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J R Davies

Publications and source records attributed to J R Davies.

At least 19 recordsLinked to original sources
PubMed1992

Release of mucus glycoconjugates by Pseudomonas aeruginosa rhamnolipid into feline trachea in vivo and human bronchus in vitro.

Pseudomonas aeruginosa colonizes the lower respiratory tracts of patients with severe bronchiectasis, including cystic fibrosis, a condition associated with increased airway mucus output. We have shown that an extract containing chloroform-soluble extracellular products of P. aeruginosa releases glycoconjugates into the cat trachea in vivo. This activity was not related to pyocyanin, a major component of the extract, but was associated with the rhamnolipids. Purified monorhamnolipid (100 micrograms/ml) released radiolabeled and periodic acid-Schiff (PAS)-reactive glycoconjugates (delta 3H = +490 +/- 70%, delta 35S = +170 +/- 40%, delta PAS = +8.6 +/- 1.7 micrograms/min; n = 6, P less than 0.02 for each). Dirhamnolipid (200 micrograms/ml) was also effective (delta 3H = +640 +/- 70%, delta 35S = +130 +/- 20%, delta PAS = +9.3 +/- 1.5 micrograms/min; n = 6, P less than 0.02 for each). Monorhamnolipid (100 micrograms/ml) also released 35S-labeled and PAS-reactive glycoconjugates from human bronchial tissue in vitro (delta 35S = +189 +/- 47%, delta PAS = +26.3 +/- 8.5 micrograms/min; n = 7, P less than 0.001 versus control tissues in which no stimulus was given). The cat tracheal glycoconjugates released by the rhamnolipids differed from those released by pilocarpine 50 microM, in having a higher 3H:35S ratio (P less than 0.001). After gel chromatography on a Sepharose CL-4B column, the void volume fractions of the glycoconjugates also had different profiles in a cesium chloride density gradient. Those released by rhamnolipid banded at 1.62 g/ml, while those released by pilocarpine banded mainly at 1.50 g/ml, with some of the higher density material also present.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1991

Mucins in cat airway secretions.

Mucous secretions were obtained from cat tracheas that had received [3H]glucose and [35S]sulphate to radiolabel mucus glycoproteins biosynthetically. Samples were collected under resting ('basal') conditions as well as after pilocarpine stimulation and were separated into gel and sol phases by centrifugation. Macromolecules were partially purified by using gel chromatography on Sepharose CL-4B, and the species that were eluted with the void volume were then separated into two major populations with isopycnic density-gradient centrifugation in CsCl. The major component from the gel phase of pilocarpine-induced secretions had a buoyant density typical of mucins and was observed as linear and apparently flexible chains by electron microscopy. Reduction of disulphide bonds gave subunits that could be further cleaved by trypsin digestion into components of approximately the same size as the high-Mr glycopeptides obtained from other mucins after this treatment. In contrast, the dominant species in the gel phase of the 'basal' secretion had a significantly higher buoyant density than expected for mucins and was largely unaffected by reduction, as studied by gel chromatography. The macromolecules were fragmented by trypsin, suggesting that they contain a polypeptide backbone. This more dense component also predominated in the sol phase both from the 'basal' secretions and from the pilocarpine-released secretions. Digestion with DNAase, chondroitin ABC lyase or heparan sulphate lyase had no effect, which shows that this component is not DNA, a dermatan sulphate/chondroitin sulphate or a heparan sulphate proteoglycan. In contrast, endo-beta-galactosidase and keratanase caused some fragmentation, suggesting that the molecules contain some linkages of the poly-(N-acetyl-lactosamine) type, although the degradation was not as extensive as expected for keratan sulphate. Treatment with alkaline borohydride resulted in extensive fragmentation of the high-Mr glycopeptides from both components, indicating that the glycans were oligosaccharides that were probably O-linked. The monosaccharide compositions of both components were consistent with that expected for mucins. The data are in keeping with the major component from the pilocarpine-stimulated gel secretions being a mucus glycoprotein and the more dense component being a mucin-like molecule, possibly related to the keratanase-sensitive material isolated from canine trachea by Varsano, Basbaum, Forsberg, Borson, Caughey & Nadel [(1987) Exp. Lung Res. 13, 157-184].

