L’infection à VIH-1 est associée à une forte déplétion des lymphocytes T CD4+ à polarisation Th17 au niveau des tissus lymphoïdes associés aux muqueuses intestinales (GALT, gut-associated lymphoid tissues). Ceci conduit à la translocation microbienne, qui est une cause d’activation immunitaire chronique et de progression de la maladie. Les cellules épithéliales (CE) jouent un rôle critique dans le maintien de l’intégrité et de l’homéostasie au niveau des muqueuses intestinales via le recrutement des cellules de l’immunité innée (e.g., neutrophiles) et adaptative (e.g., cellules Th17). Les neutrophiles produisent des molécules antivirales (e.g., défensines-) et ont la capacité de limiter la réplication virale au niveau des muqueuses. Les cellules Th17 jouent un double rôle lors de l’infection à VIH. Elles contribuent d’une part à la défense contre différents pathogènes opportunistes en augmentant, via la production d’IL-17, la capacité des CE à attirer les cellules Th17 et les neutrophiles. D’autre part, les cellules Th17 jouent un rôle délétère en tant que cibles de réplication virale et sources de cytokines pro-inflammatoires. La fréquence des cellules Th17 est diminuée dans les GALT mais pas dans les poumons des patients infectés par le VIH, suggérant qu’il existe des mécanismes différents par lesquels les cellules Th17 sont recrutées vers ces sites anatomiques. Nous avons testé l’hypothèse selon laquelle le VIH interfère avec la capacité des CE intestinales et non pas pulmonaires à produire des chimiokines (CK) responsables de l’attraction des cellules Th17 et des neutrophiles. Nous avons démontré que les CE intestinales et pulmonaires produisent des CK spécifiques pour les cellules Th17 (CCL20) et les neutrophiles (CXCL8) en réponse à des stimuli pro-inflammatoires tels que l’IL-1 et le TNF-. Le TNF- agit en synergie avec l’IL-17, un « signal de danger » récemment identifié, et augmente la capacité des CE intestinales mais pas pulmonaires à produire la chimiokine CCL20. Cette synergie s’explique par l’augmentation préférentielle de l’expression du récepteur à l’IL-17 à la surface des CE intestinales suite à la stimulation par le TNF-. L’exposition au VIH n’affecte pas la production de CCL20 et de CXCL8 par les CE intestinales, mais altère la capacité des CE alvéolaires à produire ces chimiokines en accord avec la permissivité sélective de ces dernières à l’infection par le VIH. En conclusion, nos résultats démontrent que (i) le VIH n’interfère pas directement avec la capacité des CE intestinales à recruter des cellules Th17 et des neutrophils et que (ii) la production de CCL20 par ces cellules est dépendantes de la synergie entre le TNF- et l’IL-17. Ainsi, la déplétion des cellules Th17 et la pénurie en IL-17 dans les GALT des sujets infectés pourrait causer de façon préférentielle des altérations fonctionnelles au niveau des CE intestinales, se traduisant par l’altération du recrutement des cellules Th17 en réponse au CCL20. / The HIV-1 infection is associated with a severe loss of CD4+ T-cells with Th17 polarization from the gut-associated lymphoid tissues (GALT). These alterations lead to microbial translocation, which is a cause of chronic immune activation and disease progression in HIV-infected subjects. Epithelial cells (EC) play a critical role in maintaining mucosal integrity and homeostasis in the GALT by mechanisms including recruitment of innate (e.g., neutrophils) and adaptive immunity cells (e.g., Th17 cells). Neutrophils produce antiviral molecules (e.g., -defensins) that may limit HIV replication at mucosal sites. Th17 cells play a dual role in HIV pathogenesis. Th17 cells contribute to the defence against different opportunistic pathogens by increasing the ability of epithelial cells to attract neutrophils in an IL-17-dependent manner. On the other hand, Th17 cells play a deleterious role in HIV pathogenesis as they are sites of productive viral replication and a source of pro-inflammatory cytokines. The frequency of Th17 cells is decreased in the GALT but not in the lungs of HIV-infected individuals, suggesting distinct mechanisms of Th17 recruitment in these anatomic sites in the context of HIV pathogenesis. In this manuscript we tested the hypothesis that HIV differentially interfere with the ability of intestinal but not pulmonary EC to produce chemokines that attract Th17 cells and neutrophils. We demonstrated that both intestinal and pulmonary EC produce chemokines that specifically attract Th17 cells (CCL20) and neutrophils (CXCL8) upon stimulation with the pro-inflammatory cytokines IL-1 and TNF- . TNF-α acted in synergy with IL-17, a recently identified « danger signal », and increases the capacity of intestinal but not pulmonary EC to produce CCL20. This synergistic effect can be explained by the preferential upregulation of IL-17 receptor expression on intestinal EC upon TNF- stimulation. The exposure of intestinal EC to HIV did not affect their ability to produce CCL20 and CXCL8; however, exposure to HIV altered the production of these chemokines by alveolar EC, consistent with their selective permissiveness to infection. In conclusion, our results demonstrate that (i) HIV does not interfere directly with the ability of intestinal EC to attract Th17 cells and neutrophils and that (ii) the ability of intestinal EC to recruit the Th17 cells via CCL20 production is selectively dependent on the synergy between TNF- and IL-17. Thus, the depletion of Th17 cells and the shortage in IL-17 in the GALT of HIV-infected subjects may preferentially lead to functional alterations of the intestinal barrier resulting by the alteration of Th17 recruitment in response to CCL20.

