En réponse à une infection, un signal de danger, ou de cytokines inflammatoires, les cellules dendritiques subissent un programme de maturation augmentant leur capacité à activer les lymphocytes T CD4+ et CD8+. Au cours de ce travail nous avons cherché à caractériser la reprogrammation transcriptomique des DC lors de l'infection par le cytomégalovirus murin (MCMV). Nous avons identifié un programme commun de maturation entre les différentes sous-populations de DC spléniques. Nous avons mis en évidence qu'il existe un programme transcriptomique de maturation commun à toutes les sous-populations de DC, induit par tous les stimuli examinés et évolutivement conservé au sein des mammifères. Nous avons également identifié les interférons (IFN) de type I comme des cytokines majeures promouvant la maturation des DC in vivo. La perte spécifique par les DC de la capacité à répondre aux IFN de type I entraine une diminution de la survie des souris lors de l'infection par le MCMV, révélant pour la première fois l'importance des effets intrinsèques cellulaires des IFN de type I sur les DC pour la résistance à une infection virale.Le développement puis l'utilisation d'un nouveau modèle de souris mutante ciblant la sous-population de DC XCR1+ nous a permis de mettre mis en évidence pour la première fois un rôle de ces cellules pour l'activation des lymphocytes T CD8 mémoires (Tm CD8+) dans l'infection par Listeria monocytogenes, et d'identifier les mécanismes sous-jacents. Les DC XCR1+ interagissent in situ avec les Tm CD8+. La synthèse de la chimiokine CXCL9 et la production d'interleukine-12 par les DC XCR1+ attirent et activent de façon optimale les Tm CD8+ qui produisent de l'IFN-γ. / Dendritic cells (DC) sense danger, microbial and cytokine signals that drive DC maturation which in turn allows proper activation of T lymphocytes. We characterized the gene expression program of splenic DC in vivo during murine cytomegalovirus (MCMV) infection. We identified a core set of genes commonly regulated in all subsets of mouse spleen DC. This set of genes was regulated upon DC maturation irrespective of the stimuli used and of the responding DC subsets and it was conserved between mouse and human. We identified type I interferon (IFN) as a major cytokine driving the expression of this core gene set in DC subsets. The loss of type I IFN responsiveness selectively in DC resulted in an increased mortality of mice after MCMV infection, unraveling a crucial role of cell-intrinsic responses to type I IFN in DC during a viral infection in vivo.We also developed and studied a new mouse model to target the XCR1+ DC subset in vivo. We found for the first time that XCR1+ DC promote recall of memory CD8 T cells upon secondary Listeria monocytogenes infection in vivo, and we identified the underlying mechanism. XCR1+ DC attract memory CD8 T cells through the secretion of the chemokine CXCL9. This attraction leads to an increase in the IFN-γ production by memory CD8 T cells. XCR1+ DC also induce the proliferation of memory CD8 T cells. This work significantly advanced our understanding of the in vivo functions of DC during infections.

L’ischémie/reperfusion (I/R) induit des lésions impliquées dans la physiopathologie de la transplantation cardiaque où elles contribuent à augmenter le rejet de greffe. Le stress induit par l’ischémie entraîne la libération d’ATP conduisant à l’activation de récepteurs purinergiques (P2R) dont l’expression est établie au niveau cardiaque et immunitaire. L’objectif de ce travail a été d’explorer l’effet de la signalisation P2R sur le phénotype des cellules dendritiques (DCs) et la réponse des cardiomyocytes (CM) à l’I/R. Nous avons montré que la récepteur P2Y11 (P2Y11R) avait une action immunomodulatrice sur les DCs en diminuant la sécrétion d’IL-6 et IL-12 et en inhibant la polarisation de la réponse adaptative vers Th1. Le post-conditionnement pharmacologique ciblant P2Y11R a apporté une protection efficace sur les CM en limitant le stress oxydant et en activant la PKCe connue pour inhiber l’ouverture du mPTP. Les effets protecteurs et immunomodulateurs de P2Y11R se sont confirmés in vivo en diminuant le rejet allogénique dans un modèle murin de transplantation cardiaque hétérotopique. Nos résultats suggèrent que P2Y11R pourrait être une cible thérapeutique apportant des effets bénéfiques en transplantation cardiaque. / Ischemia/reperfusion (I/R) injuries are involved in the pathophysiology of heart transplantation where they will increase graft rejection. Ischemia generates cellular stress leading to ATP release in the extracellular medium that may activate purinergic receptors (P2R) expressed by cardiomyocytes and immune cells. Therefore, these receptors may play important regulatory roles. The aim of this study was to investigate the effect of P2R signaling on dendritic cells phenotype (DCs) and cardiomyocyte (CM) response to I/R. We showed that P2Y11 receptor (P2Y11R) exhibited an immunomodulatory role in DCs by decreasing release of IL-6 and IL-12 and inhibiting polarization of the adaptive response towards Th1. Pharmacological post-conditioning targeting P2Y11R provided effective protection to CM by limiting oxidative stress and activating PKCe known to inhibit the opening of the mPTP. The protective and immunomodulatory effects of P2Y11R stimulation were confirmed in vivo by the decrease of allogeneic acute rejection in a murine model of heterotopic heart transplantation. In conclusion, our results strongly suggest that P2Y11R may be a promising therapeutic target providing beneficial effects in cardiac transplantation.

