Yes / Alterations of Endothelial cells (ECs) play a critical role in different pathogenesis of many serious human diseases, and dysfunction of the vascular endothelium is an indicator for human disorders. Endothelial dysfunction is considered to be an early indicator for atherosclerosis, which is characterised by overexpression of adhesion molecules, including intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1). Hydrogen peroxide (H2O2) released via neutrophils is an important mediator of endothelial cell function. Ambient production of superoxide anion (O2−) and subsequently H2O2 at low levels is critical for regulating endothelial cell functions and proliferation. In this study, we investigated the effects of H2O2 on the expression of adhesion molecules VCAM-1 and ICAM-1 in cultured human umbilical vein endothelial cells (HUVECs). Intracellular superoxide anion production was detected by using p-Nitro Blue Tetrazolium (NBT) assay. Our results showed that administration of 100μM of H2O2 on HUVECs for 2, 6, 12 and 24 h induced a time-dependent increase in ICAM-1 and VCAM-1 mRNA and protein expression levels with a significant increase observed from 6 h. HUVECs exposed to H2O2 exhibit increased O2−, suggesting that H2O2 induced oxidative stress may be a reasonable for atherosclerosis. This increase can be reduced by the flavonoid, N-acetyl cysteine (NAC). The modulation of endothelial cell function through this mechanism may underlie the contribution of H2O2 to the development of vascular disease.

No description available.

The discoveries, implementations and developments of NMR and MRI have had a major impact in medical imaging. Compared to other imaging modalities (PET, SPECT, CT), current MRI research helps to further and better understand the inner mechanisms of the human body in a less invasive manner. In clinical neuroimaging, perfusion MRI is of spectacular importance to study cerebrovascular diseases and cancer. However, at the moment, there is no perfusion MRI sequence that allows for a complete, non-invasive and precise quantification of microvascular flow dynamics. This work focuses on the use of the recently introduced Flow Enhanced Signal Intensity method (FENSI) to characterize and quantify vasculature at capillary level, at high and ultra high magnetic field (7 and 17.2 tesla). For that purpose, the possible quantification of blood flux with FENSI is explored in vivo. The combination of flux quantification and flow-enhanced signal (compared to Arterial Spin Labeling) can make of FENSI an ideal method to characterize in a complete non-invasive way the brain microvasculature. After removal of magnetization transfer (MT) effects, the blood flow dynamics are studied with FENSI in a very aggressive and propagative rat brain tumor model: the 9L gliosarcoma. The objective is to assess whether FENSI is suitable for a longitudinal non-invasive characterization of microvascular changes associated with tumor growth. The results obtained with FENSI are compared with literature on 9L perfusion and immuno-histochemistry. In the first paper published on FENSI, a first glance was also casted on the potential of the flow enhanced technique when applied to fMRI. The results obtained at the time were contaminated by MT effects. With the implementation of a new MT-free FENSI technique, the possibility to map the brain cerebral functioning based on a quantitative physiological parameter (CBFlux) more directly related to neuronal activity than the usual BOLD signal is within reach. At ultra high field, the influence of different anesthetics on the rat brain microvascular network and BOLD contrast is also considered. After many developments around the FENSI technique, the method is compared to classical ASL and DSC perfusion MRI sequences. The strengths and weaknesses of the FENSI method, its characteristics, 'precautions for use', and potential main applications are detailed and discussed.

L'élastographie ultrasonore est une technique d'imagerie émergente destinée à cartographier les paramètres mécaniques des tissus biologiques, permettant ainsi d’obtenir des informations diagnostiques additionnelles pertinentes. La méthode peut ainsi être perçue comme une extension quantitative et objective de l'examen palpatoire. Diverses techniques élastographiques ont ainsi été proposées pour l'étude d'organes tels que le foie, le sein et la prostate et. L'ensemble des méthodes proposées ont en commun une succession de trois étapes bien définies: l'excitation mécanique (statique ou dynamique) de l'organe, la mesure des déplacements induits (réponse au stimulus), puis enfin, l'étape dite d'inversion, qui permet la quantification des paramètres mécaniques, via un modèle théorique préétabli. Parallèlement à la diversification des champs d'applications accessibles à l'élastographie, de nombreux efforts sont faits afin d'améliorer la précision ainsi que la robustesse des méthodes dites d'inversion. Cette thèse regroupe un ensemble de travaux théoriques et expérimentaux destinés à la validation de nouvelles méthodes d'inversion dédiées à l'étude de milieux mécaniquement inhomogènes. Ainsi, dans le contexte du diagnostic du cancer du sein, une tumeur peut être perçue comme une hétérogénéité mécanique confinée, ou inclusion, affectant la propagation d'ondes de cisaillement (stimulus dynamique). Le premier objectif de cette thèse consiste à formuler un modèle théorique capable de prédire l'interaction des ondes de cisaillement induites avec une tumeur, dont la géométrie est modélisée par une ellipse. Après