Le sang provenant d’un cordon ombilical (SCO) représente une bonne source de cellules souches hématopoïétiques (CSH) pour des transplantations. Cependant, le nombre de cellules souches contenues dans ce sang est souvent insuffisant pour greffer un adulte. Le mécanisme intervenant dans la domiciliation de ces cellules au sein de la moelle osseuse (MO) est encore mal compris. On sait que l’interaction entre la chimiokine SDF-1 et le récepteur CXCR4, présent sur les cellules CD34+ de SCO, mène à la migration de ces cellules en direction de la MO. Nous pensons que l’augmentation de la proportion de cellules qui réussit à se greffer pourra pallier au problème du nombre. Les produits de dégradation, C3a et le C3desarg,, issus du système du complément, sont connus pour favoriser la réponse de cellules exprimant CXCR4 vers SDF-1. Nous avons analysé l’effet du C3adesarg, molécule non anaphylatoxique, sur la migration cellulaire vers SDF-1, de même que sur la prise de greffe des cellules CD34+ issues de SCO suite à une transplantation sur des souris NOD/SCIDyC-. Nos expériences ont démontré que le C3a ainsi que le C3adesarg augmentaient tous les deux la réponse des cellules CD34+ vers SDF-1. Toutefois, nous n’avons pas pu démontrer que ces molécules liaient directement le récepteur CXCR4. Par contre, le composé C3adesarg favorise la prise de greffe des cellules CD34+ de SCO. Il serait donc un bon candidat pour poursuivre une optimisation de ses propriétés. Nous avons également constaté que suite à une transplantation chez la souris, les cellules CD34+ de SCO subissent une hausse d’expression transitoire de leur CXCR4 environ quatre jours après la greffe. Cette hausse d’expression coïncide avec la multiplication des cellules CD34+ dans la MO. Nous avons également confirmé qu’une cellule CD34+ avec une forte expression de CXCR4 était dans un état prolifératif. Nos données suggèrent que l’interaction directe avec les cellules stromales soit responsable de cette hausse d’expression de CXCR4. / Since the first successful cord blood (CB) transplant was performed there has been a gradual increase in the use of CB for haematopoietic stem cell (HSC) transplantation, but the number of stem cells per CB is in general too low to ensure successful transplantation in adult patients. We would like to bypass the limitation of insufficient number of these cells in CB by enhancing the engraftment efficiency. The chemokine stromal-derived factor (SDF)-1, that binds to its receptor, CXCR4, plays an important and unique role in regulating the trafficking of HSC and their homing/retention in bone marrow (BM), but molecular regulatory mechanism of niches for HSC maintenance remains unclear. The complement C3 cleavage fragments, C3a and C3adesarg, modulate the responsiveness of CXCR4-expressing cell lines to SDF-1. We assessed the effect of the non anaphylatoxic complement fragment, C3adesarg, on SDF-1 responsiveness and engraftment of CB-HSC transplantation in a NOD/SCIDyC- mouse model. Complement breakdown products C3a and C3adesarg both increase the responsiveness of CD34+ cells to SDF-1. We find no evidence for direct interaction of complement fragments with CXCR4. Our data suggest that C3adesarg might contribute to optimize CB-HSC homing to bone marrow, and therefore efficacy of cord blood transplantation. We quantified the number of CXCR4 on the surface of CB-CD34+ after transplantation in mice. Our results showed that there is a transient overexpression of CXCR4 on the surface of HSC CD34+ found in the BM of NOD/SCIDyC- mice after 4-5 days post-injection. This transient overexpression correlated with multiplication of CD34+ cells in the BM. We confirm that the cells with an overexpression of CXCR4 are in a proliferation state. Our data suggested that this transient overexpression is caused by an interaction with the stomal cells.

Die vorliegende Arbeit beinhaltet die Generierung und Evaluation von Natürlichen Killerzellen, die EGFRvIII-positive und SDF-1alpha sekretierende primäre Glioblastomzellen aufspüren, erkennen und effizient abtöten können. Die Kombination der gelenkten Zytotoxizität mit einer optimierten Migration von Effektorzellen des Immunsystems wird auf Grundlage der in dieser Arbeit gewonnenen Daten als ein vielversprechender Ansatz für eine zukünftige Therapie des primären Glioblastoms vorgeschlagen.

