Cette thèse traite de la modélisation du frittage à l’échelle du Volume Élémentaire Représentatif de la pastille de matériau. L’objectif est de développer des outils numériquesde compréhension des phénomènes physiques mis en jeu lors du frittage. Le domaine d’application ciblé est la fabrication du combustible nucléaire. Une approche multi-Échelle a été mise en oeuvre. Dans un premier temps une modélisation à l’échelle d’un empilement, basée sur la méthode des Éléments Discrets, a été adoptée. Différentes études utilisant cette approche ont été proposées dans la littérature ces dernières années. Tous ces travaux utilisent une méthode discrète explicite. Si certains résultats ont pu être validés expérimentalement,une des limites vient de l’utilisation des méthodes explicites dontle pas de temps critique est très petit. Afin d’augmenter le pas de temps, la masse des particules y est augmentée artificiellement de plusieurs ordres de grandeur. Or,il a été démontré que cette pratique conduit, dans certains cas, à une diminution du réarrangement des particules au sein de l’empilement. Dans cette thèse, une méthode Éléments Discrets implicite appelée Dynamique des Contacts a été adaptée au frittage. Elle permet l’utilisation d’un pas de temps très supérieur à celui des méthodes discrètes explicites et ne nécessite pas d’augmenter artificiellement la massedes particules. La comparaison entre la Dynamique des Contacts et la Méthode des Éléments Discrets explicite montre que notre approche conduit à une représentation plus fidèle du réarrangement. Une validation expérimentale par Microtomographie X ainsi qu’une étude paramétrique sur le frittage des poudres bidispersés sont également présentées pour montrer les possibilités de l’approche discrète appliquée au frittage.La seconde partie est consacrée à une modélisation à l’échelle de deux particules parla méthode des Éléments Finis. Ce modèle repose sur une approche mécanique et vise à représenter de façon plus précise le comportement de deux particules en contact. Les diffusions au joint de grains, en surface et en volume peuvent être représentées. Pour le moment, seules les diffusions en surface et au joint de grains ont été étudiées. Si certaines optimisations restent nécessaires pour que le code soit fonctionnel, plusieurs aspects apparaissent déjà déterminants, comme la courbure de la surface à proximité du joint de grains. A l’avenir, le modèle Dynamique des Contacts du frittage pourra être complété etamélioré grâce aux éléments apportés par le modèle mécanique à l’échelle du grain. / This thesis deals with the simulation of the sintering of nuclear fuel on a pellet scale. The goal is to develop numerical tools which can contribute to a better understandingof the physical phenomena involved in the sintering process. Hence, a multi scale approach is proposed. First of all, a Discrete Element model is introduced. It aims at modeling the motion of particles on a Representative Elementary Volume scale using an original Discrete Element Method. The latter is a Non Smooth Method called Contact Dynamics. Recently, there have been numerous papers about the simulation of sintering using Discrete Element Method. As far as we know, all these papers use smooth methods. Different studies show that the results match well experimental data. However, some limits come from the fact that smooth methods use an explicit scheme which needsvery small time steps. In order to obtain an acceptable time step, the mass of particles have to be dramatically increased. The Non Smooth Contact Dynamics uses an implicit scheme, thus time steps can be much larger without scaling up the mass of particles. The comparison between smooth and non smooth approaches shows thatour method leads to a more realistic representation of rearrangement. An experimental validation using synchrotron X-Ray microtomography is then presented, followedby a parametric study on the sintering of bimodal powders that aims at showing the capacity of this model.The second part presents a mechanical model on the sub-Granular scale, using a Finite Element method. This targets a better understanding of the behavior of twograins in contact. The model is currently being developped but the first results already show that some parameters like the shape of the surface of the neck are very sensitive.In the future, the Non smooth Contact Dynamics model of sintering may be improvedusing the results obtained by the sub-Granular scale mechanical model.