validation du modèle proposé, un problème inverse est formulé permettant la quantification des paramètres viscoélastiques de l'inclusion elliptique. Dans la continuité de cet objectif, l'approche a été étendue au cas d'une hétérogénéité mécanique tridimensionnelle et sphérique avec, comme objectifs additionnels, l'applicabilité aux mesures ultrasonores par force de radiation, mais aussi à l'estimation du comportement rhéologique de l'inclusion (i.e., la variation des paramètres mécaniques avec la fréquence d'excitation). Enfin, dans le cadre de l'étude des propriétés mécaniques du sang lors de la coagulation, une approche spécifique découlant de précédents travaux réalisés au sein de notre laboratoire est proposée. Celle-ci consiste à estimer la viscoélasticité du caillot sanguin via le phénomène de résonance mécanique, ici induit par force de radiation ultrasonore. La méthode, dénommée ARFIRE (''Acoustic Radiation Force Induced Resonance Elastography'') est appliquée à l'étude de la coagulation de sang humain complet chez des sujets sains et sa reproductibilité est évaluée. / Ultrasound elastography is an emerging technology derived from the concept of manual palpation and dedicated to the mapping of biological tissue mechanical properties in a diagnostic context. Various elastographic approaches have been applied to the study of organs such as the liver, breast or prostate. All proposed techniques rely on a three-steps procedure: first, the tissue to be studied is mechanically excited, in a static or dynamic way. Induced displacements are then measured and used to estimate qualitatively or quantitatively mechanical properties of the medium. This step is called inversion. While application fields of elastography are constantly broadened, efforts are made to provide robust and accurate inversion algorithms. In this monography, theoretical and experimental works related to the development of new inversion methods dedicated to the study of mechanically inhomogeneous media in dynamic ultrasound elastography are provided. In the context of breast cancer diagnosis, a localized tumour can be assumed as a confined mechanical heterogeneity, also referred as an inclusion, which can disturb the propagation of shear waves (dynamic excitation). The first objective of this thesis is to provide a theoretical model to describe physical interactions occurring between incident shear waves and a tumour, here geometrically assumed as an ellipse. Once the theoretical model is validated, an inverse problem is formulated allowing further quantification of inclusion viscoelastic parameters. Aiming the development of realistic models, the previous work has been extended to the case of three dimensional spherical heterogeneities and adapted to the specific case of an acoustic radiation force excitation. Furthermore, the feasibility of assessing the medium rheological model (i.e., the frequency dependence of mechanical properties) is demonstrated. Finally, in the context of vascular diseases and blood coagulation, an inversion method based on the study of the mechanical resonance phenomenon induced by acoustic radiation force is proposed. The technique, termed ARFIRE (Acoustic Radiation Force Induced Resonance Elastography), is applied to human whole blood samples and the reproducibility of results is assessed.

Le Granite Passa Três est situé à l'Est de l'Etat du Paraná, au Sud du Brésil, et est allongé selon une direction NNE-SSW. Sa mise en place se fait au cœur des metapélites mesoprotérozoïques du Groupe Açungui (Province Mantiqueira). La minéralisation d’or du Granite Passa Três est composée par des veines de quartz contenant des quantités variables de fluorite, sulfures et carbonates. Les objectifs principaux de ce travail de thèse sont : de comprendre le modèle de formation du système de veines minéralisées en prenant en compte les relations entre magmatisme, hydrothermalisme, déformation et minéralogie à la fois dans l’espace et dans le temps ; la caractérisation de la nature, de la source et des conditions de dépôt des fluides ; et la caractérisation du modèle métallogénique de ce gisement singulier. Pour arriver à ces objectifs, les méthodes utilisées seront, en sus de la géologie structurale et de terrain : la pétrographie, la géochronologie U-Pb (LA-ICP-MS) sur zircon et 40Ar-39Ar sur muscovite, la microscopie électronique à balayage (MEB), la microsonde électronique, la fluorescence X (XFR), l’analyse isotopique du soufre (δ34S) et l’analyse microthermométrique et RAMAN des inclusions fluides. Les données structurales ont montré la coexistence de deux systèmes principaux de filons minéralisés, l’un N-S et l’autre E-W, avec des pendages de 60-75°W et 45-70°S, respectivement. Les deux systèmes sont interprétés comme contemporains et conjugués. Les corps minéralisés forment des géométries sigmoïdales qui résultent de l’ouverture en pull-aparts résultant de mouvements en faille normale le long de plans de glissement à faible pendage. Le fort pendage des structures minéralisées s’explique par l’enveloppe globale formée par la succession des pull-aparts. Quatre étapes minéralogiques sont à l’origine de la formation du système minéralisé : phase 1 [qtz 1 + fl], phase 2a [qtz 2 + py 2a ± or ± cpy ± aik ± fl ± sph ± musc], phase 2b [qtz 2 + py 2b + or + cpy + aik + ank ± sph ± fl ± musc] et phase 3 [qtz 3 + ank + calc + molyb ± aik ± musc ± fl]. L’or se trouve dans la forme d’or invisible et d’or natif dans des fractures qui affectent les pyrites des phases 2a et b, systématiquement associé avec la