Thesis advisor: Daniel A. Kirschner / The myelin sheath of the central nervous system contains a network of interlamellar tight junctions known as the radial component. Ablation of claudin-11, a tight junction protein, results in the absence of the radial component and compromises the passive electrical properties of the myelin sheath. Although tight junctions are known to regulate paracellular diffusion, this barrier function has not been directly demonstrated for the radial component, and some evidence suggests that the radial component may also, or instead, mediate adhesion between myelin membranes. To investigate the physical properties of claudin-11 tight junctions, we first compared fresh, unfixed Claudin 11-null and control nerves using X-ray diffraction. In Claudin 11-null tissue, we detected no changes in myelin structure, stability, or membrane interactions, which argues against the notion that myelin tight junctions exhibit significant adhesive properties. To examine myelin permeability in the absence of the radial component, we measured the kinetics of osmotic compaction and recovery in knockout and control myelin. We found that myelin lacking claudin-11 responded more rapidly to osmotic stress, indicating an increase in permeability to water and small osmolytes. To further test this hypothesis, we explored the possibility of measuring the diffusion of water through myelin using neutron diffraction, a technique that had been pioneered in myelin decades ago but was largely unused because of previous limitations in neutron technology. After establishing that present-day neutron instruments were capable of measuring diffusion in myelin, we applied this technique to samples from mice lacking claudin-11. Consistent with our X-ray diffraction studies, we found that H2O-D2O exchange was more rapid in Claudin 11-null mice compared to controls. Thus, our data indicate that the radial component serves primarily as a diffusion barrier and elucidate the mechanism by which tight junctions govern myelin function. / Thesis (PhD) — Boston College, 2016. / Submitted to: Boston College. Graduate School of Arts and Sciences. / Discipline: Biology.

Os tendões são tecidos de transição das forças contráteis geradas pelos músculos e as transferindo para os ossos, podendo assim gerar movimento. A região em que o tecido tendíneo se insere ao ósseo é chamada de junção osteotendínea ou entese. Estas junções podem ser classificadas como fibrosas ou fibrocartilaginosas, as fibrosas são aquelas em que os tendões e/ou ligamentos se inserem no eixo (diáfise) dos ossos longos e as fibrocartilaginosas a inserção ocorre nas epífises dos ossos longos ou nos ossos curtos. Este estudo tem como propósito analisar as fibras colágenas e as células presentes na junção osteotendínea, utilizando microscopia de luz com as colorações de Hematoxilina-eosina, Azocarmim, Picro-sírius e Tricromo de Masson e os aspectos ultraestruturais empregando os métodos de microscopia eletrônica de varredura e de transmissão. Foram utilizados 30 ratos Wistar machos, sendo 15 adultos com idade de 4 meses (grupo A) e 15 idosos com idade de 18 meses (grupo B), sendo utilizados 5 ratos de cada grupo para microscopia de luz, 5 para microscopia eletrônica de varredura e 5 para microscopia eletrônica de transmissão. Os membros posteriores dos ratos foram retirados, dissecados e desmineralizados com EDTA a 7% para microscopia de luz e EDTA 0,15M pH 7,4 para microscopia eletrônica de transmissão durante 4 semanas, não foi necessário desmineralizar as amostras para microscopia eletrônica de varredura. Cortes de 6 m foram realizados e montados em lâminas histológicas para análise em microscópio de luz, a técnica de criofratura em nitrogênio líquido foi realizada para análises em microscópio eletrônico de varredura e cortes de 60 nm foram realizados, montados em telas de cobre de 200 mesh (EMS) para análises em microscópio eletrônico de transmissão. O processo de envelhecimento mostrou alterações das fibrilas colágenas, em que o tipo I predomina no grupo adulto e o tipo III no idoso, diminuição na quantidade de células de fibrocartilagem, processos citoplasmáticos destas células curtos e em número reduzido e uma diminuída capacidade de síntese devido a aparelhos de Golgi menores, poucas cisternas de retículo endoplasmático granular e escassas mitocôndrias. / Tendons are transition tissues of contractile forces generated by muscles and transferring to bone, generating this way move. The region where tendon attach to bone is called bone-tendon junction or enthesis. These junctions can be classified as fibrous or fibrocartilaginous, the fibrous are those in which tendons/ligaments are attached in the shaft (diaphysis) of the long bones and fibrocartilaginous attachment occur in the epiphyses of the long bones or short bones. This study aims to analyze collagen fibers and the cells present in the bone-tendon junction, using light microscope with Hematoxylineosin, Azocarmine, Picrosirius red and Masson\'s trichrome staining and ultrastructural aspects using scanning electron microscope, transmission electron microscope methods. Thirty male Wistar rats were using, 15 adults rats at 4 months-old (A group) and 15 old rats at 18 months-old (B group), were used 5 rats of each group to light microscopy, 5 rats to scanning electron microscopy and 5 rats to transmission electron microscopy. The hind limbs of the rats were removed, dissected and demineralized using 7% EDTA solution to light microscopy and 0,15M pH 7,4 EDTA solution to transmission electron microscopy for 4 weeks, was not necessary demineralize the samples to scanning electron microscopy. Cut in slices 6 m thickness were made and mounted on histological slides to analyze in light microscope, freeze-fracture technique with liquid nitrogen was performed to analysis in scanning electron microscope and cuts in slices 60 nm thickness were made, mounted on copper grids (200-mesh - EMS) to analysis in transmission electron microscope. The aging process showed changes of the collagen fibrils, in which type I predominates on adult groups and the type III on the old group, decrease in the amount of the fibrocartilage cells, few and short cytoplasmic processes of this cells and a decreased synthesis capacity due a small Golgi apparatus, a few cisternae of rough endoplasmatic reticulum and few mitochondria.