Le tube digestif est un organe vital, conservé chez les vertébrés. Il assure la digestion des aliments, l'absorption des nutriments et l'excrétion des déchets. Une des propriétés essentielles du tube digestif est la motilité digestive, qui est définie comme l'ensemble des contractions nécessaires au transit du bolus alimentaire depuis la bouche jusqu'à l'anus. Ce processus est assuré par la coordination entre trois réseaux de cellules au sein du tube digestif, le système nerveux entérique, les cellules interstitielles de Cajal et les cellules musculaires lisses digestives. Une dysfonction dans n'importe lequel de ces trois systèmes se traduit par un désordre de la motilité digestive. La plupart de ces désordres se mettant en place au cours de la vie fétale, il est essentiel de mieux comprendre les mécanismes qui gouvernent le développement embryonnaire du tube digestif. Ainsi, la problématique globale de mon travail de thèse a été d'étudier les mécanismes moléculaires impliqués dans le développement et la différenciation des cellules musculaires lisses de l'estomac, en utilisant comme organisme modèle l'embryon de poulet. Dans un premier temps, j'ai participé à la caractérisation d'un nouveau marqueur des cellules musculaires lisses, BAPX1. Par la suite, j'ai participé à une étude sur la régulation extrinsèque du développement précoce de l'estomac par les cellules du système nerveux entérique. Cette étude a mis en évidence un rôle essentiel du système nerveux entérique dans la régulation de la voie de signalisation NOTCH pour permettre la mise en place et le maintien de l'identité de l'estomac. De plus, cette étude suggère que le système nerveux entérique est requis pour le processus de différenciation des cellules musculaires lisses de l'estomac. Enfin, pendant la majorité de mon travail de thèse, j'ai participé à l’identification de nouveaux gènes impliqués dans la régulation intrinsèque de la différenciation des cellules musculaires de l'estomac. Dans cette étude, nous avons caractérisé un nouvel acteur de ce processus et montrons que ce gène définit la population précoce des progéniteurs mésenchymateux de l'estomac et régule leur prolifération. Nous avons identifié ce gène comme un régulateur essentiel des étapes de détermination et de différenciation des cellules musculaire lisses. / The gastro-intestinal tract is a vital organ, conserved throughout the vertebrates. It is responsible for food digestion, absorption of nutrients and waste excretion. One of the most important properties of the gut is digestive motility, which is defined as all the intestinal contractions necessary for bolus transit from the mouth to the anus. This process is regulated by the coordination between three cell networks within the gut: the enteric nervous system, the interstitial cells of Cajal and the visceral smooth muscle cells. Dysfunctions in any one of these three systems result in a gastrointestinal motility disorder. Because onset of most of these diseases occurs in the fetus, a better understanding of the mechanisms involved in gastrointestinal tract development is essential. With this in mind, the main objective of my thesis was to study the molecular mechanisms that are involved in the development and differentiation of the gastric smooth muscle, using the chick embryo as a model organism. First, I participated in a study that led to the characterization of BAPX1 as a new marker of stomach smooth muscle cells. In parallel, I participated in an experimental study on the extrinsic regulation of early stomach development by the enteric nervous system. This study demonstrated that the enteric nervous system is an essential partner in the development and maintenance of the molecular identity of the stomach, through the regulation of the NOTCH pathway. Furthermore, this study suggests that the enteric nervous system is required for correct smooth muscle cell differentiation in the stomach. Finally, during most of my thesis I focused on the identification of molecular mechanisms that drive the intrinsic regulation of stomach smooth muscle differentiation. This study led to the identification of a new gene that we characterized as a new marker of stomach cells, which regulates their proliferation. Thus this gene is essential during the process of stomach smooth muscle cell determination and differentiation.