Células dendríticas (DCs) são células do sistema imunológico muito importantes no processo de indução de imunidade, capazes de conectar respostas imunes inata e adquirida e levar à ativação de células T e B. Recentemente demonstrou-se que é possível direcionar antígenos diretamente para as DCs in vivo através da administração de baixas doses de uma proteína recombinante híbrida fusionada a um anticorpo monoclonal específico para receptores presentes na superfície destas células. Dois dos anticorpos monoclonais utilizados tem a capacidade de ligar-se aos receptores endocíticos DEC205 e DCIR2 presentes na superfície de duas sub-populações distintas de DCs.Construiu-se anticorpos híbridos em fusão com os genes que codificam a proteína MSP-119 presente na superfície das formas merozoítas de P.yoelii e P.vivax. Ensaios de imunização mostraram que o anticorpo anti-DEC fusionado a qualquer das duas proteínas foi capaz de induzir principalmente uma resposta imune celular quando administrado na presença de diferentes adjuvantes. Já a resposta imune humoral foi modulada dependendo de várias combinações. / Dendritic cells (DCs) are cells of the immune system very important in the process of induction of immunity. They are able to connect innate and acquired immune responses and lead to activation of T and B cells. Recently it was shown that it is possible to target antigens directly to DCs in vivo by the administration of low doses of a recombinant hybrid protein consisting of a monoclonal antibody specific for receptors present on the surface of these cells fused with the antigen of interest. When these hybrid antibodies were injected in animals in the presence of a DC maturation stimulus, strong immune response against different antigens was obtained. Two of the monoclonal antibodies used have the ability to bind to either the DEC205 or the DCIR2 endocytic receptors present on the surface of two distinct DC sub-populations. In this work, we constructed hybrid antibodies fused with the sequence encoding the MSP-119 protein present on the surface P. yoelii and P. vivax merozoites. Most antibodies were successfully produced and maintained their ability to bind to their respective receptors. Immunization trials showed that the anti-DEC antibody fused to any of the two proteins was able to induce mainly a cellular immune response when administered in the presence of adjuvants. On the other hand, the humoral immune response depends of some combinations.

