RESUMO
The gastrointestinal tract is the largest mucosal surface in our body and accommodates the majority of the total lymphocyte population. Being continuously exposed to both harmless antigens and potentially threatening pathogens, the intestinal mucosa requires the integration of multiple signals for balancing immune responses. This integration is certainly supported by tissue-resident intestinal mesenchymal cells (IMCs), yet the molecular mechanisms whereby IMCs contribute to these events remain largely undefined. Recent studies using single-cell profiling technologies indicated a previously unappreciated heterogeneity of IMCs and provided further knowledge which will help to understand dynamic interactions between IMCs and hematopoietic cells of the intestinal mucosa. In this review, we focus on recent findings on the immunological functions of IMCs: On one hand, we discuss the steady-state interactions of IMCs with epithelial cells and hematopoietic cells. On the other hand, we summarize our current knowledge about the contribution of IMCs to the development of intestinal inflammatory conditions, such as infections, inflammatory bowel disease, and fibrosis. By providing a comprehensive list of cytokines and chemokines produced by IMCs under homeostatic and inflammatory conditions, we highlight the significant immunomodulatory and tissue niche forming capacities of IMCs.
Assuntos
Doenças Inflamatórias Intestinais , Células-Tronco Mesenquimais , Homeostase , Humanos , Imunidade nas Mucosas , Mucosa Intestinal , IntestinosRESUMO
Intestinal regulatory T cells (Tregs) are fundamental in peripheral tolerance toward commensals and food-borne antigens. Accordingly, gut-draining mesenteric lymph nodes (mLNs) represent a site of efficient peripheral de novo Treg induction when compared to skin-draining peripheral LNs (pLNs), and we had recently shown that LN stromal cells substantially contribute to this process. Here, we aimed to unravel the underlying molecular mechanisms and generated immortalized fibroblastic reticular cell lines (iFRCs) from mLNs and pLNs, allowing unlimited investigation of this rare stromal cell subset. In line with our previous findings, mLN-iFRCs showed a higher Treg-inducing capacity when compared to pLN-iFRCs. RNA-seq analysis focusing on secreted molecules revealed a more tolerogenic phenotype of mLN- as compared to pLN-iFRCs. Remarkably, mLN-iFRCs produced substantial numbers of microvesicles (MVs) that carried elevated levels of TGF-ß when compared to pLN-iFRC-derived MVs, and these novel players of intercellular communication were shown to be responsible for the tolerogenic properties of mLN-iFRCs. Thus, stromal cells originating from mLNs contribute to peripheral tolerance by fostering de novo Treg induction using TGF-ß-carrying MVs. This finding provides novel insights into the subcellular/molecular mechanisms of de novo Treg induction and might serve as promising tool for future therapeutic applications to treat inflammatory disorders.
Assuntos
Vesículas Extracelulares/imunologia , Linfonodos/imunologia , Células Estromais/imunologia , Linfócitos T Reguladores/imunologia , Animais , Linhagem Celular , Vesículas Extracelulares/genética , Vesículas Extracelulares/ultraestrutura , Fatores de Transcrição Forkhead/imunologia , Fatores de Transcrição Forkhead/metabolismo , Perfilação da Expressão Gênica/métodos , Mesentério/imunologia , Camundongos Endogâmicos BALB C , Camundongos Knockout , Microscopia Eletrônica de Varredura , Microscopia Eletrônica de Transmissão , Células Estromais/metabolismo , Células Estromais/ultraestrutura , Linfócitos T Reguladores/metabolismo , Fator de Crescimento Transformador beta/genética , Fator de Crescimento Transformador beta/imunologiaRESUMO
Adaptive immunity critically contributes to control acute infection with enteropathogenic Yersinia pseudotuberculosis; however, the role of CD4+ T cell subsets in establishing infection and allowing pathogen persistence remains elusive. Here, we assessed the modulatory capacity of Y. pseudotuberculosis on CD4+ T cell differentiation. Using in vivo assays, we report that infection with Y. pseudotuberculosis resulted in enhanced priming of IL-17-producing T cells (Th17 cells), whereas induction of Foxp3+ regulatory T cells (Tregs) was severely disrupted in gut-draining mesenteric lymph nodes (mLNs), in line with altered frequencies of tolerogenic and proinflammatory dendritic cell (DC) subsets within mLNs. Additionally, by using a DC-free in vitro system, we could demonstrate that Y. pseudotuberculosis can directly modulate T cell receptor (TCR) downstream signaling within naïve CD4+ T cells and Tregs via injection of effector molecules through the type III secretion system, thereby affecting their functional properties. Importantly, modulation of naïve CD4+ T cells by Y. pseudotuberculosis resulted in an enhanced Th17 differentiation and decreased induction of Foxp3+ Tregs in vitro. These findings shed light to the adjustment of the Th17-Treg axis in response to acute Y. pseudotuberculosis infection and highlight the direct modulation of CD4+ T cell subsets by altering their TCR downstream signaling.
