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Biomedical subjects

B Vincent

Publications and source records attributed to B Vincent.

At least 37 records · Page 2Linked to original sources
PubMed1999

Evaluation of the protective immunogenicity of the N, P, M, NV and G proteins of infectious hematopoietic necrosis virus in rainbow trout oncorhynchus mykiss using DNA vaccines.

The protective immunogenicity of the nucleoprotein (N), phosphoprotein (P), matrix protein (M), non-virion protein (NV) and glycoprotein (G) of the rhabdovirus infectious hematopoietic necrosis virus (IHNV) was assessed in rainbow trout using DNA vaccine technology. DNA vaccines were produced by amplifying and cloning the viral genes in the plasmid pCDNA 3.1. The protective immunity elicited by each vaccine was evaluated through survival of immunized fry after challenge with live virus. Neutralizing antibody titers were also determined in vaccinated rainbow trout Oncorhynchus mykiss fry (mean weight 2 g) and 150 g sockeye salmon Oncorhynchus nerka. The serum from the 150 g fish was also used in passive immunization studies with naive fry. Our results showed that neither the internal structural proteins (N, P and M) nor the NV protein of IHNV induced protective immunity in fry or neutralizing antibodies in fry and 150 g fish when expressed by a DNA vaccine construct. The G protein, however, did confer significant protection in fry up to 80 d post-immunization and induced protective neutralizing antibodies. We are currently investigating the role of different arms of the fish immune system that contribute to the high level of protection against IHNV seen in vaccinated fish.

Animals↗
PubMed1999

Confocal microscopy reveals thimet oligopeptidase (EC 3.4.24.15) and neurolysin (EC 3.4.24.16) in the classical secretory pathway.

Thimet oligopeptidase (EC 3.4.24.15; EP24.15) and neurolysin (EC 3.4.24.16; EP24.16) are closely related enzymes involved in the metabolic inactivation of bioactive peptides. Both of these enzymes were previously shown to be secreted from a variety of cell types, although their primary sequence lacks a signal peptide. To investigate the mechanisms responsible for this secretion, we examined by confocal microscopy the subcellular localization of these two enzymes in the neuroendocrine cell line AtT20. Both EP24.15 and EP24.16 were found by immunohistochemistry to be abundantly expressed in AtT20 cells. Western blotting experiments confirmed that the immunoreactivity detected in the soma of these cells corresponded to previously cloned isoforms of the enzymes. At the subcellular level, both enzymes colocalized extensively with the integral trans-Golgi network protein, syntaxin-6, in the juxtanuclear region. In addition, both EP24.15 and EP24.16 were found within small vesicular organelles distributed throughout the cell body. Some, but not all, of these organelles also stained positively for ACTH. These results demonstrate that both EP24.15 and EP24.16 are present within the classical secretory pathway. Their colocalization with ACTH further suggests that they may be targeted to the regulated secretory pathway, even in the absence of a signal peptide.

Animals↗
PubMed1999

[Exposure and good practice in helical computed tomography].

Helical CT is the imaging modality that delivers the highest exposure to patients. For each acquisition, the average radiation dose is between 20 and 30 mGy. This dose can potentially be reduced by decreasing the intensity or voltage and by increasing the pitch. Helical data can be reformatted to obtain images with smaller increment or multiplanar reconstructions, hence reducing the need for additional acquisitions. Hardware and software devices designed for dose reduction must be systematically used. Operators of CT units should be aware of the radiation dose delivered with helical CT and must carefully assess the need of each additional acquisition. Obsolete protocols such as angulation of the gantry for lumbar CT, which increases patient exposure, should no longer be used. Rational use of helical CT decreases radiation exposure and is faster, whereas improper use increases radiation exposure without added benefit.

Filtration↗
PubMed1998

Preparation of Poly(methylmethacrylate) Microcapsules with Liquid Cores.

Particles with liquid cores and solid shells have been prepared by the controlled phase separation of poly(methylmethacrylate) (PMMA) within the droplets of an oil-in-water emulsion. The oil phase of the emulsion contained poly(methylmethacrylate), a good solvent for the polymer (CH2Cl2), a poor solvent (hexadecane, decane, octanol, or tetrachloromethane), and in some cases acetone (a water soluble co-solvent) to aid emulsification. Emulsions were prepared using a Silverson high-speed stirrer, and the droplet size distributions were determined using a Coulter particle counter. Size distributions were found to be dependent on the nature of the emulsifier, the concentration of acetone in the oil phase, and the concentration of polymer in the oil phase. The good solvent was then removed under reduced pressure, causing the poly(methylmethacrylate) to phase separate within the emulsion droplets. The resultant two-phase particles were characterized by optical microscopy and scanning electron microscopy. Particle morphologies depended strongly on the nature of the non-solvent and also the emulsifier employed. Spreading coefficients were calculated from interfacial tension and contact angle measurements, and were used to account for the morphologies observed. Core/shell microcapsules were formed when hexadecane or decane was used as non-solvent, and only when polymeric emulsifiers were employed. All other combinations yielded "acorn"-shaped particles. The thickness of microcapsule walls was found to be a constant fraction of the overall capsule diameter for all microcapsule sizes and depended, as expected, on the concentration of polymer in the oil phase. Copyright 1998 Academic Press.