A paracoccidioidomicose (PCM) é uma micose sistêmica, endêmica na América Latina, causada pelo fungo dimórfico térmico Paracoccidioides brasiliensis (P. brasiliensis), cujo principal componente antigênico é a glicoproteína de 43 kDa (gp43). Diferentes formas clínicas podem ser desenvolvidas e estão diretamente associadas com vários graus de depressão da resposta imune celular. Considerando a importância das células dendríticas na interação dos sistemas imune inato e adaptativo, e na ativação de células T \"naive\", no presente trabalho estudamos se células dendríticas interagem com leveduras de P. brasiliensis, assim como seu principal componente antigênico (gp43). Foi demonstrado pela primeira vez que células dendríticas poderiam ser infectadas por leveduras de P. brasiliensis, e esse fungo permaneceu viável após fagocitose. Analisamos por citometria de fluxo a expressão das moléculas de superfície observando diminuição significativa da expressão das moléculas de MHC-II, CD80 e CD54 em células dendríticas quando estas foram incubadas com leveduras da cepa Pb18 ou com gp43. Esse resultado mostrou que a ação do P. brasiliensis em células dendríticas poderia ser mediada pela gp43. Ao analisarmos a síntese de IL-12, observamos diminuição significativa desta interleucina, quando células dendríticas ativadas com LPS, foram cultivadas na presença de leveduras de P. brasiliensis ou gp43. Esses resultados sugerem que a gp43 pode afetar várias funções das células hospedeiras, indicando que esta inibição pode ser usada pelo P. brasiliensis para reduzir a eficiência da resposta imune, facilitando assim o estabelecimento da infecção primária indivíduos suscetíveis. / Paracoccidioidomycosis, endemic in Latin America, is a progressive systemic mycosis caused by dimorphic fungus Paracoccidioides brasiliensis, where the major antigenic component is a glycoprotein of 43kDa (gp43). The infection can evolve to different clinical forms that are associated to various degrees of suppressed cell-mediated immunity. The role of dendritic cells (DCs) in P.brasiliensis infection has never been investigated. With the recognition that DCs are able to initiate response in naïve T cells and that they also participate in Th cell education the present study was undertaken to understand whether DCs interact with P. brasiliensis or gp43, as well as to elucidate possible mechanisms and consequences of this interaction. In the present report, it was demonstrated for the first time that DCs could be infected by P. brasiliensis and survive. Our results indicate that P. brasiliensis infection and purified gp43 lead to down-regulation of MHC-II and adhesion properties of immature DCs. The down-regulation was also observed in LPS-induced DCs maturation, where the expression of MHC-II, CD80, CD54 and CD40 were significantly inhibited in the presence of P. brasiliensis or gp43. These data show that the actions of P. brasiliensis on DCs could be mediated by gp43. In addition, an inhibition of IL-12 production by gp43 was observed in LPS-induced DC maturation. These results suggest that gp43 affects many functions of the host cells, indicating that this inhibition might be used by P. brasiliensis to reduce the effectiveness of the immune response, thus facilitating the establishment and fate of primary infection in susceptible host.

O sistema nervoso simpático (SNS) inerva a maioria dos órgãos linfoides e durante situações de estresse, por meio da liberação da noradrenalina de seus ramos eferentes, emitem sinais capazes de modular as repostas imunes. Nossa hipótese foi de que esta via poderia alterar a função de células dendríticas (DCs) derivadas de monócitos humanos, uma vez que receptores adrenérgicos já foram demonstrados em DCs murinas e pelo fato de que as estas células são chave na iniciação de respostas imunes adaptativas, bem como indutoras de tolerância. Desta forma, DCs diferenciadas a partir de monócitos sanguíneos provenientes de doadores saudáveis tratadas com ligantes adrenérgicos foram analisadas quanto a expressão de marcadores de membrana, atividade fagocítica, apresentação antigênica em ensaio de reação mista de linfócitos, expressão gênica de marcadores de diferenciação e ativação, bem como produção de citocinas. Os resultados revelaram que as DCs apresentam transcritos apenas para o adrenoceptor β2, e esta expressão é similar à de macrófagos, mas inferior a de linfócitos. A análise dos marcadores fenotípicos de membrana, atividade fagocítica, apresentação antigênica e produção de citocinas não mostraram alterações nas células tratadas com agonistas adrenérgicos. No entanto, o tratamento com ligantes adrenérgicos foi capaz de alterar a expressão dos genes CD40, CD80, CD83, CXCL1, TGFB1, FCGR3A, CCR7 e CCL5 em DCs e macrófagos estimulados com LPS ou TNF-α. Embora os efeitos dos ligantes adrenérgicos não tenham sido fortemente evidenciados nos testes realizados, os resultados sugerem que pequenas alterações podem ser provocadas pela ligação das catecolaminas em DCs, sugerindo que estas possam ser moduladas pelo SNS, modificando as respostas imunes / The sympathetic nervous system (SNS) innervates most of the lymphoid organs and during stress situations, by releasing norepinephrine from its efferent branches, emit signals capable of modulating immune responses. Our hypothesis was that this pathway could alter the function of human monocyte-derived dendritic cells (DCs), since adrenergic receptors have already been demonstrated in murine DCs, and by the fact that these cells are key in initiating adaptive immune responses as well as tolerance inducers. Thus, differentiated DCs from blood monocytes from healthy donors treated with adrenergic ligands were analyzed for expression of membrane markers, phagocytic activity, antigenic presentation in a mixed lymphocyte reaction assay, gene expression of differentiation and activation markers and cytokine production. The results revealed that DCs present transcripts only for the β2 adrenoceptor, and this expression is similar to that observed in macrophages, but lower than what it is found in lymphocytes. The analysis of phenotypic membrane markers, phagocytic activity, antigenic presentation and cytokine production did not revealed any changes in the cells treated with adrenergic agonists. However, treatment with the adrenergic ligands was able to alter the expression of CD40, CD80, CD83, CXCL1, TGFB1, FCGR3A, CCR7 and CCL5 genes in DCs and macrophages stimulated with LPS or TNF-α. Although the effects of adrenergic ligands have not been strongly demonstrated in the tests performed, the results suggest that small changes can be caused by the binding of catecholamines in DCs, suggesting that they can be modulated by the SNS, modifying immune responses.