O nome síndrome da cauda eqüina define as manifestações clínicas oriundas da disfunção sensorial e/ou motora causada pela lesão das raízes nervosas que formam a porção terminal da medula espinhal. É uma condição neurológica resultante da compressão, destruição ou deslocamento dos trajetos nervosos que constituem a cauda eqüina. Acomete freqüentemente os cães de raças de grande porte, como Labrador, Golden Retriever, Bernese Mountain dog, Rottweiler e Pastor Alemão, não possuindo predileção quanto ao sexo e idade. As causas são inúmeras, podendo estar relacionada a fatores adquiridos, mais comuns nas raças de grande porte; ou congênitos, os quais são mais freqüentes nas raças de pequeno porte. Em ambos os casos, a estenose lombo-sacra é a causa mais comum, correlacionando-se às alterações das partes moles e/ou tecidos ósseos no segmento lombo-sacro. Este estudo teve por objetivo realizar uma análise crítica da contribuição dos métodos de imagem, quais sejam exames: radiográfico simples e tomográfico, para avaliação do segmento lombo-sacro em 30 cães da raça Pastor Alemão. Sendo treze animais pertencentes ao grupo (A) sem manifestações clínicas e alterações radiográficas no segmento lombo-sacra; doze animais pertencentes ao grupo (B) sem manifestações clínicas com alterações radiográficas no segmento lombo-sacro e cinco animais pertencentes ao grupo (C) com manifestações clínicas e portadores de alterações radiográficas no segmento lombo-sacro. Todos os exames foram submetidos a um protocolo de avaliação tendo como principais mensurações radiográficas: ângulo lombo-sacro, ângulo formado entre as epífises de L7-S1, ângulo dos processos articulares de L7-S1, listese ventral de S1 em relação à L7, grau de intersecção do vértice dos ângulos mensurados e grau de movimento na região lombo-sacra. O deslocamento ventral de S1 em relação à L7 esteve presente na maioria dos animais nos três diferentes grupos não só na dorso-extensão, levantando a hipótese de no padrão racial estudado, certo grau de listese possa fazer parte do esperado para o segmento lombo-sacro. Foi constatado diferença estatística entre os animais, nos seguintes grupos e variáveis: ângulo formado entre as epífises de L7-S1 (grupo A e B), grau de intersecção do ângulo entre as epífises de L7-S1 (grupo A e B), ângulo lombo-sacro (grupo A e B) e grau de movimento no segmento lombo-sacro (grupo A e B, A e C). Desta maneira, o grau de movimento demonstrou-se como elemento relevante no critério de avaliação do segmento lombo-sacro. O exame tomográfico mostrou-se superior na avaliação do canal vertebral, foramens intervertebrais e processos articulares, os quais puderam ser avaliados com maior riqueza de detalhes. Concluiu-se com esta pesquisa que as duas modalidades de imagem se complementam, constituindo ferramentas importantes na avaliação clínico-cirúrgica do segmento lombo-sacro, auxiliando no diagnóstico, estabelecimento do prognóstico e da terapêutica a ser adotada. / The name cauda equina syndrome defines the clinical signs that come from the sensory and or motor neural dysfunction caused by the terminal part of the spinal cord and adjacent nerve roots damages. Its one neurological condition resulting from the compression, destruction and displacement of the nervous routes which form the cauda equina. This syndrome normally happens to large breed dogs like the Labrador retriever, Golden Retriever, Bernese mountain dog, Rottweiler and the German shepherd dogs, without preference of gender or age. There are countless causes, which can be related to acquired factors, more common in the large breed dogs; or congenital, which are more frequent in the small breed dogs. In both cases, the stenosis lumbosacral is the most frequent, correlating to the alterations of the soft tissues and/ or the bone tissues in the lumbosacral segment. The aim of this sudy was to analise critically the real contribution of diagnostic imaging (X-Ray and CT), of the lumbosacral region of 30 German shepherd dogs. There were thirteen animals that belonged to the group (A) without clinical signs and X-Ray alterations in the lumbosacral segment; twelve animals belonged to the group (B) without clinical signs, with X-Ray alterations in the lumbosacral segment; five animals belonged to the group (C) with clinical signs and they had X-Ray alterations in the lumbosacral segment. All exams were submitted to one evaluation report which had as main radiographic measurements: lumbosacral angle, endplate angle L7 S1, articular process angle L7 S1 , ventral displacement of S1 to L7, the point of intersection of the