Lesões na parte ventral do núcleo anteromedial do tálamo (AMv) interferem no processamento da memória aversiva predatória sem no entanto influenciar as respostas de defesa inatas do animal frente a um predador. O escopo deste trabalho foi entender melhor o papel do AMv e investigar se seus alvos de projeção corticais também interferem no processamento da memória aversiva. No primeiro experimento detectamos que o AMv participa da aquisição da memória aversiva. As áreas corticais pré-límbica, cingulada anterior, visual anteromedial e retroesplenial ventral, recebem e integram entre si projeções oriundas do AMv, além de enviar projeções para a amígdala e hipocampo. Estas áreas corticais estão seletivamente recrutadas durante a exposição ao predador, e observamos que lesões neuroquímicas afetaram severamente a formação da memória aversiva. Nossos dados sugerem que há um circuito de áreas corticais que está criticamente envolvido no processo mnemônico aqui abordado, e fornece as primeiras evidências para a hipótese de módulos corticais a partir do conectoma do rato. / Neurochemical lesions placed into ventral part of anteromedial thalamic nucleus (AMv) disrupt contextual, but not innate, fear responses to predatory threats. In the present investigation, we determined whether the AMv is involved in the acquisition and/or retrieval of the conditioned responses, and if its cortical targets are involved in the fear memory processing. In the first assay, we found that AMv has a critical role in the acquisition of conditioned responses. The cortical areas prelimbic (PL), anterior cingulate area (ACA), anteromedial visual area (VISam) and the ventral part of retrosplenial area (RSPv), receive projections from AMv and are recruited during predator exposure. The integrity of these cortical areas is required for the processing of the mnemonic processes here addressed. Our data corroborate current ideas on functional cortical modules, and help to elucidate how they are involved in the acquisition of fear memories related to life threatening situations.

O objetivo principal foi investigar se o osso cortical apresenta comportamento bimodular, ou seja, se os valores de módulo de elasticidade (E) dependem do tipo de carregamento que o método utiliza. Para alcançá-lo foi preciso: (1) \"calibrar\" os diferentes métodos em um material conhecidamente unimodular, para afastar a possibilidade de erros ligados à execução de um método. (2) Depurar os métodos já calibrados para torná-los exequíveis em osso cortical. O que foi feito em espécimes de resina composta, que é um material unimodular e com E parecido com o do osso cortical. Foi investigado se o método de medida interfere no valor do E obtido neste material (pois, pelas informações da literatura, existia uma dúvida consistente). Finalmente, (3) foi avaliado se o método de medida, a região anatômica e o indivíduo interferem nos valores de E em espécimes de osso cortical. Adicionalmente, (4) verificou-se a possibilidade de se encontrar e validar uma correlação entre E e dureza Knoop (KH), para medir o E de resinas compostas indiretamente. Espécimes de latão (n = 5) tiveram o seu E calculado por quatro métodos estáticos diferentes (flexão em três pontos; tração; compressão e método de Marshall), que foram calibrados até que todos medissem o valor esperado; os valores de cada método foram comparados por análise de variância de medidas repetidas (o método foi o fator vinculado e o nível de significância foi de 5%). Espécimes (n = 4) de cinco marcas diferentes de resina composta, também foram submetidos aos quatro métodos, fazendo as adaptações pertinentes (depuração), e tiveram o seu E calculado. Estes valores foram submetidos à análise de variância de medidas repetidas (o método foi o fator vinculado e o nível de significância foi de 5 %). Espécimes (n = 48) obtidos em diferentes regiões (periósteo, endósteo e \"transversal\") de fêmures de quatro bois diferentes tiveram seu E calculado através dos métodos devidamente depurados (flexão em três pontos, tração indireta e compressão). Os valores de E foram submetidos a análises de variâncias (nível de significância de 5 %) para avaliar a influência do método, da região e do indivíduo (bois diferentes) sobre os valores de E. O método experimental de tração não foi utilizado para analisar o comportamento bimodular, porque durantes os ensaios com osso cortical, ele apresentou valores com viés. Por isso, os valores de tração foram calculados de forma indireta (Eti) através de uma equação e dos valores experimentais de flexão em três pontos (Eb) e compressão (Ec). Para validar o uso de valores de Eti, eles foram comparados com os respectivos valores de tração experimental (Et) calculados previamente nos espécimes de resinas compostas por análise de variância de medidas repetidas, e não foram achadas diferenças significantes (p > 0,05). Para encontrar e validar a correlação entre E e KH, os valores médios de E em cada espécime de resina (média do E obtido nos 4 métodos em cada espécime) foram submetidos ao teste de correlação de Pearson (nível de significância de 5%) com os respectivos valores experimentais de dureza Knoop (KH) e foi obtida uma equação de regressão linear. Para validar a equação foi confeccionado um segundo grupo de espécimes de resina (de marcas diferentes das utilizadas anteriormente) que teve seu E calculado pelo ensaio de flexão (Eb) e pela equação de regressão proposta previamente (Er); verificou-se então se a correlação entre estes valores (Eb x Er) era significante (nível de significância de 5 %), e se a nova reta de regressão apresentava coeficiente angular não diferente de 1, e coeficiente constante não diferente de zero (critérios de validação). A análise de variância não apontou diferença significante (p > 0,05) entre os valores de E calculados para o metal pelos 4 métodos, e os valores obtidos não