<p></p> <p><strong><em>Background </em></strong></p> <p>Oral pharmacotherapy has the potential to offer multiple sclerosis patients improved clinical outcomes compared to traditional therapies.</p> <p><strong><em>Objectives </em></strong></p> <p>This review assesses the effects of oral therapies compared to placebo and interferon beta-1a in adults with relapsing-remitting multiple sclerosis (RRMS).</p> <p><strong><em> </em></strong></p> <p><strong><em>Search methods </em></strong></p> <p>We searched the MEDLINE, EMBASE, Cochrane Library, Web of Science (January 1980 to April 2011) and clinincaltrials.gov (April 2011) databases and reference lists of articles. The FDA website was also searched.</p> <p><strong><em>Selection criteria </em></strong></p> <p>Double-blind, placebo-controlled, randomized trials of RRMS patients who were treated with fingolimod, cladribine, laquinimod or interferon beta-1a.</p> <p><strong><em>Data collection and analysis </em></strong></p> <p>Two reviewers independently assessed articles for inclusion. Data extraction and quality assessment was completed by one reviewer and verified for accuracy. Meta-analysis and indirect treatment comparison methods were used to estimate relative measures of efficacy.</p> <p><strong><em>Results </em></strong></p> <p>Although 11 trials involving 7,127 participants were included in this review, only 2,109 (30%) and 1,738 (24%) participants contributed to the direct and indirect estimates respectively, for the primary outcome, annualized relapse rate. Oral therapy and interferon beta-1a had a significantly different rate of relapse compared to placebo (Mean difference [MD] -0.21, 95% confidence interval [CI] -0.27 to -0.16 , p < 0.00001 and MD -0.33 95% CI -0.65 to -0.01). There was a significant risk reduction of 37% and 19% in the number of patients with at least one relapse for oral therapy and interferon beta-1a compared to placebo respectively. Safety analysis favoured placebo for both sets of trials (p=0.002 and p=0.04). Indirect estimates were not significant for all three outcomes however; comparability between direct evidence was noted.</p> <p><strong><em>Conclusions </em></strong></p> <p>Oral pharmacotherapy and interferon beta-1a are effective compared to placebo in controlling relapse rate in patients with RRMS. The indirect measures of effect provide initial estimates of comparative efficacy and incorporation of future evidence will be necessary.</p> / Master of Science (MSc)

Les cytokines jouent un rôle fondamental dans la régulation des processus biologiques via la cascade de signalisation JAK-STAT. Les « Suppressors of Cytokine Signalling » (SOCS), protéines intracellulaires, inhibent la voie JAK-STAT. Plusieurs études supportent leur implication dans des maladies immunitaires, mais peu d’informations sont disponibles sur leur expression par les lymphocytes T humains. Nous postulons que les cytokines Interféron-β(IFN-β) et Interleukine-27 (IL-27), dotées d’un potentiel immuno-régulateur, ont des rôles bénéfiques via l’induction des SOCS. L’impact de l’IFN-β et l’IL-27 sur l’expression des SOCS-1 et SOCS-3 par des cellules T CD8 et CD4 humaines a été étudié en utilisant des cellules sanguines de donneurs sains. L’expression de ces régulateurs a été évaluée aux niveaux de l’ARNm par qRT-PCR et protéique par immunocytochimie. Les SOCS-1 et SOCS-3 ont été rapidement induits en ARNm dans les deux types cellulaires en réponse à l’IFN-β ou l’IL-27 et une augmentation de l’expression a été confirmée au niveau protéique. Afin de mimer les thérapies à base d’IFN-β, les cellules T ont été exposées chroniquement à l’IFN-β. Après chaque ajout de cytokine les cellules T ont augmenté l’expression du SOCS-1, sans moduler le SOCS-3. L’IL-27 a induit les SOCS-1 et SOCS-3 préférentiellement dans les cellules T CD8 ; ceci corrèle avec des résultats du laboratoire démontrant une plus petite expression des récepteurs à l’IL-27 par les lymphocytes T CD4 que les CD8. Notre projet a permis d’élucider l’expression