measured angles and the mobility of the lumbosacral junction. The ventral displacement of S1 related to L7 was present in most of the animals in the three different groups not only in the extension position, raising the hypothesis about the racial standard established, a certain degree of ventral displacement of the sacrum might be part of the expected for the lumbosacral segment. It was noticed a statistical difference among the animals in the following groups and variables: endplate angle L7 S1 (groups A and B), the point of intersection of the endplate angle L7 S1 (groups A and B), lumbosacral angle (groups A and B) and the mobility of the lumbosacral junction (groups A and B, A and C). This way, the mobility showed itself as an relevant element in the criterion of evaluation of the lumbosacral segment. The CT examination showed being superior in the evaluation of the vertebral canal, intervertebral foramen and the articular processes, which could be evaluated with a greater number of details. It was concluded with this research that the two modalities of images complement each other, becoming important tools in the clinical- surgical evaluation in the lumbosacral segment, helping in the diagnosis, prognostic and therapeutic to be adopted.

A doença granulomatosa inflamatória envolve interações coordenadas entre linfócitos, monócitos/macrófagos, células epitelióides, eosinófilos, neutrófilos e fibroblastos. A comunicação intercelular mediada por junções gap, constituídas por conexinas, é responsável pela homeostase tecidual. A Cx43 está presente em células linfóides, células mielóides, fibroblastos e outras. Assim, para compreendermos o possível envolvimento das conexinas no granuloma, nós analisamos o efeito da deleção heterozigótica de uma Gja1 (gene Cx43) na formação e desenvolvimento dos granulomas hepáticos, induzidos por ovos de Schistosoma mansoni. Para tanto, camundongos heterozigotos (Cx43+/-) e selvagens (Cx43+/+) foram infectados com cercárias de Schistosoma mansoni e avaliados nos tempos: 6, 8 e 12 semanas após a infecção. As células dos granulomas apresentaram expressão de conexina 43, notadamente após 12 semanas de infecção. As lesões dos camundongos Cx43+/- apresentaram índice de proliferação reduzido e aumento na deposição de colágeno em fases tardias da doença. Apesar desses achados, não se encontrou redução no tamanho e celularidade das lesões em comparação aos camundongos selvagens. Também não obtivemos diferenças em relação ao hemograma ou população de linfócitos esplênicos CD4, CD8 e CD19. As células peritoneais dos animais de ambos genótipos apresentaram produção de NO e H<SUB2O<SUB2 similares. Contudo, os neutrófilos e monócitos sanguíneos dos animais Cx43+/-, estimulados por PMA, apresentaram aumento significante do burst oxidativo. Concluindo, nossos resultados indicam que a deleção de um alelo do gene da Cx43 modifica claramente a evolução da doença granulomatosa, mostrando um papel desta conexina no desenvolvimento do granuloma. / Inflammatory granulomatous disease involves coordinated interactions among lymphocytes, monocytes/macrophages, epithelioid cells, eosinophils, neutrophils and fibroblasts. Intercellular communication mediated by gap junctions constituted of connexins, is responsible for tissue homeostasis. Cx43 is present in lymphoid cells, myelogenous cells, fibroblasts and others. In order to understand the possible involvement of connexins in granuloma, we analyzed the effect of the heterologous deletion of a Gja1 (Cx43 gene) on the formation and development of hepatic granulomas induced by Schistosoma mansoni eggs. Heterozigous (Cx43+/-) and wild-type (Cx43+/+) mice were infected with S. mansoni cercarie and evaluated after 6, 8 and 12 weeks. Granuloma cells express Cx43, with considerable reduction in Cx43+/- mice. Moreover, granuloma cells from Cx43v+/- mice displayed reduced proliferation and increased collagen deposition at late phases of the disease. Despite these findings, no reduction of size or cellularity of the lesions was found in comparison to wild-type mice. We didn?t find differences in relation to blood count and splenic lymphocyte population CD4, CD8 and CD19. Peritoneal cells from animals of both genotypes presented similar production of NO and H2O2. However, blood neutrophils and monocytes from Cx43+/- mice, stimulated by PMA, displayed significantly increased oxidative burst. In conclusion, our results indicate that the deletion of one allele of the Cx43 gene clearly modifies the evolution of a granulomatous disease, supporting a role for this connexin on granuloma development.

