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No. of bitstreams: 2 Dissertação - Luanna Fernandes Peixoto - 2015.pdf: 3025683 bytes, checksum: bbc78b7fe8fdd447275a7d8f73fd84b6 (MD5) license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) Previous issue date: 2015-12-15 / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior - CAPES / incidence of neurodegenerative disease (ND) is increasing with the world population aging. Among them, Alzheimer's disease (AD) is the most common of dementia in the elderly population. The AD has been linked to oxidative stress resulting in neurodegenerative disorders, such as the impairment of the antioxidant and cholinergic systems. AChE inhibitors drugs are a treatment option used for AD. However, until now, the drug treatment only temporary attenuates the AD symptoms and new drug targets, as the medicinal plants, are necessary for a better treatment of this disease. Eugenia dysenterica DC (Myrtaceae), popularly known as cagaita, is a native plant from the Cerrado region of Brazil. It is used in the folk medicine to treat diarrhea, jaundice and diabetes. Data from literature reported the presence of phenolic compounds, such as flavonoids and tannins, in the leaves of E. dysenterica. Thus, the aim of this study was to evaluate the neuroprotective effect of crude hydroalcoholic extract of E. dysenterica leaves (HEED), using a model of neurotoxicity induced by aluminium in mice and that causes similar cellular alterations of that observed in the Alzheimer’s disease. The results showed the HEED increased the mice performance in the passive avoidance test, showing the reversion of the cognitive damage promoted by aluminium. In addition, the HEED reduced the neuronal lipid peroxidation and restored the activity of superoxide dismutase (SOD), catalase (CAT) and acetylcholinesterase (AChE) enzymes in mice brain. The histological analysis showed the HEED decreased the incidence of neuronal death by necrosis. Taken together, the results here obtained suggest that the antioxidant properties of HEED are the responsible for its neuroprotective effect. / A incidência de doenças neurodegenerativas (DN) vem aumentando com o envelhecimento populacional. Dentre as DN a doença de Alzheimer (DA) é a forma mais comum de demência na população idosa. Um dos fatores envolvidos no desenvolvimento da DA é o estresse oxidativo neuronal que pode resultar em diversas desordens neurodegenerativas, como o comprometimento dos sistemas antioxidante e colinérgico. Um dos tratamentos utilizados para esta doença são os fármacos inibidores de acetilcolinesterase (AChE) porém, as terapias disponíveis apenas atenuam temporariamente os sintomas. Nesse sentido, novas terapias são necessárias para o tratamento da DA e o estudo de plantas pode ser uma fonte para obtenção de novas opções terapêuticas. Eugenia dysenterica DC (Myrtaceae), popularmente conhecida como cagaita, é uma planta encontrada em região de Cerrado, utilizada na medicina popular para tratar doenças diarreicas, icterícia e diabetes. Dados da literatura relatam a presença de compostos fenólicos, como flavonoides e taninos, nas folhas de E. dysenterica., substâncias descritas como potenciais antioxidantes e anticolinesterásicos. Dessa forma, o objetivo deste trabalho foi avaliar o efeito neuroprotetor do extrato bruto hidroalcoólico das folhas de Eugenia dysenterica (EHED) em camundongos, em um modelo de neurotoxicidade induzida por alumínio. O EHED aumentou o tempo de latência na memória de curta e longa duração no teste esquiva passiva, demonstrando a reversão do prejuízo cognitivo promovido pelo alumínio. Os testes chaminé e campo aberto demonstraram que não houve comprometimento na atividade locomotora dos animais. As análises histológicas demonstraram que o tratamento com EHED diminuiu a incidência da morte neuronal por necrose causada pelo AlCl3. Nos ensaios bioquímicos, observou-se que o EHED foi capaz de reduzir a lipoperoxidação e restaurar a atividade das enzimas antioxidantes superóxido dismutase (SOD) e catalase (CAT). Além disso, o EHED foi capaz de restaurar a atividade da AChE, revertendo os danos causados pelo AlCl3. Em conclusão, os resultados obtidos são sugestivos de que o EHED apresenta atividade neuroprotetora por apresentar propriedades antioxidantes, inibindo os efeitos deletérios centrais promovidos pelo alumínio e que se assemelham às alterações celulares descritas para a Doença de Alzheimer.