des SOCS dans deux populations de cellules T et de clarifier les mécanismes d’actions de l’IFN-β et l’IL-27. / Cytokines regulate fundamental biological processes via the JAK-STAT signaling pathway. Suppressors of Cytokine Signaling proteins (SOCS), intracellular proteins, inhibit the JAK-STAT pathway. Emerging evidence supports the involvement of SOCS in diseases of the immune system but no data is available regarding their expression in human T cells. We postulate that the cytokines Interferon-β (IFN-β) and Interleukin-27 (IL-27), both potential immuno-regulators, have beneficial roles through the induction of SOCS proteins. The impact of IFN-β and IL-27 on the SOCS-1 and SOCS-3 expression by human CD4 and CD8 T cells was assessed using peripheral blood mononuclear cells from healthy donors. We evaluated the expression of SOCS-1 and SOCS-3 at the mRNA level by qRTPCR and at the protein level by immunocytochemistry. A rapid increase of SOCS-1 and SOCS-3 mRNA levels was observed upon cytokine addition, and such upregulation was confirmed at the protein level. To mimic patients under IFN-β treatment, both T cell subsets were chronically exposed to IFN-β. We observed an increase of SOCS-1 after each stimulation but not for SOCS-3. IL-27 stimulation increased SOCS-1 and SOCS-3 mRNA levels in CD8 T cells but only slightly in CD4 T cells; these observations correlate with previous observations in our laboratory showing less IL-27 receptors on CD4 T cells than CD8 counterparts. Our project determined the distinct expression of SOCS proteins in different human T cells subsets. This study could highlight the mechanism of action of cytokines such as IFN-β and IL-27.

Os avanços científicos das últimas décadas permitiram que a compreensão do câncer evoluísse de uma visão simplista, na qual o principal motor seria uma atividade celular hiperploriferativa, para uma visão mais complexa onde o estado fisiológico geral permite a gênese e progressão tumoral. Essa evolução permite o desenvolvimento de novas abordagens terapêuticas e traz novas esperanças para o tratamento de muitos tipos de cânceres ainda extremamente deletérios. Dentro desse novo panorama, terapias que estimulem a imunidade antitumoral têm se mostrado extremamente promissoras. Nesse trabalho, procuramos investigar os efeitos antitumorais desencadeados pela combinação da indução de morte celular e imunoestimulação. Para tanto, visamos à recuperação da via de p53 (pela transferência gênica do próprio p53 ou p19) associada à transferência gênica de IFNbeta. A transferência gênica foi mediada por vetores adenovirais do sorotipo 5. Nossas observações, em um modelo murino de carcinoma pulmonar, permitem concluir que esta linhagem é sensível a morte induzida pela transferência gênica de p19 e não p53. Porém, a transferência gênica intratumoral de IFNbeta se mostrou chave no controle do crescimento do tumor primário. Destacamos, entretanto, que a associação de IFNbeta com p19 produziu efeitos imunoprotetores superiores à transferência de IFNbeta ou p19 sozinhos. Tal efeito parece ser dependente da indução de fatores quimiotáxicos e conseqüente recrutamento de neutrófilos para o sítio tumoral. O efeito da transferência gênica combinada de ambos os genes IFNbeta e p19 se mostrou ainda mais promissor quando associado à cisplatina, induzindo uma notável redução no crescimento tumoral / Scientific advances from the last decades enabled the evolution of our knowledge of cancer from a simplistic vision, in which the main motor was an excessive cell proliferation, to a more complex one, where the general physiologic state enables tumorigenesis and tumor progression. This evolution enabled the development of new therapies and brings new hopes for the treatment of several types of cancers. In this context, therapies that induce an antitumor immunity are very promising. In this work, we are investigating the antitumor effects triggered by the combination of cell death induction and immunostimulation. To this end, we aimed to