RESUMEN
The current study was designed to determine whether exposure of mice to aerosolized jet fuel (JP8 + 100) resulted in changes in the cellular distribution or immunoreactivity of the enzyme glutathione S-transferase (GST), a biomarker of toxicant exposure. Male mice were exposed to JP8 + 100 at 1000 mg/m3 or 2500 mg/m3 in aerosol for 1 h per day for 7 days and then sacrificed. The retinas were studied by immunohistochemical methods. The JP8 + 100 exposure caused a marked increase in the immunoreactivity of anti-GSTM antibodies with the radial glial cells of the retina, the Müller cells. These results are consistent with the hypothesis that JP8 + 100 acts as a toxicant to mouse retina by permitting the flux of materials across the blood-retina barrier. The findings are relevant to humans because recent studies indicate that Air Force personnel assigned to clean and maintain fuel pods may be exposed to concentrations of JP8 + 100 exceeding 1000 mg/m3.
Asunto(s)
Glutatión Transferasa/inmunología , Hidrocarburos/toxicidad , Retina/efectos de los fármacos , Retina/enzimología , Aerosoles , Animales , Anticuerpos , Biomarcadores/análisis , Inmunohistoquímica , Exposición por Inhalación , Masculino , Ratones , Neuroglía/efectos de los fármacos , Retina/citologíaRESUMEN
We analyzed protein expression in the cytosolic fraction prepared from whole lung tissue in male Swiss-Webster mice exposed 1 h/day for seven days to aerosolized JP-8 jet fuel at concentrations of 1000 and 2500 mg/m3, simulating military occupational exposure. Lung cytosol samples were solubilized and separated via large scale, high resolution two-dimensional electrophoresis (2-DE) and gel patterns scanned, digitized and processed for statistical analysis. Significant quantitative and qualitative changes in tissue cytosol proteins resulted from jet fuel exposure. Several of the altered proteins were identified by peptide mass fingerprinting, confirmed by sequence tag analysis, and related to impaired protein synthetic machinery, toxic/metabolic stress and detoxification systems, ultrastructural damage, and functional responses to CO2 handling, acid-base homeostasis and fluid secretion. These results demonstrate a significant but comparatively moderate JP-8 effect on protein expression and corroborate previous morphological and biochemical evidence. Further molecular marker development and mechanistic inferences from these observations await proteomic analysis of whole tissue homogenates and other cell compartment, i.e., mitochondria, microsomes, and nuclei of lung and other targets.
Asunto(s)
Hidrocarburos/toxicidad , Pulmón/efectos de los fármacos , Exposición Profesional , Proteoma , Animales , Electroforesis en Gel Bidimensional , Pulmón/metabolismo , Pulmón/ultraestructura , Masculino , Ratones , Ratones Endogámicos C57BL , Microscopía Electrónica , Proteínas/aislamiento & purificación , Espectrometría de Masa por Láser de Matriz Asistida de Ionización DesorciónRESUMEN
Lead is a potent neuro- and nephrotoxin in humans and a renal carcinogen in rats. Previous studies have detected lead-induced increases in the activities of specific detoxification enzymes in distinct kidney cell types preceding irreversible renal damage. While preferential susceptibility of the highly vascularized cortex to the effects of lead is clear, lead effects on the medullary region have remained unexplored. The present study was undertaken to investigate the extent to which regional renal protein expression differs and to determine which, if any, regionally distinct protein markers indicative of lead's renotoxic mechanism might be detected in kidney cortical and medullary cytosols. We examined protein expression in these two functionally and anatomically distinct regions, and identified several proteins that are differentially expressed in those regions and were significantly altered by lead. Kidney cytosols from rats injected with lead acetate (114 mg/kg, three consecutive daily injections) were separated by two-dimensional electrophoresis. Lead exposure significantly (P<0.001) altered the abundance (either or) of 76 proteins in the cortex and only 13 in the medulla. Eleven of the proteins altered in the protein patterns were conclusively identified either by matrix-assisted laser desorption/ionization mass spectrometry/electrospray ionization-mass spectrometry (MALDI-MS/ESI-MS) analysis of peptide digests, immunological methods, or by gel matching. Several of the cortical proteins altered by lead were unchanged in the medulla while others underwent similar but lesser alterations. These observations reflect the complexity of lead's nephrotoxicity and endorse the application of proteomics in mechanistic studies as well as biomarker development in a variety of toxicologic paradigms.
