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1.
Front Microbiol ; 7: 133, 2016.
Article in English | MEDLINE | ID: mdl-26904009

ABSTRACT

Shiga toxin producing Escherichia coli may damage the central nervous system before or concomitantly to manifested hemolytic-uremic syndrome symptoms. The cerebellum is frequently damaged during this syndrome, however, the deleterious effects of Shiga toxin 2 has never been integrally reported by ultrastructural, physiological and behavioral means. The aim of this study was to determine the cerebellar compromise after intravenous administration of a sub-lethal dose of Shiga toxin 2 by measuring the cerebellar blood-brain barrier permeability, behavioral task of cerebellar functionality (inclined plane test), and ultrastructural analysis (transmission electron microscope). Intravenous administration of vehicle (control group), sub-lethal dose of 0.5 and 1 ηg of Stx2 per mouse were tested for behavioral and ultrastructural studies. A set of three independent experiments were performed for each study (n = 6). Blood-brain barrier resulted damaged and consequently its permeability was significantly increased. Lower scores obtained in the inclined plane task denoted poor cerebellar functionality in comparison to their controls. The most significant lower score was obtained after 5 days of 1 ηg of toxin administration. Transmission electron microscope micrographs from the Stx2-treated groups showed neurons with a progressive neurodegenerative condition in a dose dependent manner. As sub-lethal intravenous Shiga toxin 2 altered the blood brain barrier permeability in the cerebellum the toxin penetrated the cerebellar parenchyma and produced cell damaged with significant functional implications in the test balance.

2.
PLoS One ; 8(7): e70020, 2013.
Article in English | MEDLINE | ID: mdl-23894578

ABSTRACT

Shiga toxin 2 (Stx2)-producing Escherichia coli (STEC) causes hemorrhagic colitis and hemolytic uremic syndrome (HUS) that can lead to fatal encephalopathies. Neurological abnormalities may occur before or after the onset of systemic pathological symptoms and motor disorders are frequently observed in affected patients and in studies with animal models. As Stx2 succeeds in crossing the blood-brain barrier (BBB) and invading the brain parenchyma, it is highly probable that the observed neurological alterations are based on the possibility that the toxin may trigger the impairment of the neurovascular unit and/or cell damage in the parenchyma. Also, lipopolysaccharide (LPS) produced and secreted by enterohemorrhagic Escherichia coli (EHEC) may aggravate the deleterious effects of Stx2 in the brain. Therefore, this study aimed to determine (i) whether Stx2 affects the neurovascular unit and parenchymal cells, (ii) whether the contribution of LPS aggravates these effects, and (iii) whether an inflammatory event underlies the pathophysiological mechanisms that lead to the observed injury. The administration of a sub-lethal dose of Stx2 was employed to study in detail the motor cortex obtained from a translational murine model of encephalopathy. In the present paper we report that Stx2 damaged microvasculature, caused astrocyte reaction and neuronal degeneration, and that this was aggravated by LPS. Dexamethasone, an anti-inflammatory, reversed the pathologic effects and proved to be an important drug in the treatment of acute encephalopathies.


Subject(s)
Anti-Inflammatory Agents/pharmacology , Dexamethasone/pharmacology , Lipopolysaccharides/toxicity , Motor Cortex/blood supply , Motor Cortex/drug effects , Shiga Toxin 2/toxicity , Animals , Astrocytes/drug effects , Astrocytes/pathology , Disease Models, Animal , Drug Synergism , Female , Mice , Microvessels/drug effects , Motor Cortex/pathology , Neurons/drug effects , Neurons/pathology , Neurotoxicity Syndromes/drug therapy , Neurotoxicity Syndromes/etiology , Shiga Toxin 2/isolation & purification , Shiga-Toxigenic Escherichia coli/chemistry , Specific Pathogen-Free Organisms
3.
PLoS One ; 8(1): e55812, 2013.
Article in English | MEDLINE | ID: mdl-23383285