diferiram do E conhecido do latão (ao redor de 100 GPa). As depurações necessárias foram realizadas e com elas os diferentes métodos calcularam o E dos espécimes de resina composta de modo consistente: a análise de variância encontrou diferenças significantes (p < 0,05) entre marcas (cujos E médios cobriram a faixa entre 3 e 20 GPa) e não significantes entre os 4 métodos (p > 0,05). Nos espécimes de osso cortical o valor de E foi significantemente dependente (p < 0,05) do método de obtenção (Eti > Eb > Ec). O E dos espécimes obtidos na região \"transversal\" foi menor que o E dos espécimes obtidos na região do endósteo (p < 0,05); no entanto, não houve diferença significante (p > 0,05) entre o E dos espécimes obtidos nas regiões do endósteo e periósteo e entre o E dos espécimes obtidos nas regiões do periósteo e \"transversal\". Ao considerar somente os valores obtidos no ensaio de compressão (Ec), um dos indivíduos apresentou valores de E maiores que os demais (p < 0,05). As médias de E (dos quatro métodos) em cada espécime de resina apresentaram correlação significante com os respectivos valores de KH (p < 0,05) e a equação de regressão linear: E = 0,1602 KH. Também foi significante (p < 0,05) a correlação dos valores de Eb com os respectivos valores de Er (calculados pela regressão E = 0,1602 KH); além disso, a reta de regressão linear desta nova correlação (Eb = 1,0088Er + 0,0475) apresentou coeficiente angular não diferente de 1 e coeficiente constante não diferente de zero. Pode-se concluir que: (1) Quatro métodos de medir o E apresentaram resultados semelhantes em um material metálico. (2) Estes mesmos métodos foram depurados com sucesso para sua utilização em espécimes de osso cortical ao constatar que também não interferiram no valor do E obtido em espécimes semelhantes aos de osso, confeccionados com resinas compostas. (3) Os valores de E de osso cortical dependem do método de medida, o que demonstra seu comportamento bimodular. A região anatômica e o indivíduo também afetam os valores de E dos espécimes de osso cortical (o indivíduo só foi um fator significante ao considerar somente os valores obtidos no ensaio de compressão). (4) Foi encontrada e validada uma correlação entre E e KH em resinas compostas, que permite calcular o E deste material através da regressão E = 0,1602 KH. / The main objective was to investigate whether the cortical bone has a bimodular behavior, that is, if the values of elastic modulus (E) depend on the load type used by the method. To achieve this, it was necessary: (1) \"calibrate\" the different methods in a material with well-known unimodular behavior, for excluding the possibility of systematic errors in the execution of the methods. (2) Depurate the calibrated methods to make them executable in cortical bone. This was made on specimens of composite resin, which is a unimodular material and has an E similar to that of cortical bone. It was also investigated if the method of measuring the E influences the obtained value in this material (because there was a consistent doubt in the literature). Finally (3), we assessed whether the E values in cortical bone are influenced by the measurement method, by the anatomical region and by the individual. Additionally, (4) it was verified if it would be possible suggesting and validating a correlation between E and Knoop hardness (KH), to indirectly measure the E of composite resins. Brass specimens (n = 5) had their E calculated by four different static methods (three point bending, tensile, compression and Marshall\'s method), which were calibrated until all the methods were capable of measuring the expected value; the E values of each method were compared by analysis of variance of repeated measures (the method was the linked factor and the significance level was 5%). Specimens (n = 4) of five different composite resins were also subjected to the four methods, making the relevant adjustments (depuration), and had their E calculated. These values were subjected to analysis of variance of repeated measures (the method was the linked factor and the significance level was 5%); Specimens (n = 48) obtained in different regions (periosteum, endosteum and \"transverse\") of bovine femurs from four different individual had their E calculated using the depurated methods (three point bending, indirect tensile and compression). The E values were subjected to analyzes of variance (significance level of 5%) to evaluate the influence of the method, of the region and of the individual (different cattle) in the E values. The experimental tensile method was not used to analyze the bimodular behavior because during tests on cortical bone, this method showed values with bias. Therefore, the tensile values were indirectly calculated (Eti) by a formula and by the experimental values of E calculated by three point bending (Eb) and compression (Ec). To validate the use of Eti values, they were compared with the corresponding tensile experimental values (Et) previously measured in the composite resins specimens by analysis of variance (repeated measures), and no significant difference was found (p > 0.05). For suggesting and validating the correlation between E and KH, the average values of E of each composite specimen (mean of E obtained considering the values of the four methods in each specimen) were submitted to the Pearson\'s correlation test (significance level of 5 %) with the corresponding experimental KH values and it was obtained the linear regression equation. To validate this equation, it was made a second group of composite resins specimens (with different brands of the previously used) in which the E was calculated by three point bending test (Eb) and by regression equation previously proposed (Er); then it was checked whether the correlation between these values (Eb x Er) was significant (significance level of 5 %), and