments until a treatment was observed where changes in R(e-0)/R(e-t) were < 1%. The weight at the end of this treatment was defined as "achieved dry weight (ADW)". Each R(e-0)/R(e-t) curve was fitted using a Matlab program (curve fitting toolbox) to obtain the exact weight at 20 min after beginning of the flattening of the R(e-0)/R(e-t) slope ('dry' weight estimated from BIS, DW-BIS). RESULTS: Both mean ADW (80.5 +/- 34.1 kg) and mean DW-BIS (80.6 +/- 34.1) were significantly lower than CDW (81.4 +/- 32.0 kg, p < 0.001), but there was no difference between ADW and DW-BIS. However, the average weight reduction from CDW to ADW (0.80 +/- 0.15 kg) was significantly higher than from CDW to DW-BIS (0.66 +/- 0.14 kg, p < 0.001, paired t-test). When ADW was achieved, pre-dialysis systolic blood pressure (SBP) was lower than at CDW (139.3 +/- 32.5 mmHg, vs. 129.4 +/- 33 mmHg, p < 0.05), post-HD SBP did not differ. The incidence of clinical symptoms of underhydration was similar at CDW (15%) and DW-BIS (15%), but higher at ADW (46%). CONCLUSION: Intradialytic continuous calf BIS allows the assessment of changes in extracellular calf resistance as an indicator of changes in extracellular fluid volume. Recording of a continuous R(e-0)/R(e-t) slope during dialysis appears to be a promising new tool for the prediction of dry weight in hemodialysis patients.

Analysis of Variance↗
PubMed2001

Controlling duration of contact between T cells and antigen-presenting cells.

A new method which allows precise control of the duration of contact between T cells and antigen-presenting cells (APCs) has been developed. A glass coverslip coated with poly-L-lysine, and then with T cells, was placed at the base of a cylindrical well, and the well was filled with liquid medium. A round coverslip, on which APCs were adhered, was supported on the surface of the medium by surface tension, cell-side down. By withdrawing medium from four capillary holes near the base of the well, the coverslip could be lowered to initiate contact between T cells and APCs at a defined time zero. The contact was broken at desired time points by re-introducing medium into the well in order to separate the two coverslips. Each cell type remained adherent to its original surface after separation for all contact times studied. The T cells were monitored for intracellular calcium mobilization using the fluorescent dye, Fura-2. Contact durations of less than 1 min did not trigger calcium signals. Contact durations of 3 and 5 min induced strong calcium signals. Breaking the contact caused a rapid decrease in intracellular calcium levels. This method of cell manipulation allows precise control of the duration of contact of T cells with APCs, while keeping the cells under continuous observation. The measurements so obtained provide a quantitative understanding of the dynamics of early T cell activation.

Animals↗
PubMed2001

Controlled cell deformation produces defined areas of contact between cells and ligand-coated surfaces.

A method which allows precise control of the time of initiation and the area of contact of T cells with immobilized ligands has been developed. Cells are trapped in an asymmetric film that can be quantitatively thinned by reducing the film's capillary pressure. Ligands adsorbed to the base of the apparatus are forced into close contact with the cells as the air-liquid interface is drawn down. Using interference microscopy and microbeads to indicate the film height, the amount of thinning can be controlled to within 1 microm. In this study, this system was used to produce contact areas of 182 and 356 microm2 between T cells and anti-CD3 coated surfaces. These contact areas were measured using fluorescent dye exclusion microscopy. This apparatus can be used for quantitative studies of T cell activation, as is reported in Patrick et al., J. Immunol. Method. 24:97-108, 2000.

Animals↗
PubMed2001

Modeling of early events in T cell signal transduction after controlled T cell activation by peptide major histocompatibility complex.

