Background Histone methylation is thought to be central to the epigenetic mechanisms that maintain and confine cellular identity in multi-cellular organisms. To examine epigenetic roles in cellular homeostasis, we conditionally mutated the histone 3 lysine 4 methyltransferase, Mll2, in embryonic stem (ES) cells, during development and in adult mice using tamoxifen-induced Cre recombination. Results In ES cells, expression profiling unexpectedly revealed that only one gene, Magoh2, is dependent upon Mll2 and few other genes were affected. Loss of Mll2 caused loss of H3K4me3 at the Magoh2 promoter and concomitant gain of H3K27me3 and DNA methylation. Hence Mll2, which is orthologous to Drosophila Trithorax, is required to prevent Polycomb-Group repression of the Magoh2 promoter, and repression is further accompanied by DNA methylation. Early loss of Mll2 in utero recapitulated the embryonic lethality found in Mll2-/- embryos. However, loss of Mll2 after E11.5 produced mice without notable pathologies. Hence Mll2 is not required for late development, stem cells or homeostasis in somatic cell types. However it is required in the germ cell lineage. Spermatogenesis was lost upon removal of Mll2, although spermatogonia A persisted. Conclusion These data suggest a bimodal recruit and maintain model whereby Mll2 is required to establish certain epigenetic decisions during differentiation, which are then maintained by redundant mechanisms. We also suggest that these mechanisms relate to the epigenetic maintenance of CpG island promoters.

The goal of this Master's thesis was to solve resource allocation problems in wireless networks through the implementation of a lightweight simulation platform. The spectrum and power resources of wireless networks have to be efficiently used to accommodate the growing number of wireless terminals and the massive increase of data transferred by their applications. The major problem that needs to be tackled is interference, which significantly limits the performance of wireless systems. In this thesis, the resource allocation of interest was the joint problem of scheduling and power control with Quality of Service (QoS) constraints. The Signal-to-Interference-plus-Noise Ratio (SINR) was used to quantify QoS. This thesis studied the recently proposed mixed-integer linear programming (MILP) formulation of the problem. Due to the scheduling component, the problem is inherently combinatorial and NP-hard, therefore computationally expensive and difficult to solve in tractable time. A simulation platform was implemented in order to automate and facilitate the solving process.As a starting point, wireless channels and channel modeling issues were studied. Then, the platform was implemented to simulate random instances of multi-cellular wireless networks, with several mobile stations per cell, and generate the corresponding channels. Finally, the platform was extended to use the GNU Linear Programming Kit (GLPK) API in order to optimally solve the aforementioned formulated problem for various inputs of generated channels.Tests of the simulation platform were performed to check the consistency of the results. Indeed, the output results satisfied the initial expectations regarding the SINR constraints and the formulation. Moreover, they were produced in reasonable time. An analysis of the output results was presented.This thesis resulted in a configurable and lightweight simulation platform which is able to solve the MILP-formulated resource allocation problem. The simulation platform is basic and does not cover all the aspects of multi-cellular wireless networks and wireless channels. Due to its modularity, it can be extended in a future project.

Within solid tumours, a microenvironment exists that causes resistance to chemotherapy. New drugs that target cells within this microenvironment are required, the first step in this process being the identification of new targets. The aim of this thesis was to characterise changes in the transcriptome and proteome within specific regions of multicell-tumour spheroids (MCTS), an experimental model that mimics many of the features of the tumour microenvironment. HT29 MCTS were separated by sequential trypsinisation into 3 main regions; the outer surface layer (SL) the peri-necroric region (PN) and the necrotic core (NC). Using an iTRAQ quantitative proteomics approach, the proteome of the different MCTS regions was investigated. A 2 dimensional separation approach using Agilent's OffGel system and RP-nano HPLC was incorporated prior to MS analysis. MS analysis was done using both MALDI-TOF-TOF (Bruker Ultraflex II) and ESI-Q-TOF (Agilent 6530 QTOF LC/MS) instruments. Gene expression profiles of the different MCTS were investigated and compared using Agilent's one-color oligonucleotide based microarrays. Transcriptomic and proteomic analysis identified several key differences in the proteins involved in cell metabolism between the SL and PN/NC regions. Similar metabolic changes were also noted between autophagic and normal monolayer cells. Many highlighted proteins represented established cancer associated proteins. Interestingly, a number of proteins were highlighted which have no previous association with cancer and may upon further validation, provide attractive leads for therapeutic intervention.

