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Biomedical subjects

J Boyd

Publications and source records attributed to J Boyd.

At least 181 records · Page 10Linked to original sources
PubMed1991

Proliferation in human bladder carcinoma measured by Ki-67 antibody labelling: its potential clinical importance.

Ki-67 is a monoclonal antibody which recognises a human nuclear antigen expressed in proliferating cells. The antibody was used to assess proliferation in primary human bladder tumours from 64 patients. Ki-67 index (the number of Ki-67 positive tumour cells divided by the total number of tumour cells %) was derived from 59 tumours. A wide range of Ki-67 indices were recorded, range 3.0-65.8%, mean 20.2%. The Ki-67 index correlated with known prognostic factors: T stage (P = 0.002) and histological grade (P less than 0.001), early stage disease and more differentiated tumours having lower Ki-67 indices. Patients with invasive disease (21 patients) had significantly higher Ki-67 indices than those with non-invasive disease (P = 0.01). Patients with metastatic disease at presentation (four cases) all had a Ki-67 index of greater than or equal to 29%. Ki-67 antibody staining is a simple technique for assessing the proliferation fraction than can be performed on a small amount of tissue taken at routine biopsy without prior injection of thymidine analogues.

Aged↗
PubMed1990

A systematic approach towards the complete assignment of 13C resonances for horse ferrocytochrome c.

The complete 1H-NMR assignments for horse ferrocytochrome c have been reported by Wand and colleagues and by our group at Oxford. Using these 1H assignments, we now report chemical shift assignment for 205 13C resonances arising from horse ferrocytochrome c. This is from a total of 437 13C nuclei with covalently attached protons. These chemical shift assignments have been achieved using 1H-detected two-dimensional heteronuclear 1H-13C correlation techniques. The data have been collected from samples of horse ferrocytochrome c without isotopic enrichment. The complete 13C assignments for all carbons with covalently linked protons are reported for the amino acids Ala, Thr, Val and Gly. Specific assignments are tabulated for all 49 methyl groups, for 52 of 92 alpha-carbon resonances, for 10 resonances associated with the heme group, for all aromatic side-chain 13C resonances which have covalent protons and give rise to observable cross peaks under the experimental conditions used, as well as for a number of other side chains of aliphatic amino acids.

Amino Acids↗
PubMed1990

Cellular processing of the interleukin-2 fusion toxin DAB486-IL-2 and efficient delivery of diphtheria fragment A to the cytosol of target cells requires Arg194.

We have used site-directed mutagenesis to examine the role played by Arg191, Arg193, and Arg194 of the fusion toxin DAB486-IL-2 in the intoxication of high affinity interleukin-2 receptor-bearing T-lymphocytes. These arginine residues are positioned in the proteolytically sensitive 14-amino acid loop subtended by the disulfide bond between Cys187 and Cys202 in this fusion toxin. DAB486-IL-2 was formed by the genetic substitution of the native diphtheria toxin receptor binding domain with human interleukin-2 (Williams, D.P., Parker, K., Bacha, P., Bishai, W., Borowski, M., Genbauffe, F., Strom, T.B., and Murphy, J.R. (1987) Protein Eng. 1, 493-498). We demonstrate that substitution of Arg194 with Gly results in a 1000-fold loss of DAB486-IL-2 potency. Since trypsin "nicking" of the Gly194 mutant restores biologic activity, we conclude that Arg194 is required for the cellular processing of the fusion toxin which results in the release of fragment A into the cytosol.

Animals↗
PubMed1990

Assignment of proton resonances, identification of secondary structural elements, and analysis of backbone chemical shifts for the C102T variant of yeast iso-1-cytochrome c and horse cytochrome c.

Resonance assignments for the main-chain, side-chain, exchangeable side chain, and heme protons of the C102T variant of Saccharomyces cerevisiae iso-1-cytochrome c in both oxidation states (with the exception of Gly-83) are reported. (We have also independently assigned horse cytochrome c.) Some additional assignments for the horse protein extend those of Wand and co-workers [Wand, A. J., Di Stefano, D. L., Feng, Y., Roder, H., & Englander, S. W. (1989) Biochemistry 28, 186-194; Feng, Y., Roder, H., Englander, S. W., Wand, A. J., & Di Stefano, D. L. (1989) Biochemistry 28, 195-203]. Qualitative interpretation of nuclear Overhauser enhancement data allows the secondary structure of these two proteins to be described relative to crystal structures. Comparison of the chemical shift of the backbone protons of the C102T variant and horse protein reveals significant differences resulting from amino acid substitution at positions 56 and 57 and further substitutions between residue 60 and residue 69. Although the overall folding of yeast iso-1-cytochrome c and horse cytochrome c is very similar, there can be large differences in chemical shift for structurally equivalent residues. Chemical shift differences of amide protons (and to a lesser extent alpha protons) represent minute changes in hydrogen bonding. Therefore, great care must be taken in the use of differences in chemical shift as evidence for structural changes even between highly homologous proteins.

