Biomedical subjects

R J Gilbert

Publications and source records attributed to R J Gilbert.

At least 37 records · Page 2Linked to original sources

PubMed1999

Genetic programming as an analytical tool for non-linear dielectric spectroscopy.

By modelling the non-linear effects of membranous enzymes on an applied oscillating electromagnetic field using supervised multivariate analysis methods, Non-Linear Dielectric Spectroscopy (NLDS) has previously been shown to produce quantitative information that is indicative of the metabolic state of various organisms. The use of Genetic Programming (GP) for the multivariate analysis of NLDS data recorded from yeast fermentations is discussed, and GPs are compared with previous results using Partial Least Squares (PLS) and Artificial Neural Nets (NN). GP considerably outperforms these methods, both in terms of the precision of the predictions and their interpretability.

Computational Biology↗

PubMed1999

Biomechanical basis for lingual muscular deformation during swallowing.

Our goal was to quantify intramural mechanics in the tongue through an assessment of local strain during the physiological phases of swallowing. Subjects were imaged with an ultrafast gradient echo magnetic resonance imaging (MRI) pulse sequence after the application of supersaturated magnetized bands in the x and y directions. Local strain was defined through deformation of discrete triangular elements defined by these bands and was depicted graphically either as color-coded two-dimensional strain maps or as three-dimensional octahedra whose axes correspond to the principal strains for each element. During early accommodation, the anterior tongue showed positive strain (expansive) in the anterior-posterior direction (x), whereas the middle tongue showed negative strain (contractile) in the superior-inferior direction (y). During late accommodation, the anterior tongue displayed increased positive x-direction and y-direction strain, whereas the posterior tongue displayed increased negative y-direction strain. These findings were consistent with contraction of the anterior-located intrinsic muscles and the posterior-located genioglossus and hyoglossus muscles. During propulsion, posterior displacement of the tongue was principally associated with positive strain directed in the x and y directions. These findings were consistent with posterior passive stretch in the midline due to contraction of the laterally inserted styloglossus muscle, as well as contraction of the posterior located transversus muscle. We conclude that MRI of lingual deformation during swallowing resolves the synergistic contractions of the intrinsic and extrinsic muscle groups.

Biomechanical Phenomena↗

PubMed1998

Self-interaction of pneumolysin, the pore-forming protein toxin of Streptococcus pneumoniae.

The pathogenically important cholesterol-binding pore-forming bacterial "thiol-activated" toxins (TATs) are commonly believed to be monomeric in solution and to undergo a transition on membrane binding mediated by cholesterol to an oligomeric pore. We present evidence, gained through the application of a number of biochemical and biophysical techniques with associated modelling, that the TAT from Streptococcus pneumoniae, pneumolysin, is in fact able to self-associate in solution to form the same oligomeric structures. The weak interaction leading to solution oligomerization is manifested at low concentrations in a dimeric toxin form. The inhibition of toxin self-interaction by derivatization of the single cysteine residue in pneumolysin with the thiol-active agent dithio (bis)nitrobenzoic acid indicates that self-interaction is mediated by the fourth domain of the protein, which has a fold similar to other proteins known to self-associate. This interaction is thought to have implications for the understanding of mechanisms of pore formation and complement activation by pneumolysin.

Bacterial Proteins↗

PubMed1998

The molecular mechanism of pneumolysin, a virulence factor from Streptococcus pneumoniae.

Pneumolysin, a member of the thiol-activated cytolysin family of toxins, is a virulence factor from the Gram-positive bacterium Streptococcus pneumoniae. The toxin forms large oligomeric pores in cholesterol-containing membranes of eukaryotic cells. A plethora of biochemical and mutagenesis data have been published on pneumolysin, since its initial characterization in the 1930s. Here we present an homology model of the monomeric and oligomeric forms of pneumolysin based on the recently determined crystal structure of perfringolysin O and electron microscopy data. A feature of the model is a striking electronegative surface on parts of pneumolysin that may reflect its cytosolic location in the bacterial cell. The models provide a molecular basis for understanding the effects of published mutagenesis and biochemical modifications on the toxic activity of pneumolysin. In addition, spectroscopic data are presented that shed new light on pneumolysin activity and have guided us to hypothesise a detailed model of membrane insertion. These data show that the environment of some tryptophan residues changes on insertion and/or pore formation. In particular, spectroscopic analysis of a tryptophan mutant, W433F, suggests it is the residue mainly responsible for the observed effects. Furthermore, there is no change in the secondary structure content when the toxin inserts into membranes. Finally, the basis of the very low activity shown by a pneumolysin molecule from another strain of S. pneumoniae may be due to the movements of a key domain-domain interface. The molecular basis of pneumolysin-induced complement activation may be related to the structural similarity of one of the domains of pneumolysin to Fc, rather than the presumed homology of the toxin to C-reactive protein as previously suggested.