Asunto(s)
Riñón/metabolismo , Plomo/farmacología , Proteínas/metabolismo , Secuencia de Aminoácidos , Animales , Electroforesis en Gel Bidimensional , Riñón/efectos de los fármacos , Masculino , Datos de Secuencia Molecular , Ratas , Ratas Sprague-Dawley , Espectrometría de Masa por Láser de Matriz Asistida de Ionización DesorciónRESUMEN
The rodent kidney is a target of many xenobiotics and is typified by regionally specific structure and function. This renders distinct regions of the kidney differentially susceptible to toxic exposure and effect. To characterize these differences at the proteome level, protein patterns from male rat kidney cortex and medulla cytosols were examined by two-dimensional electrophoresis (2-DE) and image analysis and prominent proteins identified immunologically or by matrix-assisted laser desorption/ionization-mass spectrometry (MALDI-MS) and electrospray/ionization-tandem mass spectrometry (ESI-MS/MS) sequence tag identification. An average of 727 protein spots were resolved and matched to the cortex cytosol reference pattern, and 716 in the medulla. Of this total, 127 proteins were found to differ in abundance (86 higher in cortex; 41 higher in medulla) (P < 0.001). Of those proteins that were detectable in both cortex and medulla, the abundance of 97 differed significantly while 30 proteins were found to be unique to one region or the other (26 in cortex, 4 in medulla). Twenty protein spots were identified and their regional differences are discussed. These results both confirm and expand our understanding of the molecular heterogeneity characterizing structurally and functionally distinct regions of the kidney and serve as a useful foundation for future nephrotoxicologic studies.
Asunto(s)
Citosol/química , Corteza Renal/ultraestructura , Médula Renal/química , Proteínas/análisis , alfa-Globulinas/análisis , Animales , Argininosuccinato Sintasa/análisis , Calbindinas , Calcineurina/análisis , Electroforesis en Gel Bidimensional , Glutatión Transferasa/análisis , Masculino , Ratas , Ratas Sprague-Dawley , Proteína G de Unión al Calcio S100/análisis , Espectrometría de Masa por Láser de Matriz Asistida de Ionización Desorción , Transcetolasa/análisisRESUMEN
The effect of acute exposure to lead acetate on the expression of glutathione S-transferase (GST) subunits and the levels of reduced and oxidized glutathione (GSH) and malondialdehyde (MDA) in rat kidney and liver was determined. The purpose of this study was to determine if GSH depletion and/or oxidative stress were responsible for changes in the expression of some or all GSTs that followed lead exposure. In kidney, all GST subunits increased following injection of lead. The level of kidney GSH was not changed at either 0.5 or 1 h after lead exposure, but increased 3, 6, 12 and 24 h after a single injection of lead. MDA levels (a marker of lipid peroxidation) did not change in kidney following lead injection. Immunohistochemical markers of oxidative stress and nitric oxide production were also unchanged by lead administration. Therefore. we conclude that the increases in GST levels in kidney following lead exposure were not dependent on oxidative stress. In liver, lead injection caused GSH depletion (61% of control 12 h after lead treatment) and increased MDA production (2.5-fold increase 6 h after lead exposure), while GSTA1, GSTA2, GSTM1 and GSTM2 did not increase. Analysis of the effects of lead on GST mRNA and GST cellular localization were performed by Northern blot and immunohistochemical techniques. Immunoperoxidase light microscopy and immunogold electron microscopy revealed that the increase in kidney GSTM1 and GSTP1 occurred in nuclei, cytoplasm and microvilli of proximal tubules. Northern blot analysis of GSTA2 and GSTP1 mRNAs showed that their increase following lead exposure was inhibited by actinomycin D, suggesting transcriptional induction. This study demonstrates that acute lead exposure causes dramatic changes in the subcellular distribution and expression of rat kidney GSTs, and that these changes are not a result of oxidative stress.