chalcopyrite et l’aikinite. L’altération associée à la minéralisation inclue des assemblages composés par muscovite/quartz/pyrite (altération du type greisen) et séricite/carbonate/clinochlore (altération phyllique). Les valeurs δ34S des pyrites (de -0.1‰ à 1.1‰) indiquant que le soufre du dépôt peut être d’origine magmatique. Cette hypothèse est en accord avec l’observation systématique, dans les parties supérieures du granite (sondage et niveaux supérieurs de la mine), de structures caractéristiques de transition magmatique-hydrothermale comme des systèmes aplo-pegmatitiques, des veines de quartz à bordure de K-feldspath, des concentrations de quartz de type stockscheider et des textures de solidification unilatérales (UST). Les résultats de géochronologie confirment cette hypothèse avec des âges U-Pb sur zircon (611.9±4.7 et 611.9±5.6 Ma pour le granite à grain moyen (GEM) et le microgranite (GEF) et 40Ar-39Ar sur muscovite (veines à bordure de K-feldspath : 612.9±2 à 608.8±2 Ma ; veines minéralisées : 611.7±2 à 608.8±2 Ma ; veines de quartz précoces : 608.4±2 Ma) très proches. Ces âges obtenus indiquent que la mise-en-place du granite, l’exsolution du fluide magmatique-hydrothermal et la formation des veines de quartz aurifères ont été réalisées pendant un écart de temps de 5 Ma, entre 613 et 608 Ma. La minéralisation (611 à 608 Ma) contemporaine de la cristallisation du granite (612 à 610 Ma), l’association de l’or avec des minéraux de bismuth (aikinite), la démonstration du contrôle structural sur la formation des veines et les évidences de transition magmatique-hydrothermale en domaine de coupole granitique montrent que le dépôt d’or du Granite Passa Três partage plusieurs similitudes avec les dépôts du type intrusion-related. / The Passa Três Granite is situated in southern Brazil (Paraná State) and presents a NNE-SSW elongated shape. This intrusion is emplaced within metapelites of the Mesoproterozoic Açungui Group (Ribeira Belt, Mantiqueira Province), between the N40E trending Morro Agudo and Lancinha faults. Gold mineralisation is composed of centimetric to metric quartz veins with fluorite, sulphides and carbonates. The main objectives of this work are i) to understand the model of formation of the mineralised veins systems taking into account the relationships between magmatism, hydrothermalism, deformation and mineralogy in space and time; ii) the characterization of the nature, source and emplacement conditions of the ore fluids; and iii) the characterization of a metallogenic model for this singular deposit. In order to reach these purposes, the methods to be applied include, beyond the structural geology and field works: petrography, U-Pb zircon (LA-ICP-MS) and 40Ar-39Ar muscovite geochronology, scanning electron microscopy (SEM), electron-microprobe analyses (EPMA), X-ray fluorescence (XRF), isotopic analysis of sulphur (δ34S), and microthermometric and Raman analysis of fluid inclusions. Structural data showed the coexistence of two major normal mineralised vein systems, one N-S and the other one E-W, with dips of 60-75ºW and 45-70ºS, respectively. Both systems are interpreted to be contemporaneous and conjugated. Orebodies form sigmoidal geometries that resulted of the opening of pull-aparts as a consequence of the normal movements along low-angle fault planes. High-angle dip of the global mineralised structures is explained by the succession of the pull-aparts. Four mineralogical stages resulted in the formation of the mineralised system: phase 1 [quartz 1 + fluorite], phase 2a [quartz 2 + pyrite 2a ± gold ± chalcopyrite ± aikinite ± fluorite ± sphalerite ± muscovite], phase 2b [quartz 2 + pyrite 2b + gold + chalcopyrite + aikinite + ankerite ± sphalerite ± fluorite ± muscovite] and phase 3 [quartz 3 + ankerite + calcite + molybdenite ± aikinite ± muscovite ± fluorite]. Gold occurs as invisible gold and as native grains within fractures that affect pyrite 2a and 2b, commonly associated with chalcopyrite and aikinite. Alteration related to the mineralisation includes muscovite/quartz/pyrite (greisen type alteration) and sericite/carbonato/clinochlore (phyllic alteration) assemblages. The δ34S values of pyrite crystals (from -0.1‰ to 1.1‰) indicate that the sulphur in this deposit may have a magmatic origin. This hypothesis agrees with the systematic observation, within the upper part of the granite (drill holes and superior levels of the mine), of structures typical of magmatic-hydrothermal transition such as aplite-pegmatite systems, quartz veins with K-feldspar border, quartz concentration of stockscheider type and unilateral solidification textures (UST). Geochronological data confirm this hypothesis with U-Pb zircon ages (611.9±4.7 and 611.9±5.6 Ma for medium grained granite facies (GEM) and microgranite (GEF), respectively) and 40Ar-39Ar muscovite dating (veins with K-feldspar border: 612.9±2 to 608.8±2 Ma; mineralised veins: 611.7±2 to 608.8±2 Ma; barren vein: 608.4±2 Ma), that are very close. These ages indicate that the granite emplacement, the magmatic-hydrothermal fluid release and the formation of gold-bearing quartz veins occur during a time lapse of approximately 5 Ma, between 613 and 608 Ma. The mineralisation (611 to 608 Ma) coeval to granite crystallization (612 to 610 Ma), the association of gold with Bi minerals (aikinite), the strong structural control for veins and magmatic-hydrothermal transition features at the roof of a small granitic intrusion suggest that the Passa Três gold deposit shares several similarities with intrusion-related gold deposits.