Fundação de Amparo a Pesquisa no Estado do Rio Grande do Sul / Animal models as reserpine are helpful to understand the pathophysiology of several diseases with involuntary movements, as Parkinson s disease (PD), and to search efficient treatments. The present study tested the effects of reserpine on behavioral alterations induced by reserpine in rats and worms, with emphasis in oxidative and dopaminergic parameters, and the effect of the antioxidant gallic acid (GA) in reserpine-exposed rats. As result, reserpine (1mg/Kg, sc, for 3 consecutive days) increased the frequency of vacuous chewing movements (VCMs) in rats in relation to controls, and maintained this increase for at least 3 days after reserpine withdrawal. Treatment with GA (4.5 , 13.5 or 40.5 mg/kg/day, po) for 3 days reverted reserpine-induced increase in VCMs, showing protective effect. Neither reserpine nor GA changed oxidative parameters (TBARS and DCFH-DA oxidation), antioxidant levels (proteic and non-proteic thiol) and the activity of Na+,K+-ATPase (total and α-subunit) in striatum and cortex. Afterward, studies were performed with Caenorhanditis elegans due its several advantages in studies of neurodegeneration and of drugs mechanism of action. L1-larval stage C. elegans were exposed to reserpine (30 ou 60 μM) for different times. Reserpine decreased the survival, development, food intake, locomotor rate on food and dopamine (DA) levels in worms and it had effect on egg laying and defecation cycles. Morphological evaluations of dopaminergic cephalic (CEP) neurons in BY200 worms (with GFP coupled to dat-1 gene) reveled neurodegeneration by: 1) decreased fluorescence intensity, 2) decreased the number of intact neurons, and 3) increased the number of shrunken somas per worm. These effects were unrelated to reserpine s effect on dat-1 gene expression. Interestingly, the reserpine effects on locomotor rate, dopaminergic CEP neurons morphology and dat-1 gene expression were reverted after reserpine withdrawal. Furthermore, reserpine decreased the survival of vesicular monoamine transporter (VMAT) and dat-1 loss-of-function mutant worms, but no of tyrosine hydroxylase (TH, cat-2) and dopaminergic receptors (dop-1, dop-2, dop-3 e dop-4) loss-of-function mutants in relation to wild-type N2 worms. Reserpine also decreased the survival of worms pre-exposed to DA; and it activated SKN-1 detoxification pathway. Moreover, no differences were found in DAT and TH immunoreactivity in striatum of rats treated with reserpine and/or GA. The GA protective effects against reserpine-induced VCMs in rats are probably not related to its antioxidant and antiapoptotic properties or monoamine oxidase (MAO) inhibition. As conclusion, the reserpine decreases DA levels though action on VMAT, and it induces neurotoxicity/neurodegeneration due probably an increase on extracellular DA contents resulted from changes on DAT function. More studies evaluating the reserpine effect on DAT and the GA mechanism of protection are necessary. / Modelos animais como o da reserpina auxiliam no entendimento da fisiopatologia de diversas doenças que se manifestam por movimentos involuntários, como a doença de Parkinson (DP), e na busca por formas de tratamento. O presente trabalho avaliou mecanismos envolvidos na indução de alterações comportamentais induzidas por reserpina em ratos e vermes com ênfase em parâmetros oxidativos e dopaminérgicos, e a ação do antioxidante ácido gálico (AG) em ratos tratados com reserpina. Como resultado, a reserpina (1mg/Kg, sc, por 3 dias consecutivos) aumentou a frequência de movimentos de mascar no vazio (MMVs) em ratos em relação ao controle, e manteve esse aumento por pelo menos 3 dias após o término das administrações da reserpina. O tratamento com AG (4,5 ou 13,5 ou 40,5 mg/kg/day, vo) por 3 dias reverteu esse aumento dos MMVs, mostrando efeito protetor. Nem reserpina nem o AG alteraram os parâmetros avaliados de dano oxidativo (TBARS e oxidação da DCFH-DA), de antioxidantes (tiol protéico e não protéico) e a atividade da Na+,K+-ATPase (total e α-subunidade) no estriado e córtex cerebral. Estudos posteriores foram realizados em Caenorhanditis elegans devido as diversas vantagens oferecidas por esse modelo animal em estudos de neurodegeneração e de investigação do mecanismo de ação de drogas. C. elegans em estágio larval L1 foram expostos a reserpina (30 ou 60 μM) por diferentes tempos. A reserpina reduziu a sobrevivência, o desenvolvimento, a ingestão de alimento, a atividade locomotora na comida e as concentrações de dopamina (DA) nos vermes, e afetou a postura de ovos e tempo entre defecações. Análise da morfologia dos neurônios dopaminérgicos cefálicos (CEP) de vermes BY200 (com GFP acoplado ao gene dat-1) indicam neurodegeneração por: 1) redução da intensidade da fluorescência, 2) redução do número de neurônios intactos e 3) aumento do número de somas atrofiados por verme em relação ao controle. Esses efeitos não estão relacionados a efeitos da reserpina na expressão do gene dat-1. Interessantemente, os efeitos da reserpina na atividade locomotora, na morfologia dos neurônios CEP e na expressão do gene dat-1 foram revertidos após a retirada dos vermes da exposição a reserpina. Em adição, a reserpina reduziu a sobrevivência de vermes deficientes do transportador vesicular de monoaminas (TVMs, cat-1) e do transportador de DA (DAT, dat-1), mas não alterou a sobrevivência de deficientes da tirosina hidroxilase (TH, cat-2) e dos receptores dopaminérgicos (dop-1, dop-2, dop-3 e dop-4) em relação aos vermes selvagens N2. A reserpina também reduziu a sobrevivência de vermes N2 pré-expostos a DA, e ativou a via de detoxificação SKN-1 dos vermes. Alterações na imunoreatividade ao DAT e a TH no estriado de ratos tratados com reserpina e/ou AG não foram encontradas. O efeito protetor do AG nos MMVs induzidos por reserpina em ratos parece não envolver sua atividade antioxidante, antiapoptótica ou na monoaminoxidase (MAO). Como conclusão, a reserpina age no TVMs causando depleção de DA, e causa neurotoxicidade/neurodegeneração dopaminérgica devido provavelmente a um acúmulo de DA no espaço sináptico resultande de uma interferência no funcionamento do DAT. Mais estudos avaliando a ação da reserpina no DAT e o mecanismo de proteção do AG são necessários.