Calcium signaling was observed in murine T cells over time, starting at a precise moment of contact with a layer of fibroblasts expressing a stimulatory major histocompatibility class II-peptide complex. The contact was controlled by a film-thinning apparatus. Intracellular calcium levels were followed with the ratiometric dye, Fura-2. The calcium response was highly synchronized and well fitted by a mathematical model. The model includes three components: a sequence of reactions occurring after T cell receptor (TCR) triggering; InsP3-mediated calcium release from intracellular stores (Meyer and Stryer, Proc. Natl. Acad. Sci. USA 85: 5051-5055, 1988); and slow changes in levels phospholipase C-gammal (PLCgammal) reflecting a decrease in receptor triggering rate. Each component in the model controls a different part of the response-the initial delay, the sharp rise, and the slow decay, respectively. Kinetic parameters determined from curve fitting were the initial delay in calcium signaling defined as the time when [PLCgammal] reached its half of its maximum (76 s), the coefficient characterizing calcium efflux from endoplasmic reticulum (ER) (2.86 microM s(-1), expressed per liter of cell volume), and a rate constant characterizing the diminishing yield of production of PLCgammal (0.00046 s(-1)) by active TCR. Only the parameter representing PLCgammal production varied much from cell to cell.

Animals↗
PubMed2000

Dependence of T cell activation on area of contact and density of a ligand-coated surface.

An apparatus which allows precise control of the time of initiation and the area of contact of cells with immobilized ligands has been developed. Cells are trapped in an asymmetric film that can be quantitatively thinned, forcing the cells into close contact with ligands adsorbed on the base of the apparatus. Using microbeads to indicate the film height, the amount of thinning can be controlled to within 1 microm, producing known contact areas between cells and the ligand-coated surface. This system was used with anti-CD3-coated surfaces of different densities to examine the effect of ligand density on T cell activation, while keeping the number of ligands presented to the cells constant. T cell activation was observed individually in each cell as intracellular calcium mobilization. In these experiments both the percent of T cell activation and the rate of calcium rise were found to depend only on the number of anti-CD3 molecules presented and not on their density. This implies that the spacing between molecules is not important in the range studied, and suggests that receptor clustering to levels higher than dimers may not be necessary for induction of calcium mobilization by anti-CD3.

Animals↗
PubMed2000

Model of structural and functional adaptation of small conductance vessels to arterial hypotension.

Vascular networks adapt structurally in response to local pressure and flow and functionally in response to the changing needs of tissue. Whereas most research has either focused on adaptation of the macrocirculation, which primarily transports blood, or the microcirculation, which primarily controls flow, the present work addresses adaptation of the small conductance vessels in between, which both conduct blood and resist flow. A simple hemodynamic model is introduced consisting of three parts: 1) bifurcating arterial and venous trees, 2) an empirical description of the microvasculature, and 3) a target shear stress depending on pressure. This simple model has the minimum requirements to explain qualitatively the observed structure in normotensive conditions. It illustrates that flow regulation in the microvasculature makes adaptation in the larger conductance vessels stable. Furthermore, it suggests that structural changes in response to hypotension can account for the observed decrease in the lower limit of autoregulation in chronically hypotensive vasculature. Independent adaptation to local conditions thus yields a coordinated set of structural changes that ultimately adapts supply to demand.

Adaptation, Physiological↗
PubMed2000

Evaluation of 11C-colchicine for PET imaging of multiple drug resistance.

UNLABELLED: Overexpression of P-glycoprotein (P-gp) can confer multiple drug resistance (MDR) phenotype on cancer cells and tumors by reducing intracellular accumulation of various cytotoxic agents. Early diagnosis of MDR in the clinic will serve to improve the efficacy of chemotherapeutic intervention and the quality of life of patients. In this article we describe use of a positron-emitting MDR tracer, 11C-colchicine (CHC), to evaluate MDR by PET imaging. Unlike existing MDR tracers such as 99mTc-sestamibi, this compound is electroneutral, with biodistribution not affected by perturbations of membrane potential. METHODS: In vitro studies showed that resistance to CHC is correlated to resistance to Taxol (paclitaxel). The results of biodistribution experiments were found to be consistent with previously reported experiments with CHC labeled with other isotopes. On the basis of in vitro experiments with a series of drug-resistant variants of the human neuroblastoma BE (2)-C cell line, a mathematic model of 11C-CHC distribution in tumors was formulated. Dynamic PET 11C-CHC imaging experiments were performed with nude rats xenografted with the BE (2)-C-sensitive and -resistant strains. Each scan was accompanied by a transmissions scan and a static FDG scan. These scans allowed improved image localization. RESULTS: We observed an approximately 2-fold difference between 11C-CHC accumulation in sensitive and resistant tumors. Imaging data were analyzed using the mathematic model, and various parameters characterizing resistance could be identified and estimated. In particular, the parameter r, proportional to the level of resistance of the tumors, was obtained. We showed that the ratio of these r parameters determined from the sensitive and resistant tumors was identical to the ratio of CHC accumulation in the corresponding sensitive and resistant cell lines used for xenografting. CONCLUSION: These in vivo experiments provided additional evidence for the indirect effect of P-gp action on CHC-to-tubulin binding, which in turn determines CHC uptake in tumors. The significance of these findings and future plans is discussed.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed1997

