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Biomedical subjects

M Perdue

Publications and source records attributed to M Perdue.

18 recordsLinked to original sources

Stress-induced decrease of the intestinal barrier function. The role of muscarinic receptor activation.

Recently the breakdown of the barrier function of the intestinal epithelium after application of an experimental psychological and physical stress protocol in rats has been observed. Not only did smaller molecules pass from the luminal to the serosal side, but so also did larger proteins with the dimensions of luminal antigens and toxins. The increased permeability for macromolecules is primarily due to a decrease of the tightness of the zonula occludens, but an increased endocytotic uptake indicates that transcytosis is increased also. From studies of model systems it can be concluded that activation of the intracellular protein kinase C route by muscarinic receptor activation or histamine receptor activation can be one of the underlying cellular pathways. The physical pathway relaying the stress from the brain to the intestinal tract appears to be the parasympathetic branch of the autonomic nervous system. The difference in reaction of different strains suggests that coping style is an important determinant of the response of the intestinal barrier to stress.

Animals↗

Production and evaluation criteria of specific monoclonal antibodies to the hemagglutinin of the H7N2 subtype of avian influenza virus.

To enhance the rapidity in diagnosing the spread of avian influenza virus (AIV) in chicken layer flocks, studies were initiated to develop more sensitive and specific immunological and molecular methods for the detection of AIV. In this study, the purification of the hemagglutinin protein (H) from field isolates of H7N2, the production of monoclonal antibodies (MAbs), and their evaluation as diagnostic reagents are reported. Hybridomas were generated by fusion of SP2/0-Ag14 myelomas and spleen cells from immunized mice. Hybridomas secreting antibodies specific for the H protein were assayed by an ELISA and cloned using limiting dilution. The MAbs produced were characterized by hemagglutination inhibition (HI), immunohistochemistry (IHC), indirect fluorescent antibody assay (IFA), Western blots, and IFA flow cytometry using various AIV subtypes (i.e., H4N2, H5N3, H7N2). Of the various MAbs assayed, 6 had consistent and reproducible results in each of the assays used. The results obtained in this investigation enhanced the usage of the MAbs to viral H protein in the surveillance of AIV in chickens.

Animals↗

Phylogenetic analysis of H7 avian influenza viruses isolated from the live bird markets of the Northeast United States.

The presence of low-pathogenic H7 avian influenza virus (AIV), which is associated with live-bird markets (LBM) in the Northeast United States, was first detected in 1994 and, despite efforts to eradicate the virus, surveillance of these markets has resulted in numerous isolations of H7 AIVs from several states from 1994 through 1998. The hemagglutinin, nonstructural, and matrix genes from representative H7 isolates from the LBM and elsewhere were sequenced, and the sequences were compared phylogenetically. The hemagglutinin gene of most LBM isolates examined appeared to have been the result of a single introduction of the hemagglutinin gene. Evidence for evolutionary changes were observed with three definable steps. The first isolate from 1994 had the amino acid threonine at the -2 position of the hemagglutinin cleavage site, which is the most commonly observed amino acid at this site for North American H7 AIVs. In January 1995 a new genotype with a proline at the -2 position was detected, and this genotype eventually became the predominant virus isolate. A third viral genotype, detected in November 1996, had an eight-amino-acid deletion within the putative receptor binding site. This viral genotype appeared to be the predominant isolate, although isolates with proline at the -2 position without the deletion were still observed in viruses from the last sampling date. Evidence for reassortment of multiple viral genes was evident. The combination of possible adaptive evolution of the virus and reassortment with different influenza virus genes makes it difficult to determine the risk of pathogenesis of this group of H7 AIVs.

Amino Acid Sequence↗

Characterization of an avian influenza A (H5N1) virus isolated from a child with a fatal respiratory illness.

An avian H5N1 influenza A virus (A/Hong Kong/156/97) was isolated from a tracheal aspirate obtained from a 3-year-old child in Hong Kong with a fatal illness consistent with influenza. Serologic analysis indicated the presence of an H5 hemagglutinin. All eight RNA segments were derived from an avian influenza A virus. The hemagglutinin contained multiple basic amino acids adjacent to the cleavage site, a feature characteristic of highly pathogenic avian influenza A viruses. The virus caused 87.5 to 100 percent mortality in experimentally inoculated White Plymouth Rock and White Leghorn chickens. These results may have implications for global influenza surveillance and planning for pandemic influenza.