Assuntos
Receptores de Antígenos de Linfócitos T/imunologia , Linfócitos T Reguladores/microbiologia , Células Th17/microbiologia , Infecções por Yersinia pseudotuberculosis/imunologia , Yersinia pseudotuberculosis/imunologia , Animais , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD4-Positivos/microbiologia , Diferenciação Celular , Feminino , Fatores de Transcrição Forkhead/imunologia , Interações Hospedeiro-Patógeno , Intestinos/imunologia , Intestinos/microbiologia , Camundongos Endogâmicos BALB C , Transdução de Sinais , Linfócitos T Reguladores/imunologia , Células Th17/imunologia , Yersinia pseudotuberculosis/fisiologiaRESUMO
Background: To successfully limit pathogen dissemination, an immunological link between the entry tissue of the pathogen and the underlying secondary lymphoid organs (SLOs) needs to be established to prime adaptive immune responses. Here, the prerequisite of CCR7 to mount host immune responses within SLOs during gastrointestinal Yersinia pseudotuberculosis infection to limit pathogen spread was investigated. Methods: Survival, bacterial dissemination, and intestinal and systemic pathology of wild-type and CCR7-/- mice were assessed and correlated to the presence of immune cell subsets and cytokine responses throughout the course of infection. Results: The CCR7-/- mice show a significantly higher morbidity and are more prone to pathogen dissemination and intestinal and systemic inflammation during the oral route of infection. Significant impact of CCR7 deficiency over the course of infection on several immunological parameters were observed (ie, elevated neutrophil-dominated innate immune response in Peyer's patches, limited dendritic cell migration to mesenteric lymph nodes [mLNs] causing reduced T cell-mediated adaptive immune responses (in particular Th17-like responses) in mLNs). Conclusions: Our work indicates that CCR7 is required to mount a robust immune response against enteropathogenic Y. pseudotuberculosis by promoting Th17-like responses in mLNs.
Assuntos
Predisposição Genética para Doença , Receptores CCR7/imunologia , Células Th17/imunologia , Infecções por Yersinia pseudotuberculosis/imunologia , Animais , Movimento Celular , Células Dendríticas/imunologia , Interações Hospedeiro-Patógeno/genética , Intestinos/imunologia , Intestinos/microbiologia , Linfonodos/imunologia , Linfonodos/microbiologia , Camundongos , Células Mieloides/imunologia , Nódulos Linfáticos Agregados/imunologia , Nódulos Linfáticos Agregados/microbiologia , Receptores CCR7/genética , Yersinia pseudotuberculosis , Infecções por Yersinia pseudotuberculosis/genéticaRESUMO
Tesmilifene, a tamoxifen analog with antihistamine action, has chemopotentiating properties in experimental and clinical cancer studies. In our previous works, tesmilifene increased the permeability of the blood-brain barrier (BBB) in animal and culture models. Our aim was to investigate the effects of tesmilifene on brain microvessel permeability in the rat RG2 glioma model and to reveal its mode of action in brain endothelial cells. Tesmilifene significantly increased fluorescein extravasation in the glioma. Short-term treatment with tesmilifene reduced the resistance and increased the permeability for marker molecules in a rat triple co-culture BBB model. Tesmilifene also affected the barrier integrity in brain endothelial cells co-cultured with RG2 glioblastoma cells. Tesmilifene inhibited the activity of P-glycoprotein and multidrug resistance-associated protein-1 efflux pumps and down-regulated the mRNA expression of tight junction proteins, efflux pumps, solute carriers, and metabolic enzymes important for BBB functions. Among the possible signaling pathways that regulate BBB permeability, tesmilifene activated the early nuclear translocation of NFκB. The MAPK/ERK and PI3K/Akt kinase pathways were also involved. We demonstrate for the first time that tesmilifene increases permeability marker molecule extravasation in glioma and inhibits efflux pump activity in brain endothelial cells, which may have therapeutic relevance. Tesmilifene, a chemopotentiator in experimental and clinical cancer studies increases vascular permeability in RG2 glioma in rats and permeability for marker molecules in a culture model of the blood-brain barrier. Tesmilifene inhibits the activity of efflux pumps and down-regulates the mRNA expression of tight junction proteins, transporters, and metabolic enzymes important for the blood-brain barrier functions, which may have therapeutic relevance.