Les matériaux III-Sb ont prouvé leur potentiel pour la réalisation de composants opto-électroniques dans des domaines aussi variés que les télécommunications ou l'environnement. Cependant, ils restent une filière quasi-inexplorée pour les systèmes photovoltaïques classiques. Dans ce projet de recherche, nous voulons démontrer que les composants à base d'antimoniures sont des candidats prometteurs pour des cellules solaires à haute efficacité et bas coût. Leurs avantages sont multiples : non seulement offrent-ils un large panel d'alliages accordés en maille et des jonctions tunnel à basse résistivité, mais ils permettent aussi une croissance directe sur substrat de Si. Nous étudions donc les briques élémentaires d'une cellule solaire multi-jonction intégrée sur Si. Tout d'abord, nous développons la croissance et fabrication de cellules homo-épitaxiales en GaSb. Les caractéristiques tension-intensité (J-V) mesurées sont proches de l'état de l'art avec une efficacité sous un soleil de 5.9 %. Puis, nous intégrons une cellule à simple jonction GaSb sur un substrat de Si par épitaxie par jet moléculaire (EJM). Les analyses de diffraction X (DRX) et de microscopie à force atomique (AFM) montrent des propriétés de structure et morphologie proches de celles reportées pour des buffers métamorphiques similaires dans la littérature. Nous adaptons alors la configuration de la cellule pour éviter la haute densité de défauts à l'interface GaSb/Si. La cellule hétéro-épitaxiale a une efficacité réduite de 0.6 %. Ce résultat est néanmoins proche des dernières avancées sur les cellules GaSb sur GaAs, et ce, malgré un désaccord de maille plus important. Enfin, nous étudions l'épitaxie d'AlInAsSb. Cet alliage pourrait en théorie atteindre une grande gamme d'énergies de bande interdite tout en restant accordé sur GaSb. Néanmoins, il souffre d'une lacune de miscibilité importante, le rendant sujet à la ségrégation de phase. Il n'y a que peu de mentions de l'AlInAsSb dans la littérature, et toutes rapportent des conditions de croissance instables et des énergies de bande interdite plus basses qu'attendues. Nous réussissons à produire des couches de bonne qualité d'AlInAsSb dont la composition en Al varie de 0.25 à 0.75 et ne présentant aucun signe macroscopique de décomposition de phase. Toutefois, l'observation au microscope à transmission électronique (TEM) révèle des fluctuations de composition nanométriques. Les données de photoluminescence (PL) sont étudiées pour déterminer les propriétés électroniques de l'alliage. Les mesures d'efficacité quantique (QE) montrent que la sous-cellule du haut limite la performance de la cellule tandem. Des modélisations numériques des courbes J-V et QE sont utilisées pour identifier des pistes d'amélioration pour chaque brique élémentaire. / III-Sb materials have demonstrated their potential for multiple opto-electronic devices, with applications stretching from communications to environment. However, they remain an almost unexplored segment for classical photovoltaic systems. In this research, we intend to demonstrate that III-Sb-based devices are promising candidates for high-efficiency, low-cost solar cells. Their benefits are two-fold: not only do they offer a wide range of lattice-matched alloys and low-resistivity tunnel junctions, but they also enable direct growth on Si substrates. We thus investigate the building blocks of a GaSb-based multi-junction solar cell integrated onto Si. First, we develop the photovoltaic growth and processing by fabricating homo-epitaxial GaSb cells. Intensity-voltage (J-V) measurements approach the state of the art with 1-sun efficiency of 5.9%. Then, we integrate a GaSb single-junction cell on a Si substrate by molecular beam epitaxy (MBE). X-ray diffraction (XRD) and atomic force microscopy (AFM) analysis show structural and morphological properties close to the best reported in the literature for similar metamorphic buffers. We further adapt the cell configuration to circumvent the high defect density at the GaSb/Si interface. The heteroepitaxial cell results in a reduced efficiency of 0.6%. Nevertheless, this performance is close the most recent advancements on GaSb heteroepitaxial cells on GaAs, despite a much larger mismatch. Last, we investigate the epitaxy of AlInAsSb. This alloy could in theory reach the widest range of bandgap energies while being lattice-matched to GaSb. However, it presents a large miscibility gap, making it vulnerable to phase segregation. AlInAsSb only counts few experimental reports in the literature, all referring to unoptimized growth conditions and abnormally low bandgap energies. We successfully grow good-quality layers with Al composition x_{Al} ranging from 0.25 to 0.75, showing no macroscopic sign of decomposition. Yet, transmission electron microscopy (TEM) observations point to nanometric fluctuations of the quaternary composition. Photoluminescence (PL) data is studied to determine the alloy's electronic properties. We eventually propose and fabricate a tandem cell structure, resulting in 5.2% efficiency. Quantum Efficiency (QE) measurements reveal that the top subcell is limiting the tandem performance. Numerical fits to both J-V and QE data indicate improvement paths for each building block.