Made available in DSpace on 2014-12-17T15:47:51Z (GMT). No. of bitstreams: 1 ClevertonHA.pdf: 1775580 bytes, checksum: 9e49f67c9b7fbb459e2b24f568db691b (MD5) Previous issue date: 2010-03-01 / Coordena??o de Aperfei?oamento de Pessoal de N?vel Superior / Some programs may have their entry data specified by formalized context-free grammars. This formalization facilitates the use of tools in the systematization and the rise of the quality of their test process. This category of programs, compilers have been the first to use this kind of tool for the automation of their tests. In this work we present an approach for definition of tests from the formal description of the entries of the program. The generation of the sentences is performed by taking into account syntactic aspects defined by the specification of the entries, the grammar. For optimization, their coverage criteria are used to limit the quantity of tests without diminishing their quality. Our approach uses these criteria to drive generation to produce sentences that satisfy a specific coverage criterion. The approach presented is based on the use of Lua language, relying heavily on its resources of coroutines and dynamic construction of functions. With these resources, we propose a simple and compact implementation that can be optimized and controlled in different ways, in order to seek satisfaction the different implemented coverage criteria. To make the use of our tool simpler, the EBNF notation for the specification of the entries was adopted. Its parser was specified in the tool Meta-Environment for rapid prototyping / Alguns programas podem ter sua entrada formalizada atrav?s de gram?ticas livres de contexto. Esta formaliza??o facilita a utiliza??o de ferramentas na sistematiza??o e na eleva??o da qualidade do seu processo de teste. Dentro desta categoria de programas os compiladores foram os primeiros a utilizar este tipo de ferramenta para a automa??o de seus testes. Neste trabalho apresentamos uma abordagem para a defini??o de testes a partir da descri??o formal das entradas do programa. A gera??o das senten?as ? realizada levando em considera??o aspectos sint?ticos definidos pela especifica??o da entrada, a gram?tica. Por quest?es de otimiza??o s?o utilizados crit?rios de cobertura para limitar a quantidade de testes sem diminuir a sua qualidade. Nossa abordagem utiliza estes crit?rios no direcionamento da gera??o de maneira a produzir senten?as que satisfa?am um crit?rio de cobertura especifico. A abordagem apresentada se baseia na utiliza??o da linguagem Lua, se apoiando fortemente em seus recursos de corotinas e constru??o din?mica de fun??es. Com estes recursos, propomos uma implementa??o compacta e simples que pode ser otimizada e controlada de formas variadas, com o intuito de buscar a satisfa??o dos diferentes crit?rios de cobertura implementados. Para tornar simples o uso de nossa ferramenta foi adotada a nota??o EBNF para a especifica??o das entradas. O seu parser foi especificado na ferramenta Meta-Environment por esta favorecer a r?pida prototipa??o

No description available.