Astma yra lėtinė kvėpavimo takų uždegimo liga. Mokslininkai neabejoja, jog 2-o tipo T limfocitai pagalbininkai bei eozinofilinis kvėpavimo takų uždegimas yra astmos patogenezės pagrindas. Tačiau šis mechanizmas ne visuomet gali paaiškinti astmos metu esančio kvėpavimo takų uždegimo bei klinikinių simptomų įvairumą, eigos ypatumus ir net skirtingą atsaką į gydymą. Eksperimentiniai tyrimai su gyvūnais parodė 17-o tipo T limfocitų pagalbininkų (Th17) svarbą alerginės astmos vystymuisi. Todėl šio mokslinio tyrimo tikslas buvo įvertinti Th17 limfocitų vaidmenį sergant alergine astma. Tyrimo metu sergantiesiems alergine astma nustatytas didesnis Th17 limfocitų kiekis periferiniame kraujyje bei didesnė interleukino (IL)-17 koncentracija serume ir indukuotuose skrepliuose, lyginant su sveikais asmenimis. Be to, bronchų provokacija su Dermatophagoides pteronyssinus alergenu sukėlė Th17 limfocitų ir IL-17 kiekio padidėjimą praėjus 7 ir 24 val.po jos, ypatingai ryškų sergantiems alergine astma su ankstyva ir vėlyva bronchų obstrukcija. Atlikto tyrimo rezultatai parodė, kad Dermatophagoides pteronyssinus alergenas sukelia vietinį ir sisteminį Th17 limfocitų imuninį atsaką kuris yra susiję su vėlyvos fazės kvėpavimo takų uždegimu. / Allergic asthma is a chronic inflammatory disease of the airways associated with the response of predominant type 2 helper T lymphocytes to an inhaled allergen. However, differences in inflammation and clinical symptoms of this disease not always can be explained by this mechanism. Recent animal model studies have shown the importance of type 17 helper T lymphocytes (Th17) in the development of allergic asthma. The role of these cells in causing allergen-induced airway inflammation as well as systemic inflammatory response in human is still not well defined. Therefore, we investigated the peripheral blood Th17 lymphocyte response to inhaled Dermatophagoides pteronyssinus (D. pteronyssinu) allergen in patients with allergic asthma. The present study has shown that patients with allergic asthma have a higher percentage of peripheral blood Th17 lymphocytes and elevated serum as well as induced sputum interleukin-17 levels compared with healthy subjects. Moreover, all studied allergic asthma patients, especially with early- and late-phase asthmatic reaction, showed an enhanced airway and systemic Th17 lymphocyte response 7 h and 24 h after bronchial challenge. We documented an enhanced local and systemic Th17 lymphocyte response to inhaled D. pteronyssinus in association with late-phase allergen-induced airway inflammation in patients with allergic asthma.