Orientadores: Sigmar Maurer Deckmann, Fernando Pinhabel Marafão / Dissertação (mestrado) - Universidade Estadual de Campinas, Faculdade de Engenharia Eletrica e de Computação / Made available in DSpace on 2018-08-07T20:09:32Z (GMT). No. of bitstreams: 1 Padua_MarceloSuzartde_M.pdf: 2971342 bytes, checksum: f838d91f23e156c63fa9582c9c5c582e (MD5) Previous issue date: 2006 / Resumo: Este trabalho visa contribuir para os estudos sobre sincronismo em um Sistema Elétrico de Potência (SEP), com enfoque em Geração Distribuída (GD). Buscar-se-á analisar as condições para operação de um gerador distribuído em paralelo com a rede elétrica, bem como desenvolver metodologias de sincronismo para tal. Muitos algoritmos propostos para aplicações em Eletrônica de Potência e Qualidade da Energia Elétrica podem ser utilizados para este propósito. Inicialmente, será apresentadas uma síntese dos tipos de geração alternativa existentes e suas características. Considerando que o Brasil, até o momento, não possui legislação específica referente à GD, serão expostas as principais normas internacionais aplicáveis ao sincronismo e à conexão do gerador, além de aspectos de segurança, como condições de ilhamento. Cada um dos algoritmos escolhidos (PLL, TDFR e FK) será estudado com o objetivo de observar sua resposta dinâmica, precisão em regime e robustez na presença de distorções harmônicas e/ou transitórios nas tensões da rede, de acordo com o ajuste dos seus parâmetros. Finalmente, serão apresentados resultados de simulação de um sistema de GD característico. Uma carga, alimentada pela rede elétrica de distribuição, passará a receber energia também de um gerador local, que será conectado em paralelo com esta rede com auxílio dos algoritmos de sincronismo estudados. Resultados experimentais confirmarão a possibilidade de implementação dessas técnicas / Abstract: This dissertation presents a contribution to the study of synchronization in Electric Power Systems, focusing Distributed Generation. The conditions for operation of a distributed generator in parallel with the grid will be analyzed and applicable methodologies of synchronism will be presented. Different algorithms proposed for Power Electronics Applications and Power Quality Analysis can be used for this intention. Initially, this work presents a synthesis of the existing types of alternative generation and their characteristics. Considering that in Brazil, up to this moment, does not exist a specific legislation for distributed generation, the international main applicable norms about synchronization and connection of the generator will be exposed, including security aspects, such as islanding conditions. Each of the chosen algorithms will be studied to observe its dynamic response, accuracy and robustness in presence of harmonic distortions or transients in the utility voltage, according to the adjusted parameters. Finally, simulation results of a characteristic industrial plant will be presented. The utility system supplies a load, which will draw power also from a local generator, connected to this grid, with the aid of the studied synchronism algorithms. Experimental results will confirm the possibility of implementation of these techniques / Mestrado / Energia Eletrica / Mestre em Engenharia Elétrica

As coronárias derivam do proepicárdio, uma estrutura formada por precursores dos constituintes de vasos coronários, células endoteliais e musculares lisas (CoSMC). In vivo observa-se um marcante atraso entre a diferenciação endotelial e a integração de CoSMC à parede do vaso. O objetivo deste trabalho foi identificar os mecanismos que inibem a diferenciação a CoSMC in vivo. Baseados na perda progressiva da expressão de raldh2, a principal enzima de síntese de ácido retinóico (AR), nós exploramos a sinalização por AR como um possível inibidor da diferenciação a CoSMC. Através de um vetor adenoviral de expressão de raldh2 e da inibição in vivo da síntese de AR nós demonstramos que a sinalização por AR bloqueia a diferenciação a CoSMC dos precursores coronários. Nós também identificamos o VEGF como um fator chave no controle da diferenciação a CoSMC. Em conjunto, nossos dados suportam o modelo que a síntese de AR e VEGF durante o desenvolvimento cardíaco foi co-optada para o bloqueio da diferenciação a CoSMC até o estabelecimento de uma vasta malha vascular. / Coronary vessels derive from the proepicardium (PE), a structure formed by precursor of coronary vessels cells, endothelial and smooth muscle cells (CoSMC). In vivo there is a clear gap between the endothelial differentiation and the integration of CoSMC into the vascular tubes. The aim of this work was to understand the mechanisms controlling the delayed in vivo CoSMC differentiation. Based on the progressive loss of expression of raldh2, the main retinoic acid (RA) synthesizing enzyme, we explored the RA signaling as a possible candidate inhibitor of CoSMC differentiation. Using a adenoviral raldh2 expression system and in vivo inhibition of RA synthesis we showed that RA signaling act as a brake to slow CoSMC differentiation in PE-derived cells. We also identified VEGF as key factor acting on the control of CoSMC differentiation. Together our results support a model that AR and VEGF synthesis during cardiac development was co-opted to block the CoSMC differentiation of coronary precursors before an extensive endothelial network of tubes is established.

