A doença hepática crônica causada pelo vírus da hepatite C (HCV) tornou-se, nos últimos anos, uma das principais comorbidades dos pacientes portadores do vírus da imunodeficiência humana (HIV) nos países desenvolvidos. Os pacientes coinfectados com HIV/HCV apresentam uma progressão mais rápida para a cirrose e as suas complicações que os pacientes monoinfectados com HCV. Embora os mecanismos responsáveis por esta evolução não estejam totalmente esclarecidos, a expressão da molécula de HLA-G, um HLA de classe Ib não clássico, que tem propriedades bem reconhecidas na regulação negativa da resposta imune, pode estar relacionada à progressão da doença hepática. Os objetivos deste trabalho foram analisar o perfil de expressão de HLA-G em tecido hepático de pacientes coinfectados HIV/HCV e identificar possíveis variáveis do hospedeiro, do HCV e do HIV que possam estar relacionadas com a expressão de HLA-G na biópsia hepática. Para isso, 57 amostras de biópsia hepática de pacientes coinfectados com HIV/HCV, nas quais a imuno-histoquímica para HLA-G foi realizada, foram analisadas retrospectivamente quanto à expressão desta molécula no tecido hepático. Avaliaram-se também outras características histopatológicas da biópsia como grau de fibrose, atividade inflamatória, deposição de ferro e gordura. Determinou-se o polimorfismo de inserção ou deleção de 14 pares de bases da região 3` não traduzida do exon 8 do gene do HLA-G, que está relacionada com a produção de RNA-mensageiro, em 43 destes pacientes, além do polimorfismo de IL-28B, relacionado com a resposta ao tratamento do HCV, em 44 deles. Características bioquímicas e virológicas, tanto do HIV quanto do HCV também foram avaliadas. O genótipo 1 do HCV foi o mais prevalente (87,75%), especialmente o subgenótipo 1a (60%). A expressão do HLA-G foi observada em 38 (66,7%) amostras de fígado, e foi mais frequente em estágios moderados e severos de fibrose do que em estágios mais leves (94,1% x 55%, P < 0,01). Não houve relação entre a expressão do HLA-G e os outros parâmetros estudados. Embora a progressão para a cirrose no contexto da coinfecção por HIV/ HCV seja um processo complexo, modulado por muitos factores, a associação da intensidade de fibrose com a expressão do HLA-G pode indicar que a expressão desta proteína desempenha um importante papel nos mecanismos que contribuem para a progressão da doença, por meio da regulação negativa da resposta imune contra o HCV na coinfecção pelo HIV. / Chronic liver disease induced by hepatitis C virus (HCV) infection has recently become one of the most common comorbidities in patients who are infected with the human immunodeficiency virus (HIV) in developed countries. HIV/HCV coinfected patients show faster progression to cirrhosis and its complications than the HCV monoinfected patients. Even though the responsible mechanisms for this evolution have not been entirely clarified yet, the expression of the HLA-G molecule, a HLA from the non-classic Ib class, with well-known properties of negatively regulating the immune response, may be related to the liver disease progression. The aims of the present work were to analyze the HLA-G expression profile in the liver micro ambience of HIV/HCV coinfected patients and to identify possible host factors, HIV or HCV, that may be related to the HLA-G expression on the liver biopsy. For this purpose, 57 liver biopsies of HIV/HCV coinfect patients, in which immunohistochemistry for HLA-G had been performed, were retrospectively analyzed according the HLA-G expression on the hepatic tissue. Other histopathological features in the liver biopsies, such as fibrosis degree, inflammatory activity, iron deposition and fat were also evaluated. The polymorphism of insertion or deletion in 14-base pairs of the 3`non-translated region of exon 8 of the HLA-G gene, which is related to the production of HLA-G messenger RNA, was evaluated in 43 of the patients. Also, the polymorphism of IL-28B, related to the response to HCV treatment, was evaluated in 44 of them. Biochemical and virological features of HIV and HCV were also evaluated. The HCV genotype 1 was the most prevalent (87.75%), especially the subgenotype 1a (60%). The expression of HLA-G was observed in 38 (66.7%) samples of the liver biopsies, and it was most frequent in moderate and severe stages of fibrosis than in the mild stages (94.1% x 55%, P < 0.01). There was no established relationship between HLA-G and other parameters studied. Although the progression to cirrhosis in the context of HIV/HCV coinfection is a complex process modulated by many factors, the association of HLA-G expression with the intensity of the liver fibrosis may indicate the protein expression play an important role in the mechanisms that contribute to the progression of the disease, through the negative regulation of the immune response against HCV setting of a coinfection with HIV.