ABSTRACT

Infection by Shiga toxin-producing Escherichia coli causes hemorrhagic colitis, hemolytic uremic syndrome (HUS), acute renal failure, and also central nervous system complications in around 30% of the children affected. Besides, neurological deficits are one of the most unrepairable and untreatable outcomes of HUS. Study of the striatum is relevant because basal ganglia are one of the brain areas most commonly affected in patients that have suffered from HUS and since the deleterious effects of a sub-lethal dose of Shiga toxin have never been studied in the striatum, the purpose of this study was to attempt to simulate an infection by Shiga toxin-producing E. coli in a murine model. To this end, intravenous administration of a sub-lethal dose of Shiga toxin 2 (0.5 ηg per mouse) was used and the correlation between neurological manifestations and ultrastructural changes in striatal brain cells was studied in detail. Neurological manifestations included significant motor behavior abnormalities in spontaneous motor activity, gait, pelvic elevation and hind limb activity eight days after administration of the toxin. Transmission electron microscopy revealed that the toxin caused early perivascular edema two days after administration, as well as significant damage in astrocytes four days after administration and significant damage in neurons and oligodendrocytes eight days after administration. Interrupted synapses and mast cell extravasation were also found eight days after administration of the toxin. We thus conclude that the chronological order of events observed in the striatum could explain the neurological disorders found eight days after administration of the toxin.


Subject(s)
Corpus Striatum/drug effects , Corpus Striatum/ultrastructure , Shiga Toxin 2/toxicity , Administration, Intravenous , Animals , Astrocytes/drug effects , Astrocytes/pathology , Blood-Brain Barrier/drug effects , Blood-Brain Barrier/pathology , Blood-Brain Barrier/ultrastructure , Corpus Striatum/pathology , Disease Models, Animal , Edema , Male , Mast Cells/pathology , Mice , Motor Activity/drug effects , Necrosis , Neurons/drug effects , Neurons/pathology , Oligodendroglia/drug effects , Oligodendroglia/pathology , Oligodendroglia/ultrastructure , Shiga Toxin 2/administration & dosage , Synapses/drug effects , Synapses/pathology , Synapses/ultrastructure
4.
J Neuroimmunol ; 222(1-2): 48-61, 2010 May.
Article in English | MEDLINE | ID: mdl-20347160

ABSTRACT

Neurological damage caused by intoxication with Shiga toxin (Stx) from enterohemorrhagic Escherichia coli is the most unrepairable and untreatable outcome of Hemolytic Uremic Syndrome, and occurs in 30% of affected infants. In this work intracerebroventricular administration of Stx2 in rat brains significantly increased the expression of its receptor globotriaosylceramide (Gb(3)) in neuronal populations from striatum, hippocampus and cortex. Stx2 was immunodetected in neurons that expressed Gb(3) after intracerebroventricular administration of the toxin. Confocal immunofluorescence of microtubule-associated protein 2 showed aberrant dendrites in neurons expressing increased Gb(3). The pro-apoptotic Bax protein was concomitantly immunodetected in neurons and other cell populations from the same described areas including the hypothalamus. Confocal immunofluorescence showed that Gb(3) colocalized also with glial fibrillary acidic protein only in reactive astrocytic processes, and not in vehicle-treated normal ones. Rats showed weight variation and motor deficits as compared to controls. We thus suggest that Stx2 induces the expression of Gb(3) in neurons and triggers neuronal dysfunctions.


Subject(s)
Brain/drug effects , Dendrites/drug effects , Neurotoxicity Syndromes/microbiology , Shiga Toxin 2/toxicity , Trihexosylceramides/agonists , Animals , Apoptosis/drug effects , Apoptosis/physiology , Biomarkers/metabolism , Brain/metabolism , Brain/pathology , Chlorocebus aethiops , Dendrites/metabolism , Dendrites/pathology , Escherichia coli Infections/complications , Glial Fibrillary Acidic Protein/metabolism , Immunohistochemistry , Injections, Intraventricular , Male , Microscopy, Confocal , Microtubule-Associated Proteins/metabolism , Nerve Degeneration/metabolism , Nerve Degeneration/microbiology , Nerve Degeneration/pathology , Neurotoxicity Syndromes/metabolism , Neurotoxicity Syndromes/pathology , Rats , Rats, Sprague-Dawley , Trihexosylceramides/metabolism , Up-Regulation/drug effects , Up-Regulation/physiology , Vero Cells , bcl-2-Associated X Protein/drug effects , bcl-2-Associated X Protein/metabolism
5.
PLoS One ; 4(9): e7065, 2009 Sep 18.
Article in English | MEDLINE | ID: mdl-19763257