restore p53 pathway (by p53 or p19 gene transfer) associated with immunostimulation by IFNbeta gene transfer. The gene transfer was mediated by Adenovectors Serotype 5. Our observations in a murine model of lung cancer showed that this cell line is sensitive to cell death induced by p19 gene transfer, but not p53. Nevertheless, intratumoral gene transfer of IFNbeta, was crucial in controlling tumor growth. Moreover, p19 and IFNbeta association induced higher immunoprotecting effects than p19 or IFNbeta alone. This effect seems to be depending on the induction of chemotactic factors, and the recruitment of neutrophils to the tumor site. The effect of combined gene transfer of p19 and IFNbeta was even more promising when associated with Cisplatine, inducing a remarkable reduction in tumor growth

Les cytokines jouent un rôle fondamental dans la régulation des processus biologiques via la cascade de signalisation JAK-STAT. Les « Suppressors of Cytokine Signalling » (SOCS), protéines intracellulaires, inhibent la voie JAK-STAT. Plusieurs études supportent leur implication dans des maladies immunitaires, mais peu d’informations sont disponibles sur leur expression par les lymphocytes T humains. Nous postulons que les cytokines Interféron-β(IFN-β) et Interleukine-27 (IL-27), dotées d’un potentiel immuno-régulateur, ont des rôles bénéfiques via l’induction des SOCS. L’impact de l’IFN-β et l’IL-27 sur l’expression des SOCS-1 et SOCS-3 par des cellules T CD8 et CD4 humaines a été étudié en utilisant des cellules sanguines de donneurs sains. L’expression de ces régulateurs a été évaluée aux niveaux de l’ARNm par qRT-PCR et protéique par immunocytochimie. Les SOCS-1 et SOCS-3 ont été rapidement induits en ARNm dans les deux types cellulaires en réponse à l’IFN-β ou l’IL-27 et une augmentation de l’expression a été confirmée au niveau protéique. Afin de mimer les thérapies à base d’IFN-β, les cellules T ont été exposées chroniquement à l’IFN-β. Après chaque ajout de cytokine les cellules T ont augmenté l’expression du SOCS-1, sans moduler le SOCS-3. L’IL-27 a induit les SOCS-1 et SOCS-3 préférentiellement dans les cellules T CD8 ; ceci corrèle avec des résultats du laboratoire démontrant une plus petite expression des récepteurs à l’IL-27 par les lymphocytes T CD4 que les CD8. Notre projet a permis d’élucider l’expression des SOCS dans deux populations de cellules T et de clarifier les mécanismes d’actions de l’IFN-β et l’IL-27. / Cytokines regulate fundamental biological processes via the JAK-STAT signaling pathway. Suppressors of Cytokine Signaling proteins (SOCS), intracellular proteins, inhibit the JAK-STAT pathway. Emerging evidence supports the involvement of SOCS in diseases of the immune system but no data is available regarding their expression in human T cells. We postulate that the cytokines Interferon-β (IFN-β) and Interleukin-27 (IL-27), both potential immuno-regulators, have beneficial roles through the induction of SOCS proteins. The impact of IFN-β and IL-27 on the SOCS-1 and SOCS-3 expression by human CD4 and CD8 T cells was assessed using peripheral blood mononuclear cells from healthy donors. We evaluated the expression of SOCS-1 and SOCS-3 at the mRNA level by qRTPCR and at the protein level by immunocytochemistry. A rapid increase of SOCS-1 and SOCS-3 mRNA levels was observed upon cytokine addition, and such upregulation was confirmed at the protein level. To mimic patients under IFN-β treatment, both T cell subsets were chronically exposed to IFN-β. We observed an increase of SOCS-1 after each stimulation but not for SOCS-3. IL-27 stimulation increased SOCS-1 and SOCS-3 mRNA levels in CD8 T cells but only slightly in CD4 T cells; these observations correlate with previous observations in our laboratory showing less IL-27 receptors on CD4 T cells than CD8 counterparts. Our project determined the distinct expression of SOCS proteins in different human T cells subsets. This study could highlight the mechanism of action of cytokines such as IFN-β and IL-27.