Background Histone methylation is thought to be central to the epigenetic mechanisms that maintain and confine cellular identity in multi-cellular organisms. To examine epigenetic roles in cellular homeostasis, we conditionally mutated the histone 3 lysine 4 methyltransferase, Mll2, in embryonic stem (ES) cells, during development and in adult mice using tamoxifen-induced Cre recombination. Results In ES cells, expression profiling unexpectedly revealed that only one gene, Magoh2, is dependent upon Mll2 and few other genes were affected. Loss of Mll2 caused loss of H3K4me3 at the Magoh2 promoter and concomitant gain of H3K27me3 and DNA methylation. Hence Mll2, which is orthologous to Drosophila Trithorax, is required to prevent Polycomb-Group repression of the Magoh2 promoter, and repression is further accompanied by DNA methylation. Early loss of Mll2 in utero recapitulated the embryonic lethality found in Mll2-/- embryos. However, loss of Mll2 after E11.5 produced mice without notable pathologies. Hence Mll2 is not required for late development, stem cells or homeostasis in somatic cell types. However it is required in the germ cell lineage. Spermatogenesis was lost upon removal of Mll2, although spermatogonia A persisted. Conclusion These data suggest a bimodal recruit and maintain model whereby Mll2 is required to establish certain epigenetic decisions during differentiation, which are then maintained by redundant mechanisms. We also suggest that these mechanisms relate to the epigenetic maintenance of CpG island promoters.

Within solid tumours, a microenvironment exists that causes resistance to chemotherapy. New drugs that target cells within this microenvironment are required, the first step in this process being the identification of new targets. The aim of this thesis was to characterise changes in the transcriptome and proteome within specific regions of multicell-tumour spheroids (MCTS), an experimental model that mimics many of the features of the tumour microenvironment. HT29 MCTS were separated by sequential trypsinisation into 3 main regions; the outer surface layer (SL) the peri-necroric region (PN) and the necrotic core (NC). Using an iTRAQ quantitative proteomics approach, the proteome of the different MCTS regions was investigated. A 2 dimensional separation approach using Agilent¿s OffGel system and RP-nano HPLC was incorporated prior to MS analysis. MS analysis was done using both MALDI-TOF-TOF (Bruker Ultraflex II) and ESI-Q-TOF (Agilent 6530 QTOF LC/MS) instruments. Gene expression profiles of the different MCTS were investigated and compared using Agilent¿s one-color oligonucleotide based microarrays. Transcriptomic and proteomic analysis identified several key differences in the proteins involved in cell metabolism between the SL and PN/NC regions. Similar metabolic changes were also noted between autophagic and normal monolayer cells. Many highlighted proteins represented established cancer associated proteins. Interestingly, a number of proteins were highlighted which have no previous association with cancer and may upon further validation, provide attractive leads for therapeutic intervention.

The liver is responsible for lipid and glucose metabolism, protein and bile synthesis and the biotransformation of xenobiotics. These functions, performed by hepatocytes, are dependent on heterotypic interactions with other liver cell types and the stratified microarchitecture of the organ. In vitro liver models provide insights into the role of each cell type and perturbations upon external stimuli. Despite the dissimilarities to in vivo and rapid dedifferentiation, most liver studies utilize hepatocyte monocultures. These models lack heterotypic interactions causing inaccurate assessments of toxicity and disease. Only a limited number of 3D hepatic models incorporate the major liver cell types, and these cultures primarily focus on the hepatocyte response. We have developed 3D liver models that include all major hepatic cell types and recapitulate the layered architecture of the organ. These models maintain hepatic functions for up to four weeks and can be used to isolate the role and response of each cell type. We used these models to study two critical aspects of the organ -- acute hepatotoxicity and liver fibrosis. There are tens of thousands of chemicals with undetermined effects on the human body. High concentrations of xenobiotics can cause acute liver damage and failure. Liver impairment can result in multiple organ failure, hepatic encephalopathy and death. Therefore, it becomes critically important to investigate hepatotoxicity in a time, cost and resource effective manner. Our 3D liver models were validated for hepatotoxicity testing with acetaminophen, a prototypic drug. We then adapted and optimized the models for high-throughput hepatotoxicity testing with automated procedures and primary human hepatic cells. Liver fibrosis and cirrhosis are well-established consequences of chronic chemical exposure, infection and alcoholism. The initiating factors, end stages and resolution of fibrosis have been extensively studied. However, there is minimal information on the role of the local microenvironment in the progression of the disease from diseased to healthy tissue. We designed 3D liver cultures with a mechanical gradient to gradually model this transition through spatial and temporal perspectives. These findings demonstrate the versatility and accuracy of these 3D hepatic models in the investigation of liver toxicity and fibrosis. / Ph. D.