Animals↗
PubMed1991

Angiotensin converting enzyme inhibitors and magnesium conservation in patients with congestive cardiac failure.

OBJECTIVE: To investigate whether angiotensin converting enzyme inhibitors reduce diuretic induced magnesium excretion in patients in congestive cardiac failure. DESIGN: Cohort analytic study. SETTING: A London district general hospital. SUBJECTS: Thirty four patients with chronic congestive cardiac failure caused by ischaemic heart disease or cardiomyopathy selected consecutively from inpatients under the care of two consultant cardiologists. Nineteen patients (group 1) on diuretics alone were compared with 15 patients (group 2) taking diuretics plus either enalapril or captopril. All drug regimens were stable for at least three months before the study. Patients with impaired renal function (plasma creatinine greater than 120 mumol/l) were excluded. INTERVENTIONS: An intravenous loading dose of magnesium sulphate was given to minimise the variability in baseline magnesium state. MAIN OUTCOME MEASURE: Total urine magnesium excretion and creatinine clearance in 24 hour urine collections. RESULTS: Plasma magnesium was similar in the two groups. However, 24 hour urine magnesium excretion was significantly lower in group 2 than in group 1. Furthermore, creatinine clearance was also significantly lower in group 2 and correlated strongly with magnesium excretion. There was no such relation in group 1. There was no difference in fractional clearance of magnesium between groups. CONCLUSION: Angiotensin converting enzyme inhibitors have an important magnesium conserving action, possibly via their effect on glomerular filtration rate.

Aged↗
PubMed1990

Evaluation of factors which affect column performance with immobilized monoclonal antibodies. Model studies with a lysozyme-antilysozyme system.

Methods are described for the automated evaluation of affinity columns by frontal boundary analysis. These methods were used to evaluate the performance of immunoaffinity columns based on antilysozyme monoclonal antibody-lysozyme immunoaffinity system. This model system enabled the effects of (i) matrix activation and (ii) the density of immobilized antibody on the change in specific activity of immobilized antibody to be quantitatively assessed. Experimental data were accumulated with carbonyldiimidazole-activated Fractogel HW65F and Trisacryl GF2000 resins and cyanogen bromide-activated Sepharose 4B. An increase in the molar ratio between the concentration of the active groups on the activated matrix and the concentration of immobilized antibody ligands did not result in significant change in the specific activity of the immobilized antibody in the immunochromatographic system. However, increased antibody density with the Fractogel HW65F resin resulted in an increase in the apparent heterogeneity of antibody binding sites for lysozyme and a significant decrease in the specific activity of the immobilized antibody. Furthermore, data from size-exclusion studies with these immunoaffinity matrices demonstrated that at high antibody densities, the accessibility of the immobilized antibody was further decreased due to steric resistance as the antigen size increased.

Animals↗
PubMed1990

Mucus glycoproteins from 'normal' human tracheobronchial secretion.