INTRODUÇÃO: A orbitopatia de Graves é uma doença autoimune cujas manifestações clínicas ocorrem por infiltração tecidual inflamatória. Apresenta estágio ativo inicial e estágio sequelar posteriormente, podendo se manifestar nas formas clínicas miogênica ou lipogênica. A inflamação da órbita pode gerar importante congestão, agravando manifestações clínicas e comprometendo o sucesso da terapia específica em alguns pacientes. O objetivo deste trabalho é a avaliação da congestão orbitária feita pelo estudo das características do fluxo sanguíneo na veia oftálmica superior, por meio do Doppler colorido de órbitas, nos grupos de pacientes com orbitopatia de graves em fase ativa, em fase sequelar miogênica e em fase sequelar lipogênica, além do grupo controle. MÉTODOS: Estudo transversal prospectivo realizado entre maio de 2006 e abril de 2008 no Serviço de Órbita do HCFMUSP. Noventa órbitas de 46 pacientes com orbitopatia de Graves foram incluídas, assim como 38 órbitas de 20 pacientes do grupo controle. Foi feito exame oftalmológico completo e avaliação pelo DCO em todos os pacientes. Pacientes com orbitopatia de Graves foram divididos em grupos com doença em atividade e doença em estágio sequelar nas apresentações miogênicas ou lipogênicas. O sentido, as velocidades máxima e mínima do fluxo sanguíneo na veia oftálmica superior, e o intervalo entre as mesmas, foram aferidos. Os achados foram comparados entre os diferentes grupos e analisados também em relação à restrição muscular, exoftalmometria, escore de atividade clínica, idade, sexo, lado estudado, comorbidades e tabagismo. RESULTADOS: Foi encontrada diferença no sentido do fluxo sanguíneo da veia oftálmica superior (p<0,001), na velocidade máxima (p=0,001), na velocidade mínima (p=0,002) e no intervalo entre a velocidade máxima e a mínima do mesmo (p<0,001) entre os grupos de estudo. A velocidade máxima do fluxo sanguíneo apresentou correlação com a restrição muscular (p=0,003), o escore de atividade clínica (p<0,001) e a exoftalmometria (p=0,015), enquanto a velocidade mínima correlacionou-se com restrição muscular (p<0,001) e escore de atividade clínica (p=0,006). CONCLUSÕES: Há diferença mensurável na velocidade e no sentido do fluxo sanguíneo da veia oftálmica superior entre pacientes dos diferentes grupos de estudo / INTRODUCTION: Graves orbitopathy is an autoimmune disease whose clinical manifestations are secondary to inflammatory tissue infiltration. It presents an initial active stage and a later latent stage, and can manifest myogenic or lipogenic clinical forms. Inflammation of the orbit can generate significant congestion, worsening clinical manifestations and limiting the success of specific therapy in some patients. The objective of this study is to evaluate orbital congestion studying the characteristics of blood flow in the superior ophthalmic vein with the color Doppler in groups of patients with Graves Orbitopathy in active stage, lipogenic latent stage and myogenic latent stage, and patients from the control group. METHODS: Transversal study made between May 2006 and April 2008 in the Orbit Department of University of São Paulo Medical School. Ninety orbits from 46 patients with Graves orbitopathy and 38 orbits from 20 patients of the control group were included. All patients underwent complete ophthalmological exams including color Doppler of the orbits. Patients with Graves orbitopathy were divided in active stage, latent lipogenic stage and latent myogenic stage groups. The direction of venous blood flow in the superior ophthalmic vein and its maximum and minimum velocities - including the interval between them - were measured. The results were compared among the groups and analyzed also in relation to muscular restriction, exophthalmometry, clinical activity score, age, sex, studied orbit side, comorbidities and use of tobacco. RESULTS: There was difference in blood flow direction of the superior ophthalmic vein (p<0,001), blood flow maximum speed (p=0,001), blood flow minimum speed (p=0,002) and blood flow speed interval (p<0,001) among the groups of this study. Blood flow maximum speed presented correlation with muscle restriction (p=0,003), clinical activity score (p<0,001) and exophthalmometry (p=0,015). Blood flow minimum speed presented correlation with muscle restriction (p<0,001) and clinical activity score (p=0,006). CONCLUSIONS: There are measurable difference in speed and direction of blood flow in the superior ophthalmic vein among patients of the different groups of this study