Os avanços científicos das últimas décadas permitiram que a compreensão do câncer evoluísse de uma visão simplista, na qual o principal motor seria uma atividade celular hiperploriferativa, para uma visão mais complexa onde o estado fisiológico geral permite a gênese e progressão tumoral. Essa evolução permite o desenvolvimento de novas abordagens terapêuticas e traz novas esperanças para o tratamento de muitos tipos de cânceres ainda extremamente deletérios. Dentro desse novo panorama, terapias que estimulem a imunidade antitumoral têm se mostrado extremamente promissoras. Nesse trabalho, procuramos investigar os efeitos antitumorais desencadeados pela combinação da indução de morte celular e imunoestimulação. Para tanto, visamos à recuperação da via de p53 (pela transferência gênica do próprio p53 ou p19) associada à transferência gênica de IFNbeta. A transferência gênica foi mediada por vetores adenovirais do sorotipo 5. Nossas observações, em um modelo murino de carcinoma pulmonar, permitem concluir que esta linhagem é sensível a morte induzida pela transferência gênica de p19 e não p53. Porém, a transferência gênica intratumoral de IFNbeta se mostrou chave no controle do crescimento do tumor primário. Destacamos, entretanto, que a associação de IFNbeta com p19 produziu efeitos imunoprotetores superiores à transferência de IFNbeta ou p19 sozinhos. Tal efeito parece ser dependente da indução de fatores quimiotáxicos e conseqüente recrutamento de neutrófilos para o sítio tumoral. O efeito da transferência gênica combinada de ambos os genes IFNbeta e p19 se mostrou ainda mais promissor quando associado à cisplatina, induzindo uma notável redução no crescimento tumoral / Scientific advances from the last decades enabled the evolution of our knowledge of cancer from a simplistic vision, in which the main motor was an excessive cell proliferation, to a more complex one, where the general physiologic state enables tumorigenesis and tumor progression. This evolution enabled the development of new therapies and brings new hopes for the treatment of several types of cancers. In this context, therapies that induce an antitumor immunity are very promising. In this work, we are investigating the antitumor effects triggered by the combination of cell death induction and immunostimulation. To this end, we aimed to restore p53 pathway (by p53 or p19 gene transfer) associated with immunostimulation by IFNbeta gene transfer. The gene transfer was mediated by Adenovectors Serotype 5. Our observations in a murine model of lung cancer showed that this cell line is sensitive to cell death induced by p19 gene transfer, but not p53. Nevertheless, intratumoral gene transfer of IFNbeta, was crucial in controlling tumor growth. Moreover, p19 and IFNbeta association induced higher immunoprotecting effects than p19 or IFNbeta alone. This effect seems to be depending on the induction of chemotactic factors, and the recruitment of neutrophils to the tumor site. The effect of combined gene transfer of p19 and IFNbeta was even more promising when associated with Cisplatine, inducing a remarkable reduction in tumor growth

Transcriptional deregulation has emerged as one of the leading causes in various human diseases. More than fifty percent of cancers arise due to frequent mutations in genes regulating transcription. Higher-order assembly via protein-protein interactions is one common mechanism of transcriptional regulation. Despite their critical role in regulating gene transcription and therapeutic relevance, detailed mechanistic understanding of these assemblies remains scarce. The primary focus of this thesis is to uncover important structural principles underlying the assembly and stability of multi-domain protein assemblies by characterizing components of the IFNβ enhanceosome and the CtBP-mediated repression complex. Using a combination of biochemical and structural analyses, I showed that the transcriptional activator C-terminal binding protein 2 (CtBP2) is a tetramer by solving the 3.6Å cryoEM structure of CtBP2. I highlighted the types of interactions that stabilize the homo-tetramer and showed the relevance of the tetramer in CtBP2 transcriptional activity. Second, I used an integrative approach to investigate the structural features leading to activation of interferon regulator factor 3 (IRF3) and its interaction with DNA. Although this work mostly focused on components of the CtBP2-mediated complex and IFNβ enhanceosome, the principles described here can be applied to other complexes. Therefore, these studies provide an overall understanding on how other macromolecular complexes regulate gene transcription. Furthermore, our structural-based analyses will provide a basis for designing drugs that can regulate gene transcription in cancer and immunological disorders.