Amino Acid Sequence↗

Pavlovian conditioning of rat mucosal mast cells to secrete rat mast cell protease II.

Antigen (egg albumin) injections, which stimulate mucosal mast cells to secrete mediators, were paired with an audiovisual cue. After reexposure to the audiovisual cue, a mediator (rat mast cell protease II) was measured with a sensitive and specific assay. Animals reexposed to only the audiovisual cue released a quantity of protease not significantly different from animals reexposed to both the cue and the antigen; these groups released significantly more protease than animals that had received the cue and antigen in a noncontingent manner. The results support a role for the central nervous system as a functional effector of mast cell function in the allergic state.

Acoustic Stimulation↗

Antigen-induced lung solute clearance in rats is dependent on capsaicin-sensitive nerves.

Chemosensitive sensory nerves have an important effector role in the control of vascular permeability in rat airways after neurogenic inflammation. To investigate whether they also have a role in antigen-induced lung inflammation, we have studied the changes in lung solute clearance (LSC) in sensitized rats after aerosol challenge with allergen and the effect of prior capsaicin-induced denervation on these changes. Sprague-Dawley rats were immunized with egg albumin (EA), using aluminum hydroxide and Bordetella pertussis as adjuvants. After 11 days, the animals were challenged for 5 min with aerosolized EA, and the clearance from the lungs of aerosolized 99mTc diethylenetriamine pentaacetic acid (99mTc-DTPA) over 7.5 min (LSC 7.5) was subsequently measured at various times after challenge as an index of epithelial permeability or integrity. Sensitized animals responded to the challenge with immediate respiratory symptoms and with an increased 99mTc-DTPA clearance rate that was detectable at 20 min (mean +/- SE LSC 7.5: baseline, 6 +/- 1%; 20 min, 17 +/- 3%; p less than 0.05), persisted at 4 h (14 +/- 1%; p less than 0.05), and returned to normal values after 24 h. Unsensitized rats exposed to EA and sensitized rats exposed to PBS or to bovine serum albumin did not show any change. Bronchoalveolar lavage failed to show significant changes of cell populations until 24 h, when an increased presence of lymphocytes, PMN, and eosinophils was observed.(ABSTRACT TRUNCATED AT 250 WORDS)

Aerosols↗

Intestinal anaphylaxis: in vivo and in vitro studies of the rat proximal colon.

The response of the rat proximal colon to an immunoglobulin E (IgE)-mediated hypersensitivity reaction was examined. Rats were sensitized to egg albumin (EA) by intraperitoneal injection, and serum titers of specific anti-EA IgE were measured at 14 days. Sensitized animals had titers of greater than or equal to 1:64, whereas no anti-EA IgE antibodies were detected in controls. Water and electrolyte absorption in the proximal colon, before and during antigen challenge, was measured by in vivo marker perfusion. Antigen challenge resulted in significant inhibition of water, Na+, Cl-, and K+ absorption in vivo. Proximal colonic tissue from sensitized and control animals was studied in Ussing chambers under short-circuited conditions. Antigen challenge of sensitized tissue resulted in significant increases in short-circuit current due to the induction of active Cl- secretion. No such changes were seen in control tissue. The abnormalities induced by antigen challenge in tissue from sensitized animals was blocked by doxantrazole (10(-3) M), a mast cell stabilizer. The findings indicate that IgE-mediated reactions in rat proximal colon to a food protein cause pertubations in water and electrolyte transport secondary to active Cl- secretion and these abnormalities appear to be due to mast cell degranulation.

Anaphylaxis↗

Inflammatory cells and the epithelium. Mast cell/nerve interactions in the lung in vitro and in vivo.

A hypothesis is presented that mast cells in and below the epithelium of the respiratory tract show functional association with nerves to form a homeostatic regulatory unit. During inflammation, mast cells may arise in situ as well as by infiltration because epithelium contains both mast cell precursors and produces factors that support their growth in vitro. Structural studies show that mast cells associate with nerves in the lung. Using a tissue culture model, we showed that sympathetic nerves formed lasting contacts with rat basophilic leukemia (RBL) cells. Electrophysiologic studies showed that nerve contact increases RBL membrane conductance, which can be mimicked by exogenous substance P (SP). Experiments with sensitized rat tracheal mucosa in Ussing chambers showed functional evidence of interaction of mast cells with SP-containing nerves: changes in short circuit current caused by antigen were blocked by the mast cell stabilizer doxantrazole and reduced by 50% by neonatal pretreatment with capsaicin. Experiments in vivo showed that lung clearance of the aerosol probe 99mTc-DTPA was increased by antigen challenge in sensitized rats. This was blocked by neonatal capsaicin treatment, again implicating SP-containing nerves. Therefore, we conclude that the functional association of mast cells with nerves is an important mechanism in regulating the local epithelial environment.