Assuntos
Barreira Hematoencefálica/efeitos dos fármacos , Permeabilidade Capilar/efeitos dos fármacos , Endotélio Vascular/efeitos dos fármacos , Antagonistas dos Receptores Histamínicos/farmacologia , Éteres Fenílicos/farmacologia , Animais , Neoplasias Encefálicas/patologia , Linhagem Celular Tumoral , Células Cultivadas , Técnicas de Cocultura , Modelos Animais de Doenças , Feminino , Glioma/patologia , Imuno-Histoquímica , Masculino , Ratos , Ratos Wistar , Reação em Cadeia da Polimerase em Tempo RealRESUMO
Under physiological and pathological conditions, extracellular vesicles (EVs) are present in the extracellular compartment simultaneously with soluble mediators. We hypothesized that cytokine effects may be modulated by EVs, the recently recognized conveyors of intercellular messages. In order to test this hypothesis, human monocyte cells were incubated with CCRF acute lymphoblastic leukemia cell line-derived EVs with or without the addition of recombinant human TNF, and global gene expression changes were analyzed. EVs alone regulated the expression of numerous genes related to inflammation and signaling. In combination, the effects of EVs and TNF were additive, antagonistic, or independent. The differential effects of EVs and TNF or their simultaneous presence were also validated by Taqman assays and ELISA, and by testing different populations of purified EVs. In the case of the paramount chemokine IL-8, we were able to demonstrate a synergistic upregulation by purified EVs and TNF. Our data suggest that neglecting the modulating role of EVs on the effects of soluble mediators may skew experimental results. On the other hand, considering the combined effects of cytokines and EVs may prove therapeutically useful by targeting both compartments at the same time.
Assuntos
Citocinas/metabolismo , Exossomos/metabolismo , Linhagem Celular Tumoral , Quimiocina CCL2/metabolismo , Análise por Conglomerados , Citocinas/genética , Humanos , Interleucina-8/genética , Interleucina-8/metabolismo , RNA Mensageiro/metabolismo , Proteínas Recombinantes/biossíntese , Proteínas Recombinantes/genética , Proteínas Recombinantes/farmacologia , Fator de Necrose Tumoral alfa/genética , Fator de Necrose Tumoral alfa/metabolismo , Fator de Necrose Tumoral alfa/farmacologia , Regulação para Cima/efeitos dos fármacosRESUMO
Peripherally-induced regulatory T cells (pTregs) expressing the retinoic acid receptor-related orphan-receptor gamma t (RORγt) are indispensable for intestinal immune homeostasis. Nuclear factor kappa family members regulate the differentiation of thymic Tregs and promote their survival in the periphery. However, the Treg intrinsic molecular mechanisms controlling the size of the pTregs in the intestine and associated lymphoid organs remain unclear. Here, we provide direct evidence that B-cell lymphoma 3 (Bcl3) limits the development of pTregs in a T cell-intrinsic manner. Moreover, the absence of Bcl3 allowed for the formation of an unusual intestinal Treg population co-expressing the transcription factors Helios and RORγt. The expanded RORγt+ Treg populations in the absence of Bcl3 displayed an activated phenotype and secreted high levels of the anti-inflammatory cytokines interleukin (IL)-10 and transforming growth factor beta. They were fully capable of suppressing effector T cells in a transfer colitis model despite an intrinsic bias to trans-differentiate toward T helper 17-like cells. Finally, we provide a Bcl3-dependent gene signature in pTregs including altered responsiveness to the cytokines IL-2, IL-6, and tumor necrosis factor alpha. Our results demonstrate that Bcl3 acts as a molecular switch to limit the expansion of different intestinal Treg subsets and may thus serve as a novel therapeutic target for inflammatory bowel disease by restoring intestinal immune tolerance.