O papel das junções intercelulares comunicantes do tipo gap e das proteínas que as compõem, as conexinas, tem sido alvo de numerosos estudos no campo da oncologia. Com a finalidade de compreender melhor os mecanismos moleculares envolvidos no processo carcinogênico e desenvolver novas armas contra o câncer, estes estudos têm se mostrado promissores, porém com muitas perguntas ainda a serem respondidas. Com o objetivo de avaliar a interferência da conexina 43 no processo carcinogênico, administramos o carcinógeno DMBA, um hidrocarboneto aromático policíclico, nas doses hebdomadárias de 1 mg durante 9 semanas, à camundongos BALB/c geneticamente modificados heterozigotos para a conexina 43 (Cx43+/-) e wild-type (Cx43+/+). O desenvolvimento de neoplasias ocorreu em 100% dos animais que receberam DMBA, porém de forma variável quanto ao tempo, tipo e número de neoplasias. No total, 6 tipos neoplásicos foram observados, incluindo neoplasias mamária, linfoma, pulmonar, gástrica, cutânea e ovariana, nesta ordem de prevalência. Com relação às neoplasias mamárias, as mamas abdominais foram as mais acometidas e o adenoacantoma foi o tipo histológico mais comum. No pulmão, estômago e pele, o tipo neoplásico mais comum em cada um foi, respectivamente, adenocarcinoma alveolar papilar, carcinoma de células escamosas e carcinoma de células escamosas queratinizante. Foi observada diferença estatística significante na incidência de tumores ovarianos, entre os grupos Cx43+/- e Cx43+/-, com maior incidência em animais Cx43+/-, indicando interferência da Cx43 neste processo carcinogênico. Apenas os animais Cx43+/- desenvolveram este tipo neoplásico, o qual foi representado exclusivamente por tumores da célula da granulosa. Não houve diferença estatística significante na incidência dos demais tumores, embora, em números absolutos, a incidência de quase todas, à exceção das neoplasias cutâneas, tenha sido maior nos Cx43+/-. O mesmo se repetiu com relação ao desenvolvimento de metástases, cujo fenótipo foi observado apenas em neoplasias mamárias e gástricas. A utilização de doses elevadas (9mg) de DMBA parece interferir na resposta, mais notadamente a pulmonar. As conexinas atuam de forma complexa e variável entre os diferentes tumores e entendimento da relação das conexinas com o câncer depende do entendimento molecular do controle da expressão das conexinas. Com este trabalho esperamos contribuir para evolução dos estudos relativos ao seu papel no processo carcinogênico e, desta forma, auxiliar no desenvolvimento de meios de previnir e combater o câncer. / The role of the intercellular communication of gap junctions and of the proteins that form these junctions, the connexins, has been the subject of numerous studies in the field of oncology. In order to better understand the molecular mechanisms involved in the carcinogenic process and develop new weapons against cancer, these studies have shown promising, but with many questions still to be answered. Aiming to evaluate the interference connexin 43 in the carcinogenic process the carcinogen DMBA, one aromatic hydrocarbon polycyclic, was administered to genetically modified BALB;c mice heterozygous for the connexin 43 (Cx43 +/-) and wild-type (Cx43 +/+). The development of cancer occurred in 100% of animals receiving DMBA, but in different timing, types and number of tumors. In total, six types of neoplasm were observed, including breast, lymphoma, lung, gastric, skin and ovarian cancers, in that order of prevalence. Regarding breast cancer, abdominal breasts were the most affected and adenoacanthoma was the most common histological type. In the lung, stomach and skin, the most common tumor type in each was, respectively, papillary alveolar adenocarcinoma, squamous cell carcinoma and keratinizing squamous cell carcinoma. There was statistically significant difference in the incidence of ovarian tumors among groups Cx43 + / - and Cx43 + / -, indicating interference of the expression of Cx43 +/- in the carcinogenic process. Only animals of the Cx43 + / - developed this tumor type, which was represented exclusively by granulosa cell tumors. There was no statistically significant difference in the incidence of other tumors, although in absolute numbers, the incidence of almost all, except for skin cancers, was higher in Cx43 + / -. The same was repeated with respect to the development of metastases, wereas observed only in breast and gastric cancers. The use of high doses (9 mg) of DMBA appears to interfere with the carcinogenic response, most notably in the lung. Connexins act in complex and variable ways among different tumors and understanding of the relationship of connexins in cancer depends on understanding the molecular control of expression of connexins. With this work we hope to contribute to the development of studies about role of connexins in the carcinogenic process and thus help in developing ways to prevent and fight cancer.