Contexte : La sclérose en plaques (SP) est une maladie chronique pouvant engendrer de nombreuses conséquences sur la qualité de vie des patients. Objectifs : L’objectif général de cette thèse est la modélisation des aspects temporels de la SP et de son traitement. Les objectifs spécifiques découlant de cette thèse sont : 1) Estimer l’effet à long terme des médicaments immunomodulateurs sur la progression et l’activité de la SP;  2) Décrire, au niveau populationnel, l’évolution de l’utilisation des soins associés à la SP et estimer l’effet de l’âge au diagnostic sur l’utilisation des soins associés à la SP ;  3) Évaluer la capacité de la méthode de trajectoire basée sur les groupes à détecter le nombre de trajectoires, la forme des trajectoires et le nombre de sujets dans chaque classe de trajectoire; 4) Proposer une nouvelle méthode pour visualiser les trajectoires longitudinales de dose et appliquer cette méthode pour un immunomodulateur utilisé dans le traitement de la SP. Méthodes : Les analyses pour les objectifs 1 et 4 ont été effectuées à partir des données de patients ayant eu un diagnostic de SP et traités à la clinique de SP du Centre hospitalier de l’Université de Montréal (CHUM). L’étude a été constituée à partir des informations colligées lors du suivi clinique des patients entre septembre 1977 et décembre 2016. Les caractéristiques socio-démographiques, l’histoire reproductive chez les femmes, la progression de la maladie, les comorbidités et médicaments ont été colligés à chaque visite médicale. Pour l’objectif 2, la cohorte de naissance québécoise sur l’immunité et la santé a été utilisée. Les individus faisant partie de cette cohorte sont nés entre 1970 et 1974 et le suivi a été réalisé jusqu’en 2014 en utilisant les bases de données médico-administratives et démographiques. Les informations sociodémographiques et celles concernant l’utilisation des services de santé pour la SP ont été obtenues pour chaque individu. Pour l’objectif 3, les données ont été générées en utilisant des scénarios basés sur la littérature. Les analyses effectuées incluent l’utilisation des modèles structurels marginaux (objectif 1), des modèles additifs généralisés (objectif 2) et des méthodes de trajectoires basées sur les groupes (objectifs 3 et 4). Résultats : Les résultats ont montré que l’utilisation de l’interféron bêta et de l’acétate de glatiramère est associée à une diminution du taux de progression d’incapacités et des poussées. Nous avons également constaté certaines différences au niveau des taux de visites chez un neurologue et chez un omnipraticien entre les personnes diagnostiquées avec la SP avant l’âge de 29 ans et celles diagnostiquées plus tard. De plus, nous avons montré l’importance d’utiliser plusieurs critères statistiques afin d’évaluer l’adéquation des groupes de trajectoires. Conclusions : L’utilisation de plusieurs variables représentant les issues dans les deux premiers objectifs nous a permis d’avoir une vue globale du traitement, des soins et de la progression de la SP. Cette thèse fait également des recommandations pour une bonne utilisation des méthodes de trajectoires et propose une alternative graphique pour visualiser les trajectoires de dose de médicaments. / Background: Multiple sclerosis (MS) is a chronic disease that can impact on the quality of life of patients in multiple ways. Objectives: The main goal of this thesis is the modelling of the temporal aspects of MS and its treatment. The specific goals of the thesis are: 1) To estimate the long-term effect of immunomodulatory drugs on the progression and the disease activity of MS; 2) To describe, at the population level, the evolution of health services related to MS and to estimate the effect of age at diagnosis on the use of health services for MS; 3) To assess the propensity of group-based trajectory modelling to detect the number of trajectories, the shape of trajectories and the number of subjects in each group of trajectories; 4) To propose a new graphical representation to visualize longitudinal patterns of medication dose and apply it to an immunomodulator used for treating MS patients. Methods: The analysis of objectives 1 and 4 were carried out using data from patients with a diagnosis of MS treated at the Montreal MS clinic, CHUM. The study was based on information collected during the clinical follow-up of patients between September 1975 and December 2016. The socio-demographic characteristics, reproductive history in women, disease progression, comorbidities and medications were collected at each medical visit. For objective 2, the Québec Birth Cohort on Immunity and Health was used. The individuals in this cohort were born