984 resultados para epsilon toxin
Resumo:
Among the goals of the Brazilian soybean improvement programmes, the breeding strategies for cultivars adapted to low latitudes have been included to extend crop areas and to increase production. Seeds of nine Brazilian soybean cultivars adapted to low latitudes were investigated regarding to their composition, and amino acid and antinutritional/toxic protein contents. Protein (394.5 ± 13.1 to 445.3 ± 8.0 g kg-1 dry matter) and oil (200.6 ± 1.2 to 232.3 ± 4.7 g kg-1 dry matter) contents showed low correlation to each other (r = -0.06). The total carbohydrate (141.7 ± 6.1 to 211.1 ± 15.0 g kg-1 dry matter) and ash contents (48.2 ± 4.2 to 52.2 ± 0.5 g kg-1 dry matter) were similar to data available for other soybean cultivars. All soybean cultivars presented low levels of tryptophan and sulphur amino acids. The lectin (1,152 to 147,456 HU kg-1 flour), trypsin inhibitor (34.45 ± 2.28 to 77.62 ± 2.63 g trypsin inhibited kg-1 flour), toxin (6,210 ± 134 to 34,650 ± 110 LD50 kg-1 flour) and urease (0.74 ± 0.02 to 1.22 ± 0.10 g kg¹ flour) presented variations in their contents amongst the cultivars. Compared to other soybean cultivars, urease was higher, the acute toxicity lower and the lectin and trypsin inhibitor contents similar to data available. In general, soybean cultivars showed similar biochemical composition to those developed in different geographic regions. The relevance of these findings to the agronomic features and to choice of soybean cultivars to be used as food or feed is discussed.
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The possibility of producing neutralizing antibodies against the lethal effects of scorpion toxins was evaluated in the mouse model by immunization with an immunogen devoid of toxicity. A toxic fraction (5 mg) from the venom of the scorpion Tityus serrulatus was entrapped in sphingomyelin-cholesterol liposomes. The liposomes were treated for 1 h at 37oC with a 1% (w/w) trypsin solution in 0.2 M sodium carbonate buffer, pH 8.3. This treatment led to a strong reduction in venom toxicity. Immunization was performed as follows: mice were injected sc with 20 µg of the liposome-entrapped toxic fraction on days 1 and 21 and a final injection (20 µg) was administered ip on day 36. After injection of the immunogen, all mice developed an IgG response which was shown to be specific for the toxic antigen. The antibodies were measured 10 days after the end of the immunization protocol. In an in vitro neutralization assay we observed that pre-incubation of a lethal dose of the toxic fraction with immune serum strongly reduced its toxicity. In vivo protection assays showed that mice with anti-toxin antibodies could resist the challenge with the toxic fraction, which killed, 30 min after injection, all non-immune control mice
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Cytotoxin production was studied in 60 Serratia marcescens strains isolated from hospitalized patients. Association of cytotoxic activity with serotype, source of isolation and presence of plasmids was also evaluated. Thirteen of the 60 S. marcescens strains produced a cytotoxic effect on Vero cells. These strains were isolated from distinct clinical sources and classified into seven different serotypes (O1:H7; O4:NM; O10:NT; O19:NM; O6,14:H4; O6,14:NM and O6,14:H1). No relationship was observed between cytotoxic activity and clinical source or serotypes of the strains. Plasmids from five cytotoxin-producing S. marcescens strains were transferred to E. coli K12/711. The transconjugants did not exhibit cytotoxicity, indicating that the cytotoxic effect is not plasmid-mediated among these strains. Although a cytotoxic activity was demonstrated in filtrates of some S. marcescens strains, further studies should be performed to assess the role of this toxin in pathogenesis
Resumo:
The oxyntic mucosa of the mouse stomach is lined with a heterogeneous population of cells that form numerous short pits continuous with long tubular glands. Tritiated thymidine radioautography has made it possible to pinpoint the origin of all cell types and to follow the differentiation/migration of different cell lineages along the pit-gland unit. The proliferating multipotent stem cells functionally anchored in the upper glandular region, the isthmus, give rise to three main lineage precursors: 1) pre-pit cells, which migrate upward to the pit while differentiating into mucus-producing pit cells; 2) pre-neck cells, which migrate downward to the glandular neck while differentiating into mucus-producing neck cells that, by approaching the glandular base, gradually change their phenotype into pepsinogen- and intrinsic