Leishmaniose visceral (LV) é uma doença crônica e potencialmente fatal causada pelas espécies Leishmania infantum/chagasi e Leishmania donovani. O desenvolvimento da resposta Th1 é classicamente associado à proteção contra esses parasitos, mas também há uma correlação positiva entre a produção de citocinas relacionadas com o padrão Th17 e a proteção contra LV por L. donovani em seres humanos. No entanto, a participação de Th17 durante a infecção por L. infantum/chagasi permanece desconhecida. O objetivo desse estudo foi avaliar a participação de Th17 e citocinas relacionadas, além do mecanismo pelo qual tais células operam durante a resposta imune do hospedeiro contra L. infantum/chagasi. Os resultados mostraram que o parasito é capaz de induzir grandes quantidades de TGF-, IL-1, IL-6 e IL-23 por células dendríticas derivadas da medula óssea (BMDC), citocinas envolvidas na indução e/ou manutenção do perfil Th17. Assim, co-cultivando células do baço de camundongos C57BL/6 naves com BMDCs infectadas com L. infantum/chagasi observou-se uma significativa produção de IL-17 por células T. Esses achados foram confirmados por experimentos in vivo onde se constatou a produção de IL-17 no fígado e no baço de camundongos WT infectados, sendo o pico de produção dessa citocina observado na 4ª e 6ª semanas após a infecção. O padrão de resposta do tipo Th17 é crítica para a imunidade protetora contra L. infantum/chagasi, uma vez que camudongos IL-17R-/-, IL-23p19-/- e IL-6-/- mostraram aumento da carga parasitária nos órgãos alvo da infecção, sendo que a susceptibilidade observada em camundongos IL-17R-/- foi associada com o aumento da produção de IL-10 por linfócitos, sugerindo que a IL-17 regula negativamente a produção de IL-10 levando ao controle da infecção causada pelo parasito. Ainda, a ausência da sinalização via IL-17R gerou uma diminuição da inflamação hepática, decorrente de uma menor capacidade proliferativa de linfócitos frente ao estímulo com conA. De maneira interessante, na ausência de IL-10 houve potencialização na produção de IL-17 por camundongos infectados, e esses foram mais resistentes à infecção, apresentando números reduzidos parasitos no baço e no fígado. Além de promover proteção através da modulação de IL-10, a IL-17 foi capaz de potencializar a produção de NO in vitro e in vivo. Juntos, nossos resultados demonstram que a L. infantum/chagasi é capaz de desencadear o padrão Th17 de resposta imune, o qual promove proteção do hospedeiro durante a infecção. / Visceral leishmaniasis (LV) is a chronic and potentially fatal disease caused by Leishmania donovani and Leishmania infantum/chagasi. The development of Th1 response is classically associated with protection against these parasites, but recent data also show that there is a positive correlation between the Th17-related cytokines production and the protection against LV by L. donovani in humans. However, the role of this CD4+ T cells subset during L. infantum/chagasi infection remains unknown. In this study, our aim was to evaluate the Th17 and related cytokines participation, besides the mechanism by which these cells playing during the L. infantum/chagasi infection.The results showed that the parasite induces high amounts of TGF-, IL-1, IL-6, and IL-23 by bone marrow-derived dendritic cells (BMDC), cytokines involved with Th17 induction and/or maintenance. Accordingly, naïve-C57BL/6 spleenocytes co-cultured with L. infantum/chagasi-infected BMDC produced significant amounts of IL-17 by TCD4+. Interestingly, IL-17 was produced in high amounts in the liver and spleen of WT infected-mice, being peaked at 4th and 6th weeks after infection. The Th17 is critical for protective immunity against L. infantum/chagasi, since that IL-17R-/-, IL-23p19-/- and IL-6-/- infected-mice showed increasing of parasite load associated with enhancement of IL-10 production in IL-17R-/- compared to WT infected-mice. Strictly, in the absence of IL-17 signaling, a smaller inflammatory infiltrate was observed in the liver. Interestingly, IL-17 production was potentiated in IL-10-/- infected-mice, and they were more resistant to infection, showing reduced parasites numbers in the spleen and liver. In addition to promoting protection through the downmodulation of IL-10, IL-17 enhanced the NO production. Together, our results demonstrate that L. infantum/chagasi trigger Th17 response that promotes the host protection during infection.