>Magister Scientiae - MSc / New therapeutic strategies are needed for a diverse array of poorly understood neurological impairments. These include neurodegenerative disorders such as Parkinson’s disease and Alzheimer’s disease, and the psychiatric disorders such as depression, anxiety and drug dependence. Popular neuropharmacotherapies have focused on dopamine (DA), serotonin (5HT), γ-aminobutric acid (GABA) and glutamate systems (Jupp & Lawrence, 2010). However recent research points to the sigma receptor (σR) as a possible neuromodulatory system. Due to its multi-receptor action, the σR can trigger several significant biological pathways. This indicates its ideal potential as a drug target to effectively minimise drug dosage and potential side effects. Currently there are a limited number of σR ligands available and few possess the selectivity to significantly show σR’s role in neurological processes. Polycyclic amines have shown notable sigma activity and provide an advantageous scaffold for drug design that can improve pharmacodynamic and pharmacokinetic properties (Banister et al., 2010; Geldenhuys et al., 2005). Aryl-heterocycle amine groups were also shown to improve σR activity (Piergentili et al., 2009).

Si l’imagerie par résonance magnétique (IRM) montre la charge lésionnelle qui reflète le caractère inflammatoire de la sclérose en plaques (SEP), il n’existe pas de marqueur permettant de prédire son évolution ou de caractériser les phénomènes de neurodégénérescence. Par conséquent, cette étude a pour objectif premièrement d’identifier des marqueurs de l’intégrité tissulaire par IRM de tenseur de diffusion (DTI), permettant de détecter les dommages tissulaires de type inflammatoire et/ou dégénératif, et deuxièmement de caractériser leur évolution par une analyse longitudinale, chez des patients de différentes formes cliniques. A cette fin, nous avons proposé une première approche régionale des substances blanche (SB) et grise (SG) sous-corticale et une deuxième approche statistique globale analysant par TBSS les variations d’anisotropie de la SB et par VBM la densité de la SG. Les résultats obtenus dans la SB montrent des variations de la fraction d’anisotropie (FA), et des diffusivités radiales et axiales reflétant respectivement une atteinte myélinique et une atteinte axonale alors que la SG présente une augmentation de la FA suggérant une atteinte dendritique neuronale. L’analyse par TBSS et VBM montre des anomalies touchant plutôt les régions sous-corticales chez les patients rémittents qui s’étendent aux régions corticales chez les patients de forme progressive. Longitudinalement, on retrouve essentiellement des changements de FA dans la SB et d’atrophie de la SG chez les patients rémittents. Ces travaux montrent que la DTI constitue une méthode sensible pour une meilleure détection et compréhension des altérations cérébrales et de leur évolution dans la SEP. / If magnetic resonance imaging (MRI) shows the inflammatory nature of multiple sclerosis (MS) lesions, there is no marker capable of predicting its evolution or characterizing neurodegeneration. Therefore, the aim of this work was first, to identify markers of tissue integrity by diffusion tensor MRI (DTI) for the detection of inflammatory and/or degenerative tissue damages, and second, to characterize their changes with time using a longitudinal analysis of patients with different clinical forms. To this end, we first proposed a regional approach based on several white (WM) and gray (GM) matter regions of interest, and second, a statistical approach for the analysis of global WM anisotropy changes (TBSS) and GM density changes (VBM). WM analysis showed variations of the fractional anisotropy (FA), and radial and axial diffusivities, reflecting myelin and axonal damage respectively, while the GM analysis showed increased FA suggesting neuronal dendritic loss. TBSS and VBM analysis showed abnormalities affecting mostly subcortical regions in patients with relapsing-remitting (RR) MS which extended to cortical regions in patients with progressive MS. Longitudinally, we mainly observed WM FA changes and GM atrophy in RR patients. This work showed that DTI is a sensitive method for the detection and a better understanding of brain alterations and their progression in MS