factor-producing zymogenic cells; 3) pre-parietal cells, which differentiate into acid-producing parietal cells in the isthmus and then undergo bipolar migration towards the pit and the glandular base. Thus, parietal cells are the only cells that complete their differentiation in the isthmus and then migrate to be scattered throughout the pit-gland unit. To determine whether parietal cells play a role in controlling decisions about cell fate within the pit-gland unit, the gastric epithelium has been examined in transgenic mice expressing the H,K-ATPase ß-subunit-1035 to +24/simian virus 40 large T antigen fusion gene. The blockade in parietal cell differentiation in these mice produces an amplification of lineage precursors, a marked depletion of zymogenic cells and an increase in pit cell census. Ablation of parietal cells in another transgenic mouse model expressing the H,K-ATPase ß-subunit-1035 to +24/diphtheria toxin fragment A fusion gene also produces amplification of lineage precursors, and similar effects on zymogenic and pit cell census. These findings strongly suggest that parietal cells produce regulatory signals that control the cellular differentiation program of both pit and zymogenic cell lineages, and would hopefully improve our ability to identify the cellular pathways leading to malignant transformation
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Toxic cyanobacteria are common in Portuguese freshwaters and the most common toxins are microcystins. The occurrence of microcystin-LR (MCYST-LR) has been reported since 1990 and a significant number of water reservoirs that are used for drinking water attain high levels of this toxin. Aquatic animals that live in eutrophic freshwater ecosystems may be killed by microcystins but in many cases the toxicity is sublethal and so the animals can survive long enough to accumulate the toxins and transfer them along the food chain. Among these, edible mollusks, fish and crayfish are especially important because they are harvested and sold for human consumption. Mussels that live in estuarine waters and rivers where toxic blooms occur may accumulate toxins without many significant acute toxic effects. In this study data are presented in order to understand the dynamics of the accumulation and depuration of MCYST-LR in mussels. The toxin is readily accumulated and persists in the shellfish for several days after contact. In the crayfish the toxin is accumulated mainly in the gut but is also cleared very slowly. In carps, although the levels of the toxins found in naturally caught specimens were not very high, some toxin was found in the muscle and not only in the viscera. This raises the problem of the toxin accumulation by fish and possible transfer through the food chain. The data gathered from these experiments and from naturally caught specimens are analyzed in terms of risk for human consumption. The occurrence of microcystins in tap water and the incidence of toxic cyanobacteria in fresh water beaches in Portugal are reported. The Portuguese National Monitoring Program of cyanobacteria is mentioned and its implications are discussed.
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Guanylate cyclases (GC) serve in two different signaling pathways involving cytosolic and membrane enzymes. Membrane GCs are receptors for guanylin and atriopeptin peptides, two families of cGMP-regulating peptides. Three subclasses of guanylin peptides contain one intramolecular disulfide (lymphoguanylin), two disulfides (guanylin and uroguanylin) and three disulfides (E. coli stable toxin, ST). The peptides activate membrane receptor-GCs and regulate intestinal Cl- and HCO3- secretion via cGMP in target enterocytes. Uroguanylin and ST also elicit diuretic and natriuretic responses in the kidney. GC-C is an intestinal receptor-GC for guanylin and uroguanylin, but GC-C may not be involved in renal cGMP pathways. A novel receptor-GC expressed in the opossum kidney (OK-GC) has been identified by molecular cloning. OK-GC cDNAs encode receptor-GCs in renal tubules that are activated by guanylins. Lymphoguanylin is highly expressed in the kidney and heart where it may influence cGMP pathways. Guanylin and uroguanylin are highly expressed in intestinal mucosa to regulate intestinal salt and water transport via paracrine actions on GC-C. Uroguanylin and guanylin are also secreted from intestinal mucosa into plasma where uroguanylin serves as an intestinal natriuretic hormone to influence body Na+ homeostasis by endocrine mechanisms. Thus, guanylin peptides control salt and water transport in the kidney and intestine mediated by cGMP via membrane receptors with intrinsic guanylate cyclase activity.
