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Biomedical subjects

J A Moore

Publications and source records attributed to J A Moore.

At least 271 records · Page 15Linked to original sources
PubMedDate not supplied

Structure-binding relationships for the interaction between a vancomycin monoclonal antibody Fab fragment and a library of vancomycin analogues and tracers.

A series of vancomycin analogues and tracers were synthesized, and their binding interactions with an anti-vancomycin Fab fragment were evaluated under mass transport limiting conditions using surface plasmon resonance detection. Differences observed in binding interactions were utilized to define the vancomycin structural elements critical for antibody recognition. Major structural regions of vancomycin shown to play an important role in anti-vancomycin Fab fragment recognition include two sugar moieties and one chlorinated phenyl ring. The N-methylleucyl residue, the carboxy terminal residue, and residues in the peptide-binding region of vancomycin have minimal impact on the anti-vancomycin Fab fragment/vancomycin binding interaction. The selection of an antibody with such binding properties plays a critical role in the development of a vancomycin immunoassay that employs stable calibrators and controls.

Antibodies, Monoclonal↗
PubMedDate not supplied

Chemoenzymatic synthesis of 3'-O-(carboxyalkyl)fluorescein labels.

A general and versatile method is described for the synthesis of fluorescent labels. Coupling of the 3'-phenol of fluorescein methyl ester with hydroxyalkyl benzyl esters, followed by benzyl ester hydrolysis, provided a series of fluorescein carboxyalkyl ethers. Use of the Mitsunobu reaction allowed for the introduction of linkers of different lengths onto the 3'-phenol of fluorescein. Chemoenzymatic benzyl ester hydrolysis was achieved with LPL-80 lipase, providing pH-independent labels useful for the preparation of fluorescent conjugates.

Fluoresceins↗
PubMedDate not supplied

Purification of oligonucleotides by high affinity, low molecular weight displacers.

High affinity, low molecular weight anionic displacers were successfully employed for the purification of antisense oligonucleotides. Several important structural characteristics were identified that contribute to the affinity of low molecular weight displacers to a hydrophilized polystyrene divinyl benzene anion exchanger. Sulfonic acid groups were found to possess higher affinity than carboxylic acid and phosphate functionalities, and nonspecific interactions (particularly hydrophobic interactions) were shown to play a major role in the retention process on this stationary phase material. Using this information, two high affinity, low molecular weight displacers were identified. These molecules are relatively inexpensive organic dyes that possess multiple sulfonic acid moieties, as well as aromatic functionalities, which increase nonspecific interactions with the stationary phase. These high affinity displacers, which can be readily detected, were then employed to displace several strongly retained antisense oligonucleotides that could not be displaced by previously established low molecular weight displacers. The displacement process resulted in very high purities of the antisense oligonucleotides. The results presented in this paper are significant in that they demonstrate that low molecular weight displacers for ion-exchange chromatography can possess equal to or greater affinities than their higher molecular weight counterparts, when nonspecific interactions with the stationary phase are exploited. In addition, the results illustrate the high resolutions possible with displacement chromatography and demonstrate an attractive technology for the process scale purification of oligonucleotides.

Chromatography, High Pressure Liquid↗
PubMedDate not supplied

Efficacy of leucocyte-poor red blood cell suspensions prepared by sedimentation in hydroxyethyl starch.

Hydroxyethyl starch (HES), which is licensed for human use as a plasma volume expander, has red blood cell sedimenting properties similar to high molecular weight dextran (HMWD). The present studies document the effectiveness of commercially available HES in preparing leukocyte-poor red blood cell suspensions (L-PRBCs) from donor whole blood and packed red blood cells stored in CPD up to 120 hours prior to processing. The effect of HES sedimentation on final white blood cell count, on erythrocyte p50 and 2,3-DPG concentration, and on short- and long-term in vivo survival of 72-hour stored donor red cells are described, along with favorable clinical experience with over 300 transfusions of HES-sedimented L-PRBCs. Since the method requires no special equipment, it is immediately available for widespread use.

Blood Sedimentation↗
PubMedDate not supplied

Purification and use of the C3d subunit C3.