between 1970 and 1974 and follow-up was carried out until 2014 using administrative health and demographic databases. Socio-demographic variables and information regarding the use of health services for MS were obtained for everyone. For objective 3, data were generated based on scenario from the literature. The analyses carried out include the use of marginal structural models (objective 1), generalized additive models (objective 2) and group-based trajectory methods (objectives 3 and 4). Results: The results showed that the use of interferon-beta and glatiramer acetate is associated with a decrease in the rate of progression of disability and relapses. We also found some differences in neurologist and general practitioner visit rates between people diagnosed with MS before age 29 and those diagnosed later. In addition, we have shown the importance of using several statistical criteria in order to assess the adequacy of groups of trajectories. Conclusions: The use of several variables for the outcomes in the first two objectives gave us a holistic view of the treatment, care and progression of MS. This thesis also makes recommendations for the good use of the trajectory modelling and proposes a graphic alternative to visualize patterns of medication dose.

Glioblastoma multiforme (GBM) is the most common and aggressive primary brain tumor. The current standard-of-care treatment including surgery, radiation and temozolomide (TMZ) chemotherapy does not prolong the survival satisfactorily. Here we have tested the feasibility, efficacy and safety of a potential gene therapy approach using AAV as gene delivery vehicle for treatment of GBM. Interferon-beta (IFNβ) is a cytokine molecule also having pleiotropic anticancerous properties. Previously it has been shown by our group that AAV mediated local (intracranial) gene delivery of human IFNβ (hIFNβ) could be an effective treatment for non-invasive human glioblastoma (U87) in orthotopic xenograft mouse model.But as one of the major challenges to treat GBM effectively in clinics is its highly invasive property, in the current study we first sought to test the efficacy of our therapeutic model in a highly invasive human GBM (GBM8) xenograft mouse model. One major limitation of using the xenograft mouse model is that these mice are immune-compromised. Moreover, as IFNβ does not interact with cross-species receptors, the influence of immune systems on GBM remains largely untested. Therefore to test the therapeutic approach in an immune-competent mouse model, we next treated a syngeneic mouse GBM model (GL261) in an immune-competent mouse (C57B6) with the gene encoding the species-matched IFNβ (mIFNβ). We also tested if combination of this IFNβ gene therapy with the current standard chemotherapeutic drug (TMZ) is more effective than any one of the therapeutic modes alone. Finally, we tested the long term safety of the AAV-mIFNβ local gene therapy in healthy C57B6 mice. Next, we hypothesized that global genetic engineering of brain cells expressing secretory therapeutic protein like hIFNβ could be more beneficial for treatment of invasive, migratory and distal multifocal GBM. We tested this hypothesis using systemic delivery of AAV9 vectors encoding hIFNβ gene for treatment of GBM8 tumor in nude mice. Using in vivo bioluminescence imaging of tumor associated firefly luciferase activity, long term survival assay and histological analysis of the brains we have shown that local treatment of AAV-hIFNβ for highly invasive human GBM8 is therapeutically beneficial at an early growth phase of tumor. However, systemic delivery route treatment is far superior for treating multifocal distal GBM8 tumors. Nonetheless, for both delivery routes, treatment efficacy is significantly reduced when treated at a later growth phase of the tumor. In syngeneic GL261 tumor model study, we show that local AAV-mIFNβ gene therapy alone or in combination with TMZ treatment can provide significant survival benefit over control or only TMZ treatment, respectively. However, the animals eventually succumb to the tumor. Safety study in the healthy animals shows significant body weight loss in some treatment groups, whereas one group shows long term survival without any weight loss or any noticeable changes in the external appearances. However, histological analysis indicates marked demyelinating neurotoxic effects upon long term exposures to mIFNβ over-expressions in brain. Overall, we conclude from this study that AAV-IFNβ gene therapy has great therapeutic potential for GBM treatment in future, but the therapeutic window is small and long term continuous expression could have severe deleterious effects on health.