O câncer de mama é um dos tumores malignos mais comuns e que afeta um grande número de mulheres da população mundial. O processo inflamatório gerado juntamente com o crescimento descontrolado das células tumorais promove um estresse metabólico no microambiente tumoral levando ao acúmulo de adenosina extracelular decorrente da hipóxia tecidual. A adenosina gerada e seus efeitos mediados via receptor A2A (A2AR) interfere em vários subtipos celulares. Neste trabalho, avaliamos o papel do receptor A2A na indução de uma resposta de células Th17 durante o carcinoma mamário experimental, visto que estas desempenham um importante papel no crescimento e na progressão do tumor de mama invasivo. Para isso, utilizamos o modelo de carcinoma mamário murino que desenvolve (4T-1) e o que não desenvolve metástase (67NR). Nossos resultados mostraram que o tumor mamário 4T-1 apresenta alta expressão do receptor A2A comparado com o tumor 67NR. A deficiência do receptor A2a preveniu o crescimento do tumor mamário 4T- 1 e de colônias de células tumorais em sítios secundários da doença, concomitante com a diminuição da resposta de perfil Th17 e do recrutamento de neutrófilos para o sitio primário. Ainda, que as células tumorais 4T-1 apresentaram uma alta expressão dos receptores de adenosina e das ectonucleotidases (CD73 e CD39), sugerindo que a via de sinalização de adenosina exerce um efeito direto nessas células. A administração de adenosina ou AMP (trifosfato de adenosina) em culturas de células tumorais 4T-1 induziu xvi um aumento da expressão de IL-6, CCL20 e CXCL1, mediadores importantes para o recrutamento de linfócitos T produtores de IL-17 e de neutrófilos. Além disso, durante o crescimento do tumor 4T-1, observamos uma alta expressão da enzima ciclooxigenase 2 (Cox-2) comparado com o tumor 67NR. No entanto, os animais deficientes geneticamente do receptor A2A apresentaram uma redução significativa da expressão de Cox-2 no microambiente tumoral comparados com os animais BALB/c. A inibição da enzima Cox-2 com indometacina ou celecoxicib (inibidor seletivo) em animais com tumor 4T-1 preveniu o crescimento do tumor primário e, consequentemente, resultou na redução da expressão de moléculas relacionadas como o perfil de resposta de células Th17 tais como IL-6, CCL20, IL-17A e Ror?t. Esses resultados indicam que o bloqueio da via de sinalização do receptor A2A interfere na indução de resposta de células Th17, abrindo novas perspectivas para o desenvolvimento de terapias alternativas para o controle de tumores invasivos / Breast cancer is one of the most common and aggressive malignant tumors, affecting a large number of women in the worldwide population. The inflammatory process generated along with the uncontrolled growth of tumor cells promotes metabolic stress in the tumor microenvironment leading to the accumulation of extracellular adenosine due to the generation of tissue hypoxia. The generated adenosine and its effects mediated by A2A (A2AR) interfere in several cell subtypes. In this study, we evaluated the role of A2A receptor in the induction of a Th17 cell response in experimental breast carcinoma, given that these cells play an important role in invasive breast tumor growth and progression. For this, we employed the murine mammary metastatic (4T-1) and nonmetastatic carcinoma (67NR) model. Our results showed that there is high expression of adenosine A2A receptor in breast tumor 4T-1 compared to 67NR. The A2AR deficiency prevented the growth of metastatic breast tumor 4T-1 and tumor cell colonies in secondary disease sites, together with a decrease in Th17 response profile and in the neutrophils recruitment to the primary site. The tumor cells 4T-1 presented high expression of adenosine receptors and ectonucleotidases (CD73 and CD39), suggesting that the adenosine signaling pathway has a direct effect on these cells. Adenosine or AMP (adenosine triphosphate) administration in tumoral 4T-1 cell cultures induced an increase in IL-6, CCL20 and CXCL1 expression, important mediators in the recruitiment of IL-17 producing T lymphocytes and neutrophils. Besides that, during 4T-1 tumor growth we observed high expression of cyclooxygenase 2 (Cox-2) compared to 67RN tumor. However A2A deficient mice showed a significative reduction in Cox-2 expression in tumoral microenvironment compared to BALB/c mice. Cox-2 inhibition with indometacine or celecoxib (selective inhibitor) in mice with 4T-1 tumor prevented primary tumor growth and, consequently, resulted in reduction of the expression of molecules related to the Th17 profile, as IL-6, CCL20, IL-17A and Ror?t. These results indicate that the blockade of the A2A signaling pathway interfere on the induction of Th17 cells response, opening new perspectives for the development of alternative therapies to the control of invasive tumors

Recentemente demonstrou-se o papel de células T produtoras de IL-17 na patogênese da esclerosa múltipla e de seu modelo, a EAE. Através da produção desta e de outras citocinas, a população chamada Th17 promove o rompimento da barreira hematoencefálica e a conseqüente infiltração de células patogênicas para dentro do SNC. Nesse contexto, em nosso trabalho utilizamos o fenômeno da tolerância oral para avaliar a capacidade deste em suprimir a resposta imune durante o modelo de EAE, mais especificamente as células Th17. Nossos dados demonstram uma diminuição de IL-17 tanto na periferia como no SNC dos animais tolerados. Além disso, detectamos menos CCL2 e IL-6 em células extraídas do CNS dia 10 pós-imunização. Não observarmos diferença na produção de IL-4,5,10, 13, IL-12p70, TNF-<font face=\"symbol\">a, e IFN-<font face=\"symbol\">g entre os grupos. Em suma, nossos resultados mostram que o fenômeno da tolerância oral é capaz de suprimir parâmetros de EAE devido a uma menor capacidade linfoproliferativa associada a uma supressão de células patogênicas Th17 tanto na periferia como no SNC. / It has recently been shown the role of IL-17 secreting cells on the pathogenesis of multiple sclerosis and also in its model, EAE. Due to the secretion of this and other cytokines, the population so called Th17, promotes the disruption of the blood-brain barrier and the following infiltration of pathogenic cells into the CNS. In this context, in our work we used the oral tolerance phenomenon to evaluate its supressive capacity, more specifically over the Th17 cells. We showed that oral tolerated mice has a diminished production of IL-17 both in the periphery and in the CNS. Futhermore, we detected lower levels of CCL2 and IL-6 also from brain and spinal cord extracted mononucear cells at day 10th post-immunization. We were not able to detect differences on IL-4,5,10, 13, IL-12p70, TNF-<font face=\"symbol\">a, e IFN-<font face=\"symbol\">g between the groups. Thus, our results show that the oral tolerance phenomenon suppresses EAE findings, mainly due to a lower lymphoprolipherative response associated to a supression over the expansion of Th17 pathogenic T cells both in the periphery and inside the CNS.