Amino Acid Sequence↗
PubMed1990

Alpha-fetoprotein in the woodchuck model of hepadnavirus infection and disease: immunochemical analysis of woodchuck alpha-fetoprotein and measurement in serum by quantitative monoclonal radioimmunoassay.

Woodchuck hepatitis virus infection of the eastern woodchuck represents a useful model for the study of hepatitis B virus infection and disease in humans, including hepatocellular carcinoma. In man, hepatocellular carcinoma is frequently detected and monitored using assays for serum alpha-fetoprotein. To study the relationship between alpha-fetoprotein and woodchuck hepatitis virus-induced hepatocellular carcinoma in the woodchuck model, we produced a monoclonal antibody to woodchuck alpha-fetoprotein and used biophysical and immunochemical methods to demonstrate its specificity and affinity (7 x 10(8) L/mol) for woodchuck alpha-fetoprotein. A competition radioimmunoassay was then developed and standardized for measuring serum alpha-fetoprotein concentrations. In the radioimmunoassay system, woodchuck alpha-fetoprotein was detected between 20 ng/ml (20% to 25% inhibition) and 8,500 ng/ml (97% to 98% inhibition). Elevated serum alpha-fetoprotein concentrations (450 to 452,000 ng/ml) were measured in 21 of 23 woodchucks in the advanced stages of woodchuck hepatitis virus-induced hepatocellular carcinoma. Serum alpha-fetoprotein was elevated above normal (greater than or equal to 450 ng/ml) as early as 3 to 11 mo before terminal hepatocellular carcinoma in 11 of 16 of the woodchuck hepatitis virus-carrier woodchucks. In a pilot study, serum alpha-fetoprotein became markedly elevated above normal in woodchuck hepatitis virus-carrier woodchucks that developed hepatocellular carcinoma but not in serologically recovered or uninfected woodchucks (i.e., without hepatocellular carcinoma). Thus, alpha-fetoprotein may provide a useful noninvasive marker in the woodchuck model for detecting and monitoring woodchuck hepatitis virus-induced hepatocellular carcinoma from earlier stages.

Animals↗
PubMed1990

Prevention of calcification of glutaraldehyde pretreated bovine pericardium through controlled release polymeric implants: studies of Fe3+, Al3+, protamine sulphate and levamisole.

Calcification is the principal cause of the clinical failure of bioprosthetic heart valves fabricated from glutaraldehyde pretreated porcine aortic valves or bovine pericardium. The present study investigated controlled-release implants for prevention of the calcification of glutaraldehyde pretreated bovine pericardium in a rat subdermal model. Either Al3+ and Fe3+ (inhibitors of the growth and dissolution rate of hydroxyapatite crystals), levamisole (alkaline phosphatase inhibitor) or protamine sulphate (charge modifier) were individually incorporated into various polymeric carriers (either silicone rubber, polyurethane or silicone rubber-polyurethane copolymer). Polymeric implants were evaluated for in vitro release kinetics, which revealed that sustained drug release was obtained from 21 d to more than 90 d from various drug matrices. In vivo efficacy was studied by co-implanting the polymeric delivery systems with glutaraldehyde pretreated bovine pericardium for 21 d using a subdermal rat model; glutaraldehyde pretreated bovine pericardium calcium levels were quantitated by atomic absorption spectroscopy in the explanted tissues. Fe3+ and Al3+ polymeric implants were the most effective for inhibiting deposition of calcium mineral. Al3+ demonstrated 82% inhibition of calcification compared to controls and Fe3+ resulted in 80% inhibition of calcification. Specific histologic staining methods showed that Fe3+ and Al3+ were localized within the devitalized cells of the explanted glutaraldehyde pretreated bovine pericardium. No adverse effects on somatic growth or recipient bone morphology were noted following controlled-release drug administration. Controlled release of protamine sulphate or levamisole did not significantly inhibit glutaraldehyde pretreated bovine pericardium calcification. It is concluded that regional controlled release of Fe3+ or Al3+ inhibits glutaraldehyde pretreated bovine pericardium calcification in the rat subdermal model without adverse effects.