A asma afeta aproximadamente 300 milhões de pessoas no mundo e é a maior causa de internação hospitalar em crianças nos países desenvolvidos. Essa doença é incurável e por vezes refratária ao tratamento em um número significativo de pacientes. As taxas de prevalência de tabagismo entre pacientes asmáticos são semelhantes aos da população em geral e o impacto da fumaça de cigarro nestes pacientes ainda é clinicamente controverso. O objetivo deste trabalho foi traçar o perfil inflamatório e analisar o papel das células dendríticas sobre a imunomodulação provocada pela exposição à fumaça do cigarro na inflamação alérgica pulmonar induzida previamente por ovalbumina (OVA) em um modelo murino. Primeiramente avaliamos in vivo a ação da fumaça de cigarro na inflamação pulmonar alérgica crônica avaliando a responsividade brônquica, o remodelamento pulmonar, a produção de anticorpos antígeno-específicos, o perfil de células inflamatórias pulmonares e sistêmicas e a produção de citocinas inflamatórias e moduladoras. Em seguida, realizamos estudo in vitro do perfil de maturação, migração e inflamatório de células dendríticas expostas a OVA e/ou a extrato de fumaça de cigarro. Nosso estudo mostrou que a sensibilização e desafios inalatórios com OVA levaram à inflamação pulmonar de característica Th2 com aumento de responsividade brônquica, remodelamento, altos níveis de IgE e de citocinas pró-inflamatórias como IL-4, IL-5 e IL-13. A exposição à fumaça de cigarro, surpreendentemente, levou a uma redução dos níveis de IL-4, IL-5 e IL-13, e simultaneamente reduziu os níveis de citocinas anti-inflamatórias como IL- 10 e TGF-beta em animais sensibilizados e desafiados com antígeno. Foi observada nestes animais, uma redução no número de eosinófilos no lavado broncoalveolar e aumento no número de neutrófilos no pulmão. A combinação da inflamação alérgica com exposição à fumaça de cigarro levou a um aumento do recrutamento e ativação de células dendríticas linfoides nos linfonodos mediastinais, que mostrou relação direta com aumento do influxo de células T CD8+ e ativação das mesmas no pulmão. A inflamação alérgica juntamente com a exposição à fumaça de cigarro, levou a uma redução no recrutamento de células dendríticas plasmocitoides além de reduzir o recrutamento de células T regulatórias. In vitro, mostramos que o extrato de fumaça de cigarro combinado ao antígeno aumenta a capacidade migratória e fagocítica do antígeno pelas BMDCs. No entanto, houve redução da expressão gênica para IL-13 neste mesmo grupo. Concluímos que neste modelo de inflamação pulmonar alérgica crônica combinada com a exposição à fumaça de cigarro leva a uma descaracterização do perfil inflamatório característico da resposta Th2 com a redução do recrutamento de eosinófilos, redução dos níveis de IL-4, IL-5 e IL-13 aliados a um aumento do número de neutrófilos, o que pode estar relacionado ao aumento do recrutamento e ativação de células dendríticas linfoides bem como de células T CD8+ e redução local de células dendríticas plasmocitoides. Mostramos ainda que a fumaça de cigarro juntamente com o antígeno leva as células dendríticas a aumentarem sua capacidade fagocítica porém, reduzir sua capacidade pró-inflamatória pela expressão gênica reduzida de IL-13 / Asthma affects approximately 300 million people worldwide and it is the major cause of hospitalization among children in developed countries. This disease is often refractory to treatment in a high number of patients. The prevalence rates of smoking among asthmatic patients are similar to the general population and the impact of cigarette smoke is also clinically controversial. The main goal of this study is to outline, in a murine model, the inflammatory profile and to analyze the role of dendritic cells on the immunomodulation caused by exposure to cigarette smoke in ovalbumin (OVA)-induced pulmonary allergic inflammation. First, we evaluated in vivo the action of cigarette smoke on chronic allergic pulmonary inflammation, evaluating the bronchial responsiveness, pulmonary remodeling, the production of antigen-specific antibodies, pulmonary and systemic inflammatory cell profile and the production of inflammatory and modulating cytokines. Next, we performed an in vitro study of the maturation, migration and inflammatory profile of dendritic cells exposed to OVA and/or cigarette smoke extract. Our study showed that sensitization and challenge with OVA led to Th2-type lung inflammation with increased bronchial responsiveness, remodeling, high levels of IgE and proinflammatory cytokines such as IL-4, IL-5 and IL-13. Exposure to cigarette smoke has surprisingly led to a reduction in levels of