Infecções por Plasmodium spp. podem acarretar em complicações pulmonares (1 a 40% dos casos), que podem resultar no desenvolvimento da síndrome do desconforto respiratório agudo (SDRA). Esta síndrome é caracterizada por inflamação aguda, lesão do endotélio alveolar e do parênquima pulmonar, disfunção e aumento da permeabilidade da barreira alvéolo-capilar pulmonar e, consequente, formação de efusão pleural. Neste sentido, os mecanismos de regulação da permeabilidade das células endoteliais e as junções interendoteliais têm papel crítico na manutenção do endotélio pulmonar. O objetivo do estudo foi determinar precocemente o desenvolvimento da SDRA associada à malária por tomografia computadorizada por emissão de fóton único (SPECT/CT), além de identificar alterações nas junções interendoteliais das células endoteliais pulmonares primárias de camundongos DBA/2 (CEPP-DBA/2), após contato com os eritrócitos parasitados de Plasmodium berghei ANKA (EP-PbA). Os nossos resultados demonstraram que é possível identificar alterações na aeração pulmonar no 5° e 7° dia após a infecção e, consequentemente, diferenciar os animais que desenvolveriam SDRA daqueles que evoluiriam para hiperparasitemia (HP). Além disso, observamos em CEPP-DBA/2 que o contato direto com EP-PbA aumenta da abertura das junções interendoteliais e da permeabilidade vascular. Assim, avaliamos a diminuição da expressão das proteínas das junções interendoteliais que contribuem para o aumento da permeabilidade vascular, por imunofluorescência e Western Blot. Apesar da SDRA ter sido identificada há mais de 50 anos, ainda não se conhece formas de diagnóstico precoce e os mecanimos efetivos de desenvolvimento desta enfermidade, que permitam um tratamento efetivo e que evite a morte do paciente. Portanto, sugere-se que a técnica de SPECT/CT seja uma importante ferramenta de diagnóstico para identificação precoce de SDRA associado a malária. / Infections by Plasmodium spp. can lead to pulmonary complications (1 to 40% of the cases), that can result in the development of acute respiratory distress syndrome (ARDS). This syndrome is characterized by the acute inflammation, injury of the alveolar endothelium and pulmonary parenchyma, dysfunction and increased permeability of the pulmonary alveolar-capillary barrier and, consequently, formation of pleural effusion. In this aspect, mechanisms of regulation of endothelial cell permeability and interendothelial junctions play a critical role in the maintenance of the pulmonary endothelium. The present study aimed to determine the early development of single-photon emission computed tomography (SPECT/CT) associated malaria ARDS in addition to identifying changes in the interendothelial junctions of the primary pulmonary endothelial cells of DBA/2 mice (CEPP-DBA/2) after contact with erythrocytes infected with Plasmodium berghei ANKA (EP-PbA). Our results demonstrated that is possible to identify changes in lung aeration on the 5th and 7th day after infection and, consequently, differentiate the animals that should develop ARDS from those that would evolve to hyperparasitemia (HP). In addition, we observed in CEPP-DBA/2 that direct contact with EP-PbA increases the opening of the interendothelial junctions and vascular permeability. Thus, we evaluated that decrease the expression of interendothelial junction proteins contribute to the increase of vascular permeability, by immunofluorescence and Western Blot. Although ARDS was identified more than 50 years ago, it is not yet known what forms of early diagnosis, and the effective mechanisms of development of this disease, that allow an effective treatment and that prevent the death of the patient. Therefore, it is suggested that the SPECT/CT technique is a valuable tool to promote the early identification of ARDS associated with malaria.

