Proteomic analysis of dorsolateral prefrontal cortex indicates the involvement of cytoskeleton, oligodendrocyte, energy metabolism and new potential markers in schizophrenia

Schizophrenia is likely to be a consequence of serial alterations in a number of genes that, together with environmental factors, will lead to the establishment of the illness. The dorsolateral prefrontal cortex (Brodmann`s Area 46) is implicated in schizophrenia and executes high functions such as working memory, differentiation of conflicting thoughts, determination of right and wrong concepts, correct social behavior and personality expression. We performed a comparative proteome analysis using two-dimensional gel electrophoresis of pools from 9 schizophrenia and 7 healthy control patients` dorsolateral prefrontal cortex aiming to identify, by mass spectrometry, alterations in protein expression that could be related to the disease. In schizophrenia-derived samples, our analysis revealed 10 downregulated and 14 upregulated proteins. These included alterations previously implicated in schizophrenia, such as oligodendrocyte-related proteins (myelin basic protein and transferrin), as well as malate dehydrogenase, aconitase, ATP synthase subunits and cytoskeleton-related proteins. Also, six new putative disease markers were identified, including energy metabolism, cytoskeleton and cell signaling proteins. Our data not only reinforces the involvement of proteins previously implicated in schizophrenia, but also suggests new markers, providing further information to foster the comprehension of this important disease. (C) 2008 Elsevier Ltd. All rights reserved.

Proliferative Defects and Formation of a Double Cortex in Mice Lacking Mltt4 and Cdh2 in the Dorsal Telencephalon

Radial glial cells (RGCs) in the ventricular neuroepithelium of the dorsal telencephalon are the progenitor cells for neocortical projection neurons and astrocytes. Here we showthatthe adherens junction proteins afadin and CDH2 are criticalforthe control of cell proliferation in the dorsal telencephalon and for the formation of its normal laminar structure. Inactivation of afadin or CDH2 in the dorsal telenceph-alon leads to a phenotype resembling subcortical band heterotopia, also known as “double cortex,” a brain malformation in which heterotopic gray matter is interposed between zones of white matter. Adherens junctions between RGCs are disrupted in the mutants, progenitor cells are widely dispersed throughout the developing neocortex, and their proliferation is dramatically increased. Major subtypes of neocortical projection neurons are generated, but their integration into cell layers is disrupted. Our findings suggest that defects in adherens junctions components in mice massively affects progenitor cell proliferation and leads to a double cortex-like phenotype.

Canine spinal cord neuron and axon myelin sheath morphometry

This inedited morphometric study has been developed from healthy canine spinal cord neuron cytoplasm and nucleus, and white matter axonal myelin sheath, from cervical, thoracic and lumbar regions. For the morphometric study, the parameters were area, perimeter, maximum and minimum diameters and roundness for neurons and myelin thickness for axon. For each parameter, 300 neurons were analysed. The results revealed that lumbar neurons had the highest mean values for the analysed parameters, indicating the presence of large neurons in this region, with large axons as a result of myelin thickness, which is proportional to axon calibre. We conclude that these morphometric results can contribute for the establishment of normal patterns, for canine spinal cord cervical, thoracic and lumbar segments.

Astrócitos imunorreativos à proteína glial fibrilar ácida (GFAP) em sistema nervoso central de equinos normais e de equinos com leucoencefalomalácia

A proteína glial fibrilar ácida (GFAP), subunidade dos filamentos intermediários do citoesqueleto celular, está presente no citoplasma de astrócitos. Técnicas imunohistoquímicas com anticorpos primários anti-GFAP são geralmente empregadas para identificar astrócitos no sistema nervoso, permitindo verificar também sua hipertrofia. Vários estudos mostram a distribuição, a morfologia e a citoarquitetura de astrócitos em várias regiões do SNC do homem e de animais de laboratório. No entanto, em animais domésticos e, especialmente em equinos, poucas informações estão disponíveis. No presente trabalho, verificou-se a densidade e a morfologia de astrócitos imunorreativos à GFAP na substância branca da córtex cerebral de equinos com leucoencefalomalácia (LEM) comparando-se esses aspectos com o de equinos normais. Animais com LEM apresentaram hipertrofia de astrócitos em áreas próximas às lesões, representada pelo aumento do corpo celular, do núcleo e dos prolongamentos citoplasmáticos. O número de astrócitos apresentou-se reduzido e a imunorreatividade foi mais acentuada. Nos animais normais, verificou-se distribuição constante de astrócitos imunorreagentes com características de fibrosos. Alterações vasculares nos animais com LEM, como por exemplo degeneração de endotélio vascular, também foram observadas, podendo estar associadas às alterações astrocíticas.