Resumo:
Tx1, a neurotoxin isolated from the venom of the South American spider Phoneutria nigriventer, produces tail elevation, behavioral excitation and spastic paralysis of the hind limbs after intracerebroventricular injection in mice. Since Tx1 contracts isolated guinea pig ileum, we have investigated the effect of this toxin on acetylcholine release, as well as its binding to myenteric plexus-longitudinal muscle membranes from the guinea pig ileum. [125I]-Tx1 binds specifically and with high affinity (Kd = 0.36 ± 0.02 nM) to a single, non-interacting (nH = 1.1), low capacity (Bmax 1.1 pmol/mg protein) binding site. In competition experiments using several compounds (including ion channel ligands), only PhTx2 and PhTx3 competed with [125I]-Tx1 for specific binding sites (K0.5 apparent = 7.50 x 10-4 g/l and 1.85 x 10-5 g/l, respectively). PhTx2 and PhTx3, fractions from P. nigriventer venom, contain toxins acting on sodium and calcium channels, respectively. However, the neurotoxin PhTx2-6, one of the isoforms found in the PhTx2 pool, did not affect [125I]-Tx1 binding. Tx1 reduced the [3H]-ACh release evoked by the PhTx2 pool by 33%, but did not affect basal or KCl-induced [3H]-ACh release. Based on these results, as well as on the homology of Tx1 with toxins acting on calcium channels (w-Aga IA and IB) and its competition with [125I]-w-Cono GVIA in the central nervous system, we suggest that the target site for Tx1 may be calcium channels.
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The present paper describes important features of the immune response induced by the Cry1Ac protein from Bacillus thuringiensis in mice. The kinetics of induction of serum and mucosal antibodies showed an immediate production of anti-Cry1Ac IgM and IgG antibodies in serum after the first immunization with the protoxin by either the intraperitoneal or intragastric route. The antibody fraction in serum and intestinal fluids consisted mainly of IgG1. In addition, plasma cells producing anti-Cry1Ac IgG antibodies in Peyer's patches were observed using the solid-phase enzyme-linked immunospot (ELISPOT). Cry1Ac toxin administration induced a strong immune response in serum but in the small intestinal fluids only anti-Cry1Ac IgA antibodies were detected. The data obtained in the present study confirm that the Cry1Ac protoxin is a potent immunogen able to induce a specific immune response in the mucosal tissue, which has not been observed in response to most other proteins.
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Innovative gas cooled reactors, such as the pebble bed reactor (PBR) and the gas cooled fast reactor (GFR) offer higher efficiency and new application areas for nuclear energy. Numerical methods were applied and developed to analyse the specific features of these reactor types with fully three dimensional calculation models. In the first part of this thesis, discrete element method (DEM) was used for a physically realistic modelling of the packing of fuel pebbles in PBR geometries and methods were developed for utilising the DEM results in subsequent reactor physics and thermal-hydraulics calculations. In the second part, the flow and heat transfer for a single gas cooled fuel rod of a GFR were investigated with computational fluid dynamics (CFD) methods. An in-house DEM implementation was validated and used for packing simulations, in which the effect of several parameters on the resulting average packing density was investigated. The restitution coefficient was found out to have the most significant effect. The results can be utilised in further work to obtain a pebble bed with a specific packing density. The packing structures of selected pebble beds were also analysed in detail and local variations in the packing density were observed, which should be taken into account especially in the reactor core thermal-hydraulic analyses. Two open source DEM codes were used to produce stochastic pebble bed configurations to add realism and improve the accuracy of criticality calculations performed with the Monte Carlo reactor physics code Serpent. Russian ASTRA criticality experiments were calculated. Pebble beds corresponding to the experimental specifications within measurement uncertainties were produced in DEM simulations and successfully exported into the subsequent reactor physics analysis. With the developed approach, two typical issues in Monte Carlo reactor physics calculations of pebble bed geometries were avoided. A novel method was developed and implemented as a MATLAB code to calculate porosities in the cells of a CFD calculation mesh constructed over a pebble bed obtained from DEM simulations. The code was further developed to distribute power and temperature data accurately between discrete based reactor physics and continuum based thermal-hydraulics models to enable coupled reactor core calculations. The developed method was also found useful for analysing sphere packings in general. CFD calculations were performed to