Journal Article↗
PubMed1998

Neuropeptide specificity and inhibition of recombinant isoforms of the endopeptidase 3.4.24.16 family: comparison with the related recombinant endopeptidase 3.4.24.15.

Endopeptidase EC 3.4.24.16 (EP24.16c, neurolysin) and thimet oligopeptidase EC 3.4.24.15 are close related members of a large family of metalloproteases. Besides their cytosolic and membrane bound form, endopeptidase EC 3.4.24.16 appears to be present in the inner membrane of the mitochondria (EP24.16m). We have overexpressed two porcine EP24.16 isoforms in E. coli and purified the recombinant proteins to homogeneity. We show here that these peptidases hydrolyse a series of neuropeptides with similar rates and at sites reminiscent of those elicited by classically purified human brain EP24.16c. All neuropeptides, except neurotensin, were similarly cleaved by recombinant endopeptidase 3.4.24.15 (EP24.15, thimet oligopeptidase), another zinc-containing metalloenzyme structurally related to EP24.16. These two EP24.16 isoforms were drastically inhibited by Pro-Ile and dithiothreitol and remained unaffected by a specific carboalkyl inhibitor (CFP-AAY-pAb) directed toward the related EP24.15. The present purification procedure of EP24.16 should allow to establish, by mutagenesis analysis, the mechanistic properties of the enzyme.

Amino Acid Sequence↗
PubMed1998

The religious ideology of the Business Roundtable.

The article is in two parts with the first part showing that the material in the New Zealand Business Roundtable documents is consistent with the contemporary, international, libertarian ideology. The second part draws parallels between this material and the characteristics shown by religious movements, including a claiming of authority from past prophets, a belief in an overarching Power, a missionary zeal to convert others, a canon of texts, a 'theodicy', a sense of bonding among believers, a 'doctrine' of humanity, and the use of ritual language. The article concludes that the documents show a self-supporting belief system built on a fallacious theoretical premise.

Administrative Personnel↗
PubMed1998

Estrogen reduces neuronal generation of Alzheimer beta-amyloid peptides.

Alzheimer's disease (AD) is characterized by the accumulation of cerebral plaques composed of 40- and 42-amino acid beta-amyloid (Abeta) peptides, and autosomal dominant forms of AD appear to cause disease by promoting brain Abeta accumulation. Recent studies indicate that postmenopausal estrogen replacement therapy may prevent or delay the onset of AD. Here we present evidence that physiological levels of 17beta-estradiol reduce the generation of Abeta by neuroblastoma cells and by primary cultures of rat, mouse and human embryonic cerebrocortical neurons. These results suggest a mechanism by which estrogen replacement therapy can delay or prevent AD.

Alzheimer Disease↗
PubMed1998

Review of counselling in a transfusion service: the London (UK) experience.

Donor (and recipient) counselling within the Transfusion Service in the UK has grown in volume and complexity over the last 10 years. The addition of new tests for donated blood and the growth of bone marrow transplantation have increased the demands on counselling staff. New initiatives, such as the HCV look-back programme, have required an extension of the skills and knowledge of staff involved in counselling.

Blood Donors↗
PubMed1998

[Acute respiratory insufficiency in patients with bronchopulmonary cancer].

Acute respiratory failure is relatively frequent in patients with bronchogenic cancer. The respiratory disorder may be related to tumor extension itself or opportunistic infection above or below the tumoral stenosis. Radiotherapy and chemotherapy can also be causal. The condition is particularly serious in patients with one lung. We discuss the different complications, their frequency and conditions of occurrence.

Acute Disease↗
PubMed1997

Emulsions from Aerosol Sprays

An electrostatic emulsification apparatus has been designed for the purpose of studying diffusion from oil droplets which have a mean size in the range of approximately 1.5-3.5 &mgr;m, with standard deviations of 40-50%. The emulsification technique involves the collection of a spray of electrically charged oil droplets onto a rotating water film which is sustained from a reservoir. In this way, emulsions with volume fractions of approximately 10(-3) are produced within several minutes at oil flow rates of around 10(-2) ml min-1. Phase-Doppler anemometry (PDA) was used to assess droplet size distributions for the sprays and emulsions. Results show that the mean emulsion droplet size was smaller than the mean spray droplet size by several orders of magnitude. At flow rates around 10(-2) ml min-1, the spray droplet size distribution was little affected by the applied potential between about -4.20 and -4.65 kV (mean droplet size between approximately 7.6 and 7.8 &mgr;m, with standard deviations of approximately 20%), whereas the mean droplet size of the corresponding emulsion decreased more rapidly with applied potential. Above an applied potential of approximately -4.30 kV, which corresponded to an emulsion droplet size below approximately 2 &mgr;m, the measured volume fraction of the emulsion decreased with respect to the volume fraction as calculated on the basis of total amount of injected oil. Copyright 1997 Academic Press. Copyright 1997Academic Press

Journal Article↗
PubMed1997

Contribution of endopeptidase 3.4.24.15 to central neurotensin inactivation.