Aluminum↗
PubMed1990

Molecular cloning and DNA sequence analysis of a diphtheria tox iron-dependent regulatory element (dtxR) from Corynebacterium diphtheriae.

Although the structural gene for diphtheria toxin, tox, is carried by a family of closely related corynebacteriophages, the regulation of tox expression is controlled, to a large extent, by its bacterial host Corynebacterium diphtheriae. Optimal yields of tox gene products are obtained only when iron becomes the growth-rate-limiting substrate. Previous studies suggest that regulation of tox expression is mediated through an iron-binding aporepressor. To facilitate molecular cloning of the tox regulatory element from genomic libraries of C. diphtheriae, we constructed a tox promoter/operator (toxPO)-lacZ transcriptional fusion in Escherichia coli strain DH5 alpha. We report the molecular cloning and nucleic acid sequence of a diphtheria tox iron-dependent regulatory element, dtxR, and demonstrate that expression of beta-galactosidase from the toxPO-lacZ fusion is regulated by dtxR-encoded protein in an iron-sensitive manner. In addition, we show that expression of the toxPO-lacZ fusion is not affected by the E. coli iron-regulatory protein Fur and that the dtxR protein does not inhibit expression of fur-regulated outer-membrane proteins.

Amino Acid Sequence↗
PubMed1990

Sleep patterns of New Zealand infants during the first 12 months of life.

The sleeping patterns of 874 infants aged 1-12 months were recorded by parents over a 6 day period. The most striking feature of the results was the wide range in total hours slept by infants of the same age, for example, the average total sleeping time per 24 h period for 4 month old infants was 14.8 h with a range of 11.0-19.3 h. Furthermore, over the 6 day period, individual infants showed wide variations in their sleeping times with a range as great as 12 h. Despite these wide variations, several clear trends emerged: from 1 to 8 months, a decrease with age of the average number of hours slept per 24 h period; and from 8 to 12 months, and a continuing shift towards a dominance of night over day sleep. The frequency of night wakings was, on average, 77%, a finding that contrasted sharply with parental perception of frequency of night waking; the transition from bassinet/carrycot to cot was most common between 3 and 4 months of age; 25% of babies slept with the light on, 9% used a dummy, and 37% sucked their fingers; first-born infants woke significantly less often at night than those with one or more sibling. These results provide an important comparative data set on the sleep patterns of infants.

Birth Order↗
PubMed1989

Determination of immunoreactive endothelin in medium from cultured endothelial cells and human plasma.

We have developed a sensitive and selective radioimmunoassay for porcine/human endothelin (ET1). The assay has a detection limit of 0.62 pg/tube and exhibits no cross-reactivity to atrial natriuretic peptide, arginine vasopressin, or angiotensin II. Procedures were developed for extraction of endothelin from human plasma samples and samples of buffer from endothelial cell incubations using C18 Sep-Pak extraction cartridges. The mean recovery following extraction was approximately 80%. Both bovine and porcine aortic endothelial cells were found to produce immunoreactive endothelin (IR-ET) with porcine cells producing 4.7 +/- 1.1 ng of IR-ET/mg cell protein after 6 hours. Human plasma samples were extracted, assayed and found to contain a mean concentration of 2.0 +/- 0.4 pg/ml of IR-ET.

Animals↗
PubMed1989

Improved resolution in 1H-detected 1H-15N correlation experiments.

The determination of protein structure by NMR is restricted at molecular masses above 10 kDa by overlapping resonances. One way of overcoming this problem is to label the protein with 15N. The conventional way to record 15N spectra is to use heteronuclear multiple-quantum coherence. We present here an alternative approach based on 15N single-quantum coherence. This is shown to have substantial advantages over the multiple-quantum method, including better F1 resolution.

Escherichia coli↗
PubMed1989

The sequence-specific assignment of the 1H-NMR spectrum of an enzyme, horse-muscle acylphosphatase.

A complete range of two-dimensional NMR experiments was used for the assignment of the 1H-NMR spectrum of horse muscle acylphosphatase. Firstly the spin systems of some easily identifiable amino acid side chains were assigned. These side chains involved all the aromatic residues and all the leucine, valine, isoleucine, threonine, alanine, proline as well as some of the glycine residues. Analysis of nuclear Overhauser enhancement spectra in our previous work had identified the sequential and long-range patterns characteristics for secondary structure elements. This result had also provided the identification of the main-chain alpha and amide proton resonances. Several of the completely assigned spin systems were then identified as being part of the secondary structure units which led, after analysis of the primary amino acid sequence, to unambiguous sequence-specific assignments. The identification and assignment of the remaining side-chain resonances was then completed and are reported here. These results provide a complete data base for the three-dimensional structure determination of this enzyme in solution.