investigate the pressure losses and heat transfer in three dimensional air cooled smooth and rib roughened rod geometries, housed inside a hexagonal flow channel representing a sub-channel of a single fuel rod of a GFR. The CFD geometry represented the test section of the L-STAR experimental facility at Karlsruhe Institute of Technology and the calculation results were compared to the corresponding experimental results. Knowledge was gained of the adequacy of various turbulence models and of the modelling requirements and issues related to the specific application. The obtained pressure loss results were in a relatively good agreement with the experimental data. Heat transfer in the smooth rod geometry was somewhat under predicted, which can partly be explained by unaccounted heat losses and uncertainties. In the rib roughened geometry heat transfer was severely under predicted by the used realisable k − epsilon turbulence model. An additional calculation with a v2 − f turbulence model showed significant improvement in the heat transfer results, which is most likely due to the better performance of the model in separated flow problems. Further investigations are suggested before using CFD to make conclusions of the heat transfer performance of rib roughened GFR fuel rod geometries. It is suggested that the viewpoints of numerical modelling are included in the planning of experiments to ease the challenging model construction and simulations and to avoid introducing additional sources of uncertainties. To facilitate the use of advanced calculation approaches, multi-physical aspects in experiments should also be considered and documented in a reasonable detail.
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Fluid particle breakup and coalescence are important phenomena in a number of industrial flow systems. This study deals with a gas-liquid bubbly flow in one wastewater cleaning application. Three-dimensional geometric model of a dispersion water system was created in ANSYS CFD meshing software. Then, numerical study of the system was carried out by means of unsteady simulations performed in ANSYS FLUENT CFD software. Single-phase water flow case was setup to calculate the entire flow field using the RNG k-epsilon turbulence model based on the Reynolds-averaged Navier-Stokes (RANS) equations. Bubbly flow case was based on a computational fluid dynamics - population balance model (CFD-PBM) coupled approach. Bubble breakup and coalescence were considered to determine the evolution of the bubble size distribution. Obtained results are considered as steps toward optimization of the cleaning process and will be analyzed in order to make the process more efficient.
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The interaction of plasminogen, tissue plasminogen activator (t-PA) and urokinase with a clinical strain of Helicobacter pylori was studied. Plasminogen bound to the surface of H. pylori cells in a concentration-dependent manner and could be activated to the enzymatic form, plasmin, by t-PA. Affinity chromatography assays revealed a plasminogen-binding protein of 58.9 kDa in water extracts of surface proteins. Surface-associated plasmin activity, detected with the chromogenic substrate CBS 00.65, was observed only when plasminogen and an exogenous activator were added to the cell suspension. The two physiologic plasminogen activators, t-PA and urokinase, were also shown to bind to and remain active on the surface of bacterial cells. epsilon-Aminocaproic acid caused partial inhibition of t-PA binding, suggesting that the kringle 2 structure of this activator is involved in the interaction with surface receptors. The activation of plasminogen by t-PA, but not urokinase, strongly depended on the presence of cells and a 25-fold enhancer effect on the initial velocity of activation by t-PA compared to urokinase was established. Furthermore, a relationship between cell concentration and the initial velocity of activation was demonstrated. These findings support the concept that plasminogen activation by t-PA on the bacterial surface is a surface-dependent reaction which offers catalytic advantages.
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In this study we characterize the presence of muscarinic acetylcholine receptors (mAChR) in the isthmo-optic nucleus (ION) of chicks by immunohistochemistry with the M35 antibody. Some M35-immunoreactive fibers were observed emerging from the retinal optic nerve insertion, suggesting that they could be centrifugal fibers. Indeed, intraocular injections of cholera toxin B (CTb), a retrograde tracer, and double-labeling with M35 and CTb in the ION confirmed this hypothesis. The presence of M35-immunoreactive cells and the possible mAChR expression in ION and ectopic neuron cells in the chick brain strongly suggest the existence of such a cholinergic system in this nucleus and that acetylcholine release from amacrine cells may mediate interactions between retinal cells and ION terminals.