Introdução: O Papilomavírus Humano está muito bem associado com diversos tipos de cânceres humanos, como câncer anogenital e oral. Alguns estudos demonstram que o aparecimento de lesões e a progressão para o câncer estão relacionados ao tipo de resposta imune do hospedeiro. Deste modo, evidências indicam que a resposta imune do hospedeiro tem um papel muito importante para o curso da infecção pelo HPV. Objetivo: Avaliar a resposta imune específica in vitro ao Papilomavírus Humano (HPV) em homens com lesões causadas por HPV e sem lesão por HPV. Material e Métodos: Foram recrutados 31 pacientes e 11 voluntários, que formaram 4 grupos de estudo; sendo 12 pacientes no Grupo A (HIV +/ HPV +); 09 pacientes no Grupo B (HIV-/HPV+); 10 pacientes no Grupo C (HIV+/ HPV-); e 11 indivíduos saudáveis no Grupo D (HIV-/HPV-). Foram realizados ensaios de cultura celular para mensurar a resposta celular específica \"in vitro\" do tipo Th1/Th2/Th17 (INF-y, IL-2, TNFalfa, IL-4, IL-10 e IL-17) sob o estímulo da vacina quadrivalente do HPV (HPV 6, 11, 16 e 18) e à proteína E7 de HPV-16. Resultados: O grupo coinfectado (HIV +/ HPV+) apresentou níveis mais elevados de citocinas, principalmente do perfil Th2, comparando-se com os dados dos demais grupos de estudo. O grupo coinfectado apresentou níveis elevados de IL-6 e IL-10 (Perfil Th2) em relação ao grupo controle (HIV-/HPV-), com significância estatística (p < 0.0001 e p < 0.0001, respectivamente). Conclusão: Foi demonstrada uma elevada produção de citocinas no grupo HPV+/HIV+, sugerindo uma forte imunomodulação pela coinfecção HIV/HPV. Entretanto, novos estudos devem ser realizados para comprovar estes dados. Além de apresentar um perfil essencialmente Th2 do grupo coinfectado, principalmente pelos níveis elevados de IL-6 e IL-10 apresentados, sugerindo que estas duas citocinas possam servir como biomarcadores para persistência viral, uma vez que, os pacientes soropositivos para HIV apresentam maior persistência de HPV, e monitorar a progressão para lesões mais graves / Introduction: Human Papillomavirus is associated with different types of human cancers, such as anogenital and oral cancer. Some studies show that the appearance of lesions and progression to cancer are related to the type of host immune response. Thus, evidence indicates that the host immune response has a role key in the course of HPV infection. Objective: To evaluate the specific immune response in vitro to HPV in men with lesions caused by HPV and without injury caused by HPV. Methods: We recruited 31 patients and 11 volunteers, who formed four groups, with 12 patients in Group A (HIV+/HPV+); 09 patients in Group B (HIV-/HPV+); 10 patients in Group C (HIV+/HPV-) and 11 healthy subjects in Group D (HIV-/HPV-). Cells culture assay was performed to measure the specific immune response \"in vitro\" Th1/Th2/Th17 (IFN-y, IL-2, TNF-alfa, IL-4, IL-10 and IL-17) under the stimulation of quadrivalent HPV vaccine (HPV 6, 11, 16 and 18) and the E7 protein of HPV-16. Results: The coinfected group (HIV+/HPV+) had higher levels of cytokines, especially Th2 profile, compared with data from the other study groups. The coinfected group showed high levels of IL-6 and IL-10 (Th2 profile) compared to the control Group (HIV- /HPV-), with statistical significance (p < 0.0001 and p < 0.0001, respectively). Conclusion: This study demonstrated a high production of cytokines in the coinfected group, suggesting a strong immunomodulation by coinfection HIV/HPV. However, further studies should be conducted to confirm these data. In addition to presenting essentially a Th2 profile, especially by high levels of IL-6 and IL-10 presented, suggesting that these two cytokines may serve as biomarkers for viral persistence, since HIV seropositive patients have a higher persistence of HPV, and monitor the progression to more serious injuries