Objectifs : Les objectifs principaux de cette thèse sont les suivants : 1) analyse de l’évolution du profil épidémiologique des patients coinfectés VIH-VHC initiant le traitement de l’hépatite C; 2) l’évaluation de l’impact des comportements de santé et des styles de vie sur l’évolution clinique de la maladie Résultats : J’ai pu mettre en évidence que le profil des patients coinfectés VIH-VHC initiant un traitement de l’hépatite C a changé en France avec l’évolution des traitements.Mes travaux ont permis de montrer que, chez les patients coinfectés VIH-VHC, une consommation élevée de café (3 tasses par jour et plus) diminue le risque de fibrose hépatique avancée. Cet effet bénéfique du café est également observé chez les patients coinfectés VIH-VHC avec une consommation élevée d’alcool. De plus, mes travaux ont mis en évidence une relation dose-dépendante entre la fréquence de consommation de cacao et la réduction du risque de fibrose hépatique avancée chez les patients coinfectés VIH-VHC. Par ailleurs, mes analyses n’ont pas mis en évidence un effet significatif de la consommation de café sur le risque de fibrose hépatique avancée chez les femmes coinfectées VIH-VHC. Enfin l’un des résultats marquant de la relation entre VHC et risque d’obésité qui est connue est que la guérison augmente davantage ce risque sur le long terme. Conclusion : Des interventions pour modifier certains styles de vie et comportements ont le potentiel de diminuer le risque de survenue ou d’aggravation de comorbidités, en particulier après la guérison VHC, un événement désormais atteignable pour tous les patients coinfectés. / Objectives: The main objectives of this thesis are: 1) analysis of the evolution of the epidemiological profile of co-infected HIV-HCV patients initiating the treatment of hepatitis C; 2) assessment of the impact of health behaviors and lifestyles on the clinical course of the diseaseResults: I was able to highlight that the profile of co-infected HIV-HCV patients initiating treatment for hepatitis C has changed in France with the evolution of treatments.My work has shown that, in HIV-HCV coinfected patients, high coffee consumption (3 cups per day or more) decreases the risk of advanced liver fibrosis. This beneficial effect of coffee is also observed in co-infected HIV-HCV patients with high alcohol consumption. In addition, my work has shown a dose-dependent relationship between the frequency of cocoa consumption and the reduced risk of advanced liver fibrosis in coinfected HIV-HCV patients. Furthermore, my analyzes did not reveal a significant effect of coffee consumption on the risk of advanced liver fibrosis in coinfected HIV-HCV women. Finally, one of the striking results of the relationship between HCV and known risk of obesity is that healing increases this risk in the long term.Conclusion: Interventions to modify certain lifestyles and behaviors have the potential to reduce the risk of developing or worsening comorbidities, particularly after HCV healing, an event now achievable for all co-infected patients.

