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Biomedical subjects

H Embaye

Publications and source records attributed to H Embaye.

11 recordsLinked to original sources
PubMed1998

Evidence for a compensatory increase in synthesis of brush border enzymes in the jejunum of Irish Setters with gluten-sensitive enteropathy.

OBJECTIVE: To determine whether biosynthesis of aminopeptidase N (ApN), alkaline phosphatase (AP), and total microvillus membrane protein is altered in Irish Setters with gluten-sensitive enteropathy (GSE). ANIMALS: A litter of 6 Irish Setters with GSE and 3 healthy Greyhounds. PROCEDURES: Explants obtained from affected dogs at 4 and 12 months were maintained in vitro and were compared with material from healthy control dogs. Biosynthesis of ApN and AP was monitored by incorporation of [35S]methionine and immunoprecipitation of these enzymes. RESULTS: Jejunal explants from affected Irish Setters had significantly higher rates of biosynthesis of total protein, microvillus membrane protein, AP, and ApN, compared with control tissue. Two forms of ApN with apparent molecular mass of 155 and 135 kd and 4 forms of AP with apparent molecular mass of 210 to 260, 150, 130, and 105 kd were identified in total membrane fractions from control and affected dogs. CONCLUSIONS: Reduced activities of ApN and AP in dogs with GSE are not attributable to decreased synthesis of these proteins and document enhanced synthesis of microvillar membrane proteins, which may be a compensatory response to enterocyte damage. The 150-kd form of AP was most prominent in tissue from the most affected dogs, probably representing an early form of this enzyme. In contrast, the 105-kd form was most intense in tissue from controls and less intense in tissue of affected dogs.

Alkaline Phosphatase↗
PubMed1997

Surface properties of diarrhoeagenic Escherichia coli isolates.

The surface properties of various Escherichia coli isolates associated with diarrhoeal illness were compared by aqueous partitioning between polyethylene glycol (PEG) and Dextran phases. Two well characterised strains of enteropathogenic E. coli (EPEC) were found to be very hydrophobic, based on the critical polymer concentration. EPEC strain E2348 cured of the EPEC adherence factor (EAF) plasmid had much reduced surface hydrophobicity. Partitioning of a series of diarrhoeagenic E. coli strains demonstrated that the majority of EAF+ EPEC strains were significantly more hydrophobic than EAF- EPEC strains. E. coli strains defined as enteroaggregative on the basis of hybridisation with a specific DNA probe showed much greater heterogeneity in their partitioning behaviour, possibly indicating that the AAF/I pili were not expressed in all strains. The E. coli K-12 strain used as a transformation host for adhesion studies had very low surface hydrophobicity but had a detectable negative charge. No alteration in these properties was observed when transformed with EPEC and recombinant plasmids known to specify adherence to tissue culture cells.

Agglutination Tests↗
PubMed1996

Growth and propagation of normal rat intestinal epithelial cells.

A combination of mild proteolytic digestion and selective growth stimulation has been used to isolate and propagate adult rat intestinal epithelial cells with a finite life span. Growth of these cells on a variety of matrices and on mesenchymal cells has resulted in the expression of brush border enzymes including sucrase-isomaltase, aminopeptidase N, and alkaline phosphatase. Examination of the cells at the electron microscopic level has revealed that although these cells express key brush border enzymes, they do not have a fully formed brush border. These findings suggest that the expression of brush border enzymes and structural proteins represent distinct stages of enterocyte differentiation that are under separate transcriptional and temporal control.

Alkaline Phosphatase↗
PubMed1995

Effects of concanavalin A and pokeweed lectins on microvillar membrane proteins during the organ culture of rabbit intestinal mucosa.

The effects of pokeweed lectin (PWL) and concanavalin A (Con A) on microvillar membrane (MVM) proteins during the organ culture of rabbit ileal explants for 24 hours were compared with the known effects of enteropathogenic Escherichia coli (EPEC). PWL resulted in the accelerated release of brush border enzymes into the culture medium, accompanied by decreased tissue activities and an increase in the total activity present in the tissue and culture medium. Con A had less effect on the release and tissue activities of brush border enzymes and the total activity was not increased. Sucrose density gradient centrifugation of the culture medium showed that MVM enzymes were predominantly particulate, consistent with their release as vesicles. Centrifugation of ileal explants showed that PWL, but not Con A, resulted in a decrease in the modal density of the brush border which was consistent with a lower glycoprotein-to-lipid ratio. These findings suggest that PWL, in common with EPEC, may cause the disruption and vesiculation of microvilli and the compensatory stimulation of MVM protein synthesis.

Animals↗
PubMed1995

Application of organ culture of small intestine to the investigation of enterocyte damage by equine rotavirus.

We used organ culture of jejunal mucosal explants obtained from ponies aged between 2 and 12 months to study enterocyte damage by group A strains of equine rotavirus. Electron microscopy of jejunal explants maintained for < or = 48 h in the presence of organ culture medium alone showed that enterocytes were structurally intact and had a densely packed brush border and overlying mucus. Similarly, examination of explants maintained in the presence of rotavirus for 48 h revealed no apparent ultrastructural abnormalities. However, obvious replication and assembly of virus in enterocytes had occurred following exposure to equine rotavirus strains H2 (three of eight occasions), trypsin-activated H2 (two of four), L338 (five of five), and EQ124 (two of two), although brush borders and intracellular organelles appeared unaltered. Despite the absence of obvious ultrastructural damage, analytical subcellular fractionation showed that rotavirus infection can result in decreased activities of microvillar membrane enzymes, with a particular loss of the main brush border peak of modal density 1.21 g/ml. These findings demonstrate that organ culture of small intestine may be applied to the investigation of rotavirus pathogenicity; they further suggest that certain strains impair enterocyte function by affecting the turnover of microvillar membrane proteins.