Asunto(s)
Regulación Enzimológica de la Expresión Génica/efectos de los fármacos , Glutatión Transferasa/biosíntesis , Riñón/metabolismo , Plomo/toxicidad , Hígado/metabolismo , Estrés Oxidativo/efectos de los fármacos , Animales , Northern Blotting , Glutatión/metabolismo , Inmunohistoquímica , Riñón/efectos de los fármacos , Riñón/enzimología , Peroxidación de Lípido/efectos de los fármacos , Hígado/efectos de los fármacos , Hígado/enzimología , Malondialdehído/metabolismo , Estrés Oxidativo/fisiología , ARN Mensajero/biosíntesis , Ratas , Ratas Sprague-Dawley , Fracciones Subcelulares/efectos de los fármacos , Fracciones Subcelulares/metabolismoRESUMEN
Glutathione S-transferases (GST) are a family of detoxification isoenzymes that catalyze the conjugation of xenobiotics and their metabolites with reduced glutathione. Lead exposure in rats is known to induce GST isoenzymes in the liver and kidney. These changes in expression have potential use as biomarkers of lead exposure. Because two-dimensional electrophoresis (2-DE) enables one to analyze both protein abundance changes and chemical changes in protein structure, 2-DE was used to determine the effect of in vivo lead exposure on GST isoform expression in rat kidney cytosols. Male Sprague-Dawley rats were exposed to inorganic lead, and proteins were separated by conventional ISO-DALT and NEPHGE-DALT techniques and blotted for immunological identification. Lead exposure caused detectable inductions in both GSTP1 and GSTM1 and quantifiable charge modification in GSTP1. These preliminary data confirm the utility of 2-D electrophoretic GST analysis as indicative of lead exposure and toxicity and support its use for further elaboration of lead's effects on renal protein expression.
Asunto(s)
Electroforesis en Gel Bidimensional , Glutatión Transferasa/análisis , Plomo/farmacología , Animales , Electroforesis en Gel Bidimensional/métodos , Masculino , Ratas , Ratas Sprague-DawleyRESUMEN
Glutathione S-transferases (GST, EC 2.5.1.18) are a family of phase II detoxification enzymes involved in the conjugation of glutathione to a highly diverse group of compounds. The purpose of this study was to evaluate the dose-response effects of lead acetate administration on the expression of rat kidney GST. Sprague-Dawley rats were injected with doses of lead acetate ranging from 0.11 to 114 mg/kg (0.3 to 300 mumol/kg) for three consecutive days and sacrificed 24 h later. Kidney GST activity, GST isoform HPLC profiles, blood lead analysis, and electron microscopy were performed. A dose of 1.1 mg/kg lead acetate resulted in a blood lead level of 26 micrograms/dl and produced a significant increase in GST activity which continued to increase with dose up to 38 mg/kg. Morphological changes were detected at 3.8 mg/kg and increasing severity of cellular damage paralleled dose, blood lead levels, and changes in body weight. Individual GST isoforms exhibited different thresholds and maxima; rGSTP1 and rGSTM1 had thresholds of 1.1 and 3.8 mg/kg, respectively, very similar rates of increase with dose, and a maximum yield that was 450% above control at a dose of 38 mg/kg for both enzymes. rGSTA1 and rGSTA3 showed similar thresholds (1.1 mg/kg) and maximal fold increase (275%) but varied in the relative response to each dose. These results indicate that renal GST increases occur at lead levels which are environmentally significant, that these changes precede cellular damage, and suggest that GST may serve as a tissue biomarker of lead exposure.