ABSTRACT

Epsilon toxin is a potent neurotoxin produced by Clostridium perfringens types B and D, an anaerobic bacterium that causes enterotoxaemia in ruminants. In the affected animal, it causes oedema of the lungs and brain by damaging the endothelial cells, inducing physiological and morphological changes. Although it is believed to compromise the intestinal barrier, thus entering the gut vasculature, little is known about the mechanism underlying this process. This study characterizes the effects of epsilon toxin on fluid transport and bioelectrical parameters in the small intestine of mice and rats. The enteropooling and the intestinal loop tests, together with the single-pass perfusion assay and in vitro and ex vivo analysis in Ussing's chamber, were all used in combination with histological and ultrastructural analysis of mice and rat small intestine, challenged with or without C. perfringens epsilon toxin. Luminal epsilon toxin induced a time and concentration dependent intestinal fluid accumulation and fall of the transepithelial resistance. Although no evident histological changes were observed, opening of the mucosa tight junction in combination with apoptotic changes in the lamina propria were seen with transmission electron microscopy. These results indicate that C. perfringens epsilon toxin alters the intestinal permeability, predominantly by opening the mucosa tight junction, increasing its permeability to macromolecules, and inducing further degenerative changes in the lamina propria of the bowel.


Subject(s)
Bacterial Toxins/metabolism , Enterotoxemia/microbiology , Intestine, Small/drug effects , Permeability/drug effects , Animals , Electrophysiology , Enterocytes/metabolism , Female , Intestinal Mucosa/drug effects , Male , Mice , Mice, Inbred BALB C , Microscopy, Electron, Transmission/methods , Rats , Rats, Wistar
6.
J Nephrol ; 21(6): 947-58, 2008.
Article in English | MEDLINE | ID: mdl-19034881

ABSTRACT

BACKGROUND: Urea transport depends on the diffusion through cell membrane and the facilitated urea transport. Two groups of urea transporters (UT-A and UT-B) have been identified in mammals, and both are involved in intrarenal recycling of urea. The aim of our study was to examine the renal urea handling in rats with chronic renal failure (CRF). METHODS: CRF rats were induced by 5/6 nephrectomy followed by a high-protein (HP) diet to increase the progressive loss of renal function for 5 months. Functional studies on water and urea handling were performed. RT-PCR, immunoblotting and immunohistochemistry were used to identify UT-A proteins in remnant kidney. RESULTS: A significant decrease in creatinine clearance consistent with development of CRF was observed. The remnant kidneys were hypertrophied, and total renal mass was increased. Urine production increased markedly, whereas urine osmolality and solute-free water reabsorption decreased significantly. Fractional urea excretion was increased reaching values of 105% -/+ 8%. UT-A protein was localized in pars recta by immunohistochemical studies, and it was identified as UT-A2 in outer medulla from remnant kidneys by RT-PCR and immunoblotting. CONCLUSION: In uremic rats, an urea transporter type UT-A2 was expressed in the pars recta, suggesting a possible relation with the fractional urea excretion increase. This expression may be a consequence of an adaptive mechanism in the handling of urea during development of CRF. Further studies will be necessary to elucidate the contribution of this mechanism to renal damage observed in the progression of CRF.


Subject(s)
Gene Expression , Kidney Failure, Chronic/genetics , Kidney Medulla/metabolism , Membrane Transport Proteins/genetics , RNA, Messenger/genetics , Animals , Disease Models, Animal , Follow-Up Studies , Humans , Immunoblotting , Immunohistochemistry , Kidd Blood-Group System , Kidney Failure, Chronic/metabolism , Kidney Failure, Chronic/pathology , Kidney Medulla/pathology , Membrane Transport Proteins/biosynthesis , Rats , Rats, Sprague-Dawley , Reverse Transcriptase Polymerase Chain Reaction , Urea Transporters
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