Investigação da capacidade intelectiva de pacientes pediátricos diagnosticados com tumores de fossa posterior

Central Nervous System are the most common pediatric solid tumors. 60% of these tumors arise in posterior fossa, mainly in cerebellum. The first therapeutic approach is surgical resection. Malignant tumors require additional strategies - chemotherapy and radiotherapy. The increasing survival evidences that childhood brain tumors result in academic and social difficulties that compromise the quality of life of the patients. This study investigated the intellectual functioning of children between 7 to 15 years diagnosed with posterior fossa tumors and treated at CEHOPE - Recife / PE. 21 children were eligible - including 13 children with pilocytic astrocytoma (G1) who underwent only surgery resection, and eight children with medulloblastoma (G2) - submitted to surgical resection, chemotherapy and craniospinal radiotherapy. Participants were evaluated by the Wechsler Intelligence Scale for Children - WISC-III. Children of G1 scored better than children of G2. Inferential tools (Mann-Whitney Ü Test) identified significant diferences (p ≤ 0.05) between the Performance IQ (PIQ) and Processing Speed Index (PSI) as a function of treatment modality; Full Scale IQ (FSIQ), PIQ and PSI as a function of parental educational level; PIQ, FSIQ, IVP and Freedom from Distractibility (FDI) as a function of time between diagnosis and evaluation. These results showed the late and progressive impact of radiotherapy on white matter and information processing speed. Furthermore, children whose parents have higher educational level showed better intellectual performance, indicating the influence of xxii socio-cultural variables on cognitive development. The impact of cancer and its treatment on cognitive development and learning should not be underestimated. These results support the need to increase the understanding of such effects in order to propose therapeutic strategies which ensure that, in addition to the cure, the full development of children with this pathology

Distribution of NADPH-diaphorase-positive neurons in the prefrontal cortex of the Cebus monkey

We studied the distribution of NADPH-diaphorase (NADPH-d) activity in the prefrontal cortex of normal adult Cebus apella monkeys using NADPH-d histochemical protocols. The following regions were studied: granular areas 46 and 12, dysgranular areas 9 and 13, and agranular areas 32 and Oap. NADPH-d-positive neurons were divided into two distinct types, both non-pyramidal. Type I neurons had a large soma diameter (17.24 +/- 1.73 pm) and were densely stained. More than 90% of these neurons were located in the subcortical white matter and infragranular layers. The remaining type I neurons were distributed in the supragranular layers. Type II neurons had a small, round or oval soma (9.83 +/- 1.03 mu m), and their staining pattern varied markedly. Type II neurons were distributed throughout the cortex, with their greatest numerical density being observed in layers II and III. In granular areas, the number of type II neurons was up to 20 times that of type I neurons, but this proportion was smaller in agranular areas. Areal density of type II neurons was maximum in the supragranular layers of granular areas and minimum in agranular areas. Statistical analysis revealed that these areal differences were significant when comparing some specific areas. In conclusion, our results indicate a predominance of NADPH-d-positive cells in supragranular layers of granular areas in the Cebus prefrontal cortex. These findings support previous observations on the role of type II neurons as a new cortical nitric oxide source in supragranular cortical layers in primates, and their potential contribution to cortical neuronal activation in advanced mammals. (c) 2006 Elsevier B.V. All rights reserved.

Fungal pyogranulomatous encephalitis in a dog with leishmaniosis

É relatado um caso de encefalite piogranulomatosa em um cão fêmea de um ano de idade, da raça Fila Brasileiro. Ao exame macroscópico do cérebro, evidenciou-se área amolecida e hemorrágica no córtex frontal direito e na superfície de corte do hemisfério esquerdo, afetando a substância branca e áreas corticais profundas. O diagnóstico de encefalite piogranulomatosa micótica multifocal foi realizado através de exame histopatológico, que mostrou a presença de macrófagos, células gigantes, focos de hemorragia e hifas septadas de coloração marrom, com distribuição difusa e invadindo a luz de vasos. A identificação de formas amastigotas no imprint de linfonodo poplíteo confirmou o diagnóstico de leishmaniose. A infecção micótica no cérebro deste cão foi relacionada com a ocorrência concomitante de leishmaniose, uma doença imunossupressora.