Dissulfeto isomerase protéica (PDIA1 ou PDI) é uma chaperona e ditiol-dissulfeto oxido-redutase residente do reticulo endoplasmático (RE). PDI é essencial à regulação da proteostase por ter função no enovelamento oxidativo de proteínas e na via de degradação associada ao RE (ERAD). Além disso, PDI interage fisicamente e regula a atividade de NADPH oxidases, e fora da célula é um regulador redox essencial à atividade de proteínas extracelulares. Este pool epi/pericelular da PDI (pecPDI) regula função de proteínas de membrana/secretadas, como integrinas, glicoproteínas gp120 do virus HIV e outras, com múltiplas funções que incluem: trombose, ativação plaquetária, adesão celular, infecção viral e remodelamento vascular. A rota de externalização da PDI permanece obscura, e seu conhecimento pode indicar mecanismos dos efeitos (fisio)patológicos da PDI. A secreção da PDI pela rota RE-Golgi foi sugerida em células endoteliais infectadas pelo vírus da dengue, células pancreáticas e tireoideanas. No entanto, uma varredura sistemática das possíveis rotas de externalização da PDI não foi previamente realizada. Neste estudo, mostramos que células endoteliais (EC) externalizam constitutivamente, por rotas distintas, dois pools de PDI, de superfície celular e solúvel, enquanto na EC não estimulada PDI não foi detectada significativamente em micropartículas. PDI externalizada corresponde a ca.1,4% do pool total de PDI celular. Tanto a PDI de superfície celular como a solúvel foram majoritariamente secretadas pela via de secreção não-convencional do tipo IV independente de GRASP. Contudo, a via de secreção clássica também contribui para externalização basal da PDI de superfície celular, mas não da solúvel basal ou estimulada por PMA, ATP e trombina indicando que todas envolvem escape do Golgi. Além disso, a externalização constitutiva da PDI de superfície em célula muscular lisa vascular também ocorre por via independente de Golgi. Externalização da PDI não foi detectavelmente mediada pela secreção não-convencional do tipo I, II, III, lisossomos secretórios, endossoma de reciclagem e transporte ativo (dependente de ATP) em EC. Considerando que chaperonas são vias essenciais de resposta a estresses, investigamos o efeito de estresse do RE e choque térmico na pecPDI. Estresse do RE não altera a PDI de superfície celular, mas aumenta PDI solúvel. Ambos os pools de PDI não foram alterados por choque térmico, embora a recuperação desse estresse diminua a secreção de PDI. Estes dados sugerem que a liberação de PDI é um processo regulado, dependente da natureza do estresse. Bloqueio da síntese de proteínas com cicloheximida não altera pecPDI, indicando que PDI recém-sintetizada não é preferencialmente externalizada e que o tráfego da PDI independe de outras proteínas recém-sintetizadas. Um aspecto importante do estudo foi indicar uma resiliência da pecPDI à modulação individual de distintas vias secretoras, consistente com uma estrita auto-regulação e possibilidade de vias sinérgicas e complementares. Estes resultados indicam que a externalização da PDI de superfície e PDI secretada possam ser externalizadas por mecanismos independentes. Estes processos compõem um processo regulado estritamente, consistente com papel homeostático da pecPDI / Protein disulfide isomerase (PDIA1 or PDI) is dithiol-disulfide oxireductase chaperone resident in the endoplasmic reticulum (ER). PDI is essential for proteostasis, due to its support of oxidative protein folding and ER-associated protein degradation (ERAD). In addition, PDI associates with NADPH oxidase(s) and regulate its activity, while outside of the cell, PDI redox-dependently modulates extracellular proteins. This epi/pericellular PDI (pecPDI) pool is known to regulate membrane/secreted proteins such as integrins, HIV glycoprotein gp120 and others, with functions that involve thrombosis, platelet function, cell adhesion, viral infection and vascular remodeling. PDI externalization route remains enigmatic and its elucidation can help understand some (patho)physiological PDI effects. An ER-Golgi route for PDI secretion has been as described on dengue virus-infected endothelial cells pancreatic and thyroid) cells. However, none of these papers addressed PDI secretion