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The triple transgenic (3xTg-AD) mouse model of Alzheimer’s disease (AD) possesses three transgenes that lead to the development of amyloid-beta (A?) plaques (APPswe, PS1M146V) and neurofibrillary tangles (and tauP301L) (Oddo et al., 2003b). Although the neuropathology of these mice has been extensively studied (Sy et al., 2011), less research has been done to investigate their working memory, emotionality, and locomotor-related behaviour. Using a cross-sectional design, male and female 3xTg-AD mice were compared to control mice (B6129SF2/J) at five ages (2-, 6-, 9-, 12-, and 15-months of age) on a battery of five tests designed to measure: anxiety- and locomotor-related behaviours (open field [OF], elevated plus maze [EPM]); depression (forced swim test [FST]); motor coordination and motor learning (rotarod); and working and reference memory (8-arm radial maze [RAM]). Additionally, the brain tissue of male and female 3xTg-AD and control mice at 2- and 15-months of age was analyzed for the presence of A? plaques and human tauP301L. 3xTg-AD mice were found to travel less and freeze more in both the OF and the EPM, engage in fewer bouts of immobility in the FST, have a longer latency to fall on the rotarod, and make more working and reference memory errors in the RAM than controls. There was no effect of age on performance in any of the tests. Intracellular A? plaques and limited human tau were present in the brain tissue of 2-month old 3xTg-AD mice. At 15-months of age, the brain tissue of 3xTg-AD mice showed extensive intra- and extracellular A? plaques as well as tauP301L staining. The presence of intracellular A? at 2-months of age supports the behavioural differences observed in the 3xTg-AD mice at 2-months of age. However, the lack of progressive behavioural change does not match the increase in neuropathology seen in the brains of the 15-month old 3xTg-AD mice. The results of the present study suggest that while the 3xTg-AD mice display similar neuropathology and some of the behavioural differences seen in individuals with AD, they also exhibit contradictory behaviours; findings that should be taken into consideration for future researchers using 3xTg-AD mice.

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Measles is one of the most contagious diseases known to mankind. Despite the availability of a safe and effective vaccine, approximately 164,000 measles-related deaths were recorded in 2008. The inherent restricted host tropism of MV means that the development of authentic rodent models will be a valuable research tool in testing new vaccines and antivirals. In addition to the receptor requirement, mouse innate immunity has been shown to inhibit MV growth. In this thesis, the contributions of several key components of the mouse innate immune system on the inhibition of MV replication were examined. The transcription factor interferon regulatory factor 3 (IRF3), which normally plays a key role in mediating innate immune signaling, contributed relatively little in inhibiting MV replication both in vitro and in vivo. In contrast, the JAK/STAT pathway and the double-stranded RNA inducible protein kinase, PKR, played more important roles in controlling virus replication. The resurgence of measles in areas where the virus was once thought to be eradicated makes the development of anti-MV treatments essential. Concurrent to the development of an animal model to better study its pathogenesis, we wanted to look at the effect of MV inhibitors on its replication. The MV fusion inhibitor, carbobenzoxy-D-phenylalanine-L-phenylalanine-glycine (ZfFG), was developed in the past to study fusion; however, its mechanism of action has not yet been elucidated. To examine this, spontaneous ZfFG-resistant mutants were generated and characterized. Mutations were found in the HRB region of the fusion (F) protein, and when these were modeled using published paramyxovirus F crystal structures, data suggested that ZfFG targeted a small pocket present between the head and stalk regions of its pre-fusion conformation. An authentic mouse model of measles developed from findings in this study may allow for in vivo efficacy testing of ZfFG in the future.

Measles is one of the most contagious diseases known to mankind. Despite the availability of a safe and effective vaccine, approximately 164,000 measles-related deaths were recorded in 2008. The inherent restricted host tropism of MV means that the development of authentic rodent models will be a valuable research tool in testing new vaccines and antivirals. In addition to the receptor requirement, mouse innate immunity has been shown to inhibit MV growth. In this thesis, the contributions of several key components of the mouse innate immune system on the inhibition of MV replication were examined. The transcription factor interferon regulatory factor 3 (IRF3), which normally plays a key role in mediating innate immune signaling, contributed relatively little in inhibiting MV replication both in vitro and in vivo. In contrast, the JAK/STAT pathway and the double-stranded RNA inducible protein kinase, PKR, played more important roles in controlling virus replication. The resurgence of measles in areas where the virus was once thought to be eradicated makes the development of anti-MV treatments essential. Concurrent to the development of an animal model to better study its pathogenesis, we wanted to look at the effect of MV inhibitors on its replication. The MV fusion inhibitor, carbobenzoxy-D-phenylalanine-L-phenylalanine-glycine (ZfFG), was developed in the past to study fusion; however, its mechanism of action has not yet been elucidated. To examine this, spontaneous ZfFG-resistant mutants were generated and characterized. Mutations were found in the HRB region of the fusion (F) protein, and when these were modeled using published paramyxovirus F crystal structures, data suggested that ZfFG targeted a small pocket present between the head and stalk regions of its pre-fusion conformation. An authentic mouse model of measles developed from findings in this study may allow for in vivo efficacy testing of ZfFG in the future.