L’inflammasome NLRP3 est un complexe multiprotéique responsable notamment de la production d’IL-1β, une cytokine inflammatoire. Les effets délétères de l’inflammasome NLRP3 ont été démontrés dans de nombreuses maladies dont le cancer. Ce travail se concentre sur les effets de NLRP3 dans le contexte du cancer.Dans un premier projet, j’ai étudié l’activation de l’inflammasome NLRP3 dans les MSDC après un traitement par chimiothérapie. Deux chimiothérapies, le 5-Fluorouracile et la Gemcitabine, sont capables d’éliminer de façon spécifique les MDSC, population de cellules immunosuppressives dont la taille augmente en cas de cancer. J’ai découvert que le 5-Fluorouracile et la Gemcitabine activaient l’inflammasome NLRP3 dans les MDSC. En effet, le 5-Fluorouracile et la Gemcitabine provoquent la perméabilisation du lysosome des MDSC, permettant la sortie de la cathepsine B, protéine lysosomale, dans le cytoplasme où elle interagit directement avec NLRP3. Cette interaction active l’inflammasome NLRP3 et la production d’IL-1β. Cette IL-1β est responsable du développement d’une nouvelle population immunosuppressive, les Th17.J’ai ensuite étudié le rôle de NLRP3 dans la différenciation des lymphocytes T CD4 Th2. Dans ces cellules, le rôle de NLRP3 s’effectue indépendamment du reste du complexe multiprotéique qui forme l’inflammasome. Après avoir été induit par la cascade de signalisation de l’IL-2, NLRP3 interagit avec IRF4 (interferon regulatory factor) et agit comme un facteur de transcription sur le promoteur du gène de l’IL-4. L’absence de NLRP3 a pour conséquence une production moins importante d’IL-4 par les Th2 qui sont alors moins fonctionnels / The inflammasome NLRP3 (NOD like receptor pyd containing 3) is a multiprotein complex notably responsible for IL-1β (interleukine-1β) production, an inflammatory cytokine. Negative effects have been observed in various diseases including cancer. My thesis focuses on the effects of NLRP3 in cancer.In my first project, I studied the NLRP3 inflammasome activation in MDSC (myeloïd derived suppressor cells) after a chemotherapy treatment. Two chemotherapies, 5-Fluorouracil and Gemcitabine, are selectively able to kill MDSC, an immunosuppressive population growing during cancer evolution. MDSC’s death restores anti-tumor immunity for a while but another immunosuppressive population is established by MDSC produced IL-1β before their disappearance. I discovered that 5-Fluorouracil and Gemcitabine trigger NLRP3 inflammasome activation in MDSC. 5-Fluorouracil and Gemcitabine induce lysosomal permeabilisation, allowing for Cathepsin B release into the cytoplasm where it directly interacts with NLRP3. That interaction activates the inflammasome and induces IL-1β production which is responsible for the development of another immunosuppressive population, called Th17 cells.I then studied the role of NLRP3 during Th2 differentiation. Here, NLRP3 actions are done independently of the other inflammasome forming proteins. After being induced by IL-2 signalization pathway, NLRP3 interacts with IRF4 (interferon regulatory factor 4) and acts as a transcription factor on the IL-4 promoter gene. Lack of NLRP3 leads to a smaller IL-4 production by Th2 cells which are consequently less functional