A doença hepática crônica causada pelo vírus da hepatite C (HCV) tornou-se, nos últimos anos, uma das principais comorbidades dos pacientes portadores do vírus da imunodeficiência humana (HIV) nos países desenvolvidos. Os pacientes coinfectados com HIV/HCV apresentam uma progressão mais rápida para a cirrose e as suas complicações que os pacientes monoinfectados com HCV. Embora os mecanismos responsáveis por esta evolução não estejam totalmente esclarecidos, a expressão da molécula de HLA-G, um HLA de classe Ib não clássico, que tem propriedades bem reconhecidas na regulação negativa da resposta imune, pode estar relacionada à progressão da doença hepática. Os objetivos deste trabalho foram analisar o perfil de expressão de HLA-G em tecido hepático de pacientes coinfectados HIV/HCV e identificar possíveis variáveis do hospedeiro, do HCV e do HIV que possam estar relacionadas com a expressão de HLA-G na biópsia hepática. Para isso, 57 amostras de biópsia hepática de pacientes coinfectados com HIV/HCV, nas quais a imuno-histoquímica para HLA-G foi realizada, foram analisadas retrospectivamente quanto à expressão desta molécula no tecido hepático. Avaliaram-se também outras características histopatológicas da biópsia como grau de fibrose, atividade inflamatória, deposição de ferro e gordura. Determinou-se o polimorfismo de inserção ou deleção de 14 pares de bases da região 3` não traduzida do exon 8 do gene do HLA-G, que está relacionada com a produção de RNA-mensageiro, em 43 destes pacientes, além do polimorfismo de IL-28B, relacionado com a resposta ao tratamento do HCV, em 44 deles. Características bioquímicas e virológicas, tanto do HIV quanto do HCV também foram avaliadas. O genótipo 1 do HCV foi o mais prevalente (87,75%), especialmente o subgenótipo 1a (60%). A expressão do HLA-G foi observada em 38 (66,7%) amostras de fígado, e foi mais frequente em estágios moderados e severos de fibrose do que em estágios mais leves (94,1% x 55%, P < 0,01). Não houve relação entre a expressão do HLA-G e os outros parâmetros estudados. Embora a progressão para a cirrose no contexto da coinfecção por HIV/ HCV seja um processo complexo, modulado por muitos factores, a associação da intensidade de fibrose com a expressão do HLA-G pode indicar que a expressão desta proteína desempenha um importante papel nos mecanismos que contribuem para a progressão da doença, por meio da regulação negativa da resposta imune contra o HCV na coinfecção pelo HIV. / Chronic liver disease induced by hepatitis C virus (HCV) infection has recently become one of the most common comorbidities in patients who are infected with the human immunodeficiency virus (HIV) in developed countries. HIV/HCV coinfected patients show faster progression to cirrhosis and its complications than the HCV monoinfected patients. Even though the responsible mechanisms for this evolution have not been entirely clarified yet, the expression of the HLA-G molecule, a HLA from the non-classic Ib class, with well-known properties of negatively regulating the immune response, may be related to the liver disease progression. The aims of the present work were to analyze the HLA-G expression profile in the liver micro ambience of HIV/HCV coinfected patients and to identify possible host factors, HIV or HCV, that may be related to the HLA-G expression on the liver biopsy. For this purpose, 57 liver biopsies of HIV/HCV coinfect patients, in which immunohistochemistry for HLA-G had been performed, were retrospectively analyzed according the HLA-G expression on the hepatic tissue. Other histopathological features in the liver biopsies, such as fibrosis degree, inflammatory activity, iron deposition and fat were also evaluated. The polymorphism of insertion or deletion in 14-base pairs of the 3`non-translated region of exon 8 of the HLA-G gene, which is related to the production of HLA-G messenger RNA, was evaluated in 43 of the patients. Also, the polymorphism of IL-28B, related to the response to HCV treatment, was evaluated in 44 of them. Biochemical and virological features of HIV and HCV were also evaluated. The HCV genotype 1 was the most prevalent (87.75%), especially the subgenotype 1a (60%). The expression of HLA-G was observed in 38 (66.7%) samples of the liver biopsies, and it was most frequent in moderate and severe stages of fibrosis than in the mild stages (94.1% x 55%, P < 0.01). There was no established relationship between HLA-G and other parameters studied. Although the progression to cirrhosis in the context of HIV/HCV coinfection is a complex process modulated by many factors, the association of HLA-G expression with the intensity of the liver fibrosis may indicate the protein expression play an important role in the mechanisms that contribute to the progression of the disease, through the negative regulation of the immune response against HCV setting of a coinfection with HIV.