Le syndrome de Down (SD), ou Trisomie 21, est l’aneuploïdie la plus fréquente chez l’humain. Le désordre génomique est tel qu’aucun traitement unique ne peut pallier à tous les symptômes (retard mental, troubles moteurs...). C’est pourquoi l’utilisation de modèles murins permet d’étudier l’impact de régions partielles du Hsa21 dans l’apparition des déficits. Mon projet de thèse s’est orienté sur un locus télomérique encadré par les gènes Abcg1 et U2af1. Mes recherches se sont focalisées sur deux modèles, Ts1Yah et Ms2Yah, dédiés à cette région. L’étude de ces lignées, combinées à d’autres modèles transgéniques, a montré la contribution de l’intervalle génique dans l’optimisation de l’apprentissage locomoteur. Dans un deuxième temps, le gène Cbs, candidat à la perte de fonction de mémoire, a permis de mettre en évidence un sauvetage fonctionnel dans une expérience à effets de doses. Cette découverte ouvre la voie à de nouvelles perspectives thérapeutiques. / The Down syndrome (DS), or Trisomy21, is the most frequent aneuploidy in human. The genomic disorder is such an extent that any single cure could overcome all the symptoms (mental retardation, motor defects…). That’s why mouse models’ using allows studies on Hsa21 regions impact in deficits apparition. My PhD project was turned toward the telomeric locus of the Hsa21 framed between Abcg1 and U2af1. We focused our research on two models, Ts1Yah and Ms2Yah, dedicated to this region. The study of those lines, combined with several other transgenic ones, showed the contribution of the interval into the optimisation of motor learning. In a second step, the Cbs gene, candidate on memory loss of function, has highlighted a functional rescue in dose-effect experiment. This discovery paves the way to new therapeutic perspectives.

L’adénocarcinome canalaire du pancréas (PDAC) est un des cancers les plus agressifs avec un taux de survie à 5 ans de moins de 5 %. Une des raisons est l’absence de traitement thérapeutique efficace. Des efforts afin d’identifier de nouvelles cibles pour le traitement du PDAC sont donc nécessaires. Il a été démontré que la dénervation du pancréas régule la progression des PDAC dans des modèles murins. De plus, on a rapporté que les axones du système nerveux périphérique (SNP) innervent les tumeurs pancréatiques, mais l'identité précise des fibres infiltrant la tumeur est inconnue.Ici, nous avons caractérisé le remodelage des principales divisions du SNP, y compris les systèmes autonomes et sensoriels, dans des modèles murins qui récapitulent la maladie humaine. Nous avons aussi commencé à caractériser l'innervation des PDAC dans des échantillons humains. Nous avons observé une augmentation de la densité des fibres sympathiques positives pour la tyrosine hydroxylase (TH) dans les lésions pré-tumorales du pancréas, alors qu'une forte densité de fibres sensorielles positives pour le peptide lié au gène de la calcitonine (CGRP) a été observée dans les PDAC. Fait intéressant, alors que dans tissus normaux les axones sympathiques et sensoriels sont principalement associés aux vaisseaux sanguins, ils sont majoritairement isolés dans les lésions pré-tumorales et les PDAC. Ces données suggèrent que la plasticité axonale survient aux stades précoces du développement tumoral pour les fibres sympathiques et à un stade plus tardif pour les fibres sensorielles. Ce travail suggère de nouvelles cibles potentielles pour le traitement des PDAC. / Pancreatic ductal adenocarcinoma (PDAC) is one of the most lethal cancers with 5-year survival rate of less than 5%. One reason to explain this poor outcome is that there has been no effective therapeutic treatment for PDAC patients. Thus, efforts to identify novel targets for PDAC treatment are required. Denervation of the pancreas has been shown to regulate PDAC progression in murine models. In addition, axons of the peripheral nervous system (PNS) have been reported to innervate pancreatic tumors, but the precise identity of the tumor-infiltrating fibers is unknown. Here we characterized the remodeling of the main divisions of the PNS, including autonomic and sensory systems, in mouse models, which recapitulate the human disease. We also started to characterize the innervation of human PDAC samples. We observed an increased density of tyrosine hydroxylase (TH)-positive sympathetic fibers in pre-tumoral lesions of the pancreas, while a high density of calcitonin gene-related peptide (CGRP)-positive sensory fibers was seen within PDAC. Interestingly, whereas in the normal tissues TH+ and CGRP+ axons were mostly associated to blood vessels, they were mainly isolated in lesions and PDAC. These data suggest that axonal plasticity occurs at the early stage of tumor development for sympathetic fibers and at the late stage for sensory fibers. This work suggests potential novel targets for the treatment of PDAC.