Assuntos
Proteína 3 do Linfoma de Células B , Diferenciação Celular , Camundongos Knockout , Membro 3 do Grupo F da Subfamília 1 de Receptores Nucleares , Linfócitos T Reguladores , Animais , Proteína 3 do Linfoma de Células B/metabolismo , Proteína 3 do Linfoma de Células B/genética , Linfócitos T Reguladores/imunologia , Camundongos , Membro 3 do Grupo F da Subfamília 1 de Receptores Nucleares/metabolismo , Membro 3 do Grupo F da Subfamília 1 de Receptores Nucleares/genética , Fatores de Transcrição/metabolismo , Fatores de Transcrição/genética , Colite/imunologia , Colite/metabolismo , Subpopulações de Linfócitos T/imunologia , Subpopulações de Linfócitos T/metabolismo , Camundongos Endogâmicos C57BL , Mucosa Intestinal/imunologia , Mucosa Intestinal/metabolismo , Intestinos/imunologia , Proteínas de Ligação a DNA/metabolismo , Proteínas de Ligação a DNA/genética , Células Cultivadas , Células Th17/imunologiaRESUMO
Numerous diseases, recently reported to associate with elevated microvesicle/microparticle (MP) counts, have also long been known to be characterized by accelerated immune complex (IC) formation. The goal of this study was to investigate the potential overlap between parameters of protein complexes (eg, ICs or avidin-biotin complexes) and MPs, which might perturb detection and/or isolation of MPs. In this work, after comprehensive characterization of MPs by electron microscopy, atomic force microscopy, dynamic light-scattering analysis, and flow cytometry, for the first time, we drive attention to the fact that protein complexes, especially insoluble ICs, overlap in biophysical properties (size, light scattering, and sedimentation) with MPs. This, in turn, affects MP quantification by flow cytometry and purification by differential centrifugation, especially in diseases in which IC formation is common, including not only autoimmune diseases, but also hematologic disorders, infections, and cancer. These data may necessitate reevaluation of certain published data on patient-derived MPs and contribute to correct the clinical laboratory assessment of the presence and biologic functions of MPs in health and disease.
Assuntos
Fenômenos Biofísicos/fisiologia , Fracionamento Celular/métodos , Micropartículas Derivadas de Células/química , Complexos Multiproteicos/farmacologia , Adulto , Idoso , Estudos de Casos e Controles , Fracionamento Celular/normas , Micropartículas Derivadas de Células/fisiologia , Feminino , Citometria de Fluxo , Humanos , Masculino , Microscopia de Força Atômica , Microscopia Eletrônica , Pessoa de Meia-Idade , Complexos Multiproteicos/química , Tamanho da PartículaRESUMO
Release of membrane vesicles, a process conserved in both prokaryotes and eukaryotes, represents an evolutionary link, and suggests essential functions of a dynamic extracellular vesicular compartment (including exosomes, microparticles or microvesicles and apoptotic bodies). Compelling evidence supports the significance of this compartment in a broad range of physiological and pathological processes. However, classification of membrane vesicles, protocols of their isolation and detection, molecular details of vesicular release, clearance and biological functions are still under intense investigation. Here, we give a comprehensive overview of extracellular vesicles. After discussing the technical pitfalls and potential artifacts of the rapidly emerging field, we compare results from meta-analyses of published proteomic studies on membrane vesicles. We also summarize clinical implications of membrane vesicles. Lessons from this compartment challenge current paradigms concerning the mechanisms of intercellular communication and immune regulation. Furthermore, its clinical implementation may open new perspectives in translational medicine both in diagnostics and therapy.