Amino Acid Sequence↗

PubMed1998

Molecular fingerprinting defines a strain of Salmonella enterica serotype Anatum responsible for an international outbreak associated with formula-dried milk.

Molecular analyses based on plasmid profile typing and pulsed-field gel electrophoresis have defined a strain of Salmonella enterica serotype Anatum associated with the consumption of a particular brand of formula-dried milk responsible for an outbreak in late 1996/early 1997 involving 15 infants and 2 relatives in the UK, and 2 infants in France. The study has demonstrated the value of laboratory-based surveillance involving identification of the outbreak strain at the molecular level coupled with food microbiology and targeted epidemiological investigations, and has highlighted the importance of rapid communication and subsequent international collaboration through the European Union-funded Salm-Net salmonella surveillance network.

Animals↗

PubMed1998

Determination of lingual myoarchitecture in whole tissue by NMR imaging of anisotropic water diffusion.

The muscular anatomy of the tongue consists of a complex three-dimensional array of fibers, which together produce the variations of shape and position necessary for deglutition. To define the myoarchitecture of the intact mammalian tongue, we have utilized NMR techniques to assess the location and orientation of muscle fiber bundles through measurement of the direction-specific diffusional properties of water molecules. Whole sheep tongues were excised and imaged with a slice-selective stimulated-echo diffusion sequence in the midline sagittal plane, and three-dimensional diffusion tensors were determined for each voxel. The derived diffusion tensors were depicted graphically as octahedra whose long axes indicate local muscle fiber orientation. Two distinct groups of midline fibers were identified: 1) in-plane sagittal fibers originating in the posteroinferior region of the tongue, radiating with a fanlike projection anteriorly and superiorly and merging with vertically oriented fibers, and 2) cross-plane (transverse) fibers, oriented at right angles to the vertically aligned fibers, predominantly in the anterior and superior regions of the tongue. Regional comparison of diffusion anisotropy revealed uniform and parallel alignment (high anisotropy) in the posteroinferior region of the tongue, corresponding to the base of the genioglossus, and less uniform, orthogonally aligned fibers (low anisotropy) in the anterosuperior region of the tongue, corresponding to the core intrinsic muscles. These data indicate that lingual myoarchitecture, determined through direction-dependent mobility of water molecules, can be depicted as discrete regions of muscle fibers, whose orientation and extent of diffusion anisotropy predict local contractility.

Animals↗

PubMed1997

Structural and functional characterisation of two proteolytic fragments of the bacterial protein toxin, pneumolysin.

Proteolytic cleavage of the bacterial protein toxin pneumolysin with protease K creates two fragments of 37 and 15 kDa. This paper describes the purification of these two fragments and their subsequent physical and biological characterisation. The larger fragment is directly involved in the cytolytic mechanism of this pore-forming protein, via membrane binding and self-association. The smaller fragment lacks ordered structure or discernible activity.

Bacterial Proteins↗

PubMed1996

Risk factors for outbreaks of infectious intestinal disease linked to domestic catering.

The epidemiology of general outbreaks of infectious intestinal disease associated with domestic catering for large numbers is described and compared with foodborne outbreaks in other settings. From 1 January 1992 to 31 December 1994, the PHLS Communicable Disease Surveillance Centre identified 101 foodborne general outbreaks of infectious intestinal disease associated with domestic catering in England and Wales (16% of all foodborne outbreaks). Salmonella species were associated with 77 of the 101 outbreaks and S. enteriditis phage type 4 accounted for 57. Small round structured viruses were implicated in five outbreaks, Clostridium perfringens in four, Bacillus cereus in two, and Campylobacter sp and Escherichia coli in one each. No pathogen was identified in 11 outbreaks. Outbreaks occurred most commonly in summer. The commonest vehicles implicated were poultry/eggs in 44 outbreaks, desserts in 13, and meat/meat products in nine. Salad/vegetables, sauces, and fish/shellfish were each implicated in eight outbreaks. Raw shell eggs were implicated in a fifth of outbreaks. Inappropriate storage was the commonest fault, reported in association with 50 outbreaks (ambient temperature for long periods before serving in 29), inadequate heat treatment was reported in 35, cross contamination in 28, an infected food handler in 11, and other faults in 14. Outbreaks associated with catering on domestic premises were independently more likely than outbreaks in other settings to be associated with salmonellas, inappropriate storage of food, and consumption of poultry, eggs, or sauces. Public health services need to direct messages about the use, preparation, and storage of food to those who cater on domestic premises.