Asunto(s)
Glutatión Transferasa/metabolismo , Túbulos Renales Proximales/efectos de los fármacos , Riñón/efectos de los fármacos , Plomo/farmacología , Animales , Peso Corporal , Cromatografía Líquida de Alta Presión , Relación Dosis-Respuesta a Droga , Glutatión Transferasa/genética , Riñón/enzimología , Riñón/patología , Túbulos Renales Proximales/patología , Plomo/sangre , Masculino , Microscopía Electrónica , Isoformas de Proteínas , Ratas , Ratas Sprague-Dawley , Factores de TiempoRESUMEN
The effects of triethyl lead chloride (TEL) on the expression of two detoxication enzyme families, glutathione S-transferases (GSTs) and NAD(P)H:quinone oxidoreductase (QR) were determined in rat liver and kidney. Fischer 344 rats were given one intraperitoneal (i.p.) injection of TEL. GST activity, GST isoenzyme levels, mRNA levels of alpha class GST isoenzymes Ya1, Ya2, and Yc1 and activity of QR were determined. Treatment of rats with TEL caused a significant increase in GST activity in kidney. In kidney, the levels of all GST subunits were significantly elevated; the largest increase was a 3.2-fold increase in GST Yb1. The levels of GST Ya1, Ya2, and Yc1 mRNA also increased after injection of TEL. In liver, TEL injection resulted in decreased GST activity and lower levels of hepatic GSTs Yb2, Yb3, Ya1, and Ya2. The largest decrease was a 40% reduction of GST Ya1. In contrast, the level of liver GST Yc1 increased from day 4 through day 14 after injection of 10 mg/kg TEL and Yp was increased 1.4-fold 4 days after injection of 12 mg/kg TEL. The levels of liver mRNAs coding for alpha class GSTs Ya1, Ya2, and Yc1 were reduced 12 h after injection of TEL. The mRNA levels of GST Ya1 and Ya2 returned to basal level while Yc1 message increased to a level higher than controls 24 h after TEL injection. The increase in Yc1 protein between days 4 and 14 is consistent with the increase in the corresponding mRNA. The activity of QR was elevated 1.5-fold in kidney and 2.7-fold in liver 14 days after the injection of TEL. This report demonstrates that administration of organic lead significantly affects GST expression and QR activity in a tissue-specific and isoenzyme-specific manner. These results indicate that GST expression and QR activity are not co-regulated.
Asunto(s)
Glutatión Transferasa/biosíntesis , Isoenzimas/biosíntesis , Riñón/efectos de los fármacos , Riñón/enzimología , Hígado/efectos de los fármacos , Hígado/enzimología , NAD(P)H Deshidrogenasa (Quinona)/biosíntesis , Compuestos Organometálicos/toxicidad , Animales , Glutatión Transferasa/efectos de los fármacos , Inyecciones Intraperitoneales , Isoenzimas/efectos de los fármacos , Riñón/metabolismo , Plomo/toxicidad , Hígado/metabolismo , NAD(P)H Deshidrogenasa (Quinona)/efectos de los fármacos , Compuestos Organometálicos/administración & dosificación , Ratas , Ratas Endogámicas F344RESUMEN
Glutathione S transferases (GST) are a family of enzymes that detoxify electrophilic xenobiotics. This enzyme family was examined in kidneys of mice exposed to mercuric chloride, a known nephrotoxin, because GST have been shown to protect cells against toxicant-induced damage and may serve as biomarkers for toxicant exposure. The purpose of this study was to determine the effect of mercuric chloride on GST activity, isoenzyme levels, and cellular localization in the kidney of Swiss Webster mice. The cellular localizations of alpha, mu, and pi class GST in the kidneys of control and mercuric chloride treated mice were studied immunohistochemically. The GST isoenzyme levels were measured by high-performance liquid chromatography. The mice treated with mercuric chloride had (1) increased amounts of GSTA1/A2 protein in kidney homogenates as compared with controls when analyzed by chromatography and electrophoresis; (2) two new isoforms of the alpha isoenzyme in kidney as demonstrated by Western blot, polyacrylamide gel electrophoresis, and high-performance liquid chromatography, and (3) increased reactivity between antibodies, against GSTA1/A2 or GSTM1 isoenzymes, and cells in the proximal and distal renal tubules as shown by immunohistochemical techniques. The authors conclude that the GSTA1/A2 may protect those cells in the proximal and distal tubules of the renal cortex from toxicant effects of mercuric chloride. This would be one general mechanism for cell protection against a wide variety of toxicants including heavy metals and halogenated aromatics.