A method is described for the preparation of C3d from fresh-frozen CPD plasma. A redissolved EDTA euglobulin precipitate formed from defibrinated plasma is chromatographed on DEAE cellulose. The C3-rich peak is further chromatographed by stepwise elution from hydroxy-apatite. The highly purified C3 and C3b so obtained are then treated with commercial C3b inactivator. G-200 sephadex filtration of this material produces a low molecular weight peak containing C3d greater than 95 per cent purity. The purified C3d, which contains negligible carbohydrate, has a molecular weight of 28,000 (based on SDS polyacrylamide gel electrophoresis). When added at a final concentration of less than 70 mug per ml, it completely inhibits the anti-C3d hemagglutinin reactivity of a potent anti-C3d antiglobulin serum but does not inhibit sera with reactivities against C3c, C4b, and C5b. Two of three rabbits hyperimmunized with purified C3d produced antisera exhibiting only anti-C3d precipitin activity. After absorption of heterologous antibodies, the antisera strongly agglutinated C3d-coated RBC, failed to agglutinate RBC glutinated C3d-coated RBC, failed to agglutinate RBC coated only with C4, and reacted weakly against strongly anti-Rh coated RBC. The latter reactivity was readily absorbed by whole IgG. Preliminary studies with 125I-labeled C3d indicate its potential value in assessing potency of anti-C3d reagents.

Animals↗
PubMedDate not supplied

Identification of a C4 Subcomponent on C3d-coated erythrocytes.

Test erythrocytes (E) used to evaluate anti-complement (C') antiglobulin sera have not been adequately standardized. This report describes a previously unrecognized C4-derived antigen (temporarily called X-Ag) found on E generally believed to be coated only with the C3d subcomponent of C3, X-Ag occurred on all E coated in vitro with C' by low ionic strength-sucrose or cold agglutinin methods and on E from ten of ten patients whose cells had been C' coated in vivo. It was not removed by incubating these cells with trypsin or fresh compatible serum. This antigen was found on "C4-only-coated" red blood cells made with normal or congenitally C2-deficient serum but not on cells similarly prepared with congenitally C4-deficient serum. It was not identified on E coated with C' via the alternate pathway, normal trypsinized cells, nor cells coated only with IgG. Absorption experiments utilizing purified complement components and subcomponents and G200 Sephadex fractions of normal human serum strongly suggest that X-Ag is a subcomponent of C4(C4d). These results show that at least one C' subcomponent other than C3d occures on both in vitro and in vivo C3d-coated erythrocytes and must be taken into account when such cells are used to evaluate antiglobulin reagents.

Absorption↗
PubMedDate not supplied

Toxicity of chlorinated phenoxy acids in combination with 2,3,7,8-tetrachlorodibenzo-p-dioxin in C57BL/6 male mice.

Male C57BL/6 mice were given feed containing various concentrations of 2,4,-dichlorophenoxyacetic acid (2,4-D),2,4,5-trichlorophenoxyacetic acid (2,4,5-T), and 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) to provide daily doses of approximately 40 mg/kg 2,4,-D, 40 mg/kg 2,4,5-T, and 2.4 micrograms/kg TCDD (group II): 40 mg/kg 2,3-D, 40 mg/kg 2,4,5-T, and 0.16 micrograms/kg TCDD (group III); and 20 mg/kg 2,4-D, 20 mg/kg 2,4,5-T, and 1.2 micrograms/kg TCDD (group IV). The chemical mixtures were chosen to approximate the concentrations of the components of Herbicide Orange. Controls (group I) were given a diet with only the corn oil vehicle added to the feed. In the treated animals, liver enlargements, hepatocellular toxicity, and reduced thymus size were found an body weight gain was significantly less than that of the control animals. Significant of complete recovery from liver and thymus toxicity was observed when the mice were returned to a control diet. The thymic toxicity and hepatotoxicity seemed to be related to TCDD levels, while the decrease in body weight gain was more closely associated with phenoxy acid concentrations. All effects attributed to the mixture were elicited by individual compounds in the mixture in previous studies.

2,4,5-Trichlorophenoxyacetic Acid↗
PubMedDate not supplied

Male fertility, sister chromatid exchange, and germ cell toxicity following exposure to mixtures of chlorinated phenoxy acids containing 2,3,7,8-tetrachlorodibenzo-p-dioxin.

Fertility and sperm number, motility, and morphology were analyzed in male C57BL/6 mice exposed to various mixtures of 2,4-dichlorophenoxyacetic acid (2,4-D), 2,4,5-trichlor-ophenoxyacetic acid (2,4,5-T), and 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) in the diet for 8 wk. Somatic cell (bone marrow) sister chromatid exchange frequencies were also evaluated in mice injected with similar chemical mixtures. The concentration of the test chemicals was such that the average daily feeding dose in the three mixtures or control was 40 mg/kg 2, 4-D, 40 mg/kg 2,4,5-T, and 2.4 micrograms/kg TCDD; 40 mg/kg 2,4-D, 40 mg/kg 2,4,5-T and 0.16 microgram/kg TCDD; 20 mg/kg 2, 4-D, 20 mg/kg 2,4,5-T, and 1.2 micrograms/kg TCDD; or no chemical added (control). No significant dose-related effects were observed in the treated mice compared to the control group.