if the new regression line showed angular coefficient no different from one, and constant coefficient no different from zero (validation criteria). The analysis of variance showed no significant difference (p > 0.05) among the E values calculated for the metal specimens by the four methods; the obtained values did not differ from the known E of the brass (about 100 GPa). The necessary adjustments (depuration) were made and then the different methods calculated the E of the composite resin specimens consistently: the analysis of variance found significant difference (p < 0.05) between composite brands (whose E mean covered the range between 3 and 20 GPa) and no significant difference among the four methods (p> 0.05). In the specimens of cortical bone, the E values were significantly dependent (p < 0.05) of the obtainment method (Eti > Eb> Ec). The E values of the \"transversal\" specimens were lower than the endosteum specimens (p < 0.05); but there was no significant difference (p > 0.05) between the E of endosteum specimens and periosteum specimens and between the E of periosteum specimens and \"transversal\" specimens. When considering just E values of the compression method, one cattle showed higher E values than the others (p < 0.05). The mean of the four methods for each specimen composite resin exhibited a significant correlation with the respective KH values (p < 0.05) and the linear regression equation: E = 0.1602 KH. It was also significant (p < 0.05) the correlation between Eb and the respective Er values (calculated by the regression E = 0.1602 KH); furthermore, the linear regression of this new correlation (Eb = 1.0088Er + 0.0475) presented angular coefficient no different from 1, and constant coefficient no different from zero. It can be concluded that: (1) Four methods of E measuring showed similar results in a metallic material. (2) The same methods were successfully depurated for their execution in cortical bone specimens, since the measurement method did not affect the obtained E value in resin composites (that is a material with E value similar to cortical bone); (3) The E values of cortical bone depend on the measurement method, which demonstrates its bimodular behavior. The anatomical region and the individual also affect the E values of the cortical bone specimens (the individual was a significant factor only when considering just the E values obtained in the compression test). A correlation between E and KH that allows the E estimation by the regression E = 0.1602 KH was suggested and validated in composite resins.

Pacientes com epilepsia farmacorresistente, frequentemente, possuem lesões extrahipocampais como etiologia. Muitas vezes, estes pacientes se beneficiam de lesionectomias para redução da ocorrência de crises epilépticas. Para que possam se submeter a este procedimento, atualmente é necessário o uso tanto da videoeletroencefalografia prolongada (VEEG) quanto da imagem de ressonância magnética do encéfalo (IRM) para delimitação apurada da zona epileptogênica, local que deve ser ressecado para controle das crises. No presente trabalho, foi estudada a acurácia diagnóstica da VEEG e da IRM na determinação da zona epileptogênica de pacientes com displasia cortical focal. Comparou-se os locais de ocorrência da zona de início ictal (VEEG) e da lesão epileptogênica (IRM) se concordantes ou discordantes com o local da cirurgia. Foram revisados os prontuários médicos de 209 pacientes, sendo o padrão de referência (local da cirurgia) e tempo de acompanhamento pós-operatório superior a 12 meses disponíveis em 43 pacientes. A VEEG apresentou sensibilidade de 85,7% (IC 95% 62,6-96,2) e especificidade de 41,1% (IC 95% 19,4-66,5), com valor preditivo positivo de 64,2% (IC 95% 44,1-80,6) e valor preditivo negativo de 70% (IC 95% 35,3-91,9). A IRM apresentou sensibilidade de 91,6% (IC 95% 71,5-98,5) e especificidade de 36,8% (IC 95% 17,2-61,3), com valor preditivo positivo de 64,7% (IC 95% 46,4-79,6) e valor preditivo negativo de 77,7% (IC 95% 40,1-96). As diferenças de sensibilidade e especificidade, áreas sob as curvas ROC e os índices de Youden não foram significativas. A concordância dos resultados da VEEG e da IRM foi moderada (k=0,599; p<0,01; IC 95% 0,468-0,730). / Patients with drug-resistant epilepsy frequently have extrahippocampal lesions as etiology. A large proportion of these patients might benefit from lesionectomy for the reduction of seizures. For surgery to be undertaken, it is usually performed both long-term videoelectroencephalography monitoring (VEEG) and magnetic resonance imaging of the brain (MRI) for the precise delimitation of the epileptogenic zone, the region that must be resected for seizure control. In the present study, the diagnostic accuracy of VEEG and MRI were studied in the localization of the epileptogenic zone in patients with focal cortical dysplasia. The seizure-onset zone (VEEG) and the region of epileptogenic lesion (MRI) were compared whereas concordant or discordant regarding surgery region. Medical charts of 209 patients were reviewed, being the reference standard (surgery region) and post-surgical follow-up longer than 12 months available in 43 patients. Videoelectroencephalography has a sensitivity of 85.7% (95% CI 62.6-96.2) and specificity of 41.1% (95% CI 19.4-66.5), with positive predictive value of 64.2% (95% CI 44.1-80.6) and negative predictive value of 70% (95% CI 35.3-91.9). Magnetic resonance imaging has a sensitivity of 91.6% (95% CI 71.5-98.5) and specificity of 36.8% (95% CI 17.2-61.3), with positive predictive value of 64.7% (95% CI 46.4-79.6) and negative predictive value of 77.7% (95% CI 40.1-96). The differences of sensitivity and specificity, areas under the ROC curves and Youden\'s indexes were not significant. The concordance between the results of VEEG and MRI was moderate (k=0.599; p<0.01; 95% CI 0.468-0.730).