Les cellules endothéliales (EC) constituent une première barrière physique à la dissémination de virus pléiotropiques circulant par voie hématogène mais leur contribution à la défense innée anti-virale est peu connue. Des dysfonctions des EC de la barrière hémato-encéphalique (BMEC) et des sinusoïdes hépatiques (LSEC) ont été rapportées dans des neuropathologies et des hépatites aiguës ou chroniques d’origine virale, suggérant que des atteintes à leur intégrité contribuent à la pathogenèse. Les sérotypes de coronavirus de l’hépatite murine (MHV), se différenciant par leur capacité à induire des hépatites et des maladies neurologiques de sévérité variable et/ou leur tropisme pour les EC, représentent des modèles viraux privilégiés pour déterminer les conséquences de l’infection des EC sur la pathogenèse virale. Lors d’infection par voie hématogène, le sérotype MHV3, le plus virulent des MHV, induit une hépatite fulminante, caractérisée par une réponse inflammatoire sévère, et des lésions neurologiques secondaires alors que le sérotype moins virulent, MHV-A59, induit une hépatite modérée sans atteintes secondaires du système nerveux central (SNC). Par ailleurs, le sérotype MHV3, à la différence du MHV-A59, démontre une capacité à stimuler la production de cytokines par la voie TLR2. Les variants atténués du MHV3, les virus 51.6-MHV3 et YAC-MHV3, sont caractérisés par un faible tropisme pour les LSEC et induisent respectivement une hépatite modérée et subclinique. Compte tenu de l’importance des LSEC dans le maintien de la tolérance hépatique et de l’élimination des pathogènes circulants, il a été postulé que la sévérité de l’hépatite et de la réponse inflammatoire lors d’infections par les MHV est associée à la réplication virale et à l’altération des propriétés tolérogéniques et vasculaires des LSEC. Les désordres inflammatoires hépatiques pourraient résulter d’une activation différentielle du TLR2, plutôt que des autres TLR et des hélicases, selon les sérotypes. D’autre part, compte tenu du rôle des BMEC dans la prévention des infections du SNC, il a été postulé que l’invasion cérébrale secondaire par les coronavirus est reliée à l’infection des BMEC et le bris subséquent de la barrière hémato-encéphalique (BHE). À l’aide d’infections in vivo et in vitro par les différents sérotypes MHV, chez des souris ou des cultures de BMEC et de LSEC, nous avons démontré, d’une part, que l’infection in vitro des LSEC par le sétotype MHV3, à la différence des variants 51.6- et YAC-MHV3, altérait la production du facteur vasodilatant NO et renversait leur phénotype tolérogénique en favorisant la production de cytokines et de chimiokines inflammatoires. Ces dysfonctions se traduisaient in vivo par une réponse inflammatoire incontrôlée et une dérégulation du recrutement intrahépatique de leucocytes, favorisant la réplication virale et les dommages hépatiques. Nous avons aussi démontré, à l’aide de souris TLR2 KO et de LSEC dont l’expression du TLR2 a été abrogée par des siRNA, que la sévérité de l’hépatite et de la réponse inflammatoire induite par le sérotype MHV3, dépendait en partie de l’induction et de l’activation préférentielle du TLR2 par le virus dans le foie. D’autre part, la sévérité de la réplication virale au foie et des désordres dans le recrutement leucocytaire intrahépatique induits par le MHV3, et non par le MHV-A59 et le 51.6-MHV3, corrélaient avec une invasion virale subséquente du SNC, au niveau de la BHE. Nous avons démontré que l’invasion cérébrale du MHV3 était associée à une infection productive des BMEC et l’altération subséquente des protéines de jonctions serrées occludine, VE-cadhérine et ZO-1 se traduisant par une augmentation de la perméabilité de la BHE et l’entrée consécutive du virus dans le cerveau. Dans l’ensemble, les résultats de cette étude mettent en lumière l’importance du maintien de l’intégrité structurale et fonctionnelle des LSEC et des BMEC lors d’infections virales aigües par des MHV afin de limiter les dommages hépatiques associés