Mucous secretions were collected from tracheas of patients undergoing minor surgery under general anaesthesia with tracheal intubation, and mucus glycoproteins were isolated by using isopycnic density-gradient centrifugation in CsCl/guanidinium chloride. 'Whole' mucins were excluded from a Sepharose CL-2B gel, whereas subunits obtained after reduction were included. Trypsin digestion of subunits afforded high-Mr glycopeptides (T-domains), which were further included in the gel. The latter fragments are heterogeneous and comprise two or three populations, as indicated by gel chromatography and ion-exchange h.p.l.c. Rate-zonal centrifugation showed that the 'whole' mucins are polydisperse in size, with a weight-average Mr of (14-16) x 10(6). The macromolecules were observed by electron microscopy, as linear and apparently flexible thread-like structures. Subunits and T-domains had weight-average contour lengths of 490 nm and 160 nm respectively. It is concluded that mucus glycoproteins are present in secretions from the healthy lower respiratory tract. The 'whole' tracheal mucins are assembled from subunits, which in turn can be fragmented into high-Mr glycopeptides corresponding to the oligosaccharide domains typically found in mucus glycoproteins. The size and macromolecular architecture of the tracheal mucins is thus similar to that observed for mucins from human cervical mucus, chronic bronchitic sputum and pig stomach, providing yet another example of this general design of these macromolecules, i.e. subunits assembled end-to-end into very large linear and flexible macromolecules.

Bronchi↗
PubMed1990

The uptake of radiolabelled precursors of mucus glycoconjugates by secretory tissues in the feline trachea.

1. We gave one of three radiolabelled precursors of mucus glycoconjugates ([3H]proline, [3H]glucose and [35S]sulphate) into the tracheas of anaesthetized cats for 3 h. In other cats [35S]sulphate was given by intravenous injection. 2. After a further 2 h, tracheas were removed and fixed. Serial actions were cut and alternate sections were stained with Haematoxylin and Eosin or prepared as unstained autoradiographs. Points on submucosal gland and surface epithelium were chosen with a grid on photomicrographs of the stained sections. Absorbance, which is proportional to autoradiographic grain density, was estimated on corresponding points on unstained autoradiographs by flying-spot microdensitometry. 3. With [3H]proline as precursor, the grain densities were greater over surface epithelium than over submucosal gland. With [3H]glucose, grain densities were greater over the surface epithelium in three cases, equal in one and greater over submucosal gland in the last. [35S]Sulphate, given either into the tracheal segment or intravenously, yielded grain densities that were greater over the submucosal glands than over surface epithelium. 4. The areas of submucosal gland the surface epithelium were estimated by point counting and the total content of radioactivity in the two structures estimated by multiplying mean absorbance by area. Ratios of the total radiolabel in surface epithelium to that in submucosal gland were consistently high when [3H]proline was the precursor and low with [35S]sulphate, given by either route. [3H]Glucose gave intermediate ratios. 5. Secretions washed from the trachea were subjected to gel-exclusion chromatography. Washings from tracheas labelled with [3H]proline contained some molecules eluting in the void volume of a Sepharose CL-4B column (suggesting a relative molecular mass of greater than 10(6) Da), but more of the radiolabel eluted in three peaks in the partially included volume. Density gradient ultracentrifugation of the void volume material gave radiolabelled peaks at densities of approximately 1.60 and 1.50 g ml-1, consistent with glycosylated proteins, as well as less dense material (less than 1.30 g ml-1), probably proteins with little or no glycosylation. 6. We discuss the justification of using these radiolabelled precursors to give relatively selective labelling of secretory products from submucosal gland and surface epithelium.

Animals↗
PubMed1989

High-performance liquid affinity chromatography with phenylboronic acid, benzamidine, tri-L-alanine, and concanavalin A immobilized on 3-isothiocyanatopropyltriethoxysilane-activated nonporous monodisperse silicas.

Nonporous, microparticulate, monodisperse silicas with particle diameters between 0.7 and 2.1 microns are introduced as stationary phases in high-performance affinity chromatography. The immobilization of m-aminophenylboronic acid, p-aminobenzamidine, tri-L-alanine, and concanavalin A onto these silicas was successfully achieved using 3-isothiocyanatopropyl-triethoxysilane as an activation reagent. Immobilized phenylboronic acid was applied to the isolation of nucleosides, nucleotides, and glycoprotein hormones such as bovine follicotropin and human chorionic gonadotropin, while immobilized benzamidine was employed for the isolation of the serine proteases thrombin and trypsin, immobilized tri-L-alanine for the separation of pig pancreatic elastase and human leukocyte elastase, and immobilized concanavalin A for the isolation of horseradish peroxidase. In all affinity chromatographic systems studied, the nonporous monodisperse silicas showed improved chromatographic performance compared to results obtained with porous silica supports using identical activation and immobilization procedures. Furthermore, frontal analysis was used as a method to evaluate the influence of experimental parameters on biological activity and accessible ligand densities. Only minor changes in bioactivity were found with the nonporous affinity supports, where accessibilities were typically higher than ca. 60%. The immobilization of affinity ligands onto porous supports as used in this and associated papers thus represents a successful general procedure for the preparation of stable matrices with fast kinetics for use in high-performance affinity chromatography.