Asunto(s)
Glutatión Transferasa/metabolismo , Isoenzimas/metabolismo , Riñón/enzimología , Cloruro de Mercurio/envenenamiento , Intoxicación por Mercurio/enzimología , Animales , Western Blotting , Cromatografía Líquida de Alta Presión , Citosol/enzimología , Electroforesis en Gel de Poliacrilamida , Glutatión Transferasa/genética , Inmunohistoquímica , Isoenzimas/genética , Masculino , RatonesRESUMEN
PURPOSE: The glutathione S-transferases (GSTs) constitute a family of cytosolic isoenzymes that are involved in the detoxication of electrophilic xenobiotics. The purpose of this investigation was to determine the concentration and cellular distribution of the various classes of cytosolic GSTs in the retina of control and triethyl lead-treated rats and thereby reveal mechanisms by which the cells are protected from damage by lead and other toxicants. METHODS: The regional and cellular distribution of cytosolic GSTs in rat retina of control and lead-treated animals was studied by immunohistochemistry. Enzyme activity was determined by a spectrophotometric assay. The GST subunit distribution of the entire retina of control and lead-treated animals was determined and quantified by reverse-phase high-performance liquid chromatography (HPLC). RESULTS: Polyclonal antibodies against mu-class Yb1 and Yb2 GSTs were primarily and strongly reactive with Müller cells and their processes. Anti-Yb1 also reacted with photoreceptor outer segments. Antibodies against two alpha-class GSTs (Ya and Yk) were strongly reactive with Müller cells and their cell processes. Antibodies against Yp and Yc GSTs were reactive with amacrine cells and their processes, and anti-Yp antibodies were reactive against retinal ganglion cells. Treatment of rats with triethyl lead caused diminished reactions of the antibodies against Yb1 and Yp GSTs and increased reactions of anti-Ya with its retinal targets, whereas the total GST activity did not change significantly. CONCLUSIONS: The positive reaction between the amacrine neuronal cells of retina and the anti-Yp and anti-Yc class antisera broadens the class of neurons that contains GST enzymes protective against toxicant insult. It also has been shown that Müller cells are strongly immuno-positive for Yb1 and Yb2 GST. Because these phagocytic cells are in contact with the vitreous fluid and proximate to pigmented epithelial layer of the eye, these GSTs may protect the cells from toxicants accumulated from this fluid.
Asunto(s)
Glutatión Transferasa/metabolismo , Compuestos Organometálicos/toxicidad , Retina/enzimología , Animales , Benzoxazinas , Cromatografía Líquida de Alta Presión , Colorantes , Electroforesis en Gel de Poliacrilamida , Técnica del Anticuerpo Fluorescente , Glutatión Transferasa/análisis , Isoenzimas/análisis , Isoenzimas/metabolismo , Plomo/toxicidad , Oxazinas , Conejos , Ratas , Ratas Endogámicas F344 , Retina/efectos de los fármacos , Retina/patologíaRESUMEN
PURPOSE: It has been shown previously that the sera, from patients with visual paraneoplastic syndrome associated with lung cancer, contain immunoglobulins that are reactive with the tumor and with photoreceptor and large retinal ganglion cells. The purpose of this study is to determine the retinal cell population that reacts with immunoglobulins in the sera of patients with melanoma-associated retinopathy. METHODS: Clinical and electrophysiologic studies were used to determine the locus responsible for the visual defect in each patient. Sera from two patients with melanoma-associated retinopathy, from a patient with herpes zoster, and from a patient who had a colon tumor were obtained. The sera were incubated with sections of retina obtained from a healthy 3-year-old child who had died of asphyxiation. The tissue sections subsequently were incubated with biotin-labeled anti-human immunoglobulin G, and then with streptavidin-labeled peroxidase. Finally, the tissue sections were developed to show peroxidase activity in the targeted retinal cells. RESULTS: Clinical and electrophysiologic studies were consistent with a defect in intra-retinal transmission distal to the photoreceptors. The immunoglobulins from the patients with the melanoma-associated retinopathy reacted selectively with the bipolar cells of the retina; approximately 30% of the bipolar cells were immunoreactive. The sera from the other two patients were not reactive with any of the retinal cells examined. CONCLUSIONS: The sera of patients with the paraneoplastic syndrome, melanoma-associated retinopathy, contain high titer immunoglobulins that are reactive only with a subset of the bipolar retinal cells. The clinical, electrophysiologic, and immunologic studies are all consistent with an intra-retinal transmission defect at the level of the bipolar cells.