Introdução: o estresse psicológico aumenta a permeabilidade intestinal em roedores e humanos, potencialmente levando a inflamação de baixo grau e aos sintomas em distúrbios gastrintestinais funcionais. No entanto, o efeito do estresse agudo combinado ao estresse da vida crônica, que mimetiza potencialmente melhor a situação humana, é desconhecido. Além disso, há poucos dados disponíveis sobre os efeitos do estresse em intestino delgado versus cólon. Métodos: ratos Wistar foram alocados em quatro protocolos de estresse: 1/ controles; 2/ estresse agudo (isolamento e movimentos limitados); 3/ Crowding stress:crônico e 4/ estresse agudo + estresse crônico. Amostras de jejuno e cólon foram colhidas para estudar a permeabilidade em câmaras deUssing, a expressão gênica de moléculas de junção firmes e a densidade de mastócitos. Níveis de corticosterona no plasma foram medidos. Principais resultados:corticosterona plasmática foi avaliada nas três condições de estresse, teve níveis mais altos na condição de estresse combinado. Permeabilidade do jejuno foi aumentada em todas as condições de estresse e correlacionada com os níveis de corticosterona. O aumento da expressão das claudinas 1, 5 e 8, daocludina e da ZO-1 foi detectado no estado de estresse agudo no jejuno. Em contraste, a permeabilidade do cólon foi aumentada no protocolo de estresse combinado, e a expressão de moléculas das junção firmes permaneceu inalterada. O aumento da densidade de mastócitos foi observado no cólon nos ratos submetidos aos estresses crônico e combinado. Conclusão e inferências:os estresses agudo, crônico e combinado influenciam diferentemente a permeabilidade intestinal, a expressão de moléculas de junção firmes e a atividade dos mastócitos, no jejuno e no cólon. Estes resultados fornecem uma visão mais aprofundada dos mecanismos de hiperpermeabilidade intestinal relacionadas ao estresse. / Background: Psychological stress increases intestinal permeability in rodents and humans, potentially leading to low-grade inflammation and symptoms in functional gastrointestinal disorders through disturbances in brain-gut axis. However, the effect of acute stress on the background of Crhonic life stress, potentially better approaching the human situation, is unknown. Moreover, only limited information is available on the effects in small intestine versus colon in animal model. Methods: Wistar rats were allocated to 4 stress protocols: 1/ sham; 2/ acute stress (isolation and limited movement); 3/ Crhonic crowding stress and 4/ acute + Crhonic stress (n = 8 per group). Jejunum and colon were harvested to study permeability in Ussing chambers, gene expression of tight junction molecules and mast cell density. Plasma corticosterone levels were measured. Key Results: Plasma corticosterone was elevated in all three stress conditions, with the highest levels in the combined stress condition. Permeability of the jejunum was increased in all stress conditions and correlated with corticosterone levels. Increased expression of claudin 1, 5 and 8, occludin and ZO-1 was detected in the acute stress condition in the jejunum. In contrast, colonic permeability was increased in the acute on Crhonic stress protocol only and the expression of tight junction molecules was unaltered. Increased mast cell density was observed in the Crhonic and acute on Crhonic stress condition in the colon only. Conclusion and Inferences: Acute, Crhonic and combined stress differentially affect intestinal permeability, expression of tight junction molecules and mast cells in the jejunum and the colon. These findings provide further insight in the mechanisms of stress-related intestinal hyperpermeability and barrier.

L'endothelium vasculaire constitue la principale barrière entre le sang et les tissus régulant le passage de macromolécules et de cellules circulantes. Longtemps considéré comme une monocouche passive, l'endothélium joue d'importants rôles dans la régulation de la pression sanguine, de l'hémostase, des réponses immunitaires et inflammatoires. L'adhérence cellule/cellule est initiée dans l'endothélium vasculaire par des interactions homophiliques entre molécules de VE-cadhérine (= jonctions adhérentes). La dynamique de la jonction et du cytosquelette est importante pour le remodelage des jonctions intercellulaires qui a lieu au cours l'angiogenèse, de la vasculogenèse et lors de la réparation de l'endothélium. C'est pourquoi la détermination des mécanismes moléculaires sous-jacents est indispensable à la comprehension de phénomènes physiopathologiques (angiogenèse et progression tumorales, inflammation...). D'après la littérature, la protéine EPLIN intervient dans la formation du complexe E-cadhérine/alpha-caténine/EPLIN et stabilise l'actine corticale. Actuellement décrite comme spécifique des modèles épithéliaux, EPLIN peut-elle intervenir dans la liaison du complexe à base de VE-cadhérine au cytosquelette d'actine? De plus, il paraît essentiel de comprendre le rôle de cette protéine dans les cellules car son expression est fortement diminuée dans la plupart des cancers alors qu'à l'inverse sa surexpression bloque la prolifération cellulaire. / The endothelium forms the main barrier regulating the passage of macromolecules and circulating cells between the blood and tissue. Historically viewed as a passive vascular lining, vascular endothelium plays important roles in the regulation of vascular pressure, hemostasis, immune and inflammatory responses. In vascular endothelium, cell/cell adhesion is mediated by homophilic interactions of VE-cadherin molecules (= adherens junctions). Cytoskeleton and junction dynamics are important for intercellular junctions remodelling that occurs during angiogenesis, vasculogenesis and endothelium repair. So, determining the underlying molecular mechanisms is essential for the comprehension of pathologic phenomena such as angiogenesis, tumor progression or inflammation. We learn from the literature that EPLIN is involved in E-cadherin/α-catenin/EPLIN complex formation and cortical actin stabilization. Usually described as a protein specific of epithelial models, we wondered if EPLIN is able to link VE-cadherin complex to actin cytoskeleton. Furthermore, it seems essential to understand its cellular role since it is downregulated in many cancers while in contrast its overexpression blocks cell proliferation.