The effects of methylene blue and oxygen concentration on the photoinactivation of Q beta bacteriophage.

Concentration effects of methylene blue (MB) and oxygen on the photoinactivation rate of Q beta bacteriophage were examined. The effect of initial virus concentration was verified on the similar f2 phage. The inactivation rate, kappa, is an increasing function of MB and O2 concentration and shows saturation with respect to MB concentration. Thus the results suggest that MB must adsorb to Q beta sites and oxygen must be present for photoinactivation to occur. The inactivation rate is independent of the initial number of phage particles present before inactivation, indicating that inactivation does not depend upon interaction among viral particles or on surface effects. The results indicate that at least two different viral phenotypes exist within the wild-type Q beta and f2 populations: one susceptible and the other resistant.

Bacteriophages↗
PubMed1996

Photo-induced inactivation of viruses: adsorption of methylene blue, thionine, and thiopyronine on Qbeta bacteriophage.

The adsorption of cationic organic dyes (methylene blue, thionine, and thiopyronine) on Qbeta bacteriophage was studied by UV-visible and fluorescence spectroscopy. The dyes have shown a strong affinity to the virus and some have been used as sensitizers for photo-induced inactivation of virus. In the methylene blue concentration range of 0.1-5 microM and at high ratios of dye to virus (greater than 1000 dye molecules per virion), the dyes bind as aggregates on the virus. Aggregation lowers the efficiency of photoinactivation because of self-quenching of the dye. At lower ratios of dye to virus (lower than 500 dye molecules per virion), the dye binds to the virus as a monomer. Fluorescence polarization and time-resolved studies of the fluorescence support the conclusions based on fluorescence quenching. Increasing the ionic strength (adding NaCl) dissociates bound dye aggregates on the virus and releases monomeric dye into the bulk solution.

Adsorption↗
PubMed1996

Detection of specific plasma proteins on surfaces by immunospecific adhesion of dyed polystyrene beads.

This paper describes and evaluates a method for quantifying the amounts of specific plasma proteins adsorbed to biomaterial surfaces. In particular, it demonstrates that macroscopic images ('stains'), that assess the spatial distribution of albumin, IgG, fibrinogen, and HMK (high molecular weight kininogen), can be obtained over areas of at least 12 cm2 using immunospecific adhesion of dyed polystyrene beads. Stain intensities, measured with a scanner and an image analysis system, were found to quantify the amount of specific protein in the solution used to coat the surfaces. Results obtained with the proposed method produced single protein isotherms for albumin, immunoglobulin G (IgG) and fibrinogen that followed Langmuir-like adsorption behavior and were similar to previously published isotherms. The HMK isotherm also exhibited Langmuir-like adsorption behavior. The proposed method also detected the presence of an expected maximum in the adsorption of fibrinogen onto glass as a function of plasma dilution. Adsorption of fibrinogen out of 6.4% plasma onto glass from a separated flow produced results indicating the quantity as well as the location of fibrinogen at the boundary of the separated region. This result confirmed the utility of the proposed method for detecting spatial distributions of specific proteins adsorbed from plasma in practical devices.

Adsorption↗
PubMed1995

Non-specific adherence of oxide particles as a means of quantifying protein adsorption on surfaces.