Asunto(s)
Anticuerpos Antineoplásicos/inmunología , Interneuronas/inmunología , Melanoma/inmunología , Síndromes Paraneoplásicos/inmunología , Retina/inmunología , Enfermedades de la Retina/inmunología , Neoplasias Cutáneas/inmunología , Anciano , Preescolar , Electrorretinografía , Femenino , Humanos , Técnicas para Inmunoenzimas , Masculino , Melanoma/complicaciones , Melanoma/patología , Persona de Mediana Edad , Síndromes Paraneoplásicos/patología , Enfermedades de la Retina/patología , Neoplasias Cutáneas/complicaciones , Neoplasias Cutáneas/patología , Pruebas del Campo Visual , Campos VisualesRESUMEN
The near-microscopic resolution of the mouse brain, by magnetic resonance imaging (MRI) at 9.4 T, permits in situ examination of the entire brain and longitudinal studies of neural development. MRI can be utilized to reveal brain structure at a resolution of 100 microns in the X, Y, and Z planes of brain, to differentiate the gray from white (myelin-rich) matter, and to reveal the ventricular compartments. The present report describes the structure of normal BALB/c mouse brain as revealed by imaging at 9.4 T and by histological stains; the structure of normal brain is compared with that from a phenylalanine hydroxylase-deficient mouse mutant line (Pah(enu2)) and those from normal littermates. The brains of patients with phenylketonuria (PKU) were reported to have demyelination and other structural abnormalities revealed by magnetic resonance imaging (MRI). Therefore, high-resolution MRI was used to examine the brain of this mutant, an animal model for the study of human phenylketonuria. Our study revealed no evidence of demyelination or other abnormalities in the brains of Pah(enu2) mice. Histologically, the mutant and normal mouse brains appear similar. This is consistent with a recent study from our laboratory which demonstrated that the histology of the brain of an untreated male patient, who died with PKU at the age of 29, was similar to control brain with the exception of changes directly related to visual blindness and seizures experienced by the patient.
Asunto(s)
Mapeo Encefálico/instrumentación , Encéfalo/patología , Procesamiento de Imagen Asistido por Computador/instrumentación , Microscopía/instrumentación , Fenilalanina Hidroxilasa/deficiencia , Adulto , Animales , Ventrículos Cerebrales/patología , Sistemas de Computación , Enfermedades Desmielinizantes/patología , Modelos Animales de Enfermedad , Femenino , Humanos , Masculino , Ratones , Ratones Endogámicos BALB C , Ratones Mutantes Neurológicos , Fibras Nerviosas Mielínicas/patología , Neuronas/patología , Fenilcetonurias/patologíaRESUMEN
The glutathione S-transferases (GSTs) constitute a family of cytosolic isoenzymes and a structurally unrelated microsomal enzyme that is involved in the detoxication of electrophilic xenobiotics. These enzymes also participate in the intracellular binding and transport of a broad range of lipophilic compounds including bilirubin, and hormones such as the glucocorticoids and thyroid hormones. The present investigation demonstrates that GSTs are present in neurons of the brainstem, forebrain, and cerebellum. An isoenzyme-specific distribution of GSTs was found in cytoplasm, nuclei, and nucleoli. The regional and cellular distribution of cytosolic GSTs in the brain was studied by immunohistochemistry, spectrophotometric enzyme assay, and reverse-phase HPLC. Polyclonal antibody against microsomal GST was strongly reactive with Purkinje cells throughout the cerebellar cortex, and with neurons in the brainstem and hippocampus. Nuclei of Purkinje cells and of neurons in the brainstem, hippocampus, and cerebral cortex were immunopositive for alpha-class GST 1-1 (YaYa), whereas alpha-class GST 2-2 (YcYc) antibody was consistently immunoreactive with the nucleolus, but not with the nucleus or soma. All alpha-class GST antibodies studied were reactive, to various degrees, with astrocytes and choroid plexus; however, ependymal cells of the subventricular zones were immunonegative. alpha-class GST 8-8 (YkYk) immunoreactivity was specifically localized to endothelial cells and/or astrocytic end feet associated with blood vessels. Reverse-phase HPLC indicated that there were also substantial regional differences in the pattern of alpha-, mu-, and pi-class GST subunit expression. For example, the thalamus/hypothalamus had the highest GST activity and greatest concentration of total GST protein and mu-class GST subunit 6 (Yb3), whereas the brainstem had the greatest concentration of pi-class GST subunit (Yp). This regional variation in GST expression may be reflective of regional differences in cell populations. In cerebellar cortex, the concentration of mu-class GST subunit 4 (Yb2) was greatest in the flocculus and lowest in the vermis. This is of clinical interest because the pattern of expression of mu-class GST subunit 4 (Yb2) in the cerebellum coincides with the known regional susceptibility of this structure to degeneration after exposure to toxic or metabolic insults. The vermis is most susceptible to these insults, whereas the lateral lobes and flocculus are most resistant.(ABSTRACT TRUNCATED AT 400 WORDS)