à l’induction d’une réponse inflammatoire exagérée et de prévenir le passage des virus au cerveau suite à une dissémination par voie hématogène. Ils révèlent en outre un nouveau rôle aggravant pour le TLR2 dans l’évolution de l’hépatite virale aigüe ouvrant la voie à de nouvelles avenues thérapeutiques visant à moduler l’activité inflammatoire du TLR2. / Endothelial cells (EC) act as a physical barrier against invasion by pleiotropic blood borne viruses but their contribution in innate antiviral defense is poorly known. Dysfunctions in blood-brain barrier EC (BMECs) and liver sinusoidal EC (LSECs) have been reported in viral neuropathologies and hepatitis, suggesting that loss of ECs integrity may contribute to the pathogenesis. Mouse hepatitis coronaviruses (MHV), differing in their ability to induce severe to subclinical hepatitis and neurological diseases and / or their tropism for ECs, are relevant viral models to study the consequences of EC infection in viral pathogenesis. Following hematogenous infection, the MHV3 serotype, the most virulent MHV, induces fulminant hepatitis, characterized by severe inflammatory response, followed by neurological damage whereas the less virulent MHV-A59 serotype induces milder hepatitis but does not invade the central nervous system (CNS). In addition, MHV3, in contrast to MHV-A59, shows ability to induce TLR2-dependent cytokine response. The attenuated MHV3 variants, 51.6-MHV3 and YAC-MHV3, are characterized by a weak tropism for LSECs and induce moderated and subclinical hepatitis respectively. Given the importance of LSECs in hepatic tolerance and the elimination of circulating pathogens, it has been postulated that the severity of hepatitis and inflammatory response induced by MHVs correlates with infection and alterations in vascular and tolerogenic properties of LSECs. Hepatic inflammatory disorders may result from differential activation of TLR2, rather than other TLRs and helicases, according to serotypes. Moreover, given the role of BMECs in preventing CNS infections, it has been postulated that secondary cerebral invasion by coronaviruses is related to infection of BMECs and subsequent breakdown of the blood-brain barrier (BBB). Through in vitro and in vivo infections of isolated BMECs, LSECs or mice with the different MHVs, we demonstrated, first, that in vitro productive infection of LSECs by the highly virulent MHV3 serotype, in contrast to 51.6- et YAC-MHV3 variants, altered their production of vasoactive factors and overthrew their intrinsic tolerogenic properties by promoting inflammatory cytokines and chemokines production. These disturbances were reflected in vivo by an uncontrolled inflammatory response and a deregulation of intrahepatic leukocyte recruitment, favoring viral replication and liver damages. We demonstrated, using TLR2 KO mice and LSECs treated with siRNA for TLR2 that the abnormal inflammatory response induced by MHV3 depended in part on preferential induction and activation of TLR2 by the virus on the surface of hepatic cells. Moreover, the severity of the primary viral replication in the liver and disorders in intrahepatic leucocyte recruitment induced by MHV3, but not by MHV-A59 and 51.6-MHV3, correlated with a subsequent brain invasion at the BBB level. Such invasion was related to productive infection of BMECs and subsequent IFN--dependent disruption of tight junction proteins occludin, VE-cadherin and ZO-1, resulting in an increase of BBB permeability and further viral entry into the CNS. Overall, the results of this study highlight the importance of structural and functional integrity of LSECs and BMECs during acute viral infections by MHVs to limit liver damages associated with viral-induced exacerbation of inflammatory response and prevent brain invasion by MHVs following viral spread through the bloodstream. They also reveal a new worsening role for TLR2 in the evolution of acute viral hepatitis paving the way for new therapies targeting TLR2-induced inflammatory activity.