Alanine↗
PubMed1989

Gamma globulin-derived standards for the determination of molecular weights, transfer, and immunodetection efficiencies in protein blotting procedures.

Molecular weight markers which are detectable using labeled antispecies antibodies or labeled Protein A have been prepared for use as standards on protein blots. The standards were prepared by the controlled reduction followed by subsequent alkylation of gamma globulin. Separate sets of standards were prepared using gamma globulins derived from human, mouse, rabbit, and sheep species. Standards were also prepared using monoclonal-derived gamma globulins from human myeloma fluid and mouse ascites fluid. Standards produced from monoclonal-derived gamma globulins produced very sharp bands on sodium dodecyl sulfate-polyacrylamide gels and proved to be excellent standards for this technique alone. However, the markers were uniquely suitable for use as standards in protein blotting procedures because their detection was achieved by the procedure used to detect the transferred antigen(s). The detection of immunoglobulin G (IgG)-derived standards on protein blots from all the species listed above was demonstrated using appropriate horseradish peroxidase (HRP)-conjugated antispecies antibodies. The use of other detection systems (biotin-labeled antibody and subsequent detection with HRP-steptavidin, HRP-Protein A) was also validated with human IgG-derived standards. Furthermore, the standards were shown to be suitable for use on both nitrocellulose and cationized nylon-based supports and could be used when adjacent samples were run under reducing conditions. Hence the gamma globulin-derived standards serve as both a control to check the adequacy of transfer and immunodetection systems and as markers which enable the molecular weights of detected antigens to be calculated.

Animals↗
PubMed1989

A tongue resection in macroglossia due to primary amyloidosis.

A lady of 62 years with longstanding macroglossia from primary amyloidosis had the anterior two-thirds of her tongue successfully resected. She had Factor X deficiency. The surgical, anaesthetic and haematological problems encountered are described.

Amyloidosis↗
PubMed1989

Natural or vaccine-induced antibody as a predictor of immunity in the face of natural challenge with influenza viruses.

A study of influenza in residential schools provided the opportunity to assess the significance of antibody as a predictor of immunity. Five hundred and fifty-six pupils from 8 schools were included in the investigations, and the outcome for these children in 27 naturally occurring outbreaks of influenza was analysed. The outbreaks comprised 5 caused by strains of influenza A H3N2, 10 caused by strains of influenza A H1N1, and 12 caused by strains of influenza B. On 8 occasions a second outbreak of the same serotype occurred in a school. There was a general correlation between the presence of antibody to the outbreak strain and protection from infection. For each of the three influenza virus serotypes the infection rate in those with no detectable antibody was approximately 80%. Those with past experience of the virus but no antibody to the outbreak strain experienced lower infection rates (62% overall) but the infection rates were lowest in those with intermediate and high level antibody to the challenge strain (18% overall). Vaccine was used by three of the schools. The effect of antibody derived from recent experience, either natural or vaccine-induced, on subsequent challenge with a drifted strain i.e. one showing antigenic drift away from the previous strain, was compared. Intermediate or high level antibody to the challenge strain in those who had experienced a recent natural infection was associated with a low infection rate (9%). A similar level of antibody produced in response to vaccination was associated with a significantly higher infection rate (23%: P less than 0.025).(ABSTRACT TRUNCATED AT 250 WORDS)

Adolescent↗
PubMed1988

Prophylactic use of amantadine in a boarding school outbreak of influenza A.