Choque séptico é caracterizado por desequilíbrio entre o transporte e o consumo de oxigênio, podendo acarretar hipóxia tecidual. A disfunção microcirculatória, característica cardinal da fisiopatologia do choque séptico, causa má distribuição de fluxo sanguíneo microvascular e, consequentemente, shunt de oxigênio, disóxia tissular e, teoricamente, diminuição no consumo de oxigênio (VO2) pela célula. No presente estudo, foi investigada a associação entre alterações microcirculatórias causadas pela sepse e o consumo de oxigênio em pacientes pediátricos. Dezessete crianças com choque séptico ressuscitadas foram estudadas em quatro momentos durante a internação na unidade de terapia intensiva (dentro de 24, 48 e 72 horas após a admissão ou diagnóstico de choque e após a resolução deste, antes da extubação traqueal). A microcirculação sublingual foi avaliada utilizando o método de imagem Sidestream dark field (SDF) e o VO2 foi calculado através da calorimetria indireta. Outras variáveis hemodinâmicas, como transporte de oxigênio, índice cardíaco, pressão arterial invasiva, lactato arterial e saturação venosa central, foram coletadas. Embora as variáveis hemodinâmicas tenham se mantido em níveis satisfatórios, graves alterações na microcirculação foram visualizadas, especialmente na densidade de vasos pequenos perfundidos (DVPP), na proporção de vasos pequenos perfundidos (PVPP) e no índice de fluxo microvascular (MFI). Foram encontradas assosciações significativas entre o VO2 e os parâmetros da microcirculação: dVO2 e dDVPP (&#946; coefficient= 6,875; p<0,001), dVO2 e dPVPP (&#946; coefficient=92,246; p<0,001) e dVO2 e dMFI (&#946; coefficient=21,213; p<0,001). Não foram encontradas correlações entre as alterações microcirculatórias e as outras variáveis. Em conclusão, este estudo mostrou que pacientes pediátricos com choque séptico apresentaram grave disfunção microvascular e que o fluxo microcirculatório alterado estava associado ao VO2, podendo estar implicado na fisiopatologia da disóxia tecidual da sepse. / Septic shock is characterized by the imbalance between oxygen delivery and consumption leading to tissue hypoxia. Microcirculatory dysfunction, a key element of septic shock pathogenesis, elicits maldistribution of microvascular blood flow and consenquently oxygen shunt, tissue oxygenation debt and, theoretically, impaired oxygen consumption (VO2). In this study, it was investigated if there is an association between microcirculatory changes and VO2 in pediatric patients with septic shock. Seventeen resuscitated patients with septic shock were studied in four moments (within 24hr, 48hr and 72hr of the admission or diagnosis of shock and after its resolution, prior to extubation). Sublingual microcirculation was evaluated using Sidestream dark field (SDF) imaging and VO2 was measured directly by indirect calorimetry. Other hemodynamic variables, like cardiac index, oxygen delivery, invasive arterial pressure, arterial lactate and central venous oxygen saturation were also recorded. Although global hemodynamic variables were within satisfactory ranges, microvascular variables were markedly altered, especially microvascular flow index (MFI), proportion of perfused small vessels (PPV) and perfused small vessel density (PVD). Significant associations between oxygen consumption and microcirculatory parameters were found: dVO2 and dPVD (&#946; coefficient= 6.875; p<0.001), dVO2 and dPPV (&#946; coefficient=92.246; p<0.001) and dVO2 and dMFI (&#946; coefficient=21.213; p<0.001). There was no correlation between microcirculatory alterations and other variables in this study. In conclusion, this study showed that pediatric patients with septic shock presented severe microcirculatory dysfunction and abnormal microvascular blood flow could be associated to oxygen consumption.