Disease Outbreaks↗

PubMed1996

An economic analysis of unilateral refusals to license intellectual property.

The intellectual property laws in the United States provide the owners of intellectual property with discretion to license the right to use that property or to make or sell products that embody the intellectual property. However, the antitrust laws constrain the use of property, including intellectual property, by a firm with market power and may place limitations on the licensing of intellectual property. This paper focuses on one aspect of antitrust law, the so-called "essential facilities doctrine," which may impose a duty upon firms controlling an "essential facility" to make that facility available to their rivals. In the intellectual property context, an obligation to make property available is equivalent to a requirement for compulsory licensing. Compulsory licensing may embrace the requirement that the owner of software permit access to the underlying code so that others can develop compatible application programs. Compulsory licensing may undermine incentives for research and development by reducing the value of an innovation to the inventor. This paper shows that compulsory licensing also may reduce economic efficiency in the short run by facilitating the entry of inefficient producers and by promoting licensing arrangements that result in higher prices.

Costs and Cost Analysis↗

PubMed1996

Rabbit sucrase-isomaltase contains a functional intestinal receptor for Clostridium difficile toxin A.

The intestinal effects of Clostridium difficile toxin A are inidated by toxin binding to luminal enterocyte receptors. We reported previously that the rabbit ileal brush border (BB) receptor is a glycoprotein with an alpha-d-galactose containing trisaccharide in the toxin-binding domain (1991. J. Clin. Invest. 88:119-125). In this study we characterized the rabbit ileal BB receptor for this toxin. Purified toxin receptor peptides of 19 and 24 amino acids showed 100% homology with rabbit sucrase-isomaltase (SI). Guinea pig receptor antiserum reacted in Western blots with rabbit SI and with the purified toxin receptor. Antireceptor IgG blocked in vitro binding of toxin A to rabbit ileal villus cell BB. Furthermore, anti-SI IgG inhibited toxin A-induced secretion (by 78.1%, P < 0.01), intestinal permeability (by 80.8%, P < 0.01), and histologic injury (P < 0.01) in rabbit ileal loops in vivo. Chinese hamster ovary cells transfected with SI cDNA showed increased intracellular calcium increase in response to native toxin (holotoxin) or to a recombinant 873-amino acid peptide representing the receptor binding domain of toxin A. These data suggest that toxin A binds specifically to carbohydrate domains on rabbit ileal SI, and that such binding is relevant to signal transduction mechanisms that mediate in vitro and in vivo toxicity.

Amino Acid Sequence↗

PubMed1996

Food poisoning: notifications, laboratory reports, and outbreaks--where do the statistics come from and what do they mean?

Three main routine sources of data on food poisoning are used in England and Wales. The first is the statutory notification system, in which clinicians notify the 'proper officer' of their local authorities of cases or suspected cases of food poisoning, and the data are collated by the Office for National Statistics. Second and third are the reporting schemes for laboratory confirmed infections and general outbreaks of infectious intestinal disease, both of which are coordinated by the PHLS Communicable Disease Surveillance Centre. This review discusses the strengths and weaknesses of the three sources.

Communicable Disease Control↗

PubMed1996

Echo-planar magnetic resonance imaging of deglutitive vocal fold closure: normal and pathologic patterns of displacement.

Abnormalities of vocal fold closure during deglutition predispose to aspiration due to impairment of airway protection. Conventional assessment of deglutitive vocal fold motion with laryngoscopy does not permit visualization through a complete adduction-abduction cycle. We determined spatiotemporal patterns of deglutitive vocal fold adduction through echo-planar magnetic resonance imaging in 15 normal volunteers and 6 patients with vocal fold paralysis. In normal volunteers, deglutitive vocal fold adduction was synchronized with laryngeal elevation, with complete vocal fold closure at the apex. Patients with unilateral vocal fold paralysis demonstrated reduced elevation and medial movement of the involved vocal fold. At maximal laryngeal elevation the uninvolved vocal fold attained a position superior to the paralyzed fold, resulting in level differences and an interglottic gap. Patients with bilateral vocal fold paralysis demonstrated reduced elevation and medial movement of both vocal folds. These findings indicate that normal and abnormal patterns of vocal fold displacement can be distinguished noninvasively through the use of echo-planar imaging.