Resumo:
Serratia marcescens cytotoxin was purified to homogeneity by ion-exchange chromatography on a DEAE Sepharose Fast Flow column, followed by gel filtration chromatography on a Sephadex G100 column. The molecular mass of the cytotoxin was estimated to be about 50 kDa. Some biological properties of the cytotoxin were analyzed and compared with well-characterized toxins, such as VT1, VT2 and CNF from Escherichia coli and hemolysin produced by S. marcescens. The sensitivity of the cell lines CHO, HeLa, HEp-2, Vero, BHK-21, MA 104 and J774 to the cytotoxin was determined by the cell viability assay using neutral red. CHO and HEp-2 were highly sensitive, with massive cellular death after 1 h of treatment, followed by BHK-21, HeLa, Vero and J774 cells, while MA 104 was insensitive to the toxin. Cytotoxin induced morphological changes such as cell rounding with cytoplasmic retraction and nuclear compactation which were evident 15 min after the addition of cytotoxin. The cytotoxic assays show that 15 min of treatment with the cytotoxin induced irreversible intoxication of the cells, determined by loss of cell viability. Concentrations of 2 CD50 (0.56 µg/ml) of purified cytotoxin did not present any hemolytic activity, showing that the cytotoxin is distinct from S. marcescens hemolysin. Antisera prepared against S. marcescens cytotoxin did not neutralize the cytotoxic activity of VT1, VT2 or CNF toxin, indicating that these toxins do not share antigenic determinants with cytotoxin. Moreover, we did not detect gene sequences for any of these toxins in S. marcescens by PCR assay. These results suggest that S. marcescens cytotoxin is not related to any of these toxins from E. coli.
Resumo:
The medical records of ten pediatric patients with a clinical diagnosis of tetanus were reviewed retrospectively. The heart rate and blood pressure of all tetanus patients were measured noninvasively every hour during the first two weeks of hospitalization. Six of ten tetanus patients presented clinical evidence of sympathetic hyperactivity (group A) and were compared with a control group consisting of four children who required mechanical ventilation for diseases other than tetanus (group B). Heart rate and blood pressure simultaneously and progressively increased to a maximum by day 7. The increase over baseline was 43.70 ± 11.77 bpm (mean ± SD) for heart rate (P<0.01) and 38.60 ± 26.40 mmHg for blood pressure (P<0.01). These values were higher and significantly different from those of the control group (group B) at day 6, which had an average heart rate increase over baseline of 19.35 ± 12.26 bpm (P<0.05) and blood pressure of 10.24 ± 13.30 mmHg (P<0.05). By the end of the second week of hospitalization, in group A the increase of systolic blood pressure over baseline had diminished to 9.60 ± 15.37 mmHg (P<0.05), but the heart rate continued to be elevated (27.80 ± 33.92 bpm, P = NS), when compared to day 7 maximal values. The dissociation of these two cardiovascular variables at the end of the second week of hospitalization suggests the presence of asymmetric cardiac and vascular sympathetic control. One possible explanation for these observations is a selective and delayed action of tetanus toxin on the inhibitory neurons which control sympathetic outflow to the heart.
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The effect of the skin secretion of the amphibian Siphonops paulensis was investigated by monitoring the changes in conductance of an artificial planar lipid bilayer. Skin secretion was obtained by exposure of the animals to ether-saturated air, and then rinsing the animals with distilled water. Artificial lipid bilayers were obtained by spreading a solution of azolectin over an aperture of a Delrin cup inserted into a cut-away polyvinyl chloride block. In 9 of 12 experiments, the addition of the skin secretion to lipid bilayers displayed voltage-dependent channels with average unitary conductance of 258 ± 41.67 pS, rather than nonspecific changes in bilayer conductance. These channels were not sensitive to 4-acetamido-4'-isothiocyanatostilbene-2,2'-disulfonic acid or tetraethylammonium ion, but the experimental protocol used does not permit us to specify their characteristics.