O tráfego de leucócitos é um processo complexo, dependente da ação de inúmeras substâncias químicas, além da perfeita interação celular. Desta forma, este estudo teve como objetivo avaliar a ação dos GCe e da ANXA1 sobre o eixo SDF-1&#945;/CXCR4 e IL-17/IL-23/G-CSF e sobre a expressão de moléculas de adesão CD18, CD49d e CD62L. Foram utilizados camundongos machos Balb/C selvagens (WT) ou ANXA1-/-. As avaliações foram realizadas em condições basais, na presença de altas concentrações de GCe e na vigência de processo inflamatório, induzidos pela administração de ACTH (5 &#181;g/animal, i.p.) ou pela injeção de LPS (100 &#181;g/kg, i.p.), respectivamente, ou na ausência da ação dos GCe, pela ação do RU 38486 (RU, 10 mg/kg, i.p.). A participação da ANXA1 e do receptor FPR2 foi avaliada pelo pré-tratamento com Ac2-26 (1 mg/Kg, i.p.) ou com BOC2 (10 &#181;g/animal, i.p.) durante 4 dias, 1 vez ao dia. A quantificação total e diferencial das células foi realizada em câmara de Neubauer e em esfregaços corados por May-Grunwald ou citometria de fluxo. As quantificações de CXCR2, CXCR4, FPR2, CD18, CD49d, CD62L e maturação granulocítica (CD11b/Ly6G) em células da medula e da circulação foram realizadas por citometria de fluxo. A expressão de ANXA1 nos tecidos do estomago e do baço foi realizada por western blotting e nas células da medula óssea e sangue circulante foi realizada por imunofluorescência. As quantificações de IL-17, IL-23, G-CSF, SDF-1&#945; e corticosterona foram realizadas por ELISA. A quimiotaxia de neutrófilos da medula óssea e sangue periférico foi ensaiada na placa de quimiotaxia com filtro de poro de 8 &#181;m. A fagocitose de neutrófilos apoptóticos por macrófagos da medula óssea foi avaliada por ensaio in vitro. Para verificar os efeitos do ACTH na migração de neutrófilos no processo inflamatório, foi empregado o modelo de bolsa de ar (100 &#181;g/mL; LPS); e o comportamento dos leucócitos circulantes de animais tratados com ACTH foi avaliado pela técnica de microscopia intravital. Os resultados obtidos, que estão apresentados em quatro temáticas, mostraram que: 1) neutrófilos da medula óssea e sangue periférico expressam ANXA1 no citoplasma e membrana, bem como o receptor FPR2, constitutivamente, e a expressão de ambos é regulada pelos GCe. A ANXA1, via receptor FPR2 expresso em células da medula óssea, controlam a maturação neutrofílica e o tráfego destas células da medula óssea para o sangue. A ANXA1, via interação ao FPR2, controla o clearance de neutrófilos do sangue para a medula óssea, modulando o eixo SDF-1&#945;/CXCR4; 2) A administração do ACTH causa neutrofilia e os neutrófilos circulantes são ANXA1+, CD18+, CD49d+, CD62L+, mostrando que injeção do ACTH in vivo altera o fenótipo destas células na circulação. Estas modificações alteram o comportamento dos neutrófilos na circulação, bem como a migração para a bolsa de ar na vigência de inflamação e para os tecidos de clearance. Estes efeitos podem ser dependentes, pelo menos em parte, da inibição de migração orientada, já que quimiotaxia frente ao fMLP ou ao SDF-1&#945; estavam reduzidas. Ainda, o clearance de neutrófilos é reduzido em animais tratados com o ACTH pela menor atividade fagocítica e secretora dos macrófagos medulares; 3) Animais tratados com RU 38486 e ANXA1-/- mobilizam granulócitos da medula óssea para o sangue circulante e, deste compartimento para o foco de inflamação com maior intensidade que o observado em animais controles. O eixo IL-17/IL-23/G-CSF parece estar envolvido na granulopoiese e na mobilização de neutrófilos para o sangue durante a inflamação, mas não é alvo de ação da ANXA1 e o GCe nesta etapa do processo inflamatório. Adicionalmente, foi observado que na vigência de peritonite, as moléculas de adesão, CD49d e CD62L estão envolvidas no processo de migração de neutrófilos da medula óssea para o sangue. Os resultados aqui obtidos permitem concluir que os GCe e a ANXA1 são relevantes para granulopoiese e tráfego dos neutrófilos da medula óssea em condições fisiológicas e na vigência de processo inflamatório. Ainda, em conjunto com os dados da literatura, os nossos resultados podem sugerir a participação da ANXA1 dos GCe na plasticidade fenotípica dos neutrófilos de acordo com os estímulos a que são submetidos, e podem auxiliar na compreensão dos novos conceitos sobre a produção, tempo de vida, localização e funções de neutrófilos. / The traffic leukocytes is a complex process dependent on the action of severals chemical mediators, in addition to perfect cell