The tridecapeptide, neurotensin elicits naloxone-insensitive analgesia after its intracebroventricular administration in mice. We used this central pharmacological effect to assess the putative contribution of the endopeptidase 3.4.24.15 to central inactivation of the peptide. By means of combinatorial chemistry, we previously designed the first potent endopeptidase 3.4.24.15 inhibitor. This agent, Z-(L,D)Phe psi(PO2CH2)(L,D)Ala-Lys-Met (phosphodiepryl 21), is shown here to behave as a fully specific endopeptidase 3.4.24.15 inhibitor, as demonstrated by the absence of effect on a series of other exo- and endopeptidases belonging to various classes of proteolytic activities present in murine brain membranes. Furthermore, central administration of phosphodiepryl 21 drastically prolongs the forepaw licking latency of mice tested on the hot plate and injected with sub-maximally active doses of neurotensin. Altogether, our results demonstrated that, in addition to endopeptidase 3.4.24.16, endopeptidase 3.4.24.15 likely contributes to the physiological termination of the neurotensinergic message in murine brain.

Analgesia↗
PubMed1997

Adsorption of Lead Ions onto N -Isopropylacrylamide and Acrylic Acid Copolymer Microgels

The interaction of hydrolyzable lead ions with thermosensitive microgel dispersions of N -isopropylacrylamide modified with a range of acrylic acid comonomer concentrations has been investigated. The hydrodynamic diameter of the microgel particles was observed, by dynamic light scattering, to reversibly alter due to changes in either temperature or pH, or due to the presence of lead ions. The hydrodynamic diameter of all the microgel particles decreases with increasing temperature. However, upon increasing the pH, the anionic microgel particles increase in diameter at a fixed temperature, while in the presence of Pb(II) at pH 5, the hydrodynamic diameter of the anionic microgel particles decreases. The size of the homopolymer microgel does not vary with pH, nor in the presence of Pb(II). The adsorption isotherms of hydrolysable Pb(II) with the microgel particles were established as a function of pH. Lead ion adsorption was observed to not significantly alter with temperature and was demonstrated to be completely reversible to pH adjustment.

Journal Article↗
PubMed1997

Examination of the role of endopeptidase 3.4.24.15 in A beta secretion by human transfected cells.

1. We have taken advantage of our recent development of highly potent and specific phosphinic inhibitors of endopeptidase 3.4.24.15 to examine the putative contribution of the enzyme in the secretion of A beta by HK293 transfected cells overexpressing the wild type and the Swedish (Sw) double mutated form of beta APP751. 2. First, we showed that HK293 cells contain a peptidase activity, the inhibition profile of which fully matches that of purified endopeptidase 3.4.24.15. Second, we established that the treatment of HK293 cells with specific phosphinic inhibitors leads to about 80% inhibition of intracellular endopeptidase 3.4.24.15 activity, indicating that these inhibitors penetrate the cells. 3. Metabolic labelling of wild type and Sw beta APP751-expressing cells, followed by immunoprecipitation of A beta-containing peptides, revealed the secretion of A beta and the intracellular formation of an A beta-containing 12 kDa product. 4. A beta secretion by Sw beta APP751 transfected cells was drastically enhanced when compared to cells expressing wild type beta APP751. This production was not affected by endopeptidase 3.4.24.15 inhibitors in either cell type. This correlates well with the observation that endopeptidase 3.4.24.15 does not cleave recombinant baculoviral Sw beta APP751, in vitro. 5. Our previous data indicated that endopeptidase 3.4.24.15 activity was reduced in the parietal cortex of Alzheimer's disease affected brains and that the enzyme probably participated, in this brain area, to the catabolism of somatostatin 1-14. However, the present work indicates that endopeptidase 3.4.24.15 does not seem to behave as a beta-secretase in HK293 transfected cells. Therefore, it is suggested that endopeptidase 3.4.24.15 could participate in the symptomatology, but probably not in the aetiology of Alzheimer's disease.

Alzheimer Disease↗
PubMed1997

Effect of a novel selective and potent phosphinic peptide inhibitor of endopeptidase 3.4.24.16 on neurotensin-induced analgesia and neuronal inactivation.