In vitro, la formation de structures de type tubulaire avec des cellules endothéliales de veine ombilicale humaine (HUVEC) a été étudiée en combinant la fonctionnalisation de la chimie des matériaux et le développement de la géométrie tridimensionnelle. Le polycarbonate (PC) a été utilisé comme modèle pour le développement de l'échafaud. Le film de polysaccharide naturel, basé sur un dépôt alternatif couche par couche (LbL) d’acide hyaluronique (HA) et de chitosane (CHI), a d’abord été appliqué sur une surface PC et caractérisé en termes de croissance d’épaisseur microscopie à balayage lascar (CLSM). Cette première fonctionnalisation se traduit par un revêtement complet de la couche PC. Une biofonctionnalisation supplémentaire avec un peptide adhésif (RGD) et deux peptides angiogénétiques (SVV et QK) a été étudiée, immobilisant ces peptides sur le groupe carboxylique de HA précédemment déposé, en utilisant la chimie bien connue du carbodiimide. La version marquée de chaque peptide a été utilisée pour caractériser l’immobilisation et la pénétration des peptides dans les couches de polyélectrolytes, aboutissant à une greffe réussie avec une pénétration complète dans toute l’épaisseur du LbL. Des tests in vitro ont été effectués à l'aide de cellules HUVEC pour évaluer leur efficacité d'adhésion et leur activité métabolique sur la LbL avec et sans immobilisation de peptides, ce qui a permis d'améliorer l'activité préliminaire lorsque des combinaisons de peptides sont utilisées. Enfin, les micro-canaux PC (μCh) ont été développés et caractérisés pour la première fois, et les autres expériences ont été réalisées sur un micromètre de 25 μm de largeur, fonctionnalisé avec une architecture (HA / CHI) 12,5 (PC-LbL) avec des peptides RGD et QK -RGD + QK) ou avec des peptides RGD et SVV (PC-RGD + SVV). Notre première expérience de tubulogénèse a montré de manière surprenante la formation de structures de type tubulaire déjà après 2h d'incubation en utilisant la combinaison double-peptides, mais uniquement avec PC-RGD + QK. Les tubes étaient également présents après 3 et 4 heures de culture. L'expérience de co-culture avec des péricytes humains dérivés du placenta (hPC-PL) montre comment la stabilisation des tubes a été améliorée après 3 et 4 heures également pour l'échantillon de PC-RGD + SVV. Globalement, notre matériel bio-fonctionnel avec les peptides PC-RGD + QK et PC-RGD + SVV permet la formation d'une structure de type tubulaire à la fois dans une expérience de monoculture et de co-culture. / In vitro, tubular-like structures formation with human umbilical vein endothelial cells (HUVECs) was investigated by combining material chemistry functionalization and three-dimensional geometry development. Polycarbonate (PC) was used as a template for the development of the scaffold. Natural polysaccharide’s film based on alternate layer-by-layer (LbL) deposition of hyaluronic acid (HA) and chitosan (CHI), was first applied to PC surface and characterized in terms of thickness growth both, in dry conditions using ellipsometry, and confocal lascar scanning microscopy (CLSM). This first functionalization results in a complete coating of the PC layer. Further biofunctionalization with one adhesive peptide (RGD) and two angiogenetic peptides (SVV and QK) was investigated, immobilizing those peptides on the carboxylic group of HA previously deposited, using the well-known carbodiimide chemistry. The labeled version of each peptide was used to characterize the peptides’ immobilization and penetration into the polyelectrolytes layers, resulting in a successful grafting with complete penetration through the entire thickness of the LbL. In vitro tests were performed using HUVECs to assess their adhesion efficiency and their metabolic activity on the LbL with and without peptide immobilization, resulting in a preliminary improved activity when peptide-combinations is used. Finally, PC micro-channels (μCh) were first developed and characterized, and the rest of the experiments were performed on μCh of 25μm width, functionalized with (HA/CHI)12.5 architecture (PC-LbL) with RGD and QK peptides (PC-RGD+QK) or with RGD and SVV peptides (PC-RGD+SVV). Our first tubulogenesis experiment surprisingly showed the formation of tubular-like structures already after 2h of incubation using the double-peptides combination but only using PC-RGD+QK the tubes were present also after 3 and 4 hours of culture. The co-culture experiment with human pericytes derived from placenta (hPC-PL) demonstrates how the stabilization of the tubes was improved after 3 and 4 hours also for the PC-RGD+SVV sample. Globally our bio-functional material with PC-RGD+QK and PC-RGD+SVV peptides allow the formation of tubular-like structure in both mono and co-culture experiment.