Plus de 170 millions d'individus sont infectés par le VHC dans le monde et 37 millions par le VIH. La coinfection VIH/VHC est fréquente et représente un élément clé de la prise en charge des patients infectés par le VIH. Depuis l'arrivée des HAART, les maladies du foie sont devenues la cause principale de mortalité chez les patients coinfectés VIH/VHC. L'évolution naturelle et le pronostic de l'hépatite C sont plus sévères en cas de coinfection par le VIH du fait d'une fibrose accélérée et d'une évolution rapide vers la cirrhose et ses complications. Certains facteurs accélérant la fibrose hépatique sont clairs aujourd'hui comme: l'absence de recours au traitement anti-VHC, la réplication active du VHC et la consommation excessive d'alcool. De plus, il existe de plus en plus de preuves que les variants génétiques contribuent à la fibrose hépatique chez les patients monoinfectés par le VHC, mais cet aspect a été peu étudié dans la coinfection VIH/VHC.Durant ma thèse, j'ai eu accès aux données d'un échantillon de 494 patients coinfectés génotypés issu de la cohorte ANRS CO13 HEPAVIH. L'histoire naturelle du VIH et du VHC y est renseignée de manière très détaillée et le suivi clinique des patients permet d'avoir des informations précises sur l'état de fibrose hépatique. J'ai pu alors réaliser deux études d'association « génome-entier » pour identifier des polymorphismes associés à la sévérité de la fibrose à l'aide de données complètes de 292 patients. La première étude a mis en évidence une association entre la quantification de l'élasticité hépatique par Fibroscan® et un locus, également répliqué dans la monoinfection par le VHC. Cette association a permis d'identifier deux gènes impliqués dans des mécanismes de maintien de structure et de signalisation cellulaire (CAV3) mais aussi dans la réplication du VHC (RAD18). La seconde étude a identifié deux associations significatives en comparant deux groupes de scores METAVIR (F0F1F2 vs F3F4), en particulier dans le gène CTNND2 qui est impliqué dans un réseau d'interaction associé à des mécanismes moléculaires lié à des maladies hépatiques.Ces deux études sont en cours de publication dans des revues scientifiques internationales à comité de lecture. Ces nouvelles perspectives dans la compréhension des mécanismes de fibrose dans le contexte de la coinfection VIH/VHC pourraient aider à l'identification de nouvelles cibles pour la création de médicaments ou de tests diagnostiques afin d'améliorer les soins des patients. / Over 170 million people worldwide are infected by HCV and 37 million by HIV. Both viruses share the same modes of transmission, and HIV/HCV coinfection is common and represents a key element in the management of patients infected with HIV. Since the appearance of HAART, liver diseases have become the leading cause of death in HIV/HCV coinfected patients. The natural history and prognosis of hepatitis C are more severe in case of coinfection with HIV due to accelerated rate of fibrosis progression and rapid progression to cirrhosis and its complications. Factors accelerating liver fibrosis are known today such as the lack of recourse to anti-HCV treatment, active HCV replication and excessive alcohol consumption. There is increasing evidence that genetic variants contribute to liver fibrosis in HCV monoinfection, but this aspect has been little studied in HIV/HCV coinfection.I have exploited the genotype information from 494 coinfected patients from the cohort ANRS CO13 HEPAVIH. These patients are very-well documented regarding the history of their HIV/HCV infection and are very carefully followed-up, especially regarding the status of liver fibrosis. I have performed two genome-wide association studies to identify polymorphisms associated with the severity of fibrosis from complete data of 292 patients. The first study has dealt with the quantification of liver stiffness by Fibroscan® and an association with the 3p25 region has been identified, also replicated in monoinfection HCV. Two genes involved in cell signaling and structure of holding mechanisms (CAV3) but also in HCV replication (RAD18) appear as good candidates. The second study has unraveled two significant associations by comparing the METAVIR score group (F0F1F2 vs F3F4), especially in the CTNND2 gene implicated in a network of interactions with molecular mechanisms involved in liver diseases.These results are under publications in peer-review international scientific journals. These new insights into the molecular mechanisms of liver fibrosis in patients with HIV/HCV co- infection may help to define new targets for drug development or new diagnostic tests, to improve patient care.