2,4,5-Trichlorophenoxyacetic Acid↗
PubMedDate not supplied

Development and viability of offspring of male mice treated with chlorinated phenoxy acids and 2,3,7,8-tetrachlorodibenzo-p-dioxin.

Male C57BL/6 mice were given feed containing various concentrations of 2,4-dichlorophenoxyacetic acid (2,4-D), 2,4,5-trichlorophenoxyacetic acid (2,4,5-T) and 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) such that their diets contained daily doses of approximately 40 mg/kg 2,4,-D, 40 mg/kg 2,4,5-T, and 2.4 micrograms/kg TCDD (group II); 40 mg/kg 2,4-D, 40 mg/kg 2,4,5-T, and 0.16 microgram/kg TCDD (group III); or 20 mg/kg 2,4-D, 20 mg/kg 2,4,5-T, and 1.2 micrograms/kg TCDD (group IV). Controls (group I) were given a diet with only the corn oil vehicle added to the feed. At the conclusion of an 8-wk dosing period, treated males cohabited with untreated virgin C57BL/6 female mice. Females were randomly assigned either to undergo teratological examination at 18 d of gestation, when the fetuses were examined for external, visceral, and skeletal malformations, or to be segregated and allowed to deliver their pups. Live and dead offspring and birth weight were recorded and viability was monitored until postnatal d 21. In both cases, development and survival of offspring in the test groups, whose sire had been treated with the mixture of phenoxy acids and TCDD, were not significantly different from those in the control group, whose sire had received untreated feed.

2,4,5-Trichlorophenoxyacetic Acid↗
PubMedDate not supplied

Accuracy of computational hemodynamics in complex arterial geometries reconstructed from magnetic resonance imaging.

PURPOSE: Combining computational blood flow modeling with three-dimensional medical imaging provides a new approach for studying links between hemodynamic factors and arterial disease. Although this provides patient-specific hemodynamic information, it is subject to several potential errors. This study quantifies some of these errors and identifies optimal reconstruction methodologies. METHODS: A carotid artery bifurcation phantom of known geometry was imaged using a commercial magnetic resonance (MR) imager. Three-dimensional models were reconstructed from the images using several reconstruction techniques, and steady and unsteady blood flow simulations were performed. The carotid bifurcation from a healthy, human volunteer was then imaged in vivo, and geometric models were reconstructed. RESULTS: Reconstructed models of the phantom showed good agreement with the gold standard geometry, with a mean error of approximately 15% between the computed wall shear stress fields. Reconstructed models of the in vivo carotid bifurcation were unacceptably noisy, unless lumenal profile smoothing and approximating surface splines were used. CONCLUSIONS: All reconstruction methods gave acceptable results for the phantom model, but in vivo models appear to require smoothing. If proper attention is paid to smoothing and geometric fidelity issues, models reconstructed from MR images appear to be suitable for use in computational studies of in vivo hemodynamics.

Carotid Arteries↗
PubMedDate not supplied

Computational blood flow modeling based on in vivo measurements.

Study of the relationship between hemodynamics and atherogenesis requires accurate three-dimensional descriptions of in vivo arterial geometries. Common methods for obtaining such geometries include in vivo medical imaging and postmortem preparations (vessel casts, pressure-fixed vessels). We sought to determine the relative accuracy of these methods. The aorto-iliac (A/I) region of six rabbits was imaged in vivo using contrast-enhanced magnetic resonance imaging (MRI). After sacrifice, the geometry of the A/I region was preserved via vascular casts in four animals, and ex situ pressure fixation (while preserving dimensions) in the remaining two animals. The MR images and postmortem preparations were used to build computer representations of the A/I bifurcations, which were then used as input for computational blood flow analyses. Substantial differences were seen between MRI-based models and postmortem preparations. Bifurcation angles were consistently larger in postmortem specimens, and vessel dimensions were consistently smaller in pressure-fixed specimens. In vivo MRI-based models underpredicted aortic dimensions immediately proximal to the bifurcation, causing appreciable variation in the aorto-iliac parent/child area ratio. This had an important effect on wall shear stress and separation patterns on the "hips" of the bifurcation, with mean wall shear stress differences ranging from 15% to 35%, depending on the model. The above results, as well as consideration of known and probable sources of error, suggests that in vivo MRI best replicates overall vessel geometry (vessel paths and bifurcation angle). However, vascular casting seems to better capture detailed vessel cross-sectional dimensions and shape. It is important to accurately characterize the local aorto-iliac area ratio when studying in vivo bifurcation hemodynamics.