Assuntos
Micropartículas Derivadas de Células/fisiologia , Exossomos/fisiologia , Doenças Autoimunes/diagnóstico , Doenças Autoimunes/metabolismo , Biomarcadores/metabolismo , Micropartículas Derivadas de Células/química , Micropartículas Derivadas de Células/metabolismo , Exossomos/química , Exossomos/metabolismo , Humanos , Neoplasias/diagnóstico , Neoplasias/metabolismo , Tamanho da Partícula , Proteoma/metabolismoRESUMO
Experimental and clinical evidence for T cell involvement in the pathology of rheumatoid arthritis (RA) is compelling, and points to a local dysregulation of T cell function in the inflamed joint. Nitric oxide (NO) has been shown to regulate T cell function under physiological conditions, but overproduction of NO may contribute to lymphocyte dysfunction characteristic of RA. Several investigations in patients with RA have documented evidence of increased NO synthesis, but these studies have focused largely on macrophage-derived NO and its impact on innate immune and inflammatory responses. In this study, we set out to explore the contribution that T cells make to NO production. We find that T cells from RA patients produce >2.5 times more NO than healthy donor T cells (p<0.001). Although NO is an important physiological mediator of mitochondrial biogenesis, mitochondrial mass is similar in RA and control T cells. In contrast, increased NO production is associated with increased cytoplasmic Ca(2+) concentrations in RA T cells (p<0.001). In vitro treatment of human peripheral blood lymphocytes, or Jurkat cells with TNF increases NO production (p=0.006 and p=0.001, respectively), whilst infliximab treatment in RA patients decreases T cell derived NO production within 6 weeks of the first infusion (p=0.005). Together, these data indicate that TNF induced NO production in T lymphocytes may contribute to perturbations of immune homeostasis in RA.
Assuntos
Artrite Reumatoide/metabolismo , Óxido Nítrico/biossíntese , Linfócitos T/metabolismo , Artrite Reumatoide/imunologia , Cálcio/metabolismo , Células Cultivadas , Citoplasma/metabolismo , HumanosRESUMO
Increased susceptibility to infectious diseases is a hallmark of the neonatal period of life that is generally attributed to a relative immaturity of the immune system. Dendritic cells (DCs) are innate immune sentinels with vital roles in the initiation and orchestration of immune responses, thus, constituting a promising target for promoting neonatal immunity. However, as is the case for other immune cells, neonatal DCs have been suggested to be functionally immature compared to their adult counterparts. Here we review some of the unique aspects of neonatal DCs that shape immune responses in early life and speculate whether the functional properties of neonatal DCs could be exploited or manipulated to promote more effective vaccination in early life.
Assuntos
Células Dendríticas/imunologia , Vacinação/métodos , Vacinas/imunologia , Adulto , Fatores Etários , Animais , Animais Recém-Nascidos , Infecções Bacterianas/imunologia , Infecções Bacterianas/microbiologia , Infecções Bacterianas/prevenção & controle , Humanos , Imunogenicidade da Vacina , Recém-Nascido , Camundongos , Vacinas/administração & dosagem , Viroses/imunologia , Viroses/microbiologia , Viroses/prevenção & controleRESUMO
Adaptive immunity is essentially required to control acute infection with enteropathogenic Yersinia pseudotuberculosis (Yptb). We have recently demonstrated that Yptb can directly modulate naïve CD4+ T cell differentiation. However, whether fully differentiated forkhead box protein P3 (Foxp3+) regulatory T cells (Tregs), fundamental key players to maintain immune homeostasis, are targeted by Yptb remains elusive. Here, we demonstrate that within the CD4+ T cell compartment Yptb preferentially targets Tregs and injects Yersinia outer proteins (Yops) in a process that depends on the type III secretion system and invasins. Remarkably, Yop-translocation into ex vivo isolated Foxp3+ Tregs resulted in a substantial downregulation of Foxp3 expression and a decreased capacity to express the immunosuppressive cytokine interleukin-10 (IL-10). Together, these findings highlight that invasins are critically required to mediate Yptb attachment to Foxp3+ Tregs, which allows efficient Yop-translocation and finally enables the modulation of the Foxp3+ Tregs' suppressive phenotype.
RESUMO
Conventional dendritic cells (cDCs) are versatile activators of immune responses that develop as part of the myeloid lineage downstream of hematopoietic stem cells. We have recently shown that in mice precursors of cDCs, but not of other leukocytes, are marked by expression of DNGR-1/CLEC9A. To genetically deplete DNGR-1-expressing cDC precursors and their progeny, we crossed Clec9a-Cre mice to Rosa-lox-STOP-lox-diphtheria toxin (DTA) mice. These mice develop signs of age-dependent myeloproliferative disease, as has been observed in other DC-deficient mouse models. However, despite efficient depletion of cDC progenitors in these mice, cells with phenotypic characteristics of cDCs populate the spleen. These cells are functionally and transcriptionally similar to cDCs in wild type control mice but show somatic rearrangements of Ig-heavy chain genes, characteristic of lymphoid origin cells. Our studies reveal a previously unappreciated developmental heterogeneity of cDCs and suggest that the lymphoid lineage can generate cells with features of cDCs when myeloid cDC progenitors are impaired.
