Biobanco “AZORBIO”: organização de recursos biológicos para a investigação

Dissertação de Mestrado em Ciências Biomédicas.INTRODUÇÃO: Os biobancos são essenciais em investigação, por possuírem coleções de amostras e dados armazenados de forma organizada. O biobanco Açores (AZORBIO) do Serviço Especializado de Epidemiologia e Biologia Molecular (SEEBMO) possui uma coleção de material biológico e dados associados de doentes açorianos com diversas patologias. A Litíase Urinária (LU) é uma entidade clínica de etiologia multifatorial, cujo desenvolvimento resulta de um processo complexo que envolve vários mecanismos fisiopatológicos. OBJETIVO: Melhoria das condições de colheita, acondicionamento, transporte, receção, processamento e controlo de qualidade das amostras e dados destinados ao AZORBIO, através da aplicação de novos procedimentos padronizados. METODOLOGIA: Foi realizada uma revisão da literatura sobre normas e leis aplicáveis aos biobancos e foram verificadas as condições do AZORBIO. Foi elaborado um termo de consentimento informado e um questionário e foram estabelecidos procedimentos operacionais padrão para que todo o circuito de colheita, receção, processamento e armazenamento das amostras de LU seja uniformizado. Todos os procedimentos foram posteriormente aferidos com as primeiras amostras de LU recebidas no AZORBIO. As amostras de DNA e RNA foram submetidas a controlo de qualidade para verificar a concentração e pureza, integridade e funcionalidade. RESULTADOS E CONCLUSÃO: O AZORBIO apresenta as instalações e os equipamentos necessários para o seu funcionamento. A documentação produzida (consentimento informado, folha informativa, questionário e procedimentos operacionais padronizados) permitiu uniformizar todo o circuito para as amostras de LU. As amostras de DNA e RNA obtidas, através dos procedimentos realizados, apresentaram boa qualidade. Com este trabalho o AZORBIO, num futuro próximo, poderá garantir amostras e dados de qualidade de LU e consequentemente contribuir para o melhoramento dos cuidados de saúde e redução da morbilidade desta doença. Para além disso, os procedimentos aqui propostos poderão servir de base para outras patologias de interesse para o AZORBIO.INTRODUCTION: The biobanks are essential in research, by having collections of samples and data stored in an organized manner. The biobank Azores (AZORBIO) of the Specialized Service of Epidemiology and Molecular Biology (SEEBMO) has a collection of biological material and associated data of Azorean patients with different pathologies. The pathology of interest, chosen for this thesis was the Urinary lithiasis (LU), a multifactorial clinical entity that results from several pathophysiologic mechanisms. OBJECTIVE: The application of new standardized procedures for collection, packaging, transportation, reception, processing, storage and quality control of the samples and data for the AZORBIO. METHODOLOGY: Literature review of rules and laws applicable to biobanks. An informed consent form and a specific LU questionnaire were performed and standard operating procedures for the entire circuit were established. All procedures were subsequently assessed with the first ten samples received on AZORBIO. Nucleic acid samples were subjected to quality control to verify the concentration and purity, integrity and functionality. RESULTS AND CONCLUSION: AZORBIO has the facilities and equipment necessary for its operation. The documentation produced (informed consent, informative document, questionnaire and standard operating procedures) contributed for the standardization of the entire circuit for samples of LU. Although the overall circuit assessment showed good quality results, minor changes were suggested to improve the final outcome. With this work the AZORBIO in the near future, will ensure good LU quality samples and data contributing to the improvement of health care and reduce morbidity of this disease. In addition, the procedures proposed here can be adapted to other pathologies of interest to AZORBIO

Transtorno mental orgânico após meningoencefalite pneumocócica com sintomas semelhantes aos do autismo

Universidade Federal de São Paulo (UNIFESP)Universidade Federal do TocantinsUNIFESPSciEL