Animals↗
PubMedDate not supplied

Interferon reduces hepatic drug metabolism in vivo in mice.

The effects of two highly purified human leukocyte interferons (IFN-A and IFN-AD) on drug-metabolizing capacity in mice have been investigated. IFN-AD was found to produce significant changes in antipyrine half-life, assessed by analysis of 14CO2 exhalation rates following 14C-antipyrine administration. By contrast, IFN-A, which has considerably less antiviral potency than IFN-AD, was found to have no effect on antipyrine half-life. The administration regimen was found to markedly alter the effects seen with IFN-AD. When IFN-AD was given as single daily doses (5 X 10(7) units/kg/day X 3 days), the half-life of antipyrine increased by a mean of 40% (from 21.0 to 28.9 min). However, when a smaller daily dose (3 X 10(7) units/kg/day) was given as a continuous infusion, the antipyrine half-life increased by more than 3-fold (from 20.8 to 68.5 min) after 3 days of administration. Continued infusion for a further 3 days produced no additional change in antipyrine half-life. These results demonstrate that human leukocyte interferons can significantly inhibit hepatic metabolic activity in vivo.

Animals↗
PubMedDate not supplied

Pharmacokinetics and tissue distribution of recombinant human tumor necrosis factor-alpha in mice.

The serum pharmacokinetics and the major organs of accumulation of recombinant human tumor necrosis factor-alpha (rHuTNF) were determined in BDF1 mice after intravenous and intramuscular administration. Serum concentrations of immunoreactive protein were determined by enzyme-linked immunosorbent assay, and radioactivity was quantitated by beta and gamma scintigraphy. The serum pharmacokinetics of labeled and unlabeled rHuTNF were identical when administered by the intravenous route. After intravenous doses of 165 to 320 micrograms/kg, the clearance was 2.9-3.6 ml/hr, the initial volume of distribution was 1.4-1.6 ml (70-80 ml/kg), and the half-life was 18.5-19.2 min. Intramuscular administration of 320 micrograms/kg resulted in a peak serum concentration of 112 ng/ml. The time of the peak concentration was 1 hr, and the bioavailability of the intramuscular dose was 12%. The data suggest that the disposition of this protein may be biexponential. If this is the case, the terminal phase would appear to account for less than 1% of the total AUC. Since serum concentrations in the terminal phase are at the sensitivity limit of the assay, a single half-life is reported. 125I-Labeled and metabolically labeled 3H-rHuTNF were used to examine tissue distribution. After intravenous 125I-rHuTNF administration, the rank order of accumulation of the 125I-radiolabel in the major organs (per cent dose per organ over 1440 min) was: liver greater than kidney greater than lung greater than heart greater than spleen. This rank order of accumulation was confirmed by intravenous 3H-rHuTNF administration.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMedDate not supplied

Failure of propranolol to prevent chronic hyperthyroid induced cardiac hypertrophy and multifocal cellular necrosis in the rat.

The effect of excess thyroid and thyroid plus propranolol on right and left ventricular structure was examined. Perfusion fixed hearts from untreated adults and those treated for 10 weeks with thyroid, propranolol, and thyroid plus propranolol were embedded in plastic and sectioned for light and electron microscopy. Hyperthyroidism stimulated a significant increase in heart rate, oxygen consumption, and systolic blood pressure. Propranolol was effective in normalizing the heart rate of hyperthyroid rats, but did not affect oxygen consumption or blood pressure. Both left and right ventricular weight increased similarly in thyroid and thyroid-propranolol treated animals, but right ventricular mass increased more than left ventricular mass in both groups. The majority (21 of 24) of right ventricles from thyroid and thyroid-propranolol treated animals had multifocal areas of histologically detected fibrosis. Left ventricular fibrosis was absent or minimal in both hyperthyroid groups. Neither left nor right ventricular fibrosis was observed in control or propranolol only treated animals. We conclude that beta adrenergic stimulation is unnecessary for thyroid induced cardiomegaly, and excess thyroid hormones induce right ventricular fibrosis which is not prevented by propranolol.

Animals↗