IL-4, IL-5 and IL-13, and simultaneously reduced levels of anti-inflammatory cytokines such as IL-10 and TGF-beta in animals sensitized and challenged with the antigen. We also observed a reduction in the number of eosinophils in bronchoalveolar lavage fluid and an increase in the number of neutrophils in the lung of these animals. The combination of allergic inflammation with exposure to cigarette smoke led to increased recruitment and activation of lymphoid dendritic cells in the mediastinal lymph nodes, which showed to be direct related with increased activation and influx of CD8+ T cells in lung. Allergic inflammation combined with cigarette smoke led to a decrease of plasmacytoid dendritic cells a well as regulatory T cells. In vitro, we showed that cigarette smoke extract combined with antigen increased migratory and phagocytic capacity of BMDCs. However, there was a reduction of IL-13 gene expression in this same group. We conclude that in this model of chronic pulmonary allergic inflammation combined with exposure to cigarette smoke leads to a mischaracterization of the characteristic inflammatory profile of the Th2 response with the reduction of eosinophil recruitment, reduction of levels of IL-4, IL-5 and IL-13 allied to increased number of neutrophils, which is related to increased recruitment and activation of lymphoid dendritic cells as well as CD8+ T cells and local decrement of plasmacytoid dendritic cells. We further show that cigarette smoke combined with antigen increases dendritic cell phagocytic capacity however, reduces its pro-inflammatory capacity by the reduced gene expression of IL-13

As células dendríticas (DCs) são críticas no processo de indução de imunidade e tolerância periférica. Estas células apresentam inúmeros receptores em sua superfície e podem ser classificadas em diferentes subpopulações de acordo com a expressão dos mesmos. Anticorpos monoclonais dirigidos contra diferentes receptores celulares vêm sendo utilizados com sucesso no tratamento de patologias como câncer. Nos últimos dez anos, demonstrou-se que é possível enviar antígenos diretamente para o receptor DEC205 presente na superfície das DCs. Quando isso acontece na ausência de estímulo inflamatório, o resultado é a indução de tolerância imunológica. Quando o antígeno é enviado a estas mesmas células na presença de um estímulo inflamatório, o resultado é a indução de uma forte resposta imunológica. Geramos um anticorpo <font face=\"Symbol\">aDEC205 em fusão com a proteína 2 de superfície dos amastigotas (ASP-2) de Trypanosoma cruzi e estudamos o efeito da administração deste anticorpo in vivo na presença de diferentes adjuvantes em duas linhagens distintas de camundongos. Observamos que ambas as linhagens de camundongos não foram capazes de gerar anticorpos, mas a resposta imune celular foi suficiente para reduzir a parasitemia quando desafiados com tripomastigotas cepa Y de Trypanosoma cruzi. Camundongos A/J foram capazes de mediar apresentação cruzada. Mapeamos um peptídeo que é reconhecido por camundongos BALB/c. Camundongos imunizados com Poly (I:C) como estímulo de maturação mostrou-se superior ao CpG. / As part of the innate immune system, dendritic cells (DCs) are critical in the process of induction of immunity and peripheral tolerance. These cells offer many receptors on their surface and can be classified in different subsets according to the expression there of. Monoclonal antibodies directed against different cellular receptors have been used successfully to treat diseases such as cancer. In the last ten years has been demonstrated that antigen can be sent directly to the receiver DEC205 present on the surface of DCs in vivo. When this happens in the absence of concomitant inflammatory stimulus, the result is the induction of immunological tolerance. Moreover, when the antigen is sent to the same cells in the presence of an inflammatory stimulus, the result is to induce a strong immune response. In this study we generated an antibody <font face=\"Symbol\">aDEC205 fusion protein with the amastigote surface protein 2 (ASP-2) from Trypanosoma cruzi and study the effect of administration of this antibody in vivo in the presence of different adjuvants in two distinct strains of mice. We observed that both strains of mice were unable to generate antibodies, but cellular immune response was sufficient to reduce the parasitemia when challenged with trypomastigotes Y strain of Trypanosoma cruzi. The mice A/J were able to mediate cross-presentation. In contrast, we mapped a peptide that is recognized by BALB/c mice. In counterpart mice immunized with Poly (I:C) as maturation stimuli was superior to CpG.