1. We have examined a series of novel phosphinic peptides as putative potent and selective inhibitors of endopeptidase 3.4.24.16. 2. The most selective inhibitor, Pro-Phe-psi(PO2CH2)-Leu-Pro-NH2 displayed a Ki value of 12 nM towards endopeptidase 3.4.24.16 and was 5540 fold less potent on its related peptidase endopeptidase 3.4.24.15. Furthermore, this inhibitor was 12.5 less potent on angiotensin-converting enzyme and was unable to block endopeptidase 3.4.24.11, aminopeptidases B and M, dipeptidylaminopeptidase IV and proline endopeptidase. 3. The effect of Pro-Phe-psi(PO2CH2)-Leu-Pro-NH2, in vitro and in vivo, on neurotensin metabolism in the central nervous system was examined. 4. Pro-Phe-psi(PO2CHH2)-Leu-Pro-NH2 dose-dependently inhibited the formation of neurotensin 1-10 and concomittantly protected neurotensin from degradation by primary cultured neurones from mouse embryos. 5. Intracerebroventricular administration of Pro-Phe-psi(PO2CH2)-Leu-Pro-NH2 significantly potentiated the neurotensin-induced antinociception of mice in the hot plate test. 6. Altogether, our study has established Pro-Phe-psi(PO2CH2)-Leu-Pro-NH2 as a fully selective and highly potent inhibitor of endopeptidase 3.4.24.16 and demonstrates, for the first time, the contribution of this enzyme in the central metabolism of neurotensin.

Analgesia↗
PubMed1997

Stably transfected human cells overexpressing rat brain endopeptidase 3.4.24.16: biochemical characterization of the activity and expression of soluble and membrane-associated counterparts.

We recently cloned endopeptidase-24.16 (neurolysin; EC 3.4.24.16), a neurotensin-degrading peptidase likely involved in the physiological termination of the neurotensinergic signal in the central nervous system and in the gastrointestinal tract. We stably transfected human kidney cells with the pcDNA3-lambda 7aB1 construction bearing the whole open reading frame encoding the rat brain peptidase. Transfectants displayed endopeptidase-24.16 immunoreactivity and exhibited QFS- and neurotensin-hydrolyzing activities, the biochemical and specificity properties of which fully matched those observed with the purified murine enzyme. Cryoprotection experiments and substrate degradation by intact plated cells indicated that transfectants exhibited a membrane-associated form of endopeptidase-24.16, the catalytic site of which clearly faced the extracellular domain. Transfected cells were unable to secrete the enzyme. Overall, our experiments indicate that we have obtained stably transfectant cells that overexpress an enzymatic activity displaying biochemical properties identical to those of purified endopeptidase-24.16. The membrane-associated counterpart and lack of secretion of the enzyme were clearly reminiscent of what was observed with pure cultured neurons, but not with astrocytes. Therefore, the transfected cell model described here could prove useful for establishing, by a mutagenesis approach, the structural elements responsible for the "neuronal" phenotype exhibited by the enzyme in transfected cells.

Animals↗
PubMed1996

Distinct properties of neuronal and astrocytic endopeptidase 3.4.24.16: a study on differentiation, subcellular distribution, and secretion processes.

Endopeptidase 3.4.24.16 belongs to the zinc-containing metalloprotease family and likely participates in the physiological inactivation of neurotensin. The peptidase displays distinct features in pure primary cultured neurons and astrocytes. Neuronal maturation leads to a decrease in the proportion of endopeptidase 3.4.24.16-bearing neurons and to a concomitant increase in endopeptidase 3.4.24.16 activity and mRNA content. By contrast, there is no change with time in endopeptidase 3.4.24.16 activity or content in astrocytes. Primary cultured neurons exhibit both soluble and membrane-associated endopeptidase 3.4.24.16 activity. The latter behaves as an ectopeptidase on intact plated neurons and resists treatments with 0.2% digitonin and Na2CO3. Further evidence for an association of the enzyme with plasma membranes was provided by cryoprotection experiments and electron microscopic analysis. The membrane-associated form of endopeptidase 3.4.24.16 increased during neuronal differentiation and appears to be mainly responsible for the overall augmentation of endopeptidase 3.4.24.16 activity observed during neuronal maturation. Unlike neurons, astrocytes only contain soluble endopeptidase 3.4.24.16. Astrocytes secrete the enzyme through monensin, brefeldin A, and forskolin-independent mechanisms. This indicates that endopeptidase 3.4.24.16 is not released by classical regulated or constitutive secreting processes. However, secretion is blocked at 4 degrees C and by 8 bromo cAMP and is enhanced at 42 degrees C, two properties reminiscent of that of other secreted proteins lacking a classical signal peptide. By contrast, neurons appear unable to secrete endopeptidase 3.4.24.16.

Animals↗