Asunto(s)
Encéfalo/enzimología , Glutatión Transferasa/metabolismo , Isoenzimas/metabolismo , Neuroglía/enzimología , Neuronas/enzimología , Animales , Encéfalo/citología , Cromatografía Líquida de Alta Presión/métodos , Inmunohistoquímica , Concentración Osmolar , Ratas , Distribución TisularRESUMEN
The glutathione S-transferases (GSTs) are a family of isoenzymes involved in the detoxication of a variety of electrophilic xenobiotics. The present investigation demonstrates that GST activity and the concentration of cytosolic GSTs in cerebellar cortex of Gunn rats were increased in hyperbilirubinaemic animals compared with non-jaundiced controls. Age-dependent and region-specific increases in GST isoenzymes were seen in three regions of the cerebellar cortex of jaundiced Gunn rats, whereas GST concentrations were not altered in the brainstem, thalamus/hypothalamus, cortex or liver. Cytosolic GST activity was increased 1.3-fold in the flocculus and lateral hemispheres of 20-day-old and 1.7-fold in the flocculus, lateral hemispheres and vermis of 60-day-old jaundiced (jj; homozygous) Gunn rats compared with non-jaundiced (Jj; heterozygous) Gunn rats. H.p.l.c. was used to determine the GST subunit protein concentrations in cytosolic fractions isolated from liver and brain regions of jaundiced and non-jaundiced animals. In all regions of the cerebellum from 20-day-old animals, the levels of Alpha-class GST subunits 2 (Yc1; 3.0-fold) and 8 (Yk; 2.0-fold) were increased in jaundiced rats. In 60-day-old animals, the concentrations of Alpha-class GST subunits 2 (Yc1; 5.0-fold) and 8 (Yk; 3.0-fold), Mu-class subunit 11 (Yo; 2.5-fold) and Pi-class subunit 7 (Yp; 2.0-fold) were increased in all regions of cerebellar cortex of jaundiced animals. In cerebellum of 10-, 20- and 60-day-old non-jaundiced and jaundiced Gunn rats, the flocculus had the highest concentration of Mu-class GST subunit 4 (Yb2) and vermis the lowest; hyperbilirubinaemia increased the concentration of subunit 4 (Yb2; 3- to 5-fold) in the flocculus and lateral hemispheres, but not the vermis, of 20- and 60-day-old rats. Intraperitoneal injection of sulphadimethoxine, a long-acting sulphonamide which displaces bilirubin from its albumin-binding sites and increases the bilirubin levels in tissues, further increased the already elevated concentrations of GST subunits in the lateral regions of cerebellar cortex of hyperbilirubinaemic rats. For example, the concentration of subunit 4 (Yb2) was increased 2.2-fold (compared with non-jaundiced controls) in Gunn rats injected with saline and 7.4-fold in rats injected with 100 mg of sulphadimethoxine/kg body weight. In contrast, GSTs in the vermis of jaundiced animals were not affected by sulphadimethoxine injection. Sulphadimethoxine had no effect on GST concentrations in lateral regions and vermis of heterozygous (Jj) Gunn rats.(ABSTRACT TRUNCATED AT 400 WORDS)
Asunto(s)
Corteza Cerebelosa/enzimología , Glutatión Transferasa/metabolismo , Hiperbilirrubinemia/enzimología , Isoenzimas/metabolismo , Envejecimiento/metabolismo , Animales , Encéfalo/enzimología , Corteza Cerebelosa/efectos de los fármacos , Cromatografía Líquida de Alta Presión , Citosol/enzimología , Hígado/enzimología , Masculino , Ratas , Ratas Gunn , Sulfadimetoxina/farmacologíaRESUMEN
This report describes the morphological changes observed in the brain of an untreated 27-year-old man with phenylketonuria, cortical blindness, and seizures. Golgi-Kopsch silver, cresyl violet, and hematoxylin and eosin stains were used to study cell structure and organization of the cerebellum, the lateral geniculate nuclei, the visual cortex, frontal cortex, and hippocampus. Extensive neuronal losses occurred in the right lateral geniculate nucleus (LGN), the visual cortex, and hippocampus. The left LGN, cerebellum, and frontal cortex retained neuronal components; there was a reduction in the number of dendritic processes on the Purkinje cells of the PKU subject. The loss of neurons in the LGN and occipital cortex is related to the blindness and the neuronal loss in the hippocampus is related to seizure activity.