Halicephalobus gingivalis (H.deletrix) in the brain of a horse

Um eqüino macho, com 10 anos, Mangalarga, apresentou uma infecção por um nematódeo rabditiforme no cérebro. Os sinais clínicos limitaram-se ao fato de o animal andar em círculos e apresentar paralisia do lado direito. O exame histológico do cérebro revelou acentuada gliose e discreto edema intersticial. O infiltrado inflamatório mononuclear perivascular era composto por poucas camadas de linfócitos, plasmócitos, macrófagos e raros eosinófilos, associados aos nematódeos rabditiformes. Áreas de malácia e trajetos com esferóides axonais são vistos ao redor de vasos e do agente etiológico, sendo mais evidentes na substância branca. Nas meninges, o infiltrado inflamatório foi moderado e associado a parasitas perivasculares. A identificação do nematódeo foi baseada no exame histológico do cérebro do cavalo.

Vitamin E responsive neurological signs in ostriches (Struthio camelus).

Encephalomalacia in birds is commonly related to vitamin E deficiency. To the best of our knowledge, in ostriches there are no reports of neurological signs associated to nutritional deficiencies. Fourteen ostriches aged from 1 day to 3 weeks old were sent to necropsy and showed apathy, progressive weight loss, twisting or S-shaped flexure of the neck and mortality. Gross changes were mild and consisted of congested blood vessels besides edematous aspect of the cerebrum and cerebellum. Microscopic examination revealed congested meningeal vessels. Capillaries were evident in cortex and white matter of the encephalon. Focal spongy vacuolation and reactive gliosis were observed within the white matter of cerebellum. Four birds that presented neurological signs were inoculated by the subcutaneous route in the neck with 50 mg of vitamin E (alfatocoferol acetate), followed by clinical evaluation. These birds were fully recovered in a period of 24 h. Macroscopic and microscopic lesions associated to the clinical signs and to the therapeutic improvement provided by vitamin E administration are strong evidences of nutritional encephalomalacia (vitamin E deficiency).

Fetal and maternal lesions of cyanide dosing to pregnant goats

The present work evaluated the pathological effects in both dams and their fetuses of cyanide ingestion during pregnancy using goat as animal model. From the Day 24 of pregnancy, three pregnant goats were dosed by gavage with 3.0 mg KCN/kg bw/day, and two others received only tap water. All dams were euthanized and necropsied at Day 120 of pregnancy. The fetuses were examined carefully for gross abnormalities. Determinations of cyanide and thiocyanate were performed in maternal and fetal blood and in amniotic fluid. Samples of several organs were collected for histopathologic evaluation. No clinical changes were seen in any animal throughout the pregnancy. No malformations or dead fetuses were observed: however, placenta from one treated goat presented increased cotyledon surface area occupied by vascular lumina. Histological lesions in KCN-treated dams consisted of vacuolation of hepatocytes and thyroid follicular cells, increased number of vacuoles on thyroid follicular colloid. and spongiosis of cerebral, internal capsule, and cerebellar peduncles tracts. Fetuses from these mothers showed vacuolation of hepatocytes and thyroid follicular cells, and spheroids in the cerebellar white matter. Levels of cyanide and thiocyanate were higher in maternal than fetal blood, which suggests that these substances were largely but not freely transferred from mothers to fetuses. (C) 2009 Elsevier B.V. All rights reserved.

Optimizing optimum-path forest classification for huge datasets

Traditional pattern recognition techniques can not handle the classification of large datasets with both efficiency and effectiveness. In this context, the Optimum-Path Forest (OPF) classifier was recently introduced, trying to achieve high recognition rates and low computational cost. Although OPF was much faster than Support Vector Machines for training, it was slightly slower for classification. In this paper, we present the Efficient OPF (EOPF), which is an enhanced and faster version of the traditional OPF, and validate it for the automatic recognition of white matter and gray matter in magnetic resonance images of the human brain. © 2010 IEEE.