No description available.

Dans le but de gérer la consommation et ainsi que la production efficace des énergies renouvelables, l’utilisation et augmentation de l’efficacité des systèmes de conversion d’énergie est devenue indispensable. Dans ce contexte, les transistors en nitrure de gallium (GaN HFETs) pour une densité de puissance commutée très importante, offrent des nouvelles possibilités et ils tirent vers le haut la gestion efficace des énergies renouvelables. Toutefois, cette nouvelle possibilité passe par un pilotage efficace des composants et une encapsulation et des interconnexions optimales. Ces travaux de thèse étudient et analyse les avantages et inconvénients d'une nouvelle structure de conversion multi cellulaire et ceux d’un driver spécifique multivoies monolithique et synchronisé pour les composants GaN, appliqué dans ce contexte précis. Ce manuscrit de thèse est composé de quatre chapitres. Après une étude bibliographique, le positionnement du convertisseur DAB (Dual Active Bridge) parmi les autres structures de conversion DC à l’aide d’une nouvelle méthodologie de comparaison (FOM topologique – Figure de Mérite) est présenté dans le premier chapitre. Afin de diminuer les contraintes de conversion d’énergie, une étude est amenée dans le chapitre 2 sur les principaux défis et enjeux d’une solution générique à travers de réalisation d’un réseau de convertisseurs. Le chapitre 3 présente la partie importante d’expérimentation et d’optimisation de la cellule élémentaire à base de convertisseurs DAB des points de vue fonctionnels mais aussi et surtout structurels. L’étude de l’isolation galvanique de la structure de conversion DAB reste l'objectif principal à développer pour démontrer le potentiel de remplacer le transformateur par une isolation capacitive. La conception de la puce de commande dédiée aux nouveaux transistors GaNs, les résultats pratiques des performances sont présentés dans le dernier chapitre. Certaines comparaisons du driver QGD (Quad Gate Driver) avec les autres solutions de transfert d’ordres de commande sont également discutées. La mise en œuvre du circuit de commande dans un convertisseur DAB afin de valider le fonctionnement de QGD est introduite dans les perspectives. / In order to improve the management and the production efficiency in renewable energy, power electronics systems have become important contributors. In this context, gallium nitride transistors (GaN HFETs) provide new opportunities for high power density, high switching speed and they pull up the effective management of renewable energies. However, this new opportunity requires effective gate drivers and optimal packaging and assembly. This thesis will introduce the general approach of a new architecture for multi cellular conversion and a monolithic multichannel and synchronized drive for GaN components, which are applicable in our specific context. This thesis is composed of four chapters. A bibliographic section is presented in first chapter. A new comparative methodology has been developed in this chapter in order to benchmark the DAB (Dual Active Bridge) converter with respect to other DC converters. In the second chapter, a generic solution (converter grid) has been explained in order to reduce the energy conversion constraints. Chapter 3 presents the important parts of the experimentation and the optimization of DAB converter. High frequency transformer replacing by capacitors is the main objective of this section. The design of the Quad Gate Driver (QGD) IC dedicated to GaNs transistors control in H bridge configuration and the results of their performances are presented in the last chapter. Some comparisons of this approach with other signal transfer solutions are also discussed. The implementation of the QGD in a full bridge transistor converter is introduced into the perspective section.