Genetic variants associated with ectopic calcifications

Diffuse idiopathic skeletal hyperostosis (DISH) is a common skeletal disorder characterized by the presence of new bone formation in ligaments and entheses. DISH can co-exist with Chondrocalcinosis (CC) and it has been suggested that both diseases share the same pathogenic mechanism. To date, two genes, COL6A1 and FGF2, have been shown to have a weak positive association with DISH susceptibility. The main objective of this thesis was to investigate the genetic basis of the DISH/CC disease, making use of Next Generation Sequencing technology, association and expression studies, in a group of DISH/CC samples from the Azores biobank. Two regulatory variants in the RSPO4 gene were significantly more frequent in controls than in DISH/CC patients. These may protect against the DISH/CC phenotype, possibly by altering gene expression of the RSPO4 gene. Using whole exome sequencing we identified a significant association between the DISH/CC disease and a genetic variant in BMP4 (rs17563), a gene involved in endochondral bone formation. Another of the candidate genes associated with DISH/CC was ABCC6 that is of relevance in ectopic calcification disorders. Although inconclusive, the expression studies performed in human cartilage tissue indicated overexpression of ABCC6 in DISH and CC patients relative to the controls, raising the hypothesis that this gene may be involved in calcium pyrophosphate formation in DISH and CC. A comparative approach using teleosts revealed that the abcc6 gene is expressed in skin but was not associated with ectopic calcification of the scales. Furthermore, comparative genomics revealed the abcc6 has only been retained in the genome of bony vertebrates. In summary, I identify for the first time potential gene variants that protect (RSPO4) or predispose (BMP4) to DISH/CC. The relevance of the ABCC6 gene in this phenotype remains to be proven. It is unlikely that one major gene is responsible for DISH/CC and instead it appears to be a polygenic disease.A hiperostose idiopática difusa do esqueleto (DISH) é uma doença musculoesquelética comum caracterizada pela formação óssea de novo em ligamentos e enteses. A DISH pode coexistir com a condrocalcinose e por isso tem sido sugerido que ambas partilham o mesmo mecanismo patogénico. COL6A1 e FGF2 são os dois genes de suscetibilidade conhecidos com uma ligação genética fraca à DISH. O objetivo principal desta tese foi investigar a genética da DISH/CC, utilizando a sequenciação de nova geração e estudos de associação e expressão, num grupo de amostras de doentes com DISH/CC do AZORBIO. Duas variantes na região reguladora do gene RSPO4 são significativamente mais frequentes nos controlos do que nos doentes DISH/CC. Estas variantes podem afetar a expressão do gene, conferindo proteção à doença. Utilizando a sequenciação exómica identificamos uma associação significativa entre a DISH/CC e a variante genética rs17563 no gene BMP4, um gene diretamente envolvido na formação óssea endocondral. Outro gene candidato estudado foi o ABCC6, que parece ser relevante em doenças caracterizadas por calcificações ectópicas. Embora inconclusivos, os estudos de expressão em tecidos de cartilagem humana mostraram que o gene ABCC6 apresenta expressão superior nos doentes DISH e CC em relação a um doente controlo, levantando a hipótese de que este aumento de expressão poderá estar envolvido com a deposição de cristais de pirofosfato de cálcio nestes doentes. Uma abordagem comparativa utilizando teleósteos revelou que o gene abcc6 está expresso na pele dos peixes, mas não está associado com a calcificação ectópica das escamas. Além disso, nos resultados da genómica comparativa o gene abcc6 só foi encontrado no genoma de vertebrados ósseos, indicando que este gene poderá estar envolvido em inovações específicas dos vertebrados. Concluindo, foi identificado pela primeira vez potenciais variantes genéticas que protegem (RSPO4) ou predispõem (BMP4) à DISH/CC. A relevância do gene ABCC6 neste fenótipo necessita de ser provada. É pouco provável que um único gene esteja envolvido no aparecimento de DISH/CC, e por isso a doença parece ser poligénica.O trabalho foi co-financiado pelo Fundo Regional da Ciência e Tecnologia dos Açores através do projeto M3.1.2/F/023/201

FORMAÇÃO TERRITORIAL DO TRIÂNGULO MINEIRO (MG): a cidade de Uberlândia como centro comercial

Este trabalho visa compreender a atual configuração do Triângulo Mineiro (MG) por meio da análise do processo de ocupação e dinamismo comercial da cidade de Uberlândia. A área passou por significativos processos de transformações ligados a ocupação humana, até se constituir como importante centro comercial brasileiro. Verifica-se a importância de algumas cidades, como é o caso de Uberlândia, como um nó da rede urbana para o Triângulo Mineiro pautada nas relações comerciais destacando-se o setor atacadista. A área sempre foi destaque como passagem para importantes centros, primeiramente ligando Goiás ao litoral, e a partir da década de 1960, se destacava como conexão entre Brasília ao Rio de Janeiro e São Paulo. A cidade de Uberlândia se destacou na região triangulina, já que concentrou as redes rodoviárias e ferroviárias no município ligando aos mais importantes centros do Brasil. Com essa posição estratégica, a cidade se desenvolveu economicamente aproveitando sua localização para o setor atacadista. Novas relações se estabelecem, já que Uberlândia se destacou como centro de distribuição para as cidades da região, e todo país. FORMATION TERRITORIAL TRIÂNGULO MINEIRO (MG): The Uberlândia city as comercial center AbstractThis work aims to understand the current configuration of Triangulo Mineiro (MG) through an analysis of the process of occupation and the commercial dynamism of its citie of Uberlândia. The area has undergone significant processes of transformation linked to human settlement, to be constituted as important Brazilian shopping center. There is the importance of some cities, such as Uberlandia, as a node of the urban network for the Mineiro Triangle guided in trade relations highlighting the sector atacadista. The area was always highlighted as a gateway to important centers, first linking Goiás to the coast, and from the 1960s, it stood out as the connection between Brasília to Rio de Janeiro and São Paulo. The city of Uberlândia stood out in triangulina region as focused road and railway networks linking the city to the most important centers of Brazil. With this strategic position, the city developed economically taking advantage of its location to the wholesale sector. New relationships are established, since Uberlandia stood out as a distribution center for the region's cities, and the whole country