routes in a systematic fashion. Here, we show that endothelial cells (EC) constitutively externalize, through different routes, two PDI pools, a cell-surface and a secreted one, while in nonstimulated ECs PDI was not significantly detected in microparticles. Externalized PDI corresponds to < 2% of total cellular PDI pool. Both cell-surface and soluble PDI were predominantly externalized through unconventional type IV GRASP-independent pathway(s). However, the classical secretory pathway also contributes to basal cell-surface, but not soluble, PDI externalization, as PMA, ATP or thrombin-stimulated secretion also involve Golgi bypass. Furthermore, constitutive cell-surface PDI externalization in vascular smooth muscle cells also occurs in a Golgi-independent way. PDI externalization was not detectably mediated by non-conventional type I, II and III secretion routes, secretory lysosomes, recycling endosomes and ATP dependent active transport in EC. Since chaperones are essential for cellular stress response, we assessed the effects of ER stress and heat-shock on pecPDI. ER stress did not affect cell-surface PDI but increased the soluble pool. Both PDI pools were unaltered by heat shock, while stress recovery decreased PDI secretion. These data suggest that PDI release is finely tuned and dependent on the type of stress. Blockade of protein synthesis with cycloheximide did not change pecPDI levels, suggesting that newly-synthesized PDI is not preferentially externalized and that PDI traffic does not require newly-synthesized proteins. An important aspect of the study was the evidence for pecPDI resilience to individual modulation of distinct secretion routes, consistent with strict auto-regulation and possible synergic or complementary pathways. Overall, our data suggest that cell-surface and secreted PDI pool externalization are regulated through independent mechanisms, which in both cases involve Type IV non-conventional routes, with some minor contribution of Golgi-dependent secretory pathway. These patterns compose a strictly regulated process, consistent with an important homeostatic role for pecPDI

Lesão do DNA mitocondrial (mtDNA) promove disfunção desta organela, contribuindo para a gênese do envelhecimento e fisiopatologia de doenças como aterosclerose e diabetes. A mitocôndria é a principal fonte quantitativa de espécies reativas de oxigênio (ROS) em células, e o complexo NAD(P)H oxidase a principal fonte de ROS envolvidas na sinalização celular. A possível inter-relação entre estas duas importantes vias produtoras de ROS não está definida. O objetivo deste estudo foi investigar o perfil de alterações na expressão e atividade da NAD(P)H oxidase de células musculares lisas vasculares (VSMC) em resposta a perturbações mínimas da função mitocondrial análogas às esperadas em doenças crônico-degenerativas vasculares. Inicialmente, validamos modelo in vitro de disfunção mitocondrial induzida por incubação de VSMC com brometo de etídio (24 - 72 h). Lesões mínimas do mtDNA foram documentadas por alterações nos produtos de amplificação (PCR) da região repetitiva da D-loop e redução da taxa de consumo de oxigênio total em ~15% vs. basal (p<0,05). Este grau de lesão não foi suficiente para induzir alterações morfológicas evidentes ou apoptose, e foi associado ao retardo de 25 - 30% no aumento de população celular induzido por soro fetal bovino. Nestas condições, não se detectou aumento da produção basal de superóxido ou mudanças nos níveis de glutationa, óxidos de nitrogênio, ou da atividade superóxido dismutase. A produção basal de peróxido de hidrogênio aumentou ~15%. Após disfunção mitocondrial, houve significativo aumento (30 - 45%) na atividade basal do complexo NAD(P)H oxidase em fração de membrana de VSMC. Entretanto, a ativação da oxidase pela AII, conhecido agonista da oxidase vascular, foi essencialmente abolida, indicando dependência funcional da ativação da oxidase com a integridade da mitocôndria. Em sintonia com esses dados, na condição basal, ocorreu aumento de expressão da isoforma Nox4 da oxidase, enquanto o aumento do mRNA da Nox1 normalmente visto após AII foi minimizado. Por outro lado, o aumento da atividade da NADPH oxidase causado pelo estressor do RE tunicamicina (indutor