O nervo alveolar inferior (Nai), o mais espesso ramo do nervo mandibular (V par craniano), está funcionalmente relacionado à inervação da mandíbula e dentes inferiores, além de parte dos tecidos moles circunjacentes. Seu estudo morfológico, sobretudo em mandíbulas adultas, foi sistematicamente realizado, contudo, sem ainda haver um consenso definitivo para o seu padrão de ramificação. Desse modo, considerando-se a importância em se demonstrar um modelo de constituição básica para o Nai, realizou-se o presente estudo em fetos humanos da 19ª a 36ª semana de vida intrauterina. Foram utilizados 86 hemimandíbulas, de ambos os sexos, em que se avaliou o padrão morfológico intraósseo do Nai e de seus ramos, seus envoltórios conjuntivos e suas relações com as estruturas vasculares, utilizando-se de microdissecção e microscopia eletrônica de varredura (MEV). Relativamente ao padrão morfológico do segmento intramandibular do Nai, foram propostos 4 tipos de ramificação: Tipo I (16%), onde o Nai emite 3 ramos principais, os nervos alveolares inferiores posterior (Naip), médio (Naim) e anterior (Naia); Tipo II (34%), semelhante ao anterior, porém ausente o Naim; Tipo III (30%), semelhante ao primeiro, porém ausentes Naim e o Naia; e o Tipo IV (20%), a forma clássica do Nai, onde somente o Naia se ramifica a partir dele. Verificou-se uma série de conexões entre todos os feixes nervosos, sobretudo entre o Naip e o Nai, sendo que os ramos dentais e peridentais partem, em sua maioria, do Naip, Naim e Naia. Sobre os envoltórios conjuntivos, verificou-se que se espessam à medida que o feto se desenvolve, notando também a presença de uma bainha neurovascular comum a envolver o feixe vasculonervoso alveolar inferior. Quanto ao posicionamento da artéria alveolar inferior, adjacente ao forame da mandíbula, cruza o feixe nervoso, em sua maioria, pela face medial (41%) ou pela lateral (55%), para logo em seguida alcançar sua posição definitiva sobre a face superior do feixe nervoso. Quanto às veias alveolares inferiores, em número de 1 a 3, posicionam-se posteromedialmente às outras estruturas do feixe vasculonervoso. Para a veia de Serres, observada em todos os espécimes e envolta por um canal ósseo, propôs-se a terminologia veia paramandibular. / The inferior alveolar nerve (Nai), the thickest branch of the mandibular nerve (V pair of cranial nerve), is functionally related to jaw and lower teeth innervation as well as, being part of the surrounding smooth tissue. The Nai morphology has been systematically studied in mandible of adults however, no definitive consensus has been reached in relation to its branched pattern. Therefore, this study was been performed in 19 to 36 week-gestation fetuses, due to the importance of determining a basic model of Nai as available data refers only to its arrangement in adults. Micro dissection and Scanner Electron Microscopy (MEV) were performed on 86 human hemi-mandibles of both sexes in order to evaluate the morphological intraosseous pattern of Nai and its branches, its connective tissue and their relationship with vascular structures. In connection with the morphological pattern of intramandibular segment of Nai, four types of branches were proposed: Type I (16%) where Nai give rise to 3 main branches: posterior inferior alveolar nerve (Naip), medium (Naim), and anterior (Naia); Type II (34%), similar to Type I but Naim was absent; Type III (30%), similar to the first, however Naim and Naia were absent; and type IV (20%),the classic form of Nai, from which only Naia was raised. A series of connections was verified among all of the nervous bundles, especially between Nai and Naip. besides dental and periodontal branches commonly arising from Naip, Naim and Naia. In relation with the connective tissues, it was shown that they became thicker as the fetuses are developing; at this stage it was also noted one neurovascular sheath covering nerve bundles and blood vessels. Relatively to the inferior alveolar artery position, adjacent to mandible foramen, it can, frequently, cross the nerve bundle through medial (41%) or lateral surface (55%), reached after its final position on the superior surface of nerve bundle; the inferior alveolar veins (1 to 3), are posterior-medially positioned to the neurovascular bundle other structures. The terminology paramandibular vein was proposed to the Serres vein observed in all specimens studied which was surrounded by bone channel.