Could disulfiram be a new treatment for crack cocaine dependence?: a pilot study

Universidade Federal de São Paulo (UNIFESP)Universidade Federal do TocantinsSecretaria Municipal de Saúde de Palmas Centro de Atenção Psicossocial para Álcool e DrogasUNIFESPSciEL

Whole-genome-wide linkage and IBS/IBD studies

Twelve families with exuberant and early-onset calcium pyrophosphate dehydrate chondrocalcinosis (CC) and diffuse idiopathic skeletal hyperostosis (DISH), hereafter designated DISH/CC, were identified in Terceira Island, the Azores, Portugal. Ninety-two (92) individuals from these families were selected for whole-genome-wide linkage analysis. An identity-by-descent (IBD) analysis was performed in 10 individuals from 5 of the investigated pedigrees. The chromosome area with the maximal logarithm of the odds score (1.32; P=0.007) was not identified using the IBD/identity-by-state (IBS) analysis; therefore, it was not investigated further. From the IBD/IBS analysis, two candidate genes, LEMD3 and RSPO4, were identified and sequenced. Nine genetic variants were identified in the RSPO4 gene; one regulatory variant (rs146447064) was significantly more frequent in control individuals than in DISH/CC patients (P=0.03). Four variants were identified in LEMD3, and the rs201930700 variant was further investigated using segregation analysis. None of the genetic variants in RSPO4 or LEMD3 segregated within the studied families. Therefore, although a major genetic effect was shown to determine DISH/CC occurrence within these families, the specific genetic variants involved were not identified.publishersversionpublishe

Emissions simulation by coupling chemical equilibrium and reduced kinetics for gasoline/ethanol mixture in IC engines / Simulação de emissões acoplando equilíbrio químico e cinética reduzida para mistura de gasolina / etanol em motores CI

The analysis of the combustion engine is one on which fails the thermodynamic equilibrium hypotheses since the variables and the system constants are fast changing with time therefore you need a special treatment for this situation. The chemical equilibrium thermodynamics indicates start and end of the process but doesn't reveal the time that the phenomenon occurs. The chemical kinetics has information on the time. The methodology of chemical equilibrium calculations was based on mass balances and equilibrium relations generating a nonlinear system with twenty-two incognita and twenty-two algebraic equations. To solve the nonlinear system, it was chosen an iterative method by Newton-Raphson associated with a new methodology for the determination of the initial estimative of the system avoiding the non-convergence of the system. It was possible to calculate the concentrations of twenty-one chemical species generated in the combustion of various fuels. It was made a comparison between the results from other softwares, finding compatible results for concentrations of emissions generated. After the validation of the chemical equilibrium routine, it was developed the chemical kinetics routine with six chemical equations for the prediction of species concentrations (NO and CO). A simulation of an IC engine was developed using the coupling of the methodology of chemical equilibrium and kinetics. In the engine simulation model developed, as the crank angle increases, it was done the integration of species chemical compositions with coupling chemical equilibrium and kinetics by derivatives addition.  The resulting non-linear system with twenty-one differential equations was integrated by a fourth order Runge-Kutta method.  Made some experiments on a Flex-Fuel engine with mixtures gasoline/ethanol: E25 (25% anhydrous ethanol), H30 (30% hydrous ethanol), H50 (50% hydrous ethanol), H80 (80% hydrous ethanol) and H100 (100% hydrous ethanol). The results obtained in the computer simulation of the engine were compatible with these experimental data

The V410L knockdown resistance mutation occurs in island and continental populations of Aedes aegypti in West and Central Africa