de Nox4) foi também abolido pela disfunção mitocondrial, entretanto, ocorreu aumento do mRNA da Nox4, indicando que as alterações funcionais da oxidase nesta situação não decorrem apenas de mudanças da expressão. Dissociação semelhante entre expressão e atividade ocorreu após exposição de 72 horas ao EtBr (i.e., durante adaptação). Nesta, ocorreu maior expressão do mRNA de Nox1 e Nox4 com AII, sem aumento da atividade da oxidase em membranas. Incubação do EtBr por 24 horas não induziu per se aumento consistente nos índices de estresse do RE e induziu inversão do padrão do tráfego subcelular da dissulfeto isomerase protéica (PDI), uma chaperona redox descrita recentemente como reguladora da NADPH oxidase. Após 72 horas de incubação com EtBr, a expressão de chaperonas marcadoras de estresse do RE foi bastante diminuída e o tráfego da PDI teve o padrão restaurado. Demonstramos por microscopia confocal evidências preliminares de possível co-localização entre Nox1 e mitocôndria. Estes dados sugerem uma relevante inter-relação funcional entre mitocôndria e complexo NAD(P)H oxidase, associada pelo menos a alterações de expressão e/ou tráfego subcelular de subunidades catalíticas e reguladoras desse complexo. / Mitochondrial DNA (mtDNA) damage induces dysfunction of this organelle, contributing to the genesis of aging and to the pathophysiology of diseases such as atherosclerosis and diabetes. Mitochondria are the main quantitative source of reactive oxygen species (ROS) in cells, while NAD(P)H oxidase complex is a major source of cell signaling-associated ROS. The possible crosstalk between these two relevant sources of ROS is unclear. The aim of this study was to investigate changes in activity and/or expression of vascular smooth muscle cell (VSMC) NAD(P)H oxidase in response to minor perturbations of mitochondrial function similar to those expected to occur in chronic degenerative vascular diseases. Initially, we validated an in vitro model of mitochondrial dysfunction in VSMC, through incubation with ethidium bromide (24 - 72 h). Minimal mtDNA damage after EtBr was shown by distinct amplification patterns (at PCR) of D-loop repetitive region and by ~ 15% oxygen consumption decrease vs. basal (p<0.05). Such mtDNA damage was not sufficient to induce morphologic changes or apoptosis, whereas serum-stimulated increase in cell number was prevented by 25-30%. Under those conditions, baseline superoxide production, as well as levels of glutathione or nitrogen oxides or superoxide dismutase activity were unchanged. Baseline hydrogen peroxide production increased ~15%. VSMC membrane fraction NADPH oxidase activity was increased by 30-45% after mitochondrial dysfunction. However, oxidase activation due to AII (100 nM, 4h) was markedly abrogated, indicating that A-II-driven oxidase activation requires integrity of mitochondrial function. Accordingly, there were increases in baseline mRNA expression of Nox4 oxidase isoform, while the expected increase in Nox1 by AII was minimized. On the other hand, the NADPH oxidase activity induced by the endoplasmic reticulum stressor tunicamycin (Nox4 inducer) after mitochondrial dysfunction was abrogated, however simultaneously with increased Nox4 mRNA, thus indicating that the observed functional alterations in the oxidase complex in these conditions cannot be associated only to mRNA expression changes. After VSMC EtBr incubation for 72 h, similar dissociation between expression and activity was observed, with increase in Nox 1 and Nox4 mRNA by AII, without parallel increase in membrane fraction oxidase activity. Although there was little change in ER stress markers after 24h EtBr, protein disulfide isomerase (PDI), a redox chaperone recently described by us as a novel NAD(P)H oxidase regulator, exhibited a reversal of its subcellular traffic pattern. After 72 h EtBr, the expression of ER markers was strongly decreased and normal PDI traffic was restored. Confocal microscopy suggested possible co-localization between Nox1 and mitochondria. These results suggest a functionally relevant crosstalk between mitochondria and NADPH oxidase complex associated at least to changes in expression and/or subcellular traffic of catalytic or regulatory subunits of this complex.