Les maladies auto-immunes, qui touchent plus de 5% de la population, sont induites par une perte de la tolérance aux antigènes du Soi. Ces pathologies, généralement multifactorielles, résultent de l’effet combiné de plusieurs allèles de susceptibilité et de différents facteurs environnementaux. Les agents infectieux ont été tout particulièrement incriminés, mais les mécanismes en jeu restent encore mal élucidés. Les lymphocytes B, qui jouent un rôle central dans la pathogénie de nombreuses maladies auto-immunes, sont susceptibles d’être activés selon différents mécanismes au cours d’un processus infectieux et cette activation peut englober des cellules autoréactives. On ne sait cependant pas si cette activation peut entraîner la production d’auto-anticorps pathogènes de forte affinité et d’isotype IgG à partir du pool de cellules productrices d’auto-anticorps naturels de faible affinité, qui sont présentes de façon constitutive dans le répertoire B de l’individu sain. Nous avons mis au point un nouveau modèle murin knock-in pour des lymphocytes B présentant une affinité intermédiaire pour leur auto-antigène, la protéine HEL2X mutée (Hen-Egg Lysozyme). Ce modèle autoréactif d’affinité intermédiaire SWHEL X HEL2X, élaboré sur un fond génétique non autoimmun, permet de suivre le processus de maturation d’affinité des cellules B anti-HEL en présence de leur auto-antigène HEL2X au cours de l’infection chronique par la bactérie Borrelia burgdorferi. L’infection induit au niveau ganglionnaire une prolifération ainsi qu’une activation lymphocytaire B incluant des cellules anergiques. Certains clones autoréactifs sont capables de gagner les centres germinatifs ganglionnaires, de commuter vers l’isotype IgG et présentent des mutations somatiques au niveau de la région variable de la chaîne lourde de leur immunoglobuline, dans la zone d’interaction avec HEL2X, indiquant un processus de sélection par l’auto-antigène. Malgré un taux augmenté d’auto-anticorps d’isotype IgM, ces animaux ne produisent pas de plasmocytes capables de sécréter des auto-anticorps d’isotype IgG. Nos observations suggèrent l’existence de mécanismes de tolérance périphérique intrinsèques mis en place en particulier au niveau du centre germinatif. Un premier point de contrôle va éliminer les lymphocytes B autoréactifs ayant commuté de classe et présentant des mutations somatiques leur conférant une affinité augmentée pour l’auto-antigène tandis qu’un second point de contrôle va empêcher la différenciation en plasmocytes IgG+.Chez l’individu non prédisposé génétiquement, des mécanismes pourraient ainsi permettre de prévenir le développement d’une auto-immunité pathogène au cours d’un épisode infectieux. / Autoimmune diseases, affecting more than 5% of the population, reflect a loss of tolerance to selfantigens. These multifactorial diseases result from the combined effect of several susceptibility alleles and different environmental factors. Infectious agents have been particularly incriminated but there is no clear understanding of the underlying mechanisms. B lymphocytes, that appear central to the pathogenesis of several autoimmune diseases, may be activated by several mechanisms during infectious processes and this activation can encompass autoreactive cells. Whether or not the lattercan induce the production of high-affinity pathogenic IgG isotype auto-antibodies from the naturally present low-affinity self-reactive B cells is still unknown. To gain further insight into this question, we created a new intermediate affinity autoreactive mouse model called SWHEL X HEL2X. In these mice, knock-in B cells express a B cell receptor highly specific for Hen-Egg Lysozyme (HEL) that recognizes HEL2X mutated auto-antigen with intermediate affinity. This model, generated on a non-autoimmune-prone genetic background, allows the following of anti-HEL B cells affinity maturation process in presence of their auto-antigen during Borrelia burgdorferi chronic bacterial infection. The infection leads to lymph nodes lymphoproliferation and B cell activation including anergic cells. Some autoreactive clones are able to form germinal centers, toswitch their immunoglobulin heavy chain and to introduce somatic mutations in the heavy chain variable regions on amino-acids forming direct contacts with HEL2X, suggesting an auto-antigen-driven selection process. Despite increased levels of IgM autoantibodies, infected mice are unable to generate IgG autoantibody secreting plasma-cells. These observations suggest the existence of intrinsic peripheral tolerance mechanisms operating mainly at the level of germinal centers. The first checkpoint eliminates switched autoreactive B cells with increasing affinity mutations while a secondcheckpoint avoids IgG+ plasma-cell differentiation. Thus, in genetically non predisposed individuals, tolerance mechanisms may be set-up to prevent the development of pathogenic autoimmunity during the course of an infection.

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