The extensive use of insecticides for vector control has led to the development of insecticide resistance in Aedes aegypti populations on a global scale, which has significantly compromised control actions. Insecticide resistance, and its underlying mechanisms, has been investigated in several countries, mostly in South American and Asian countries. In Africa, however, studies reporting insecticide resistance are rare and data on resistance mechanisms, notably knockdown resistance (kdr) mutations, is scarce. In this study, the recently described V410L kdr mutation is reported for the first time in old world Ae. aegypti populations, namely from Angola and Madeira island. Two additional kdr mutations, V1016I and F1534C, are also reported for the first time in populations from Angola and Cape Verde. Significant associations with the resistance phenotype were found for both V410L and V1016I individually as well as for tri-locus genotypes in the Angolan population. However, no association was found in Madeira island, probably due to the presence of a complex pattern of multiple insecticide resistance mechanisms in the local Ae. aegypti population. These results suggest that populations carrying the same kdr mutations may respond differently to the same insecticide, stressing the need for complementary studies when assessing the impact of kdr resistance mechanisms in the outcome of insecticide-based control strategies.publishersversionpublishe

Urolithiasis Bioresource

Urolithiasis is frequent and raises significant health care burden in a working-age population. Its prevalence, in the Azores archipelago, is currently unknown but it is thought to be higher than the overall estimated prevalence. The Azores Biobank (AZORBIO) Urolithiasis samples have been collected in accordance with standard operation procedures (SOPs) to ensure high quality. Each donor provided 30 ml whole blood and 9 ml of urine. If possible, we preferred blood collected from fasting individuals and first morning urine samples. Aliquots of plasma, serum, DNA, RNA and urine, are stored at -80°C freezers. This collection of samples, and data, will be used to investigate the genetic and/or environmental risk factors associated with the disease in this geographical area.</p

Cough peak flow in preschoolers: success rate and test-retest reproducibility

La evaluación de la función pulmonar en niños en edad preescolar es importante pues hay pocos estudios que investigan los test de función pulmonar en este grupo. Este estudio tuvo el propósito de evaluar la tasa de éxito y la reproducibilidad test-retest del pico de flujo de tos (PFT) en una muestra con niños entre 4 y 6 años de edad. Se estudió el PFT en 44 participantes saludables (26 chicos y 18 chicas) elegidos a través de un cuestionario ATS-DLD-78-C para detectar la presencia de enfermedades respiratorias de base y exposición ambiental. Se utilizó el medidor del pico de flujo espiratorio (Piko-I Electronic Peak Flow Meter, Pulmonary Data Services, USA) para medir el PFT. Se definió la tasa de éxito como el porcentaje de niños capaces de realizar el test según los criterios de aceptabilidad y de reproducibilidad. Para evaluar la reproducibilidad test-retest tras tres semanas se reevaluaron 10 niños según el cálculo de la muestra. Se analizó la reproducibilidad a través del test t pareado, considerando significativoÉ importante avaliar a função pulmonar em pré-escolares. Poucos estudos relacionados aos testes de função pulmonar nessa população estão disponíveis. O objetivo deste estudo foi avaliar a taxa de sucesso e reprodutibilidade teste-reteste do pico de fluxo da tosse (PFT) em uma amostra de crianças com idade entre 4 e 6 anos. O PFT foi estudado em 44 crianças saudáveis (26 meninos e 18 meninas), selecionadas de acordo com o questionário ATS-DLD-78-C, utilizado para detectar a presença de doenças respiratórias de base e exposição ambiental. O medidor de pico de fluxo expiratório (Piko-I Electronic Peak Flow Meter, Pulmonary Data Services, USA) foi usado para mensurar o PFT. A taxa de sucesso foi definida como a porcentagem de crianças capazes de realizar o teste de acordo com os critérios de aceitabilidade e reprodutibilidade. Para avaliar a reprodutibilidade teste-reteste, 10 crianças (de acordo com o cálculo amostral) foram reavaliadas após três semanas. A reprodutibilidade teste-reteste foi avaliada pelo teste t pareado, considerando significativo pIt is important to evaluate lung function in preschoolers. There are few studies related to pulmonary function tests with this part of the population. The purpose of this study was to evaluate the success rate and test-retest reproducibility of the cough peak flow (CPF) in a sample with children between four and six years old. The CPF was tested in 44 healthy children (26 boys, 18 girls), selected according to the ATS-DLD-78-C questionnaire, used to detect the presence of common respiratory diseases or induced by environmental exposure. An expiratory peak flow meter (Piko-I Electronic Peak Flow Meter, Pulmonary Data Services, USA) was used to measure CPF. The success rate was defined as the percentage of children able to perform the test according to the acceptability and reproducibility criteria. To evaluate the test-retest reproducibility, 10 children (according with the sample calculation) were reevaluated after three weeks. The study was approved by the Ethics Research Committee of the Institution. The test-retest reproducibility was evaluated by the paired t-test, considering a significance of