La génomique d'association cherche à déterminer des liens entre le génome et des traits ou phénotypes, notamment dans le contexte de maladies. Aujourd'hui, les études les plus fréquentes en génomique d’association sont les études génome entier, qui analysent autant de variants du génomes (SNPs) que possible, sans avoir de préjugé a priori sur leur fonction biologique. Cependant, les méthodes de génotypage utilisées pour ces études ne permettent pas toujours d'obtenir des informations précises dans une région hypervariable comme la région HLA, qui joue un rôle crucial dans l'immunité, et les variants génétiques de ces régions sont souvent prédits par des approches bioinformatiques. J'ai durant ma thèse créé un nouvel outil, HLA-Check, permettant d'évaluer, à partir des génotypes obtenus par puce de génotypage, la plausibilité de données d'allèles HLA d'un même individu, et démontré que cette technique permettait d'identifier plus précisément les individus dont les allèles HLA avaient été mal caractérisés afin de les retyper ou de les écarter de l'étude. Un article documentant cet outil a été publié dans BMC Bioinformatics. J'ai également effectué une étude d'association génome entier sur le déclenchement de la cirrhose chez les patients co-infectés par le VIH (virus de l'immunodéficience humaine) et le VHC (virus de l'hépatite C). La coinfection par ces deux maladies est fréquente en raison de modes d’infection similaires, et l'infection par le VIH stimule l'activité du VHC et accélère la fibrose du foie puis sa cirrhose, causant la mort des patients co-infectés. Mon étude porte sur 306 patients co-infectés issus de la cohorte ANRS CO-13 HEPAVIH. J'ai ainsi pu mettre en évidence trois signaux associés avec le déclenchement de la cirrhose, dont deux ont un lien pertinent avec les maladies hépatiques (gene CTNND2 et gene MIR7-3HG). L'identification de ces nouveaux variants devrait permettre une meilleure compréhension des mécanismes moléculaires de la cirrhose, et contribuer au développement de nouvelles stratégies diagnostiques ou thérapeutiques. L’article documentant cette étude est en cours de publication. / Association genomics aims at finding links between the genome and some traits or illnesses. Today, the most frequent studies in this field are genome wide association studies (GWAS), which analyze as many genome variants (mainly Single Nucleotide Polymorphisms) as possible, without any a priori on their biological function. However, genotyping methods used in these studies may be insufficient to get reliable information in higly variable regions such as the HLA which plays a crucial role in immunity, and the genetic variants of such regions are often predicted using bioinformatics approaches. During my PhD, I have created a new tool, HLA-Check, that allows to rate the plausibility of HLA alleles from the genotypes obtained from genotyping chips. I also assesses its performances and showed that it was able to point out individuals with a wrong HLA typing, in order to retype them or remove them from the study. An article documenting this tool was published in BMC Bioinformatics. I have also performed a genome-wide association study on cirrhosis outbreak in individuals coinfected with HIV (human immunodeficiency virus) and HCV (hepatitis C virus). Because of similar infection routes (blood-related), co-infection with those two viruses are frequent, and the infection by HIV enhances HCV activity and increases liver fibrosis leading to cirrhosis and death of co-infected patients. Our study has dealt with 306 co-infected patients from the ANRS CO-13 HEPAVIH cohort. I could point out three statistically significant signals, two of them being highly relevant for their involvement in liver diseases (gene CTNND2 and gene MIR7-3HG). The identification of these new variants should lead to a better understanding of the molecular mechanisms involved in cirrhosis, and should contribute to the rational developement of new diagnostic or therapeutic strategies. A publication is under way.