Acid Anhydride Hydrolases↗
PubMed1989

Identification and description of beta-structure in horse muscle acylphosphatase by nuclear magnetic resonance spectroscopy.

Nuclear magnetic resonance spectra of acylphosphatase were searched for signs of beta-structure, i.e. characteristic nuclear Overhauser enhancement patterns displayed in the two-dimensional spectra, typical chemical shifts, coupling constants and slow 2H-H exchange. The results provided identification of the main-chain resonances of amino acid residues involved in the beta-structure. The full sequential assignment of this region was gained by identification of some amino acid spin systems and their alignment with the primary sequence. The assignment of the side-chains was virtually completed subsequently and a list produced of nuclear magnetic resonance (n.m.r.) constraints derived from the spectra. The beta-structure consists of a beta-sheet with four antiparallel chains, one attached parallel chain, three tight turns and a beta-bulge. The conformation of the beta-sheet was determined by distance geometry calculation using the n.m.r. constraints (174 intraresidual, 107 sequential and 226 long-range distances, 32 torsion angles, phi, and 28 hydrogen bonds) as input. Observation of some interactions between the sheet and previously identified alpha-helical regions made it possible to give an outline of the three-dimensional structure of the enzyme.

Acid Anhydride Hydrolases↗
PubMed1989

Obstructive airways disease in non-smoking subjects with pigeon fanciers' lung.

The aim of the study was to assess the degree of airways involvement in non-smoking subjects with extrinsic allergic alveolitis resulting from pigeon-derived antigens. Two hundred and eight pigeon fanciers completed a questionnaire of symptoms, performed spirometry on a Vitalograph Compact spirometer and had IgG antibody to pigeon gamma-globulin measured. Eighty fanciers had the acute form of pigeon fanciers' lung (PFL) and these subjects were less likely to be current smokers (P less than 0.001). Subjects with PFL had a higher prevalence of chronic bronchitis (P less than 0.001), large airways (P = 0.014) and peripheral airways obstruction (P less than 0.001). The prevalence of chronic bronchitis and peripheral airways obstruction increased significantly as the level of antibody to pigeon gamma-globulin rose. Peripheral airways obstruction, as manifest by a reduced FEF25-75%, was the most common abnormality found. Airways disease is an important feature of the acute form of PFL and consists of a hypersecretory disorder in the form of chronic bronchitis and of obstructive airways disease particularly affecting the peripheral airways.

Adolescent↗
PubMed1989

Chronic simple bronchitis in pigeon fanciers. Relationship of cough with expectoration to avian exposure and pigeon breeders' disease.

The prevalence of chronic bronchitis and its relationship to antigenic exposure was studied in a population of pigeon fanciers. Two hundred and eighty seven fanciers completed a questionnaire regarding symptoms and circumstances of avian exposure and had IgG antibody to pigeon gammaglobulin measured by an enzyme linked immunosorbent assay. Chronic bronchitis occurred in 26.2 percent of nonsmoking fanciers and increased in prevalence as antibody levels rose (p less than 0.001). Although significantly more common (p less than 0.005) in the 85 (29.6 percent) fanciers who also had typical delayed symptoms of pigeon breeders' disease (PBD), chronic bronchitis was the only manifestation of the disease in 24 (8.4 percent) of the population surveyed. Chronic bronchitis was not related to the intensity or duration of avian exposure suggesting that host factors are more important in its pathogenesis. Chronic bronchitis must be considered to be an integral part of the clinical spectrum of PBD.

Adolescent↗
PubMed1989

Isoenzyme analysis of human and animal isolates of Giardia duodenalis from British Columbia, Canada.

Isoenzyme patterns of 32 isolates of Giardia duodenalis, obtained from 6 beavers and 11 humans from British Columbia, plus 15 other isolates were evaluated using thin-layer starch-gel electrophoresis. We attempted to use 12 enzymes; 9 gave reproducible and interpretable results. The isoenzyme patterns of the isolates were classified into 12 groups with 17 (53%) of the 32 isolates confined to 1 group. The other 11 groups each comprised only 1 or 2 isolates. There was no obvious correlation between clinical symptoms and isoenzyme patterns. Our findings suggest that beavers, like humans and gerbils are receptive to organisms with many different isoenzyme patterns.

Animals↗