La sclérose latérale amyotrophique (SLA) est une maladie neurodégénérative fatale caractérisée par une perte progressive et sélective des neurones moteurs. La SLA est incurable et il n’existe aucun traitement efficace pour les personnes atteintes de cette maladie. Environ 90% des cas sont sporadiques tandis que 10% sont familiaux, et les patients décèdent généralement deux à cinq ans après l'apparition des premiers symptômes. De nombreuses anomalies génétiques sont associées à la SLA, incluant des mutations dans les protéines FUS, C9orf72, SOD-1 et TDP-43. Le laboratoire a développé un modèle transgénique de Caenorhabditis elegans surexprimant la protéine humaine mutante TDP-43(Q331K) dans les neurones moteurs GABAergiques. Nous avons également obtenu par mutagénèse et CRISPR-Cas9 des modèles physiologiquement représentatifs du nématode basés sur des mutations dans tdp-1, l'orthologue de TARDBP chez le C. elegans. L'objectif est de caractériser ces modèles et de déterminer s'ils peuvent récapituler certains aspects phénotypiques clés de la SLA, tels que des déficits moteurs et une neurodégénérescence dépendante de l'âge générant une paralysie. L’hypothèse est que le modèle TDP-1 pourra refléter plus précisément l’expression physiologique du gène dans la maladie humaine grâce à la mutation dans un gène endogène, l’absence de surexpression et l’expression ubiquitaire de la protéine TDP-1. Les résultats montrent que les modèles TDP-43/TDP-1 ont des déficits moteurs, une transmission synaptique altérée et une neurodégénérescence liée à l’âge. Cependant, seule la mutation dans TDP-43 semble avoir un effet sur la durée de vie. Ces modèles procurent différentes expressions physiologiques des protéines mutantes et donc, des phénotypes de niveaux d'intensité variables. Ils constitueront des outils utiles pour élucider de nouveaux mécanismes pathogéniques de la SLA ainsi que de bons candidats pour le criblage de médicaments et le développement de stratégies thérapeutiques. / Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease characterized by a progressive and selective loss of motor neurons. ALS is incurable and there are no effective treatments available for people living with the disease. About 90% of the cases are sporadic whereas 10% are familial, and patients usually die two to five years after symptom onset. Many gene defects are associated with ALS, including mutations in genes encoding FUS, C9orf72, SOD-1 and TDP-43 proteins. We have developed a transgenic Caenorhabditis elegans model expressing human mutant TDP-43(Q331K) in GABAergic motor neurons. We have also obtained by mutagenesis and CRISPR-Cas9 physiologically accurate models based on mutations in tdp-1, the C. elegans ortholog of TARDBP. Our objective is to characterize these models and determine if they can recapitulate key aspects of the disease such as motor deficits and age-dependent neurodegeneration causing paralysis. We believe that the TDP-1 model will reflect more precisely the physiological expression of the gene in the human disease because of its mutation in an endogenous gene, the absence of overexpression and ubiquitous protein expression. Our results show that both TDP-43 and TDP-1 models have motor deficits, synaptic transmission impairments and age-dependent neurodegeneration. However, only the TDP-43 mutation seems to have an effect on lifespan. These models provide different physiological expression of mutant proteins and thus phenotypes of varying intensity levels. They will be useful tools to elucidate new pathogenic mechanisms of ALS as well as being good candidates for drug screening and developing therapeutic strategies.

No description available.