La transplantation d’hépatocytes est un procédé séduisant pour remplacer les cellules déficientes dans un foie anatomiquement normal. Dans les maladies métaboliques héréditaires hépatiques (MMHH), la thérapie cellulaire présente un potentiel espoir thérapeutique. Le remplacement d'un pourcentage restreint (5-10%) d’hépatocytes déficients par des hépatocytes normaux pourrait rétablir durablement la fonction métabolique. Les résultats des essais cliniques de transplantation d'hépatocytes génétiquement modifiés ou non sont moins concluants, montrent une prise de greffe insuffisante et, dans la plupart des études, un effet thérapeutique transitoire. L’efficacité limitée de la transplantation d’hépatocytes isolés dans le traitement des MMHH semble en partie liée au faible pourcentage de la masse hépatocytaire reconstituée par les hépatocytes définitivement greffés et fonctionnels. De nombreux modèles animaux ont été développés pour étudier les facteurs pouvant augmenter le nombre et le pourcentage d’hépatocytes transplantés et greffés. Cependant, la majorité de ces modèles ne sont pas transposables en clinique car ils présentent des risques importants ou mal évalués pour les patients. Les principaux objectifs de ce travail ont été d’étudier des moyens peu invasifs pour induire une régénération hépatique et une prise de greffe hépatocytaire significatives dans le but de développer une nouvelle approche de transplantation d’hépatocytes génétiquement modifiés ex vivo pour le traitement de l’hypercholestérolémie familiale. L’effet d’une embolisation portale partielle (EPP) réversible sur la prolifération hépatocytaire et la régénération hépatique a été évalué chez le macaque. A la différence de l’EPP par un produit non résorbable, il ne s’agit pas d’une approche potentiellement délétère à long terme. Une obstruction veineuse plus complète a été provoquée en utilisant le Curaspon®, une gélatine biodégradable, en forme de poudre. Nous avons démontré pour la première fois dans la littérature l’efficacité d’une EPP réversible à induire une importante prolifération hépatocytaire et régénération hépatique. Nos données suggèrent qu’une occlusion portale initiale et temporaire est suffisante pour déclencher les mécanismes responsables d’une régénération hépatique dans le foie non-embolisé. L’utilisation du Curaspon® en poudre peut être considérée comme la forme la plus évoluée d’EPP : très distale, résorbable, qui dure suffisamment pour induire l’hypertrophie hépatique. Cette technique pourrait être indiquée dans des situations cliniques nécessitant une régénération hépatique de courte durée (ex. le traitement des cancers du foie en plusieurs étapes) ou dans des cas qui ne nécessitent pas une résection hépatique, comme la transplantation d’hépatocytes pour le traitement de MMHH. Ces résultats nous ont permis d’évaluer cette approche dans notre protocole préclinique de thérapie génique pour le traitement de l’hypercholestérolémie familiale chez le primate, par autotransplantation d’hépatocytes génétiquement modifiés ex vivo par un vecteur lentiviral. Nous avons démontré que l’EPP réversible induit une régénération hépatique du foie non-embolisé et améliore notablement les résultats de la transplantation d’hépatocytes isolés génétiquement modifiés exprimant la GFP. Seize semaines après la transplantation, les hépatocytes transduits et greffés exprimaient le transgène contrôlé par le promoteur apo-AII humain. Notre protocole a montré pour la première fois chez un gros animal que l’EPP par un produit résorbable entraine avec des conditions de sécurité optimales une repopulation hépatique importante par des hépatocytes transduits par un vecteur lentiviral, et ceci même à distance de la transplantation hépatocytaire. Les résultats encourageants de ces travaux nous ont ouvert la voie pour avancer sur notre projet préclinique et envisager la réalisation d’une étude clinique de phase I/II pour le traitement de l’hypercholestérolémie familiale. / Hepatocyte transplantation is an attractive process for replacing deficient cells in an anatomically normal liver. In metabolic liver diseases, cell therapy could be an interesting alternative to orthotopic liver transplantation. The replacement of a small percentage (5-10%) of deficient hepatocytes by normal hepatocytes could restore the metabolic defect at a long term. Data from clinical studies of hepatocyte autotransplantation or allotransplantation, genetically modified or not, provided poor results, insufficient cell engraftment in the liver parenchyma and, in the majority of cases, a transient therapeutic effect. The limited efficacy of hepatocyte transplantation in metabolic liver diseases is mainly due to the poor percentage of engrafted and finally functional hepatocytes. Numerous animal models have been developed in order to study the factors that could increase the number and the percentage of transplanted and engrafted hepatocytes. However, the majority of these models cannot be used in patients since they present important risks for them. The aim of this work was to evaluate less invasive procedures for inducing liver regeneration and significant hepatocyte engraftment in order to develop a new approach of transplantation of ex vivo genetically modified hepatocytes for the treatment of familial hypercholesterolemia. The effect of reversible portal vein embolization (PVE) on liver regeneration and hepatocyte proleferation was evaluated in monkeys. In contrast to PVE by a permanent embolizing agent, reversible PVE has not a long term deleterious effect on embolized liver. A more complete venous occlusion was obtained by using the powdered form of an absorbable gelatin sponge (Curaspon®). We showed for the first time in the literature the safe and successful use of reversible PVE for inducing significant hepatocyte proliferation and liver regeneration. Our data support that an initial occlusion of the portal branch, even if not permanent, is sufficient to start the mechanisms of liver regeneration in the contralateral lobe. Embolization with Curaspon® powder could be considered to be the ultimate form of embolization: very distal, reversible and lasting sufficiently in order to induce substantial liver hypertrophy. Our findings suggest that this method could reliably be used for clinical purposes, particularly in situations in which short-term regeneration is required (i.e. multi-step management of hepatic malignancies) or in cases where resection of the liver is not finally necessary, such as in hepatocyte transplantation for the treatment of metabolic liver diseases. These promising results on reversible PVE allowed us to evaluate this approach in our preclinical study of gene therapy for the treatment of familial hypercholesterolemia in macaques. Our protocol consisted of an autotransplantation of ex vivo genetically modified hepatocytes by a lentiviral vector. We showed that reversible PVE induces liver regeneration of the non-embolized liver segments and improves considerably hepatocyte transplantation of genetically modified cells expressing Green Fluorescent Protein (GFP). Sixteen weeks after transplantation, transduced engrafted hepatocytes expressed the transgene, which was under control of the human apo-AII promoter. Our protocol showed for the first time in a big animal that PVE by an absorbable agent leads safely to an important and long-term repopulation of the liver by lentivirally transduced hepatocytes. The extremely encouraging results of this work opened our way advancing in our preclinical study and preparing a phase I/II clinical trial for the treatment of familial hypercholesterolemia based on our protocol of autotransplantation of ex vivo genetically modified hepatocytes by a lentiviral vector.