interaction. Therefore, this study aimed to evaluate the effect of GCe and ANXA1 on SDF-1&#945;/CXCR4 and IL-17/IL-23/G-CSF and on the expression of adhesion molecules CD18, CD49d and CD62L. Balb/C wild type and ANXA1-/- male mice were employed. The analysis were performed at physiological conditions, in the presence of high concentrations of GCe and during of inflammatory process induced by ACTH administration (5 &#181;g/animal, i.p.) or LPS injection (100 &#181;g/kg, i.p.), respectively or in the absence of GCe action, by the action of RU 38486 (RU, 10 mg/kg , i.p.). The involvement of the receptor FPR2 and ANXA1 was assessed by pre-treatment with Ac2-26 (1 mg/kg, i.p.) or BOC2 (10 &#181;g/animal, i.p.) for 4 days, once a day. The quantification of total and differential cell was performed in a Neubauer chamber and stained smears by May-Grunwald and flow cytometry. Quantification of expression of CXCR2, CXCR4, FPR2, CD18, CD49d, CD62L and granulocytic maturation (CD11b/Ly6G) in the bone marrow and circulation were performed by flow cytometry. The expression of ANXA1 on tissues was performed by western blotting and on cells from bone marrow and blood by immunocytochemistry. Quantification of IL-17, IL-23, G-CSF, SDF-1&#945; and corticosterone were performed by ELISA. The chemotaxis of neutrophils from the bone marrow and blood was tested in the chemotaxis chamber with filter pore of 8 microns. The phagocytosis of apoptotic neutrophils by bone marrow macrophages was assessed by in vitro assay. To investigate the effects of ACTH in the migration of neutrophils in the inflammatory process, the model employed was air pouch (100 &#181;g/ ml, LPS), and the behavior of circulating leukocytes from animals treated with ACTH were evaluated by intravital microscopy. The results obtained, which are presented in three sections, showed that: 1) neutrophils from the bone marrow and blood expressed ANXA1 in the cytoplasm and membrane, as well as FPR2, constitutively and the expression of both is regulated by GCe. The ANXA1 via FPR2 receptor expressed in bone marrow cells, controls the neutrophilic maturation and traffic of these cells from the bone marrow into the blood. The ANXA1 via interaction to FPR2 controls the clearance of neutrophils from the blood to the bone marrow by modulating the SDF-1&#945;/CXCR4 axis; 2) the administration of ACTH induces neutrophilia and the circulating neutrophils are ANXA1+, CD18+, CD49d+ and CD62L+, showing that the injection of ACTH in vivo alters the phenotype of these cells in the blood. These modifications alter the behavior of neutrophils in the blood, as well as the migration to the air pouch in the presence of inflammation and to the tissue clearance, and these effects may be dependent, at least in part, on inhibition of migration oriented events, as chemotaxis in response to fMLP or SDF-1&#945; were reduced. Further, the clearance of neutrophils is reduced in animals treated with ACTH due to the lower phagocytic and secretory activity of medullary macrophages; 3) Animals treated with RU 38486 and ANXA1-/- mobilize granulocytes from bone marrow into the blood, and from this compartment to the focus of inflammation with higher intensity than that observed in the control group. The axis IL-17/IL-23/G-CSF seems to be involved in granulopoiesis and mobilization of neutrophils into the blood during inflammation, but it is not the target of action of ANXA1 and GCe at this step of inflammatory process. Additionally, it was observed that in the presence of peritonitis, the adhesion molecules, CD49d and CD62L are involved in the migration of neutrophils from the bone marrow into the blood. The results obtained allow concluding that the GCe and ANXA1 are relevant to the granulopoiesis and the traffic of neutrophils from bone marrow under physiological conditions and in the presence of inflammation. Furthermore, together with literature data, the data presented here may suggest the involvement of ANXA1 the GCe in phenotypic plasticity of neutrophils according to the stimuli that are submitted, and may support to understand the new concepts of production, half-life, location and function of neutrophils.