Orientadores: IsciaTeresinha Lopes Cendes, Fábio Rossi Torres / Dissertação (mestrado) - Universidade Estadual de Campinas, Faculdade de Ciências Médicas / Made available in DSpace on 2018-08-21T05:32:00Z (GMT). No. of bitstreams: 1 Avansini_SimoniHelena_M.pdf: 4063977 bytes, checksum: fd585dcd7befa8b76aa15db5e445d1d2 (MD5) Previous issue date: 2012 / Resumo: A displasia cortical focal (DCF) é uma malformação do córtex cerebral humano que ocorre na fase de proliferação e diferenciação neuronal e está frequentemente associada com a refratariedade das crises epilépticas. É designada como um espectro de anormalidades da estrutura laminar do córtex, associada com características citopatológicas que incluem neurônios gigantes, dismórficos e células em balão e, sua etiologia é pouco conhecida. Os microRNAs (miRNAs) são uma classe de RNAs de fita simples não codificadores de proteínas que regulam a expressão gênica pós-transcricional. Há evidências que indicam que eles estão envolvidos em importantes processos do sistema nervoso e que os mesmos podem ter um papel nas DCF. A elucidação das vias moleculares desta malformação pode permitir uma melhor compreensão dos mecanismos subjacentes e levar a novas estratégias de tratamento, melhora na conduta clínica e identificação de novos alvos terapêuticos, bem como a descoberta de biomarcadores que possam ser associados ao diagnóstico, prognóstico e resposta ao tratamento. Com isso, o objetivo deste trabalho foi investigar o padrão de expressão dos miRNAs em tecidos com DCF obtidos através de cirurgia para o controle de crises refratárias, verificar os prováveis genes alvos para esses miRNAs diferencialmente expressos e comparar a assinatura molecular baseada nos miRNAs em tipos distintos de DCF do tipo 2. Foi utilizado para isso o RNA total de 17 pacientes com DCF e de 20 controles oriundos de autópsia. Os experimentos de microarranjos de miRNAs revelaram 39 miRNAs diferencialmente hipoexpressos e um miRNA hiperexpresso. Utilizouse a técnica de qPCR para validação desses miRNAs e foi possível identificar uma diferença na expressão de três miRNAs: hsa-miR-31, hsa-miR-34a e hsa-let-7f em pacientes com DCF tipo 2 em relação ao grupo controle. Além disso, o hsa-miR-31 foi identificado como um possível biomarcador para o subtipo 2b de DCF. Na busca por genes alvos foi encontrado hiperexpresso o NEUROG2. Também foi verificada a desregulação do gene DICER1, encontrado hipoexpresso, o que justifica a predominância de miRNAs com expressão diminuída encontrados. E por fim, observou-se que o padrão diferencial de expressão dos três miRNAs e os dois genes identificados em nosso estudo fornecem subsídios importantes para esclarecer os mecanismos moleculares envolvidos na falha da diferenciação neuroglial em DCF tipo 2 / Abstract: Focal cortical dysplasia (FCD) is a malformation of human cerebral cortex that occurs during proliferation and neuronal differentiation frequently associated with drug-resistant epilepsy. It is designated as a spectrum of abnormalities of laminar structure of the cortex, associated with cellular abnormalities that consist of giant and dysmorphic neurons and balloon cells; however, its etiology is poorly understood. MicroRNAs (miRNAs) are small noncoding RNAs which regulate post-transcriptional gene expression. There is evidence that they are involved in important processes in the nervous system and that they may play a role in FCD. The elucidation of the molecular pathways involved in FCD may allow a better understanding of the underlying mechanisms and may lead to new treatment strategies, improvement in clinical management and identification of new therapeutic targets, as well as the discovery of biomarkers that may be associated with the diagnosis, prognosis and response to treatment. Thus, the aim of this study was to investigate miRNAs expression pattern in tissue with FCD obtained at surgery for control of refractory seizures. In addition, we aimed to identify target genes for these miRNAs differentially expressed, as well as to compare the molecular signature based on miRNAs in different types of FCD. We used total RNA isolated from brain tissue obtained after surgery for the treatment of medically refractory seizures from 17 patients with DCF and 20 controls from autopsy. Microarray analysis revealed 39 miRNAs differentially downregulated and only one miRNA overexpressed. Decreased expression of three miRNAs was confirmed by qPCR when patients with type 2 FCD were compared with controls: hsa-miR-31, hsa-miR34a and hsa-let-7f. In addition, we found that hsa-miR-31 could be a potential biomarker for type 2b FCD. In the search for target genes, NEUROG2 was found upregulated and DICER1 was found underexpressed, which explains the predominance of miRNAs with decreased expression. Finally, we observed that the differential pattern of expression of three miRNAs, and the two genes identified in our study provide important information which may help to clarify the molecular mechanisms involved in the failure of neuroglial differentiation in type 2 FCD / Mestrado / Neurociencias / Mestra em Fisiopatologia Médica