Submitted by Ana Lúcia Torres (bfmhuap@gmail.com) on 2017-09-29T23:29:42Z No. of bitstreams: 2 license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) DISSERTAÇAO RENATO GOMES.pdf: 1085453 bytes, checksum: 52c34bec90105d63a6d951a99405c02d (MD5) / Approved for entry into archive by Ana Lúcia Torres (bfmhuap@gmail.com) on 2017-09-29T23:31:18Z (GMT) No. of bitstreams: 2 license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) DISSERTAÇAO RENATO GOMES.pdf: 1085453 bytes, checksum: 52c34bec90105d63a6d951a99405c02d (MD5) / Made available in DSpace on 2017-09-29T23:31:18Z (GMT). No. of bitstreams: 2 license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) DISSERTAÇAO RENATO GOMES.pdf: 1085453 bytes, checksum: 52c34bec90105d63a6d951a99405c02d (MD5) Previous issue date: 2012 / Santa Casa de Misericórdia de Vitória. Escola Superior de Ciências / Objetivo: Comparar a acuidade visual em pacientes com oclusão de ramo de veia central da retina (ORVCR), antes e após tratamento combinado de injeção intravítrea de bevacizumabe com fotocoagulação a laser. Método: Uma análise retrospectiva de pacientes com ORVCR que foram tratados com injeção intravítrea de bevacizumabe (1,25mg/0,05ml) seguida de fotocoagulação a laser na área afetada. Os pacientes passaram por avaliação oftalmológica completa, incluindo medida da acuidade visual (AV) com tabela de Snellen no momento do diagnóstico e nas visitas de acompanhamento. Foram excluídos prontuários incompletos ou com erros de preenchimentos e prontuários de pacientes com outras alterações oculares que pudessem interferir nas medidas da acuidade visual. Resultados: Foram analisados 26 prontuários (11 homens e 15 mulheres) de pacientes com diagnóstico de ORCVR no Serviço de Oftalmologia da Santa Casa de Vitória durante todo o ano de 2010. Todos os pacientes foram acompanhados por 6 meses. O tempo médio de baixa da acuidade visual antes do diagnóstico foi de 21,2 dias (1-140 dias). Com acuidade visual inicial média de 0.74 LogMAR (p<0.001) e com melhora média após 6 meses do início do tratamento para 0.32 LogMAR (p<0.001), representando assim um ganho médio de 0.42 LogMAR (p<0.001) ou 19.4 letras de ETDSR (p<0.001). Foram utilizados testes não paramétricos a fim de permitir mais generalização aos resultados evitando a premissa de normalidade. O teste de Wilcoxon foi utilizado para verificar a variação da acuidade visual inicial e final. Não foram observadas complicações oculares ou sistêmicas durante o acompanhamento. Conclusão: Houve melhora da acuidade visual comparando pré o pós tratamento combinado de injeção intravítrea de bevacizumabe com fotocoagulação a laser. A estabilidade do ganho obtido no tratamento combinado com a ausência de complicações demonstra o potencial do uso combinado de injeção intravítrea de bevacizumabe com fotocoagulação a laser no tratamento de ORVCR / Purpose: Compare the visual acuity (VA) in patients with branch retinal vein occlusion (BRVO), before and after been treated with a combination of intravitreal injection of bevacizumab followed by laser photocoagulation. Method: A retrospective analyses of patients with BRVO that underwent a treatment of intravitreal injection of bevacizumab (1,25mg/ 0,05ml) followed by laser photocoagulation on the affected area. Patients went through complete eye examination, including visual acuity measure with Snellen chart on the diagnostic visit and on the follow-ups. Uncompleted and miswritten charts ware excluded as well as charts of patients with others ocular affections that could interfere in visual acuity measure Results: 26 charts (11 male and 15 female) of patients with BRVO from the Ophthalmological Department of Santa Casa de Misericórdia Hospital during 2010 were analyzed. All patients underwent a 6 months follow-up. The average VA loss before diagnostics was 21.2 days (1-140). The average VA was 0.74 LogMAR (p<0,001) with improvement after 6 month of treatment to 0.32 LogMAR (p<0,001), representing a average gain of 0.42 LogMAR (p<0,001) or 19.4 ETDRS letters (p<0,001). Non parametric tests ware used to allow more generalization of the results avoiding normality assumption. The Wilcoxon was applied to verify de VA variation during de study. No eye or systemic complications ware observed during the study. Conclusions: Visual acuity gain occur comparing before and after treatment with bevacizumab intravitreal injection followed by laser photocoagulation. The stability of the gain combined with a absences of complications demonstrate de potential of combined intravitreal bevacizumab and laser photocoagulation in patients with BRVO