Acinetobacter baumanii fait aujourd’hui partie des bactéries les plus problématiques dans le monde. Responsable de nombreux pics épidémiques d’infections nosocomiales auxquelles sont associés de forts taux de mortalité, cette bactérie puise sa pathogénie dans de multiples caractéristiques qui lui permettent ainsi d’échapper au système immunitaire de l’hôte et à la plupart des traitements actuels. Capable d’adhérer à de multiples surfaces, A. baumanii persiste dans l’environnement hospitalier à travers un mode de vie communautaire au sein duquel ses capacités de survie sont exacerbées. Chez les espèces du genre Acinetobacter, le mode de vie communautaire peut prendre deux formes distinctes : le biofilm et la pellicule. Dans la première partie de cette thèse, nous avons cherché à discriminer ces deux modes de vie, chez la souche ATCC 17978, par une analyse protéomique à large échelle. Nous avons confirmé la présence de nombreux marqueurs communs aux deux communautés (transporteurs, systèmes de sécrétion, d’acquisition d’ions, adhésines et pili) et mis en exergue des systèmes spécifiquement reliés à la formation du biofilm (pilus Fim, T2SS, T1SS/pompe A1S_0535-38, LPS/LOS, motif capsulaire) et à celle de la pellicule (Gac). Grâce à l’étude de la souche A. baumannii SDF en mode biofilm, qui présente un génome plus compact, nous montrons que très peu de mécanismes moléculaires sont partagés par les deux souches étudiées. Ce résultat témoigne de la difficulté quant au développement d’un traitement dirigé contre les biofilms A. baumannii. Dans une deuxième partie, nous avons testé deux approches pour prévenir et éradiquer les biofilms à A. baumannii. La première a ciblé le Quorum Sensing (QS), système de communication essentielle à la coordination des cellules. Nous avons pu montrer que les acides gras mono-insaturés (acide palmitoléique et acide myristoléique), au même titre que la virstatine, limitait la formation de communautés à A. baumannii en inhibant l’expression du régulateur abaR nécessaire au QS. Dans une seconde stratégie, nous avons finalement évalué l’action antibactérienne et antibiofilm d’un nouveau composé d’origine naturelle : la squalamine. Dans cette étude, nous montrons pour la première fois qu’A. baumannii est capable d’entrer dans un état de dormance (persistant/VBNC) pour survivre à de fortes doses de ciprofloxacine, mais que la squalamine est capable d’éradiquer ces cellules persistantes grâce à des concentrations inférieures à la concentration hémolytique. / Today, Acinetobacter baumannii is one of the most problematic pathogens in the world. This bacterium is responsible for worldwide epidemic outbreaks associated with dramatic mortality rates. It possesses high capacities to evade the immune host system and to resist to numerous available antibacterial agents. A. baumannii is also able to persist into hospital environment due to high adhesion abilities which induce community development. This process is also associated to an enhanced survival rate. In Acinetobacter genus, community modes of lif can take two forms : biofilm and pellicle. In this study on the strain ATCC 17978, we tried to discriminate these two lifestyles by a large scale proteomic analysis. We have confirmed the presence of many common community markers (transporters, ion acquisition secretion systems, adhesins and pili) and highlighted systems specifically related to biofilm (pilus Fim, T2SS, T1SS / pump A1S_0535-38, LPS / LOS, capsular pattern) and pellicle communities. Furthermore the proteomic analysis of an avirulent A. baumannii strain, SDF, in biofilm allowed to highlight peculiar metabolic pathways, specific adhesion determinants but very few markers shared by ATCC 17978. This demonstrated the difficulty in developing a treatment directed against A. baumannii biofilm. Then, we tested different approaches to prevent and eradicate biofilms. The first one targeted the Quorum Sensing system (QS), an essential communication system for cell coordination. We have showed that monounsaturated fatty acids (palmitoleic acid and myristoleic acid), like virstatin prevent the community formation of A. baumannii by inhibiting the expression of the abaR regulator required for QS. In a second strategy, we have evaluated the antibacterial and antibiofilm activity of a new natural compound : the squalamine. We showed for the first time that if ciprofloxacin treatment was able to induce a dormancy population (persistent/VBNCs) in A. baumannii, squalamine was able to eradicate this population of dormant cells.