Animals↗

Acute and chronic models for hypersecretion of intestinal mucin.

Mucin secretion was examined in three functional models relevant to human disease, using rat small intestinal rings or in situ loops, [3H]glucosamine precursor labelling, gel chromatography and a specific radioimmunoassay for mucin. As a model for acute bacterial secretory diarrhoea, tissues were exposed to cholera toxin for up to 4 h. Both stored and newly synthesized radioactive glycoproteins were secreted in amounts twofold to threefold above control levels. Immunoreactive mucin secretion increased fivefold to eightfold. Other agents known to raise cAMP levels did not stimulate mucin secretion, suggesting that cholera may release mucin by a non-cAMP-dependent mechanism. Sepharose 2B chromatography indicated that secreted mucin was smaller in size than intracellular mucin and had compositional differences suggestive of 'immaturity' or protein contamination. In chronically (seven days) reserpinized rats, used as a model of glycoprotein abnormalities relevant to cystic fibrosis, mucin secretion increased twofold to threefold, but the most prominent abnormality was a marked increase in [3H]glucosamine incorporation into all tissue glycoproteins. On purification, the intracellular mucin of reserpine-treated rats had the same composition as mucin from control rats, but the former was smaller in size and had a higher specific radioactivity. Mucin hypersecretion in reserpinized rats may therefore be secondary to a primary and chronic hyperstimulation of mucin biosynthesis. A model of intestinal 'anaphylaxis' or immune-mediated diarrhoea was created in Hooded Lister rats by immunizing with egg albumin (10 micrograms) and challenging with the same antigen in intestinal loops 14 days later. After 4 h, total protein, DNA and brush border sucrase were increased in the lumen. Enhancement of mucin secretion did not occur, however, and therefore does not seem to be a particular feature of the pathophysiology of this model.

Anaphylaxis↗

Postnatal development of glucose transport in the proximal small intestine of the rabbit.

Developmental change in intestinal solute transport was examined by studying glucose transport in rabbit jejunal tissue at age intervals after birth. Net D-glucose absorption was greater in jejunum from suckling rabbits (10-14 days old) compared to older animals when studied in Ussing-type short-circuited chambers, and jejunal tissue slices from these younger animals accumulated more intracellular D-glucose. Net fluxes of 3-0-methyl-D-glucose were also greater in jejunum from the younger age group, excluding this effect being due to substrate metabolism. Increased permeability of jejunal epithelium in immature animals was evidenced by larger unidirectional fluxes and higher conductance. The findings suggest that the developing jejunum, although more permeable, has an increased capacity for active sugar transport, a pattern differing from that seen with most other intestinal absorptive functions.

3-O-Methylglucose↗

Intestinal glucose transport in acute viral enteritis in piglets.

1. We studied intestinal glucose transport in pigs during the acute and convalescent phases of an invasive viral enteritis, transmissible gastroenteritis. 2. When diarhoea was severe 40 h after experimental infection, net absorption of glucose, Na+ and water, measured by marker perfusion in the jejunum, was reduced; the enhancement of Na+ and water absorption in response to increasing perfusate glucose concentrations up to 120 mmol/l was diminished compared with the response observed in control and convalescent pigs. 3. Measured in vitro, 40 h after infection, unidirectional fluxes of 3-O-methyl-D-glucose across the jejunal epithelium were reduced and net absorption of the sugar was obliterated. Phlorizin (0.05 mmol/l), which completely inhibited net 3-O-methyl-D-glucose absorption in control tissue, had no significant effect on transmissible gastroenteritis jejunum. 4. Our data suggest that in this invasive viral enteritis, which closely resembles human rotavirus enteritis, glucose absorption is impaired as a result of defects in both active and passive glucose flux. 5. Differences between the mechanisms of viral diarrhoea, demonstrated by our study and those of the enterotoxigenic diarrhoeas, should be taken into consideration in formulating active therapeutic measures for children with acute viral diarrhoea.

3-O-Methylglucose↗

Fire in the OR.

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Accident Prevention↗