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Ceguera/complicaciones , Encéfalo/patología , Fenilcetonurias/complicaciones , Convulsiones/complicaciones , Ceguera/patología , Humanos , Masculino , Neuronas/patología , Fenilcetonurias/patologíaRESUMEN
Murine trisomy 16 (mts16) placentas and fetuses, 17-day gestation age, were examined histologically and by magnetic resonance imaging at 9.4 T and compared with control littermate tissues. Placentas were studied by immunohistochemical methods, at 15-days gestational age, for expression of the major histocompatibility complex (MHC) class I H-2Kk cell surface marker. Immunohistochemical studies revealed a markedly increased expression of the MHC marker H-2Kk on cells in the labyrinth of the placenta of mts16. There were differences between the magnetic resonance (MR) images of the trisomic and normal placentas, which may be correlated with the increased expression of H-2Kk in the mts16 placental labyrinth. The decidual and labyrinthine components of the normal placentas showed similar high signal intensities (SI) while in trisomic placentas a marked high SI was characteristic only of the decidual region on proton spin density images. The MRI also revealed a smaller cerebellum in the ts16 fetuses. The potential effects of the compromised structure of the placental labyrinth and the overexpression of the H-2Kk marker on the mts16 neural and placental dysgenesis are discussed.
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Anomalías Congénitas/genética , Antígenos H-2/metabolismo , Placenta/inmunología , Trisomía , Animales , Biomarcadores , Anomalías Congénitas/inmunología , Anomalías Congénitas/patología , Modelos Animales de Enfermedad , Síndrome de Down/genética , Femenino , Antígenos H-2/genética , Imagen por Resonancia Magnética , Ratones , Ratones Endogámicos BALB C , EmbarazoRESUMEN
Neuronal cells from murine trisomy 16 fetuses have increased levels of class I MHC H-2Kk. To determine whether this increased level of H-2Kk protein product resulted from an increased synthesis of mRNA, a 33 base antisense cDNA probe complementary to a region in exon 2 of the H-2Kk sequence (nucleotide 392-424) was synthesized. This probe was used to examine, by in situ hybridization and immunohistochemistry, the neural distribution of H-2Kk mRNA and protein product. A marked elevation of the H-2Kk mRNA and protein were localized in mts16 neuronal populations that were susceptible to dysgenesis. The results implicate the expression of the H-2Kk in the neuropathology of mts16 and its human counterpart, Down syndrome.
Asunto(s)
Encéfalo/inmunología , Mapeo Cromosómico , Antígenos H-2/genética , Antígenos de Histocompatibilidad Clase I/genética , Neuronas/inmunología , ARN Mensajero/genética , Trisomía , Animales , Encéfalo/embriología , Sondas de ADN , Femenino , Feto , Edad Gestacional , Masculino , Ratones , Ratones Endogámicos BALB C , ARN Mensajero/metabolismo , Translocación GenéticaRESUMEN
Clinical, magnetic resonance imaging (MRI), and serologic studies were performed on 11 patients with diffuse central nervous system (CNS) systemic lupus erythematosus and 8 patients with focal CNS lupus. MRI of patients with diffuse clinical disease showed symmetrically distributed areas of increased signal intensity in the subcortical white matter; these resolved after treatment with high-dose methylprednisolone. These patients' sera contained elevated levels of antineurofilament antibodies. Patients with focal CNS lupus had areas of increased signal intensity and atrophic changes in regions corresponding to the major cerebral vessels. These MRI abnormalities did not improve after treatment with high-dose steroids. The sera of patients with focal CNS lupus had elevated levels of cardiolipin and lupus anticoagulant but normal levels of antineurofilament antibody. Our findings suggest that results of a combined clinical, MRI, and serologic evaluation of patients with CNS lupus may predict the response of patients to high-dose steroid therapy.