L’Hypertension Pulmonaire Post-Embolique (HPPE) est une maladie rare et grave résultant d'une obstruction mécanique progressive des artères pulmonaires provoquant une augmentation progressive de la PAPm (>25 mmHg). Cette pathologie a un traitement chirurgical, l’endartériectomie pulmonaire. Bien que cette maladie ait un traitement, les mécanismes qui impliquent la formation et la persistance de thrombus conduisant à un remodelage vasculaire fibreux, restent à ce jour inconnus. La pathogénèse de l’HPPE présente de grandes similitudes avec celles de l’HTAPi.Ce travail a pu mettre en évidence le phénotype anomal de la cellule endothéliale provenant de séquestres de patients ayant une HPPE ainsi que les anomalies moléculaires à l’origine de cette dysfonction. Plusieurs anomalies moléculaires sont à l’origine de ces perturbations fonctionnelles dont notamment une production et une libération anormale d’ICAM-1 et de FGF-2. Ce travail montre le rôle clé de la protéine ICAM-1 grâce à une boucle autocrine qui va maintenir cette dysfonction ainsi que l'association du génotype KE à la maladie (polymorphisme SNP Exon 6 E469K). Ce travail montre également le rôle essentiel que joue la protéine FGF-2 et le recrutement de progéniteurs endothéliaux dans la néo-vascularisation observée dans les thrombus de patients souffrant d’HPPE. / Chronic ThromboEmbolic Pulmonary Hypertension (CTEPH) is a rare and serious disease resulting from progressive mechanical obstruction of the pulmonary arteries causing a gradual increase in the mPAP (> 25 mmHg). This pathology has a surgical treatment: Pulmonary Endarterectomy. Mechanisms that involve the formation and persistence of thrombus leading to a fibrous vascular remodeling remain unknown. The pathogenesis of CTEPH is very similar to idiopathic PAH.This work has been able to demonstrate the abnormal phenotype of endothelial cells taken from thrombus of CTEPH and molecular anomalies in the origin of this dysfunction. Several molecular abnormalities are the cause of these functional disturbances including production and abnormal release of ICAM-1 and FGF-2. This work shows the key role of ICAM-1 protein through an autocrine loop maintains this dysfunction and the association of genotype KE disease (SNP Exon 6 E469K).This work also showed the important role of FGF-2 protein and the recruitment of endothelial progenitor cells in neovascularization observed in thrombus of CTEPH patients.

L’AMPc est un second messager exerçant un rôle vasculoprotecteur majeur, par ses effets relaxants et inhibiteurs de la prolifération et de la migration cellulaires. Les concentrations intracellulaires d’AMPc sont finement régulées par leur synthèse via les adénylates cyclases et leur dégradation par les phosphodiestérases (PDEs). Nous avons évalué l’impact de l’environnement cellulaire sur la voie de signalisation couplée au récepteur β-adrénergique (β-AR/AMPc/PDE) dans les cellules musculaires lisses vasculaires (CMLVs), ainsi que les altérations potentielles de celle-ci en situation pathologique d’insuffisance cardiaque (IC).Notre première étude montre que dans les CMLs d’aorte de rat en culture, adoptant un phénotype synthétique, la voie de signalisation β-AR/AMPc/PDE est hautement modulée par la densité cellulaire Ainsi, une faible densité cellulaire est associée à une régulation négative de l’expression fonctionnelle du récepteur β1-AR, à une activité hydrolytique des PDEs-AMPc plus faible et à des concentrations d’AMPc intracellulaire plus élevées que celles observées dans des cellules confluentes.Notre deuxième étude montre que dans l’aorte de rat, l’IC est associée à une dysfonction endothéliale (DE), une hyperréactivité aux agents contractants et une altération de la fonction et de l’expression des PDEs-AMPc. Nos résultats suggèrent que l’altération de la voie du NO/GMPc suite à la DE conduit à une hyper-activation de la PDE3, qui masque la fonction de la PDE4 et altère la relaxation β-AR.L’ensemble de ce travail met en évidence le rôle critique de l'environnement cellulaire dans le contrôle de la voie β-AR/AMPc/PDE des CMLVsMots clés : Muscle lisse vasculaire, récepteur β-adrénergique, AMPc, phosphodiestérases, densité cellulaire, insuffisance cardiaque / CAMP is second messenger which plays a prominent vasculoprotective role by its relaxing effects and inhibition of cell proliferation and migration. Intracellular cAMP level is regulated by its synthesis by adenylate cyclase and its degradation by phosphodiesterases (PDEs). We evaluated the influence of cellular environment on signaling pathway coupled to β-adrenoceptors (β-AR/cAMP/PDE) on vascular smooth muscle cells (VSMCs), as well as potential alterations in heart failure (HF).The first study showed that in cultured rat aortic SMCs exhibiting synthetic phenotype, the β-AR/cAMP/PDE signaling pathway is highly modulated by the cellular density.Thus, the low density state being associated to a downregulation of the β1-AR, a lower cAMP-PDE activity and a higher basal [cAMP]i compared to confluent cells.Our second study showed that in rat aorta, HF is associated with endothelial dysfunction, hyper-reactivity to contractile agents and an alteration of function and expression of cAMP-PDEs. Our results suggest that NO/cGMP pathway alteration following the ED in HF leads to hyper-activation of PDE3 which masks PDE4 function and alters β-adrenoceptor relaxationThus, our works highlights the critical role of the cellular environment in controlling the vascular β-AR signaling.Keywords: Vascular smooth muscle, β-adrenoceptor, cAMP, phosphodiesterases, cellular density, heart failure.