Le glioblastome (GBM) est la forme de cancer du cerveau la plus courante et la plus meurtrière. Sa nature diffusive entraine une impossibilité d’élimination complète par chirurgie. Une récidive de la tumeur chez ≥ 90% des patients peut être provoqué par des cellules GBM résiduelles se trouvant près du bord de la cavité de résection. Un implant pouvant libérer de manière durable la protéine SDF-1α, qui se lie aux récepteur CXCR4 à la surface des cellules GBM, peut être utile pour induire le recrutement des cellules GBM résiduelles, permettre leur élimination sélective et finalement réduire la récurrence de la tumeur. Dans ce travail, le SDF-1α a été initialement encapsulé dans des nanoparticules à base d'acide poly-lactique-co-glycolique (PLGA). Une efficacité d'encapsulation élevée (76%) a pu être obtenue en utilisant un processus simple de séparation de phase. Les nanoparticules chargées de SDF-1α ont ensuite été incorporées dans un scaffold à base de chitosan par électrofilage pour obtenir des implants nanofibreux imitant la structure de la matrice extracellulaire du cerveau. Une étude de libération in vitro a révélé que l'implant pouvait fournir une libération prolongée de SDF-1α jusqu'à 35 jours, utile pour établir un gradient de concentration de SDF-1α dans le cerveau et induire une attraction des cellules GBM. Une étude de biocompatibilité in vivo à 7 jours a révélé des signes d'inflammation locale sans aucun signe visible de détérioration clinique chez les sujets animaux. Une étude à 100 jours visant à confirmer l'innocuité in vivo des implants avant de passer aux études d'efficacité dans un modèle de résection GBM approprié est actuellement en cours. / Glioblastoma (GBM) is the most common and lethal form of brain cancer. The diffusive nature of GBM means the neoplastic tissue can not be removed completely by surgery. Often, residual GBM cells can be found close to the border of the resection cavity and these cells can multiply to cause tumor recurrence in ≥90% of GBM patients. An implant that can sustainably release chemoattractant molecules called stromal cell-derived factor-1α (SDF-1α), which bind selectively to CXCR4 receptors on the surface of GBM cells, may be useful for inducing chemotaxis and recruitment of the residual GBM cells. This may then give access to selective killing of the cells and ultimately reduce tumor recurrence. In this work, SDF-1α was initially encapsulated into poly-lactic-coglycolicacid (PLGA)-based nanoparticles. A high encapsulation efficiency (76%) could be achieved using a simple phase separation process. The SDF-1α-loaded nanoparticles were then incorporated into a chitosan-based scaffold by electrospinning to obtain nanofibrous implants that mimic the brain extracellular matrix structure. In vitro release study revealed that the implant could provide sustainedSDF-1α release for 5 weeks. The gradual SDF-1αrelease will be useful for establishing SDF-1α concentration gradients in the brain, which is critical for the chemotaxis of GBM cells. A 7-day in vivo biocompatibility study revealed evidence of inflammation at the implantation site without any visible signs of clinical deterioration in the animal subjects. A long-term study (100 days) aiming to confirm the in vivo safety of the implants before proceeding to efficacy studies in a suitable GBM resection model is currently underway.

Die vorliegende Arbeit beinhaltet die Generierung und Evaluation von Natürlichen Killerzellen, die EGFRvIII-positive und SDF-1alpha sekretierende primäre Glioblastomzellen aufspüren, erkennen und effizient abtöten können. Die Kombination der gelenkten Zytotoxizität mit einer optimierten Migration von Effektorzellen des Immunsystems wird auf Grundlage der in dieser Arbeit gewonnenen Daten als ein vielversprechender Ansatz für eine zukünftige Therapie des primären Glioblastoms vorgeschlagen.