La spondyloarthrite (SpA) est un rhumatisme inflammatoire chronique fréquent avec une prévalence de 0,43% en France, fortement associée à HLA-B27. À l’heure actuelle, il n’existe aucun traitement curatif et les mécanismes physiopathologiques impliqués restent méconnus. Afin de mieux comprendre les mécanismes immunologiques impliqués dans le développement de la SpA, nous avons étudié deux populations cellulaires clé, les cellules dendritiques (DC) et les lymphocytes T (LT) CD4+, chez le rat transgénique pour le HLA-B27 et la β2 microglobuline humaine (rat B27) qui développe spontanément tous les symptômes de la SpA. Il a été démontré que l’accumulation de lymphocytes T helper producteurs d’interleukine 17 (IL-17) pathogénique (lymphocyte Th17), et plusieurs défauts fonctionnels des cellules dendritiques (DCs) sont corrélés avec le développement de la SpA chez les rats B27.Nous nous sommes tout d’abord intéressés aux lymphocytes T régulateurs (Treg), dont le rôle est d’empêcher l’établissement d’une réponse immune pathogène pour l’hôte, chez le rat B27. Nous avons découvert que les Treg de rats B27 présentent un phénotype pro-inflammatoire (surexpression d’IL-17 et sous-expression d’IL-10 anti-inflammatoire), lié à la surexpression de la molécule ICOS. De plus, la sévérité des signes cliniques chez les rats B27 n’exprimant pas ICOS (rats B27 ICOS KO) est diminuée comparé aux animaux HLA-B27 sauvages. Cette protection partielle est corrélée à une réduction de la proportion de lymphocytes Th17. Ces résultats mettent en lumière le rôle majeur d’ICOS dans la physiopathologie de la SpA du rat.La deuxième partie de ce travail s’est concentrée sur les conséquences de la sous-expression d’IL-27 par les DC de rats B27, cytokine connue pour inhiber le développement des Th17. Nous avons observé que l’addition d’IL-27 exogène permet de diminuer la production d’IL-17 et d’augmenter la synthèse d’IL-10 anti-inflammatoire par les LT différenciés (T effecteurs et Treg) et les LT naïfs de rats B27 différenciés in vitro. De façon intéressante, l’IL-27 réduit également la synthèse d’IL-17 par les LT CD4+ circulants de patients atteints de SpA.Ces travaux démontrent pour la première fois le rôle clé de l’IL-27 et d’ICOS dans le contrôle de l’inflammation chez le rat B27 et suggèrent fortement que ces deux molécules sont de nouvelles cibles thérapeutiques prometteuses dans la SpA. / Spondyloarthritis (SpA) is a frequent chronic rheumatic inflammatory disorder with a prevalence of 0.43% in France and closely associated to HLA-B27. To date, there is no curative treatment and pathophysiological mechanisms involved in this pathology remain elusive. To better understand these mechanisms, we studied two crucial cell populations, dendritic cells (DC) and CD4+ T cells in rats transgenic for HLA-B27 and human β2 microglobulin (B27 rats) which spontaneously develop a phenotype closely resembling human spndyloarthritis. Previous studies demonstrated that accumulation of pathogenic IL-17 producing T cells (Th17 cells) and several function defects of DCs are correlated with SpA development in B27 rats.First, we focused on regulatory T cells, whose role is to prevent the establishment of pathogenic immune responses. We discovered that Treg from B27 rats have a pro-inflammatory phenotype. They overexpress IL-17 and underexpress anti-inflammatory IL-10, linked to ICOS overexpression. Furthermore, B27 rats knock-out for ICOS (B27 ICOS KO rats) have reduced severity of clinical symptoms compared to B27 ICOS WT rats. This protective effect is correlated with a reduced proportion of Th17 cells. These results highlight the crucial role of ICOS in rat SpA physiopathology.In the second part of this work we studied the consequences of IL-27 underexpression by B27 DC, a cytokine known to inhibit Th17 development. Addition of exogenous IL-27 reduces IL-17 and increases IL-10 productions by differentiated T cells (Teff and Treg) and by naive T cells polarized in vitro. Interestingly, IL-27 also reduces IL-17 production by circulating CD4+ T cells isolated from blood of SpA patients.This work demonstrate for the first time the key role of IL-27 and ICOS in the control of inflammation in B27 rats and highly suggest that these molecules may be new promising therapeutic targets in SpA.