Zymographic patterns of MMP-2 and MMP-9 in the CSF and cerebellum of dogs with subacute distemper leukoencephalitis

Distemper leukoencephalitis is a disease caused by the canine distemper virus (CDV) infection. It is a demyelinating disease affecting mainly the white matter of the cerebellum and areas adjacent to the fourth ventricle; the enzymes of the matrix metalloproteinases (MMPs) group, especially MMP-2 and MMP-9 have a key role in the myelin basic protein fragmentation and in demyelination, as well as in leukocyte traffic into the nervous milieu. To evaluate the involvement of MMPs during subacute distemper leukoencephalitis, we measured the levels of MMP-2 and MMP-9 by zymography in the cerebrospinal fluid (CSF) and in the cerebellum of 14 dogs naturally infected with CDV and 10 uninfected dogs. The infected dogs presented high levels of pro-MMP-2 in the CSF and elevated levels of pro-MMP-2 and pro-MMP-9 in the cerebellar tissue. Active MMP-2 was detected in the CSF of some infected dogs. As active MMP-2 and MMP-9 are required for cellular migration across the blood-brain barrier and any interference between MMPs and their inhibitors may result in an amplification of demyelination, this study gives additional support to the involvement of MMPs during subacute distemper leukoencephalitis and suggests that MMP-2 and MMP-9 may take part in the brain inflammatory changes of this disease. © 2013 Elsevier B.V.

Correlação entre imagens de tomografia computadorizada e cortes anatômicos do encéfalo de cães

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

NADPH-diaphorase activity in area 17 of the squirrel monkey visual cortex: neuropil pattern, cell morphology and laminar distribution

We studied the distribution of NADPH-diaphorase activity in the visual cortex of normal adult New World monkeys (Saimiri sciureus) using the malic enzyme "indirect" method. NADPH-diaphorase neuropil activity had a heterogeneous distribution. In coronal sections, it had a clear laminar pattern that was coincident with Nissl-stained layers. In tangential sections, we observed blobs in supragranular layers of V1 and stripes throughout the entire V2. We quantified and compared the tangential distribution of NADPH-diaphorase and cytochrome oxidase blobs in adjacent sections of the supragranular layers of V1. Although their spatial distributions were rather similar, the two enzymes did not always overlap. The histochemical reaction also revealed two different types of stained cells: a slightly stained subpopulation and a subgroup of deeply stained neurons resembling a Golgi impregnation. These neurons were sparsely spined non-pyramidal cells. Their dendritic arbors were very well stained but their axons were not always evident. In the gray matter, heavily stained neurons showed different dendritic arbor morphologies. However, most of the strongly reactive cells lay in the subjacent white matter, where they presented a more homogenous morphology. Our results demonstrate that the pattern of NADPH-diaphorase activity is similar to that previously described in Old World monkeys.

Methylmercury intoxication and histochemical demonstration of NADPH-diaphorase activity in the striate cortex of adult cats

The effects of methylmercury (MeHg) on histochemical demonstration of the NADPH-diaphorase (NADPH-d) activity in the striate cortex were studied in 4 adult cats. Two animals were used as control. The contaminated animals received 50 ml milk containing 0.42 µg MeHg and 100 g fish containing 0.03 µg MeHg daily for 2 months. The level of MeHg in area 17 of intoxicated animals was 3.2 µg/g wet weight brain tissue. Two cats were perfused 24 h after the last dose (group 1) and the other animals were perfused 6 months later (group 2). After microtomy, sections were processed for NADPHd histochemistry procedures using the malic enzyme method. Dendritic branch counts were performed from camera lucida drawings for control and intoxicated animals (N = 80). Average, standard deviation and Student t-test were calculated for each data group. The concentrations of mercury (Hg) in milk, fish and brain tissue were measured by acid digestion of samples, followed by reduction of total Hg in the digested sample to metallic Hg using stannous chloride followed by atomic fluorescence analysis. Only group 2 revealed a reduction of the neuropil enzyme activity and morphometric analysis showed a reduction in dendritic field area and in the number of distal dendrite branches of the NADPHd neurons in the white matter (P<0.05). These results suggest that NADPHd neurons in the white matter are more vulnerable to the long-term effects of MeHg than NADPHd neurons in the gray matter.