The individual behaviour of prokaryotic organisms such as bacteria often gives rise to complexity that is commonly associated with multicellular behaviour. The transition from unicellular to multicellular behaviour occurs in response to chemical signals, called autoinducers, which bacteria generate and receive internally within a given population. These autoinducers control the gene expression necessary for the emergence of group-behaviour-phenotype. This phenomenon is called quorum sensing (QS). An example of the quorum sensing control of gene regulation has been the luminescence (lux) operon in Vibrio fischeri. The luxI and ainS quorum signalling systems work in conjunction to regulate luminescence in V. fischeri. LuxI and AinS are acyl-synthases that catalyse the production of the autoinducers C6-HSL and C8-HSL respectively. These autoinducers bind to LuxR, a transcriptional activator of the lux operon, which activates expression of the lux genes causing an increase in luminescence. It was shown that quorum signalling also affects motility and biofilm formation in bacteria. However, the evidence with respect to these phenotypes is conflicting and inconclusive, the reason being the state of quorum is ambiguously defined. It is not properly known whether the observed collective behaviour is purely a result of physical crowding of bacteria, or that both chemical signalling and crowding contribute to this phenomenon. This work attempts to address these issues by studying luminescence, motility, and biofilm, a diverse set of behaviours, yet closely linked to each other in V. fischeri-squid symbiosis. We studied the luminescence response of V. fischeri to both endogenous and externally added signals at per-cell and population level. Experiments with ES114, a wild-type strain of V. fischeri, and ainS mutant showed that (i) luminescence per cell does not mutually correlate with the cell-density, indicating that bacteria do not show greater response to the signal at higher densities; (ii) the activity of the lux signalling circuit shows a strong dependence on the growth stage, (iii) the cells do not show enhanced growth, i.e., they do not derive fitness benefits at higher densities in response to the signal. We anticipated that the culture with a higher cell-density should exhibit greater per-cell-luminescence. However, we found that the luminescence curve of the culture with lower density crosses that of the cultures with higher densities during the exponential phase. Kinetic modelling of the luxI mRNA expression showed that the expression profile qualitatively agrees with the luminescence trend observed in the cultures, supporting the observation that growth-phase plays a major role in regulating the luminescence gene expression. We also studied the effect of autoinducers on motility of V. fischeri. V. fischeri uses flagella to move into the inner crypts of the light organ of the squid. The bacterium secretes autoinducers, encounters secretions of the light organ, and slows down during the final stage of colonization process. Studies have shown that flagellar elaboration is repressed as a consequence of ainS signalling. However, those studies were soft-agar migration assays and carried out with the mutant strain of ainS. We measured real-time planktonic motility of ES114 and the signalling mutant strains of V. fischeri in response to autoinducers added exogenously at different concentrations. We found that the autoinducers do not affect the motility of the strains. We also showed that reduction in motility is purely a consequence of physical crowding of bacteria, and chemical signalling may not be involved in the process. It was shown that reduction in motility leads to biofilm formation. Motile bacteria must lose flagella in order to form biofilm, and signalling controls biofilm formation in many species. Our study on motility showed that reduction in motility occurs because of physical crowding in V. fischeri. Hence, we explored the possibility that physical crowding might lead to formation of biofilm rather than signalling in this species. We quantified exopolysaccharide production by crystal violet assay, which revealed that planktonic cells produce exopolysaccharides, in addition to biofilm cells. The study revealed that V. fischeri cells always produce exopolysaccharides irrespective of their physiological state. We examined the effect of signalling on biofilm in ES114 and the mutant strains using gene-expression analysis. We quantified the expression of various genes involved in biofilm formation and found that both ES114 and the mutants expressed rscS and sypP indicating that exopolysaccharide production is not under the control of autoinducers. Therefore, we hypothesized that biofilm formation in V. fischeri may be a result of physical agglomeration of cells. Our observations indicate that the state of quorum is inadequately defined and there is no direct measure of the underlying process. Multicellular behaviour in V. fischeri is regulated by a complex interplay of cell-density, signalling, and other factors such as the growth phase of the culture, indicating that the state of quorum employs different mechanisms to regulate various phenotypes. Our study reveals that QS is an intricate process, and the accepted mechanisms for QS are incomplete at best.