This paper reports quantification of a method for measuring amounts of protein adsorbed to a surface; the method is especially useful for revealing macroscopic spatial patterns of adsorption. The experiments tested the effectiveness of iron oxide suspensions adsorbed onto the adsorbed protein to indicate, in separate trials, the amount of either human plasma fibrinogen or human serum albumin (HSA) present on glass slides. Corresponding trials, using radioactively labeled proteins, were performed to calibrate the amount of either albumin or fibrinogen adsorbed onto similar slides out of solutions of varying bulk concentrations. The oxide deposits were quantified using a scanner and an image analysis program. The isotherms produced from the collected data indicate a continuous, monotonic correlation between light absorbed by adherent oxide and surface concentration of protein. The same correlation applies to albumin and fibrinogen when surface concentrations are expressed in weight units. These results confirm that patterns of oxide deposition correspond to patterns of protein deposition and show clearly how qualitative observations, such as those previously reported, can be made quantitative with scanning and digital image analysis.

Adsorption↗
PubMed1994

Adsorption of proteins out of plasma onto glass from a separated flow.

Perturbations in the adsorption of plasma proteins caused by flow separation were studied quantitatively. An instrument was constructed that causes flow to separate over approximately half the width of a standard microscope slide and the pattern of protein deposition in and near the separated flow was observed by staining the slide with black iron oxide. The slide was mounted at the edge of a Couette flow field established between two concentric cylinders, the outer of which was rotating. The slide was located on the stationary, inner cylinder just downstream of a rectangular bar that causes the flow to separate. After exposure to dilute plasma injected upstream of the bar, the slide was removed and stained with oxide suspension. The resulting, visible pattern was scanned through a video camera and analyzed to yield relative values of stain density that could be quantified. The oxide patterns suggest that proteins were deposited onto the slide less rapidly in and just downstream of the separated flow region than farther downstream. At a shear rate of 6.61 s-1, corresponding to a velocity of 1.32 cm s-1 0.2 cm above the point of flow separation, overall amounts of adsorbed proteins increased with exposure time in the range 3-30 min with the exception of a period from 10 to 11 min when all data show a temporary decrease. In calibration experiments, oxide failed to adhere to slides exposed to purified albumin but adhered copiously to slides exposed to purified fibrinogen. These results suggest that the oxide patterns following plasma exposure are attributable primarily to fibrinogen and that the temporary decrease in the separated flow experiments is attributable to the displacement of fibrinogen by a less stainable protein, conjecturally high molecular weight kininogen and factor XII. This study yields quantitative information confirming earlier findings that were less controlled and non-quantitative. It confirms the hypothesis that the sequence of protein deposition from dilute plasma to glass surfaces is delayed in regions of separated flow.

Adsorption↗
PubMed1993

The effects of pressure and flow on hemolysis caused by Bio-Medicus centrifugal pumps and roller pumps. Guidelines for choosing a blood pump.

Two Bio-Medicus BP-50 centrifugal pumps and two roller pumps were tested simultaneously with porcine blood at 21 degrees +/- 1 degree C in four in vitro circuits to determine the effect of four combinations of flow and pressure conditions on blood damage. Flows of 300 ml/min (1/4-inch inner-diameter tubing in the roller pump) and 1775 ml/min (1/2-inch inner-diameter tubing in the roller pump) and pressure differences across the pump (delta P = outlet pressure--inlet pressure) of 215 mm Hg (n = 6) and 345 mm Hg (n = 5) were examined. The index of hemolysis (milligrams plasma hemoglobin per 100 L blood pumped) for the BP-50 pump was higher at a flow of 300 ml/min than at a flow of 1775 ml/min (p < 0.0002). At 300 ml/min, the index of hemolysis for the BP-50 pump tended to be higher at 345 mm Hg than at 215 mm Hg (mean +/- standard error of the mean, 135 +/- 22 versus 88 +/- 9, p = 0.059). At 1775 ml/min, there was no difference in the index of hemolysis for the BP-50 pump between 215 and 345 mm Hg (37 +/- 7 versus 29 +/- 5, p = 0.32). With the roller pump, the index of hemolysis was higher at a flow of 300 ml/min than at a flow of 1775 ml/min (p < 0.036), but there was no difference in the indexes of hemolysis between 215 and 345 mm Hg at 300 ml/min (60 +/- 9 versus 61 +/- 11, p = 0.93) or at 1775 ml/min (40 +/- 6 versus 36 +/- 6, p = 0.61). Comparison between the two types of pumps showed that the index of hemolysis was significantly higher for the BP-50 than for the roller pump at a flow of 300 ml/min and a delta P of 215 mm Hg (88 +/- 9 versus 60 +/- 9, p = 0.009), as well as at a flow of 300 ml/min and a delta P of 345 mm Hg (135 +/- 22 versus 61 +/- 11, p = 0.001). At a flow of 1775 ml/min, there was no difference in the index of hemolysis between the two pumps at either pressure condition.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1991