Adult↗

PubMed1995

Respiratory phase resetting and airflow changes induced by swallowing in humans.

1. Relationships between the timing of respiration and deglutition were studied in thirty awake healthy subjects at rest. Deglutition was monitored by submental electromyography, pharyngeal manometry and videofluoroscopy. Respiration was recorded by measurement of oronasal airflow and chest wall movement. Three types of deglutition were studied: injected bolus swallows, spontaneous swallows, and visually cued swallows of boluses previously placed in the mouth. 2. The effect of each swallow on respiratory rhythm was characterized by measurement of cophase, defined as the interval between the onset of deglutitive submental EMG activity to the onset of subsequent rescheduled inspirations. Cophase was determined for swallows initiated at different phases of the respiratory cycle. In all subjects deglutition caused phase resetting of respiratory rhythm. Cophase was largest for swallows initiated near the the inspiratory-expiratory (E-I) transition and smallest for swallows initiated near the expiratory-inspiratory (E-I) transition. The pattern of respiratory resetting by deglutition was topologically classified as type 0. This pattern was shown for swallows induced by bolus injection or visual cue, and for spontaneous swallows. 3. The incidence of spontaneous deglutition was influenced by the position of the swallow in the respiratory cycle. Few spontaneous swallows were initiated near the E-I transition whereas most occurred from late inspiration to mid-expiration. 4. Deglutition caused an abrupt decrease in airflow leading to an interval of apnoea, followed by a period of expiration. The duration of deglutition apnoea for spontaneous swallows was shorter than that for 5 ml bolus swallows, and was unaffected by the respiratory phase of swallow initiation. The period of expiration after swallowing was longest for swallows initiated at the I-E transition, and shortest for E-I swallows. 5. The intervals between bolus injection and the onset of deglutition apnoea, and the timing of swallowing events, were not significantly altered by the phase in the respiratory cycle at which swallowing was exhibited. 6. To quantify the relationship between bolus flow and respiration, we determined the latencies between cessation of inspiratory airflow and arrival of the bolus at the larynx (alpha), and between laryngeal bolus departure and resumption of inspiratory airflow (delta). Both values were dependent upon the respiratory phase of swallowing. The lowest values for alpha and delta were found for early-inspiratory and late-expiratory swallows, respectively. 7. We conclude that swallowing causes respiratory phase resetting with a pattern that is characteristic of the strong perturbations of an attractor-cycle oscillator.(ABSTRACT TRUNCATED AT 400 WORDS)

Adult↗

PubMed1995

Dynamic magnetic resonance imaging of vocal cord closure during deglutition.

BACKGROUND & AIMS: Vocal cord closure is instrumental in airway protection during deglutition. Conventional imaging of vocal cord closure and reopening during deglutition requires invasive and nonphysiological methods. The aim of this study was to characterize the biomechanical properties of normal vocal cord adduction/abduction during deglutition using echoplanar magnetic resonance imaging, a technique that has the capability of imaging soft-tissue motion with real-time temporal resolution. METHODS: The movements of laryngeal and vocal cord structures during swallowing were determined in 11 normal volunteers by single slice and axially reformatted multislice coronal echoplanar images. RESULTS: During swallowing, the larynx ascended to peak elevation, maintained peak elevation for a discrete interval, and descended to its resting position. Vocal cord adduction occurred with a symmetric tent-like configuration at the midpoint of maximal laryngeal elevation, whereas vocal cord abduction occurred at the midpoint of laryngeal descent. Spatial analysis of vocal cord configuration during adduction determined that the vocal cords attained an initial parallel configuration during ascent, followed by closure at peak laryngeal elevation. CONCLUSIONS: These results show that the vocal cords adduct and abduct synchronously with laryngeal ascent and descent, respectively, during deglutition. Echoplanar magnetic resonance imaging constitutes a novel tool used to assess clinical abnormalities of deglutitive laryngeal function.

Adult↗

PubMed1995

Salmonella enteritidis phage type 4 isolates more tolerant of heat, acid, or hydrogen peroxide also survive longer on surfaces.