L’Hypertensions Artérielle Pulmonaire (HTAP) résulte de l’obstruction progressive des artères pulmonaires de petits calibres due à un remodelage de la paroi vasculaire ainsi qu’à une vasoconstriction. Cette thèse repose sur 3 piliers de la physiopathologie de l’HTAP : l’inflammation, l’épigénétique et la susceptibilité génétique de la maladie.Le premier projet démontre que les cellules dendritiques des patients atteints d’HTAP sont moins sensibles à l’action immunomodulatrice des glucocorticoïdes et orientent la réponse des lymphocytes T vers une réponse Th17, souvent impliquée dans les maladies autoimmunes.Le second projet a pour but de comparer le profil de méthylation des cellules endothéliales d’artères pulmonaires (CE-AP) de patients atteints d’HTAP comparé aux CE-AP de patients contrôles. Nous avons mis en évidence des clusters de gènes ayant un profil de méthylation différents entre les CE-HTAP comparé aux CE-CTR. Un gène différentiellement méthylé s’est dégagé durant l’étude bioinformatique : ABCA1 (ATP binding cassette 1). L’altération de son expression prédite par l’étude bioinformatique a été validée chez l’homme et le modèle de rat monocrotaline.Enfin, le dernier projet décrit la caractérisation d’un nouveau modèle animal de susceptibilité génétique à l’HTAP liée à des mutations dans le gène Bmpr2 chez le rat, au niveau hémodynamique, histologique, vasculaire, moléculaire et électrophysiologique. Nous avons démontré que ce modèle reproduit des schémas physiopathologiques pertinents vis-à-vis de la maladie humaine, ces résultats font de ce modèle un nouveau outil d’étude de la physiopathologie de l’HTAP. / Pulmonary arterial hypertension (PAH) is characterized by a progressive pulmonary arterial obstruction due to pulmonary vascular remodeling of distal arterioles as well as abnormal vasoconstriction. This project allowed to study three biological process clearly establish to be implicated in physiopathology of PAH.The first project demonstrated a dendritic cells dysfunction and a Th17 immune polarization of idiopathic PAH patients.The objective of the second project was to study the epigenetic variations in pulmonary endothelial cells (PEC) through a specific pattern of DNA methylation. We identified clusters of probes that discriminates controls and PAH patients. During bioinformatics study, ABCA1 (ATP binding cassette 1) gene was emphasized. Alteration of ABCA1 expression predicted during bioinformatics study has been validated in human and in monocrotaline rat model.The last project described the validation of a new PAH model by a hemodynamic, histological, vascular, molecular and electrophysiological characterization of heterozygous rat mutated to Bmpr2 gene. Whole functional and molecular dysregulation define this animal model like a useful tool in the study of BMPRII signaling alteration in PAH physiopathology.

We study one-parameter families of quasi-periodically forced monotone interval maps and provide sufficient conditions for the existence of a parameter at which the respective system possesses a non-uniformly hyperbolic attractor. This is equivalent to the existence of a sink-source orbit, that is, an orbit with positive Lyapunov exponent both forwards and backwards in time. The attractor itself is a non-continuous invariant graph with negative Lyapunov exponent, often referred to as ‘SNA’. In contrast to former results in this direction, our conditions are C² -open in the fibre maps. By applying a general result about saddle-node bifurcations in skew-products, we obtain a conclusion on the occurrence of non-smooth bifurcations in the respective families. Explicit examples show the applicability of the derived statements.