Alzheimer’s disease (AD) is a degenerative disorder where the distribution of pathology throughout the brain is not random but follows a predictive pattern used for pathological staging. While the involvement of defined functional systems is fairly well established for more advanced stages, the initial sites of degeneration are still ill defined. The prevailing concept suggests an origin within the transentorhinal and entorhinal cortex (EC) from where pathology spreads to other areas. Still, this concept has been challenged recently suggesting a potential origin of degeneration in nonthalamic subcortical nuclei giving rise to cortical innervation such as locus coeruleus (LC) and nucleus basalis of Meynert (NbM). To contribute to the identification of the early site of degeneration, here, we address the question whether cortical or subcortical degeneration occurs more early and develops more quickly during progression of AD. To this end, we stereologically assesses neurone counts in the NbM, LC and EC layer-II in the same AD patients ranging from preclinical stages to severe dementia. In all three areas, neurone loss becomes detectable already at preclinical stages and is clearly manifest at prodromal AD/MCI. At more advanced AD, cell loss is most pronounced in the NbM > LC > layer-II EC. During early AD, however, the extent of cell loss is fairly balanced between all three areas without clear indications for a preference of one area. We can thus not rule out that there is more than one way of spreading from its site of origin or that degeneration even occurs independently at several sites in parallel.

Huntington's disease (HD) is an inherited autosomal dominant neurodegenerative disorder caused by an unstable expansion of the CAG repeat sequence within the huntingtin gene. Huntingtin associates with ubiquitin-proteasome system that ensures degradation of particular proteins including β-catenin which is an important molecule whose equilibrated degradation is necessary for the proper functioning of the Wnt signaling pathway. The binding of β-catenin to the destruction complex is altered in HD, leading to the toxic stabilization of β-catenin. The main goal of my thesis was to determine whether the accumulation of β-catenin due to the presence of mutant huntingtin is also characteristic of Liběchov minipigs, a large animal model of Huntington's disease stably expressing N-truncated human mutant huntingtin. Using immunoblot and specific antibodies, we have revealed age-dependent accumulation of mutant huntingtin in transgenic minipigs. Unlike endogenous huntingtin, no decrease of the level of mutant huntingtin was observed in the striatum of transgenic animals. Surprisingly, this was followed by a decrease of phosphorylated β-catenin. Nevertheless, our results demostrate the accumulation of β-catenin in mesenchymal stem cells isolated from the oldest boars during ontogenesis. Furthermore, we have revealed a...

The role of metal accumulation in the pathophysiology of neurodegenerative diseases has been a hot topic in recent years due to the possibility of its treatment by chelating agents. Although the mechanisms of neurodegeneration are well known, the role of metal accumulation is still unclear. The main limitation are unsatisfactory methods for in vivo metal imaging; the most widely used technique is magnetic resonance imaging (MRI). Our aim was to assess the possibility of using transcranial sonography (TCS) in differential diagnosis of neurodegenerative diseases and to further explore the underlying factors of echogenicity. In the first study, using TCS fusion with MRI, we focused on location verification of the commonly assessed structures (substantia nigra and nucleus lentiformis) and exclusion of possible focal structural changes affecting the echogenicity in WD and PD patients. Moreover, obtained MRI were used for semi-quantitative comparison with TCS images. Although TCS has been confirmed to be highly beneficial in differential diagnosis of Wilson's disease and it should be recommended as a screening method for extrapyramidal patients with atypical course of the disease, the direct relationship between TCS and metal deposits could not be proven. The obtained results from the ultrasound fusion...

Pro les of cognitive de cit and the use of a short neuropsychological battery in di erent types of dementia Eva Bolceková Abstract This work focuses on cognitive de cit pro les and the use of a short neuropsychological battery in patients with di erent types of dementia. Increasing prevalence of dementia syndromes highlights the need of their e cient diagnostics in clinical-psychological practice. Theoretical part of this work presents an overview of neurodegenerative diseases with em- phasis on their neuropsychological presentation. We describe the cogni- tive domains and their neuropsychological examination. We address in detail the Repeatable Battery for the Assessment of Neuropsychological Status (RBANS) and its experimental Czech version. Research part of the work is based on neuropsychological examinations of 311 patients with cognitive de cit and 118 healthy controls. Patients' group is comprised of subjects with Alzheimer's disease, mild cognitive impairment, vascular cognitive impairment, Lewy Body disease, frontotemporal lobar degene- ration and patients with depression. We assess their cognitive pro les, present sensitivity, speci city, positive and negativne predictive values of RBANS scores for the groups and compare the results of control subjects with the original normative sample. We...