In a comparative study of different Salmonella enteritidis phage type 4 isolates we found that those isolates with enhanced heat tolerance also survived better than isolates that were heat sensitive either at pH 2.6, in 10 mM H2O2, or on surfaces. Culture to the stationary phase increased the heat tolerance of all isolates and the acid and H2O2 tolerance of heat-tolerant isolates. With heat-sensitive isolates, however, extended culture had no impact on survival in H2O2 and only a marginal impact on acid tolerance. The growth phase had no appreciable impact on the surface survival of any of the isolates.

Animals↗

PubMed1995

Involvement of Ras-related Rho proteins in the mechanisms of action of Clostridium difficile toxin A and toxin B.

Toxins A and B of Clostridium difficile are responsible for pseudomembranous colitis, a disease that afflicts a substantial number of hospitalized patients treated with antibiotics. A major effect of these proteins is the disruption of the actin cytoskeleton. Recently, I. Just, G. Fritz, K. Aktories, M. Giry, M. R. Popoff, P. Boquet, S. Hegenbarth, and C. von Eichel-Streiber (J. Biol. Chem. 269:10706-10712, 1994) implicated Rho proteins as cellular targets of C. difficile toxin B, since pretreatment of cells or purified Rho with toxin prevented subsequent ADP-ribosylation of Rho by exoenzyme C3. Moreover, they showed that overexpression of Rho proteins in cells suppressed cell rounding normally associated with exposure of cells to C. difficile toxin B. Here we expand these findings by showing directly that Rho proteins are covalently modified by both C. difficile toxins A and B. In addition, we demonstrate that the stability of toxin-modified Rho in NIH 3T3 cells is dramatically reduced. Finally, we show that C. difficile toxins A and B do not have similar effects on the closely related Rac and CDC42 GTP-binding proteins.

3T3 Cells↗

PubMed1995

Clostridium difficile toxin B activates calcium influx required for actin disassembly during cytotoxicity.

The principal cellular response to Clostridium difficile toxin B, a protein toxin associated with antibiotic-associated colitis, is the disassembly of actin microfilaments. Although receptor-activated signal transduction mechanisms have been proposed to mediate these effects, the intracellular events that precede actin breakdown are unknown. In NIH-3T3 fibroblasts, toxin B induced an elevation of intracellular calcium possessing either a slow (minutes) or fast (seconds) rise time, followed by a sustained elevation of calcium concentration. Subcellular analysis of steady-state calcium distribution after toxin B demonstrated that the increase of calcium was homogeneous throughout the cytosol and did not vary based on the kinetics of the initial calcium rise. All calcium responses were blocked by substitution with calcium-free buffer or buffer containing lanthanum chloride, indicating that the rise in calcium was attributable to calcium influx from the extracellular space. Quantitatively similar responses were observed in primary cultured gastric smooth muscle and AR42J pancreatic tumor cells, suggesting that toxin-induced calcium signal transduction was conserved between cell types. The morphological response to toxin B consisted of sequential dissociation of the actin cytoskeleton from membrane attachments, retraction of actin stress fibers from the periphery to the perinuclear region, loss of fibre alignment, and cell rounding. The actin reorganization associated with toxin B was blocked by incubation of cells in calcium-free media or the clamping of intracellular calcium with cell-permeant calcium chelating agents. These results demonstrate that the calcium influx activated by C. difficile toxin B is a necessary condition for the breakdown of filamentous actin associated with cytotoxicity.

3T3 Cells↗

PubMed1994

Substitution of arginine 719 for glutamic acid in human plasminogen substantially reduces its affinity for streptokinase.

In isolation human plasminogen possesses no enzymatic activity, yet upon formation of an equimolar complex with the bacterial protein streptokinase, it acquires a plasminogen activator function. The region(s) of plasminogen and of streptokinase which mediate complex formation has (have) not been previously published. Here it is reported that a single-residue substitution (Arg719-->Glu) in the serine protease domain of full-length Glu-plasminogen substantially reduces its affinity for streptokinase. The plasminogen variant displays no other significant differences from the wild-type molecule with respect to activation by two-chain urokinase-type plasminogen activator, recognition by monoclonal antibodies, or ability to undergo conformational change. It is concluded that Arg719 in human plasminogen is an important determinant of the streptokinase binding site, although further sites are likely to contribute both to the affinity of plasminogen for streptokinase and to mechanisms by which the active site is formed within the complex.

Animals↗