Orientador: Iscia Teresinha Lopes Cendes, Fábio Rossi Torres / Tese (doutorado) - Universidade Estadual de Campinas, Faculdade de Ciências Médicas / Made available in DSpace on 2018-08-21T06:16:28Z (GMT). No. of bitstreams: 1 Tsuneda_SimoneSayuri_D.pdf: 5548129 bytes, checksum: b0213ba3907f298ca16dddb2df837b62 (MD5) Previous issue date: 2012 / Resumo: A polimicrogiria (PMG) é uma malformação do córtex cerebral causada por falhas no seu desenvolvimento, caracterizando-se por um número excessivo de pequenos giros e laminação anormal, dando à superfície cortical uma aparência irregular e grosseira. A gravidade de suas manifestações clínicas se relaciona diretamente com a extensão da malformação e das regiões cerebrais afetadas, sendo que a presença de lesões bilaterais ou unilaterais extensas indica um pior prognóstico. Uma das síndromes de polimicrogiria mais frequentes e, consequentemente, mais bem descritas clinicamente, é a polimicrogiria perisylviana bilateral (PPB). Essa forma de polimicrogiria atinge a região que tange a fenda Sylviana, podendo apresentar-se tanto unilateralmente quanto em ambos os hemisférios. O padrão de herança da PPB foi descrito inicialmente como ligada ao cromossomo X por Borgatti et al. em 1999. Já em 2000, Guerreiro et al. confirmaram o padrão de herança consistente com herança ligada ao cromossomo X, mas ainda nenhum gene havia sido identificado como responsável pelo distúrbio. Nosso grupo recentemente mapeou uma nova região candidata para a PPB em Xq27.1-q27.3, e esta tese se propôs a avaliar essa região através da técnica de sequenciamento em larga escala aliada à tecnologia de captura para o cromossomo X. Os resultados apontaram como potenciais patogênicos os genes MAGEC1, UBE2NL, além da região do gene SPANXC, todos localizados na região candidata, mas uma avaliação mais detalhada levantou a hipótese de uma relação complexa entre as alterações encontradas no gene MAGEC1 e o quadro clínico dos pacientes. Além da análise da região Xq27.1-q27.3, considerando o grande número de genes de microtúbulo que tem sido relacionado a malformações do córtex cerebral, esse trabalho também avaliou pacientes esporádicos e famílias com histórico de PPB realizando triagem de mutações nas regiões codificantes dos genes AFF2, SLITRK2 e SLITRK4, localizados na região candidata, nos genes de microtúbulo TUBA1A, TUBB2B e TUBA8, além dos genes SRPX2 e WDR62, presentes em trabalhos na literatura de malformações corticais. A triagem foi realizada utilizando as técnicas de DHPLC e de sequenciamento utilizando a técnica de Sanger por eletroforese capilar. Foi encontrada uma alteração potencialmente patogênica no gene AFF2. As alterações identificadas neste estudo que resultam em troca de aminoácidos foram avaliadas utilizando as ferramentas in silico MutPred, SNPs&GO, Polyphen 2, Panther e SIFT, de forma a fornecer mais informações a respeito de seu potencial patogênico. Além disso, as variantes inéditas identificadas nesse trabalho foram estudadas em uma amostra de indivíduos normais (grupo controle). Com esses dados foi possível sugerir que algumas dessas variantes encontradas possuem potencial patogênico que deve ser futuramente investigado através de estudos funcionais / Abstract: Polimicrogyria (PMG) is a cortical malformation caused by failures during the brain cortex development process and is characterized by an excessive number of small gyri, resulting in an irregular cortical surface. The severity of its clinical manifestations is directly related to the extension of the tissue abnormalities. Bilateral Perisylvian Polimicrogyria (BPP) is the most comum and, consequently, a very well described syndrome that affects the cortex surrounding the Sylvian fissures in both hemispheres. The genetic pattern for BPP was initially described by Borgatti et al. as an X-linked pattern, confirmed by Guerreiro et al. in 2000, but with no specific gene identified. We have recently described a candidate site for BPP at the Xq27.1-27.3 region and, in this project, we proposed to evaluate this site through next generation sequencing technology combined with capture technology. Our results suggest that MAGEC1 and UBE2NL genes, or the SPANXC gene area might be related to the pathogeny in this case, however a further analysis brought up the hypothesis of a complex relation between the MAGEC1 mutations and the clinical manifestations in each different patient. Considering recurrent description of relations between microtubule genes and cortex malformations, we also performed the evaluation of exon regions of eight selected genes from sporadic patients and BPP families through DHPLC and sequencing. The analysis focused on AFF2, SLITRK2 and SLITRK4 genes, located at the identified site, microtubule genes TUBA1A, TUBB2B and TUBA8, and SRPX2 and WDR62 genes, also related to cortical malformations. As a result from this screening, we identified a potentially pathogenic mutation in gene AFF2. All non-synonymous SNPs were evaluated using the in silico tools MutPred, SNPs&GO, Polyphen 2, Panther and SIFT, providing further insights for their analysis. A control group of individuals was analyzed for the presence of the non-described SNPs. These data suggest a pathogenic potential for these genetic alterations that must be investigated through function studies / Doutorado / Fisiopatologia Médica / Doutora em Ciências