Le développement de vaccins à ARNm est en plein essor dans le domaine de la vaccinologie. Un des défis majeurs de ces approches est de parvenir à transporter et délivrer les ARNm dans le cytoplasme des cellules dendritiques (DC) pour permettre la production d’antigène et l’activation des réponses immunitaires.L’objectif de ce travail a porté sur la conception et l’évaluation de nouvelles stratégies de vectorisation des ARNm par des nanoparticules de poly(acide lactique) (NP-PLA). Une stratégie basée sur l’adsorption des ARNm à la surface des NP-PLA par l’intermédiaire du LAH4-L1, un peptide cationique amphipathique, a été développée. Des polyplexes formés suite à la condensation de l’ARNm par le LAH4-L1 ont été adsorbés sur les NP-PLA pour former des nanocomplexes. L’intérêt des polyplexes et nanocomplexes pour le ciblage des DC et l’activation des réponses immunitaires in vitro a été évalué. Dans des tests de transfection, ces formulations ont induit une expression efficace d’ARNm modèles dans des DC. La présence des NP-PLA dans les formulations semble favoriser l’expression des ARNm. Le traitement avec des inhibiteurs a révélé que les polyplexes et nanocomplexes semblent internalisés par phagocytose et endocytose clathrine-dépendante, et s’échappent des endosomes par un mécanisme dépendant de la pompe à protons v-ATPase. Suite à la transfection de DC dérivées de monocytes (moDC), nous avons montré que nos formulations stimulent les récepteurs de l’immunité innée et induisent une réponse pro-inflammatoire. Cette activation est associée à la maturation des moDC, à la présentation de peptides antigéniques sur le CMH-I et -II et à la sécrétion de cytokines et chémokines impliquées dans l’immunité adaptative. Ces données soulignent l’intérêt des NP-PLA associées au LAH4-L1 pour vectoriser des ARNm, cibler des DC et activer les réponses immunitaires. Dans le contexte du VIH-1, ce type de réponse pourrait aider le système immunitaire à contrôler la charge virale / MRNA-based vaccines currently raise a growing interest in vaccinology. However, the transport and delivery of mRNAs to DC cytoplasm in order to induce antigen production and immune responses remains challenging. The objective of this thesis concerns the design and evaluation of novel strategies to vectorize vaccine mRNAs by poly(lactic acid) nanoparticles (PLA-NPs). We developed a strategy based on mRNA adsorption onto PLA-NPs using, as intermediate, LAH4-L1, an amphipathic cationic peptide. To do this, mRNAs were condensed by LAH4-L1 to form polyplexes which was then adsorbed onto PLA-NPs in a second step to form nanocomplexes. The LAH4-L1/mRNA polyplexes and PLA-NP/LAH4-L1/mRNA nanocomplexes ability to target DCs and induce immune responses in vitro was evaluated. We showed that formulations induce an efficient transfection of mRNA in DCs in vitro. The addition of PLA-NPs in formulations seems to increase sustained expression of mRNAs. DC treatment by inhibitors revealed that polyplexes and nanocomplexes are taken up by phagocytosis and clathrin-dependent endocytosis, and escape endosomes by a v-ATPase-dependent mechanism. Transfection of monocyte-derived DCs (moDCs) showed that LAH4-L1/mRNA polyplexes and PLA-NP/LAH4-L1/mRNA nanocomplexes trigger innate-sensing activation with pro-inflammatory responses. This activation is associated with moDCs maturation, MHC-I and MHC-II presentation, and the secretion cytokines and chemokines involved in adaptive immunity.These data highlight the interest of these new platform formulations to vectorize mRNAs, target DCs and induce immune responses, which in the context of HIV-1, could help the immune system to control the viral load