Alkaline Phosphatase↗
PubMed1992

Effects of enteropathogenic Escherichia coli on microvillar membrane proteins during organ culture of rabbit intestinal mucosa.

This study examines the effects of an enteropathogenic Escherichia coli on microvillar membrane proteins during organ culture of rabbit ileal explants. Explants maintained with enteropathogenic E coli showed brush border effacement affecting approximately 50% of enterocytes, and where enteropathogenic E coli were closely adherent to the enterocyte surface microvilli were apparently being shed as vesicles. The microvillar membrane enzymes alkaline phosphatase, aminopeptidase N and alpha-glucosidase were released into the culture medium during organ culture, and this process was significantly enhanced by enteropathogenic E coli. This increased loss of microvillar membrane enzymes into the culture medium was associated with decreased tissue activities of microvillar membrane enzymes in enteropathogenic E coli infected ileal explants compared with control. For aminopeptidase N in particular, however, total enzyme activities in the tissue plus culture medium were increased comparing enteropathogenic E coli with control, suggesting that there might be an increase in the rate of synthesis of certain microvillar membrane proteins. Reorientating sucrose density gradient centrifugation of culture medium showed that alkaline phosphatase, aminopeptidase N and alpha-glucosidase were predominantly associated with particles of peak modal density 1.19 g/ml in both groups, confirming that enteropathogenic E coli accelerate release of microvillar membrane enzymes as vesicles. Analytical fractionation of ileal explants showed that enteropathogenic E coli resulted in a loss of microvillar membrane enzyme activities from the main brush border peak of modal density 1.21 g/ml present in controls. The density of the remaining smaller and lighter peak increased from 1.19 g/ml to 1.23 g/ml after homogenisation in digitonin, confirming association of these proteins with cholesterol containing membranes and not endoplasmic reticulum. These findings suggest that enteropathogenic E. coli accelerate the normal shedding of microvillar membrane proteins as vesicles, and may stimulate a compensatory increase in microvillar membrane protein synthesis.

Alkaline Phosphatase↗
PubMed1990

Histopathology of oesophagogastric lesions in pigs.

The development of parakeratotic lesions and ulcers of the oesophagogastric epithelium was studied in pigs from a variety of sources and of ages ranging from 10 to 22 weeks. In general, the severity, as assessed by gross appearance, paralleled the depth to which parakeratotic change extended into the epithelium. However, some early degenerative change and erosion was observed in epithelium graded as macroscopically normal. Separation and erosion usually occurred beneath a band of epithelial cells with cytoplasmic pallor and nuclear degeneration underlying the parakeratotic layers. These findings are discussed in relation to the possible role of diet and gastric acidity in the pathogenesis of the lesions.

Animals↗
PubMed1990

Attaching effacement of the rabbit enterocyte brush border is encoded on a single 96.5-kilobase-pair plasmid in an enteropathogenic Escherichia coli O111 strain.

An enteropathogenic Escherichia coli (EPE) O111 serotype a,b,H- strain carried the following four plasmids: pLV501 (96.5 kilobase pairs [kbp]) specifying resistance to chloramphenicol, tetracycline, and kanamycin; pLV502 (8 kbp) specifying ampicillin resistance; pLV503 (1.9 kbp) specifying streptomycin resistance; and pLV504 (80 kbp) with no resistance markers. This EPEC attached to HEp-2 cells to produce localized clumps of bacteria (localized adhesion) and attached intimately to the enterocyte surface, leading to loss of the brush border (attaching effacement). Plasmid pLV501 was also found to specify the ability to produce localized adhesion on HEp-2 cells and attaching effacement in a rabbit ileal explant model system. Restriction maps showed considerable dissimilarities between pLV501 and pMAR-2, an EPEC plasmid carrying the EPEC adherence factor (EAF) genes. Furthermore, pLV501 did not hybridize with the EAF probe, whereas pLV504 did. There was sequence homology between pLV501 and large plasmids in all seven other well-characterized EPEC, only five of which hybridized with the EAF probe. These findings indicate that pLV501 carries at least one of the genes responsible for production of the brush border damage characteristic of EPEC.

Animals↗
PubMed1989

Interaction of enteropathogenic Escherichia coli 0111 with rabbit intestinal mucosa in vitro.

A model system using rabbit intestinal mucosal explants has been developed to examine the characteristic ultrastructural damage to the brush border induced by enteropathogenic Escherichia coli 0111. In this model, as in others, bacterial adherence to the microvillous membranes occurred in two morphologically distinct stages. Initial attachment of enteropathogenic strains of E. coli to ileal mucosa appeared to be a goblet cells and the mucous layer covering the microvilli. The next stage involved binding of enteropathogenic strains of E. coli to the bases of the microvilli that became elongated and vesiculated. Eventually, large areas of brush border effacement occurred with close apposition between bacterial and enterocyte membranes, leading to cup and pedestal formation. With a relatively large inoculum of bacteria (10(8) cfu/ml) these changes occurred within 4 h, but even with much lower inocula (10(5) cfu/ml) localized areas of damage were seen within 8 h. Although the bacteriostatic antibiotic tetracycline (700 mg/L) inhibited bacterial replication, it did not prevent the characteristic damage produced by enteropathogenic strains of E. coli. Enteropathogenic strains of E. coli 0111 were able to produce attaching effacement to gastric, duodenal, jejunal, ileal, and colonic mucosa.

Animals↗