Neural entrainment quite recently became considered an important mechanism used by the brain to process stimuli with periodic qualities, such as the frequency and duration time of signals reaching sensory organs. An increasing amount of data strongly implies that the brain might be using neural entrainment as a mechanism to either directly process speech and/or to facilitate speech interpretation. Neural entrainment is therefore a promising marker to use for research of speech perception. This literature review aims to summarize the most recent findings within this area with the end-goal to be used as a basis for designing an EEG experiment intended to analyze speech perception as a means to distinguish human voices.    For this reason, data was collected from the scientific databases Europe PMC, Academic Search Premier, PsycINFO, PubMed, Scopus and Web of Science, where the keywords “EEG” + either the phrase “neural entrainment”, “neural oscillation”, or “cortical oscillation” were used to gather articles. Inclusion and exclusion criteria were then applied and the data was analyzed with the intention to answer the following research questions: “is it possible to observe neural entrainment to human voice/speech using EEG?”, “if so, what are the possibilities to use such neural entrainment as a marker for differentiating human voices from each other?” and “what is the nature of the mechanisms used by the brain to attain this entrainment?”. The resulting data from the articles indicated that, in order to yield reliable results when investigating neural entrainment to speech, the technique for analysis of brain activity could be done with EEG, a number of participants between 15-30 persons is enough, the spectral bands of interest are delta (&lt;3 Hz), theta (4-8 Hz), beta (15-35 Hz) and gamma (&gt;40 Hz), the method of analysis could be looking at both frequency and amplitude in the speech envelope, and finally the anatomical areas for investigating the brain’s ability to distinguish human voices using speech entrainment could be either areas within the auditory cortex or prefrontal areas involved in behavioral responses to speech processing.

Une meilleure compréhension des relations entre la structure complexe de l'os cortical et ses propriétés mécaniques est nécessaire à l'évaluation de la qualité osseuse. Les méthodes conventionnelles ex vivo de mesure de l'élasticité à l'échelle du millimètre ont des limitations liées à l'anisotropie du tissu, à son inhomogénéité et à la petite taille des échantillons. Au contraire, la spectroscopie par résonance ultrasonore (RUS) est bien adaptée à la mesure de petits échantillons anisotropes. Cette méthode estime l'élasticité à partir des fréquences de résonance de l'échantillon, et l'amortissement à partir de la largeur des pics de résonance. Son application à l'os était considérée difficile, du fait de l'amortissement important des modes de vibration, qui induit un recouvrement des pics de résonance et complique la mesure des fréquences. Pour surmonter cette difficulté, des adaptions de la méthode - dans la mesure, le traitement du signal et l'estimation des propriétés du matériau - ont été proposées. Elles ont été validées sur de l'os cortical et sur des échantillons de polymère et de matériau composite imitant l'os. La précision de la méthode a été démontrée, ainsi que sa capacité à mesurer tous les termes du tenseur d'élasticité à partir d'un seul échantillon. De plus, une nouvelle formulation Bayésienne de l'inversion apporte une solution automatique à un problème qui nécessitait une stratégie fastidieuse d'essai-erreur ou de complexes modifications du dispositif expérimental. Finalement, l'application à une grande collection d'échantillons de tibias humains démontre que la méthode RUS pourrait être utilisée en routine pour la mesure de la viscoélasticité de l'os. / Deep understanding of the structure-function relationships of cortical bone in the context of bone quality assessment is still missing. Currently available methods to measure millimeter-scale elasticity ex vivo have limitations arising from theanisotropy of the tissue, its heterogeneity, and the small size of the specimens.Resonant ultrasound spectroscopy is particularly suitable for the measurement of small anisotropic specimens. This method estimates elasticity from the free resonant frequencies of a specimen, and damping from the width of the resonant peaks. Its application to cortical bone was considered challenging because of the high damping of the vibrations modes, which causes overlapping of the resonant peaks and prevents a direct measurement of the resonant frequencies. To overcome the difficulty,adaptations of all the steps of RUS – measurement, signal processing and inverse estimation of the material properties – have been introduced. Validation of each step of the procedure has been achieved by application to several test samples, including a cortical bone specimen and bone-mimicking composite and polymer specimens.RUS was shown to be precise and accurate, with the advantage of providing the complete stiffness tensor from the measurement of a single specimen. Additionally, an original Bayesian formulation of the inversion provides an automated solution toa problem that was previously solved by tedious trial-and-error procedures or complex additions to the basic experimental setup. Finally, the application to a large collection of human tibiae specimens demonstrates that RUS can be considered a routine method to characterize the viscoelasticity of bone.