Cette thèse propose un nouveau paradigme de représentation d’applications pour les machines virtuelles, capable d’abstraire l’architecture des systèmes informatiques. Les machines virtuelles actuelles reposent sur un modèle unique de représentation d’application qui abstrait les instructions des machines et sur un modèle d’exécution qui traduit le fonctionnement de ces instructions vers les machines cibles. S’ils sont capables de rendre les applications portables sur une vaste gamme de systèmes, ces deux modèles ne permettent pas en revanche d’exprimer la concurrence sur les instructions. Or, celle-ci est indispensable pour optimiser le traitement des applications selon les ressources disponibles de la plate-forme cible. Nous avons tout d’abord développé une représentation « universelle » d’applications pour machine virtuelle fondée sur la modélisation par graphe flux de données. Une application est ainsi modélisée par un graphe orienté dont les sommets sont des unités de calcul (les acteurs) et dont les arcs représentent le flux de données passant au travers de ces sommets. Chaque unité de calcul peut être traitée indépendamment des autres sur des ressources distinctes. La concurrence sur les instructions dans l’application est alors explicite. Exploiter ce nouveau formalisme de description d'applications nécessite de modifier les règles de programmation. A cette fin, nous avons introduit et défini le concept de « Représentation Canonique et Minimale » d’acteur. Il se fonde à la fois sur le langage de programmation orienté acteur CAL et sur les modèles d’abstraction d’instructions des machines virtuelles existantes. Notre contribution majeure qui intègre les deux nouvelles représentations proposées, est le développement d’une « Machine Virtuelle Universelle » (MVU) dont la spécificité est de gérer les mécanismes d’adaptation, d’optimisation et d’ordonnancement à partir de l’infrastructure de compilation Low-Level Virtual Machine. La pertinence de cette MVU est démontrée dans le contexte normatif du codage vidéo reconfigurable (RVC). En effet, MPEG RVC fournit des applications de référence de décodeurs conformes à la norme MPEG-4 partie 2 Simple Profile sous la forme de graphe flux de données. L’une des applications de cette thèse est la modélisation par graphe flux de données d’un décodeur conforme à la norme MPEG-4 partie 10 Constrained Baseline Profile qui est deux fois plus complexe que les applications de référence MPEG RVC. Les résultats expérimentaux montrent un gain en performance en exécution de deux pour des plates-formes dotées de deux cœurs par rapport à une exécution mono-cœur. Les optimisations développées aboutissent à un gain de 25% sur ces performances pour des temps de compilation diminués de moitié. Les travaux effectués démontrent le caractère opérationnel et universel de cette norme dont le cadre d’utilisation dépasse le domaine vidéo pour s’appliquer à d’autres domaine de traitement du signal (3D, son, photo…) / This thesis proposes a new paradigm that abstracts the architecture of computer systems for representing virtual machines’ applications. Current applications are based on abstraction of machine’s instructions and on an execution model that reflects operations of these instructions on the target machine. While these two models are efficient to make applications portable across a wide range of systems, they do not express concurrency between instructions. Expressing concurrency is yet essential to optimize processing of application as the number of processing units is increasing in computer systems. We first develop a “universal” representation of applications for virtual machines based on dataflow graph modeling. Thus, an application is modeled by a directed graph where vertices are computation units (the actors) and edges represent the flow of data between vertices. Each processing units can be treated apart independently on separate resources. Concurrency in the instructions is then made explicitly. Exploit this new description formalism of applications requires a change in programming rules. To that purpose, we introduce and define a “Minimal and Canonical Representation” of actors. It is both based on actor-oriented programming and on instructions ‘abstraction used in existing Virtual Machines. Our major contribution, which incorporates the two new representations proposed, is the development of a “Universal Virtual Machine” (UVM) for managing specific mechanisms of adaptation, optimization and scheduling based on the Low-Level Virtual Machine (LLVM) infrastructure. The relevance of the MVU is demonstrated on the MPEG Reconfigurable Video Coding standard. In fact, MPEG RVC provides decoder’s reference application compliant with the MPEG-4 part 2 Simple Profile in the form of dataflow graph. One application of this thesis is a new dataflow description of a decoder compliant with the MPEG-4 part 10 Constrained Baseline Profile, which is twice as complex as the reference MPEG RVC application. Experimental results show a gain in performance close to double on a two cores compare to a single core execution. Developed optimizations result in a gain on performance of 25% for compile times reduced by half. The work developed demonstrates the operational nature of this standard and offers a universal framework which exceeds the field of video domain (3D, sound, picture...)