L'optique non linéaire sur silicium a pris son essor en raison des nombreuses perspectives d'applications à l'optoélectronique en circuit intégré. Pour observer des effets non-linéaires sans travailler à des puissances trop élevées, il faut utiliser des résonateurs à très haut facteur de confinement optiques (Q/V). Les microcavités à cristal photonique bidimensionnel sont une technologie mature et planaire pour réaliser de tels résonateurs sur silicium. Au cours de cette thèse, nous avons travaillé sur une application des microcavités à cristal photonique à la détection télécom. Le silicium est en effet transparent dans cette plage de longueurs d'onde, sauf si on atteint des densités de puissance élevées, auquel cas l'absorption à deux photons intervient. Le principe du détecteur repose sur l'exaltation de absorption à deux photons grâce à la microcavité en cristal photonique. La collection des porteurs ainsi générés est assurée par une jonction latérale métal-semiconducteur-métal (MSM). Nous avons d'abord étudié numériquement la viabilité du concept du détecteur sous deux aspects : collection des porteurs libres à travers le cristal photonique et influence des métallisations sur le facteur de qualité. Les modèles standards pour le courant d'obscurité et le photocourant dans les photodétecteurs MSM ont été étendus pour tenir compte du cristal photonique. La fabrication d'une jonction MSM dans le cristal photonique a fait l’objet d’un travail approfondi en salle blanche de l’IEF. La mesure du courant circulant dans le dispositif a permis de mettre en évidence un photocourant résonnant. On retiendra que la réponse peut alors atteindre 90 mA/W et que la bande-passante est supérieure au GHz. Outre la démonstration du détecteur en elle-même, des résultats originaux ont été obtenus. Nous avons montré qu'il est possible de contrôler les densités de porteurs dans les microcavités à cristal photonique en jouant sur la polarisation externe. Enfin, le détecteur est un moyen de mesurer certaines grandeurs essentielles de la physique des microcavités sur silicium, comme l’absorption linéaire résiduelle ou la résistance thermique de la cavité. / Silicon non linear optics is of considerable interest to the scientific community because of its applications to integrated optoelectronics. In order to observe non linear phenomena at a reasonable power, one has to work with very high confinement factor (Q/V) optical resonators. As they are now a fully mastered planar technology, two-dimensional photonic crystal microcavities are an efficient way of actually obtaining such resonators in silicon. In this thesis, an application of photonic crystal microcavities to telecom wavelength detection is demonstrated. Bulk silicon is transparent at these wavelengths, except when working at very high power density. Only then, two-photon absorption (TPA) becomes significant. In our detector, TPA is enhanced in the microcavity. A metal-semiconductor-metal (MSM) junction then ensures very fast carrier collection. We studied the physics of the detector and focused on two aspects: collection of carrier in a photonic crystal, and impact of the metal on the optical quality factor. Standard models for dark current and photocurrent in MSM junctions were adapted to the photonic crystal case study. Fabrication of the photonic crystal junction was carefully undertaken and optimized in the clean room of the IEF laboratory. The current circulating in the fabricated device resonates as the same wavelength as the optical cavity. Response can be as high as 90 mA/W, and the optical-electrical bandwidth is larger than 1 GHz. In addition to the demonstration of the detector, some original results were obtained. It is possible to control the carrier concentration in photonic crystal microcavities by tuning the external polarisation of the MSM junction. Finally, the detector allows one to measure important parameters of the physics of the cavity, such as the residual linear absorption coefficient, and the thermal resistance.