Exossomos (Exo) originados de células dendríticas (DCs) carregam moléculas associadas à apresentação antigênica. Neste trabalho procurou-se estabelecer se Exo de DCs seriam capazes de conferir imunogenicidade às células tumorais. Os Exo isolados de culturas de DCs expressavam as moléculas HLA-ABC, HLA-DR, CD86, CD11c, CD81, CD54 e CD18. Estes foram então adicionados às células da linhagem humana de adenocarcinoma mamário, SK-BR-3, as quais passaram a expressar as moléculas HLA-DR, CD86 e CD11c. As células tumorais modificadas pelos Exo induziram a produção de IL-6 e IL-10, detectados no sobrenadante das co-culturas destas com linfócitos T. Estas células tumorais também induziram aumento do número de linfócitos produtores de IFN-<font face=\"Symbol\">g, pré-sensibilizados contra antígenos tumorais, e aumento da expressão de SOCS3 nestes. Em conclusão, nossos resultados mostram que, Exo de DCs alteram o fenótipo de células tumorais, modificando sua interação com linfócitos T, sem induzir nas mesmas capacidade de ativar respostas proliferativas ou citotóxicas de linfócitos T in vitro. / Exosomes (Exo) originated from dendritic cells (DCs) contain molecules involved in antigen presentation. The present work sought to determine if Exo from DCs would be able to transfer immunogenicity to tumor cells. Exo isolated from DCs cultures carried HLA-ABC, HLA-DR, CD86, CD11c, CD81, CD54 and CD18. These Exo were added to cultures of the human breast adenocarcinoma cell line, SK-BR-3, which gained expression of HLA-DR, CD86 and CD11c. Tumor cells modified by Exo induced IL-6 and IL-10 production, detected in the supernatant of their co-cultures with T lymphocytes. These tumor cells also induced an increase in the frequency of IFN-<font face=\"Symbol\">g-producing T lymphocytes, pre-sensitized against tumor antigens, and an increased expression of SOCS3. In conclusion, our results show that, Exo from DCs affect the phenotype of tumor cells, modifying their interaction with T lymphocytes, without inducing the ability to activate cytotoxic or T cell proliferative responses in vitro.

As células dendríticas (DCs) são as principais células apresentadoras de antígeno do sistema imune. No entanto, trabalhos têm demonstrado seu envolvimento na manutenção da tolerância imunológica. As células T CD4+CD25+Foxp3+ possuem a capacidade de suprimir respostas imunes. Neste estudo avaliamos as alterações ocorridas na população de células T CD4+CD25+Foxp3+ após co-cultura de células de linfonodo com DCs. Nossos resultados demonstram que após a co-cultura há um aumento da população de células CD4+CD25+Foxp3+ de maneira independente do estado de ativação das DCs ou da presença de antígenos exógenos. No entanto, o aumento observado é maior quando DCs imaturas são incubadas com antígenos exógenos. Notamos ainda que há presença de TGF-ß em todas as condições experimentais em que observamos aumento da população de células CD4+CD25+Foxp3+. Nossos dados sugerem ainda que este aumento se deve à proliferação das células T CD4+CD25+Foxp3+. / Dendritic cells (DCs) are the most important antigen-presenting cells of the immune system. However, DCs have also been implicated in maintaining immunologic tolerance. CD4+CD25+Foxp3+ T lymphocytes are known as cells with regulatory properties. In this study we evaluated the changes in the CD4+CD25+Foxp3+ T cell population after co-culture of lymph-node cells with DCs. Our results show an increase in the CD4+CD25+Foxp3+ T cell population after co-culture and occurs regardless of the activation state of DCs and the presence of exogenous antigens; however it is greater when immature DCs are previously pulsed with exogenous antigen. We also noticed that TGF-? is present in all cultures conditions in which the CD4+CD25+Foxp3+ T cell population increases. Our data also suggests that the increase of the CD4+CD25+Foxp3+ T cell population may be due to the proliferation of these cells.