Is the Vroman effect of importance in the interaction of blood with artificial materials?

The successive displacement of plasma proteins adsorbed to artificial surfaces (biomaterials) is well documented, mostly by specially designed experiments that stretch out the effect in time and space. Analysis of displacement has been focused principally on molecular events on the adsorbing surface. In this paper attention is directed rather to the antecedent transport phenomena necessary to deliver successive proteins to a surface. The different limitations on protein arrival fixed by the total quantity present and by the rates of transport of super-sufficient quantities are distinguished. The transport perspective is then used to ask, and partly answer, the question: Can protein displacement be responsible for patterns of thrombus formation and cellular adhesion that are seen on the blood-wetted surfaces of devices found in medical practice: artificial organs and vascular prostheses? Calculations and a small amount of preliminary data suggest that such patterns may form when blood is introduced into these devices, particularly in the neighborhood of boundary shapes that cause separated flows.

Adsorption↗
PubMed1990

Errors in estimates of peritoneal fluid volume.

Inherent limitations in the suitability of drainage volumes for monitoring intraperitoneal fluid volume have resulted in the frequent use of indicator dilution techniques, but little attention has been given to confirming the adequacy of the estimates that volume markers provide. In a series of experimental exchanges in rats, volume estimates were compared based on the dilution of blue dextran and hemoglobin with direct collections of surgically exposed intraperitoneal fluid. Significant systematic and random errors in the indicator dilution volume estimates were observed. The systematic errors appeared to be due to the rapid removal of a fixed amount of marker from peritoneal fluid, while the random errors were caused by the rapid appearance of a variable amount of endogenous chromogen. The behavior of the markers observed in this study was not consistent with the assumptions commonly used to analyze volume transport in peritoneal dialysis.

Animals↗
PubMed1988

The use of polyelectrolytes as osmotic agents for peritoneal dialysis.

Various small and large molecules have been studied as osmotic agents to replace dextrose in peritoneal dialysis. Macromolecules are attractive because of their slow absorption from intraperitoneal solutions; however, it has been assumed that they are only marginally effective as osmotic agents unless they function as polyelectrolytes at physiological pH. In experimental exchanges conducted in rats we measured volume changes induced by Gelifundol (5.5% oxypolygelatin) and Ringers lactate to which was added either nothing, 4.25% dextrose, or 5% albumin. In the control exchanges using Ringers lactate, intraperitoneal fluid volume remained unchanged for eight hours. The volume changes induced by 4.25% dextrose were complete within two hours and resulted in a two-thirds increase over the amount of fluid administered. In both series in which polyelectrolytes were used volume transport was sustained throughout an eight hour dwell. With 5% albumin the total increase in fluid volume was about 40% of that installed, while Gelifundol caused fluid volume to double. Qualitatively similar results were obtained in transport studies conducted in vitro. Physical studies of the oxypolygelatin solutions indicated that the fixed charges per liter were comparable to those in the albumin solutions. Thus the different volume transport the two proteins induced could not be attributed to Donnan effects. However, since the molecular weight of albumin is triple that of Gelifundol the van't Hoff pressures of the two macromolecules can explain the observed differences in volume transport. These results suggest that neutral macromolecules deserve further study as potential osmotic agents for peritoneal dialysis.

Albumins↗