Dans le but d’enrichir la modélisation virtuelle d’êtres humains et de mieux comprendre la biomécanique de certaines parties du squelette, ce travail propose une analyse comparative des propriétés histologiques et mécaniques de deux os appendiculaires fonctionnellement opposés : l’humérus et le fémur. La campagne a été réalisée à partir d’échantillons provenant de quatre SHPM embaumés (Sujet Humain Post-Mortem), de sexe masculin. Une étude géométrique en 3D a débuté l’expérimentation. Elle a été suivie par une analyse histomorphométrique de 153 photographies réalisées à partir de la face antéromédiale du cortex diaphysaire, à quatre niveaux de hauteur et à trois niveaux de profondeur. Des essais mécaniques ont ensuite été effectués sur 28 éprouvettes d’os cortical non congelé, provenant du même site anatomique. L’expérimentation s’est déroulée sur machine conventionnelle de traction. Elle comportait des essais en traction/compression et des essais de cyclage en traction dans le domaine élastique, à la vitesse de 0,05 mm/mn, jusqu’à rupture. Une loi d’endommagement a également été élaborée. Ces travaux ont montré que ces deux os offrent un comportement différent. L’humérus s’avère être moins résistant et plus raide que le fémur. Son endommagement intervient plus rapidement. Les valeurs mécaniques relevées sont en rapport avec la densité et la taille des ostéones actifs, ainsi qu’avec les caractéristiques de la porosité Haversienne. Les différences de comportement mécanique relevées s’expliquent par l’adaptation microscopique du tissu osseux cortical aux contraintes subies par un os porteur ou non-porteur. Les variations interindividuelles observées sont fonction de son état physiologique. / With the aim of enriching the virtual modelisation of human beings and understanding better the biomechanics of some parts of the skeleton, this work proposes a comparative analysis of histological and mechanical attributes of two functionally opposed appendicular bones: femur and humerus. The campaign has been done with samples coming from four embalmed PMHS (post mortem human subjects) of the male gender. A 3D geometric study started the experiment. It was followed by an histomorphometric analysis of 153 pictures carried out on the anteromedial face of the diaphyseal cortex at four levels of height and three levels of depth. Mechanical tests were then done on 28 specimens of non frozen cortical bone coming from the same anatomic site. The experiment took place on a conventional traction machine. It consisted of traction/compression tests and cycling tests under traction in the elastic zone, at the speed of 0.05 mm/mn until yield point. A damage law has also been elaborated. These studies have shown that these two bones offer a different behaviour. The humerus bone turns out to be less resistant and stiffer than the femur. It is damaged more quickly. The mechanical values noted are related to the density and the size of active osteons and also to the characteristics of Haversian porosity. The difference of mechanical behavior noticed can be explained by the microscopic adaptation of the cortical bone tissue to the stresses undergone by the bearing and non bearing bones. The inter-individual variations observed are linked to the physiological state of this tissue.

Le développement du cortex cérébral se déroule selon des étapes bien définies qui sont essentielles à la formation d’un cerveau fonctionnel. La perturbation de l’une ou plusieurs de ces étapes peut conduire à des malformations neuro-développementales, responsables de différents troubles cognitifs, d’épilepsies ou encore de déficience intellectuelle. De nombreuses mutations dans des gènes codant pour les tubulines ou bien les kinésines, sont retrouvées chez des individus présentant diverses anomalies neuro-développementales. Bien que les kinésines soient impliquées dans le développement cortical, les mécanismes fonctionnels par lesquels elles conduisent aux malformations demeurent encore méconnus. Mon travail de thèse identifie la kinésine Kif21b, jusqu’alors peu connue, comme étant essentielle au développement cortical. Nous montrons que Kif21b régule la migration neuronale dans le cortex et identifions quatre variants chez des individus présentant des malformations neuro-développementales. Nous montrons que l’expression ectopique des variants chez la souris et le poisson zèbre récapitulent les phénotypes observés chez ces patients. / The development of the cerebral cortex is a highly regulated process that is crucial for the establishment of functional cortical networks. Disruption of one or several of these steps can lead severe neurodevelopmental disorders that are associated with intellectual disabilities, epilepsies and cognitive impairment. Over the past few years, several genetic mutations in genes encoding either tubulin or microtubule-associated motors such as kinesins, have been found in individuals with neurodevelopmental disorders. Although kinesins have been found to be essential for a proper cortical development, the exact functions of kinesins in these processes are still poorly understood. My work clearly identified Kif21b, a poorly-known kinesin, as a novel key regulator of cortical development both in mouse and human. We show that Kif21b regulates both radial and tangential migration of cortical neurons, and identify four KIF21B variants in individuals presenting neurodevelopmental disorders. We show that ectopic expression of variants recapitulate phenotypes both in mice and zebrafish.