<p>Understanding the mechanisms that control T cell function and differentiation is crucial to develop new strategies to modulate immune function and prevent autoimmune and inflammatory disease. The balance between effector (Teff; Th1, Th2 and Th17) and regulatory (Treg) T cells is critical to provide an appropriate, but not excessive, immune response and therapies to induce Treg or inhibit Teff are likely promising treatment strategies. It has recently become clear that T cell metabolism is important in both T cell activation and differentiation. T cells undergo a metabolic reprogramming upon activation and not all differentiated T cell subsets utilize the same metabolic fuels or programs.</p><p>These metabolic differences are not trivial, as T cell metabolism is tightly</p><p>regulated and dysregulation can lead to cell death or reduced immunity. An</p><p>understanding of the metabolic differences between Teff and Treg may lead to a new direction for treating inflammatory diseases by modulating the Teff:Treg balance through metabolic inhibition. Previous studies have shown that Teff express higher levels of the glucose transporter Glut1 than Treg, however the role of Glut1, and importantly, the cell-intrinsic role of glucose metabolism in T cell differentiation and inflammation was not previously examined. The work presented here examines the role of Glut1 in T cell differentiation. We show that effector CD4 T cells were dependent on Glut1 for proliferation and function both in vitro and in vivo. In contrast, Treg were Glut1-independent and capable of suppressing colitis in the absence of Glut1 expression.</p><p>Additionally, previous studies have shown broad metabolic differences between Teff and Treg, however the specific metabolic profiles of Teff and Treg are poorly understood. Here, Teff and Treg metabolism is examined to test if dependence on distinct metabolic pathways will allow selective targeting of different T cell populations. We show that pyruvate dehydrogenase kinase 1 (PDHK1) is differentially expressed in the T cell subsets and inhibition of PDHK1 selectively suppresses Th17 and promotes Treg differentiation and function. Because Teff and Treg have distinct metabolic profiles, we hypothesized that the Treg-­specific transcription factor FoxP3 may drive the Treg oxidative metabolic program. We therefore examined the role of FoxP3 in T cell metabolism and determined that FoxP3 promotes glucose and lipid oxidation and suppresses glycolytic metabolism. Importantly, we show that promoting glycolysis with transgenic expression of Glut1 inhibits Treg suppressive capacity. Together, these data suggest that FoxP3 drives an oxidative metabolic program that is critical to Treg function. Overall, this work examines the metabolic phenotypes and regulation of Teff and Treg and potential metabolic targets that could be used to treat autoimmune and inflammatory disease.</p> / Dissertation

Control strategies against the parasitic nematode Teladorsagia circumcincta are problematic under current sheep management systems. Infection with the parasite, particularly in young lambs, results in significant production losses therefore sustainable worm control is being sought. It has been established that variation in resistance to T. circumcincta is under genetic control and the development of resistance is an acquired characteristic and has an immunological basis. This project investigated the immunological response to infection, of lambs with predicted resistance or susceptibility to T. circumcincta. Specifically, the study aimed to identify immune response-associated genes that were differentially-expressed in resistant and susceptible lambs and attempted to identify mutations in these genes. This study was part of a long term project that aims to identify genetic marker/s to aid in marker-assisted selection (MAS) for resistance to T. circumcincta. Real time reverse transcription-quantitative polymerase chain reaction (real time RTqPCR) was performed on abomasal mucosa and lymph nodes from 55 lambs used in a previous experiment. The lambs had been either trickle-infected with 2,300 infective larvae every two days over three months (infected resistant/susceptible, n=45) or sham-dosed (non-infected control, n=10). Lambs were ranked in relation to faecal egg count (FEC) and adult worm count (AWC) at post mortem; zero or low FEC (resistant) to high FEC (susceptible). Histopathology showed only mild pathological changes in the abomasal mucosa of resistant lambs but heavy lymphocytic inflammatory infiltration in the mucosa and submucosa of infected susceptible animals. Measurements of a range of cytokine transcripts and cell markers associated with the four major CD4+ T cell subsets identified IL6, IL21, and IL23A as significantly increased by at least two-fold in abomasal lymph nodes and abomasal mucosa of susceptible lambs in comparison to resistant animals. Highly significant (P<0.02) positive correlations were found between IL6 (ρ=0.35), IL21 (ρ=0.54) and IL23A (ρ=0.38) transcript levels and AWC. Similarly, there were highly significant (P<0.01) positive correlations between FEC and IL6 (ρ=0.41), IL21 (ρ=0.65) and IL23A (ρ=0.31). In contrast, significant negative correlation (P<0.04) between IL23A with IgA antibody levels (ρ=-0.31) was found. There was also a significant positive correlation (P<0.03) of TGFB1 levels with AWC (ρ=0.42) and FEC (ρ=0.32) in the abomasal mucosa. These data suggests that susceptibility to T. circumcincta is linked to the activation of the inflammatory TH17 T cell subset and that this chronic inflammatory response was inappropriate to clear worm infection. High resolution melt analysis failed to identify single nucleotide polymorphisms in the coding regions of IL21 and IL21R. This is the first report of the involvement of TH17 response in GI worm infection in sheep. Similar gene expression studies involving the known upstream and downstream players of the TH17 response could be done.