Amantadine was used in a boarding school to control an outbreak of influenza A H3N2. Of 859 pupils 79% took amantadine and almost all of them completed the course (100 mg per day for 15 days). While amantadine was being taken the number of clinical cases of influenza was considerably fewer than that predicted on the basis of previous outbreaks of influenza A at the school. However, during the month following the course of amantadine, the outbreak continued with many clinical cases confirmed by virus isolation. The advantages and limitations of amantadine prophylaxis are discussed.

Adolescent↗
PubMed1986

Infection with influenza A H1N1. 1. Production and persistence of antibody.

Three outbreaks of influenza caused by influenza A H1N1 occurred in a boys' boarding school in 1978, 1979 and 1983. The serological response to infection with variants of the H1N1 virus was studied by radial haemolysis and haemagglutination inhibition after primary infection and reinfection. The persistence of this antibody was also studied. Infection in 1978 resulted in the production of persistent antibody to both the haemagglutinin and neuraminidase of the homotypic strain. Antibody which cross-reacted with later variants of the virus was less frequently produced, the peak response was delayed and such antibody persisted less well. Infection in 1979 resulted in a similar response to that observed in 1978 after primary infection. Reinfection resulted in a broad response in all cases. In 1983 all infected boys produced antibody which reacted with the homotypic strain but only approximately one-third of primary infections produced antibody which reacted with the A/USSR/92/77 strain. The neuraminidase of the A/USSR strain failed to detect one third of the primary infections. Reinfection again resulted in a broad response.

Adolescent↗
PubMed1986

Infection with influenza A H1N1. 2. The effect of past experience on natural challenge.

Following its reintroduction in 1978 influenza A H1N1 spread widely in the child population. By the autumn of 1979, 75% of 11-year olds entering a boys' boarding school had detectable antibody. The protective effect of previous experience could be assessed during two outbreaks in the school. In the first outbreak in 1979, 90% of those known to have been infected in the previous year were protected against reinfection. In 1983 after strains of the H1N1 subtype had undergone antigenic drift a large outbreak occurred. It was estimated that past infection conferred protection against clinical influenza in 55%. Where past infection resulted in the presence of antibody which reacted with the outbreak strain the attack rate was further reduced. A large number of sub-clinical infections was detected in all groups.

Adolescent↗
PubMed1986

The effect of tobacco smoke upon airway secretion in the cat.

We have collected secretions from a segment of trachea in the neck of cats anaesthetized with chloralose. At the start of experiments sodium [35S]sulphate and [3H]glucose were given into the segment to radiolabel secretions by biosynthesis. Subsequently, changes in output of radiolabelled macromolecules were used to indicate changes in mucin secretion rate. Cigarette smoke, diluted to 10% by volume, was passed through the larynx or through the tracheal segment, or given into the lower airway. Control insufflations of air were given by the same routes. When cigarette smoke was passed through the larynx, it did not change the secretion rate of radiolabelled mucins into the tracheal segment. Cigarette smoke, given directly into the tracheal segment, stimulated the output of 35S-labelled mucins. Cutting the autonomic nerves to the segment failed to alter the response to cigarette smoke, but ganglion blockade with pentolinium abolished it. Cigarette smoke given into the lower airways and lungs stimulated the output of radiolabelled mucins into the segment. Denervation of the segment failed to prevent this, but ganglion blockade abolished the response. Measurements of plasma nicotine concentrations in some experiments showed that nicotine was absorbed from smoke in the lower airway and also suggested some absorption of nicotine from the tracheal segment. We conclude that nicotine, absorbed either directly into the tracheal wall or via the blood, can stimulate the autonomic ganglia which innervate airway submucosal mucous glands. This mechanism outweighed the effect of airway irritant reflexes in these experiments.

Animals↗