Objectifs : Le bruxisme survenant au cours du sommeil est un trouble du mouvement caractérisé par du grincement de dents et l’activité rythmique des muscles masticateurs (ARMM). Le bruxisme/ARMM est souvent associé à des mouvements du corps et des à éveils corticaux. Une séquence d’activation précède le ARMM/bruxisme. Ces événements incluent une augmentation des variables suivants : l’activité sympathique (-4 minutes), les activités encéphalographique (-4 second), le fréquence cardiaque, l’amplitude de la respiration (-1 second) et l’activité des muscle suprahyoïdiens (-0.8 second). La présente étude a examiné l’association entre le bruxisme et les changements de la pression artérielle. Méthodes: Dix sujets avec le bruxisme (5 hommes, 5 femmes, âge moyen = 26 ± 1,8) ont complétés 3 nuits de polysomnographie qui comprenait l'enregistrement non invasive de la pression artérielle. La première nuit a servi de dépistage et d’habituation au laboratoire. L'analyse a été réalisée sur les deuxièmes et troisièmes nuits enregistrements. Seuls les épisodes de bruxisme isolés survenant au cours du stade 2 du sommeil ont été utilisés pour l’analyse, pour un total de 65 épisodes. Les mesures des pressions systolique et diastolique ont été prises 20 battements avant et 23 battements après l'apparition de chaque épisode bruxisme lors du sommeil. Les épisodes de bruxisme ont été classés comme suit: 1) bruxisme avec éveil cortical; 2) bruxisme avec mouvement du corps (MC), 3) bruxisme avec éveil cortical et MC. Une quatrième catégorie, bruxisme seul, a également été analysée, mais utilisée comme donnée préliminaire puisque la catégorie se composait de seulement 4 épisodes de bruxisme. Résultats: Les deux pressions systolique et diastolique ont augmenté avec les épisodes de bruxisme. Cette augmentation a été statistiquement significative pour la pression systolique et diastolique pour les épisodes de bruxisme avec éveil cortical et/ou MC (p ≤ 0,05). L’augmentation moyenne de la pression (systolique / diastolique ± SE) a été : 28,4 ± 2,4/13,2 ± 1,5 mm Hg pour le bruxisme avec éveil cortical; 30,7 ± 1,6/19.4 ± 2.3 mm Hg pour bruxisme avec MC; 26.4 ± 2,8 / 14,6 ± 2.0mm Hg pour bruxisme avec éveil cortical et MC; 22,9 ± 5,2/12,4 ± 3,3mm Hg pour les épisodes de bruxisme seuls. Conclusion: Le bruxisme du sommeil est associé à des hausses de la pression artérielle pendant le sommeil. Cette hausse est supérieure dans les épisodes de bruxisme associés à un éveil cortical et / ou MC, qui sont souvent associés avec les événements bruxisme. Ces résultats sont en accord avec nos observations antérieures, où le bruxisme est précédé par une augmentation de l'activité sympathique et de la tachycardie sinusale. / Objectives: Sleep Bruxism (SB) is a movement disorder identified by tooth grinding and rhythmic masticatory muscle activity (RMMA). It is often associated with body movements and sleep arousals. Increases in autonomic sympathetic activities that characterize sleep arousal precede SB. These events include an augmentation of the following variables: sympathetic cardiac activity (-4 minutes), electroencephalography frequencies (-4 seconds), heart rate and respiratory amplitude (-1 seconds), and suprahyoid muscle activity (-0.8 seconds). This study examined whether these sympathetic activities are associated with significant changes in arterial blood pressure (BP). Methods: Ten subjects with SB (5 male; 5 female; mean age ± standard error = 26 ± 1.8) underwent 3 nights of full polysomnography that included non-invasive beat to beat BP recording. The first night served as a screening and habituation night. Analysis was performed on second and third night recordings. Overall analysis was based on single SB episodes occurring in stage 2 sleep only, for a total of 65 episodes. Systolic and diastolic BP measurements were taken from a window of 20 beats before and 23 beats after onset of each SB episode. SB episodes were categorized as: 1) SB + cortical arousal; 2) SB + body movement (BM); 3) SB + cortical arousal + BM. A fourth category, SB alone, was also analysed but used as preliminary data since the category consisted of only 4 episodes. Results: Both systolic and diastolic BP increased with SB episodes. This increase was significant for both systolic and diastolic BP for SB events with cortical arousal and/or BM (p≤0.05). The average BP surges (systolic/diastolic ± SE) were: 28.4 ± 2.4/13.2 ± 1.5mm Hg for SB + cortical arousal; 30.7 ± 1.6/19.4 ± 2.3mm Hg for SB + BM; 26.5 ± 2.8/14.6 ± 2.0mm Hg for SB + cortical arousal + BM; 22.9 ± 5.2/12.4 ± 3.3mm Hg for SB episodes occurring alone. Conclusion: Sleep bruxism is associated with blood pressure fluctuations during sleep. This BP surge is greater in SB episodes associated with cortical arousal and/or BM, which often co-occur with SB events. These results are congruent with our previous observations, where SB is preceded by a rise in sympathetic activity and sinus tachycardia.