Assuntos
Células Dendríticas/imunologia , Lectinas Tipo C/imunologia , Receptores Imunológicos/imunologia , Animais , Células Dendríticas/efeitos dos fármacos , Toxina Diftérica/farmacologia , Lectinas Tipo C/genética , Camundongos , Receptores Imunológicos/genéticaRESUMO
Gut-draining mesenteric lymph nodes (mLNs) are important for inducing peripheral tolerance towards food and commensal antigens by providing an optimal microenvironment for de novo generation of Foxp3+ regulatory T cells (Tregs). We previously identified microbiota-imprinted mLN stromal cells as a critical component in tolerance induction. Here we show that this imprinting process already takes place in the neonatal phase, and renders the mLN stromal cell compartment resistant to inflammatory perturbations later in life. LN transplantation and single-cell RNA-seq uncover stably imprinted expression signatures in mLN fibroblastic stromal cells. Subsetting common stromal cells across gut-draining mLNs and skin-draining LNs further refine their location-specific immunomodulatory functions, such as subset-specific expression of Aldh1a2/3. Finally, we demonstrate that mLN stromal cells shape resident dendritic cells to attain high Treg-inducing capacity in a Bmp2-dependent manner. Thus, crosstalk between mLN stromal and resident dendritic cells provides a robust regulatory mechanism for the maintenance of intestinal tolerance.
Assuntos
Células Dendríticas/imunologia , Tolerância Imunológica/imunologia , Linfonodos/imunologia , Células Estromais/imunologia , Animais , Animais Recém-Nascidos , Microambiente Celular/genética , Microambiente Celular/imunologia , Células Dendríticas/metabolismo , Fatores de Transcrição Forkhead/imunologia , Fatores de Transcrição Forkhead/metabolismo , Perfilação da Expressão Gênica , Tolerância Imunológica/genética , Linfonodos/metabolismo , Linfonodos/transplante , Mesentério/imunologia , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Camundongos Knockout , Camundongos Transgênicos , Células Estromais/metabolismo , Linfócitos T Reguladores/imunologia , Linfócitos T Reguladores/metabolismoRESUMO
Peripheral inflammation can aggravate local brain inflammation and neuronal death. The blood-brain barrier (BBB) is a key player in the event. On a relevant in vitro model of primary rat brain endothelial cells co-cultured with primary rat astroglia cells lipopolysaccharide (LPS)-induced changes in several BBB functions have been investigated. LPS-treatment resulted in a dose- and time-dependent decrease in the integrity of endothelial monolayers: transendothelial electrical resistance dropped, while flux of permeability markers fluorescein and albumin significantly increased. Immunostaining for junctional proteins ZO-1, claudin-5 and beta-catenin was significantly weaker in LPS-treated endothelial cells than in control monolayers. LPS also reduced the intensity and changed the pattern of ZO-1 immunostaining in freshly isolated rat brain microvessels. The activity of P-glycoprotein, an important efflux pump at the BBB, was also inhibited by LPS. At the same time production of reactive oxygen species and nitric oxide was increased in brain endothelial cells treated with LPS. Pentosan polysulfate, a polyanionic polysaccharide could reduce the deleterious effects of LPS on BBB permeability, and P-glycoprotein activity. LPS-stimulated increase in the production of reactive oxygen species and nitric oxide was also decreased by pentosan treatment. The protective effect of pentosan for brain endothelium can be of therapeutical significance in bacterial infections affecting the BBB.
Assuntos
Encéfalo/efeitos dos fármacos , Endotélio Vascular/efeitos dos fármacos , Lipopolissacarídeos/farmacologia , Poliéster Sulfúrico de Pentosana/farmacologia , Membro 1 da Subfamília B de Cassetes de Ligação de ATP/metabolismo , Animais , Barreira Hematoencefálica , Encéfalo/irrigação sanguínea , Endotélio Vascular/citologia , Potenciais da Membrana/efeitos dos fármacos , Poliéster Sulfúrico de Pentosana/farmacocinética , RatosRESUMO
The immune system is a complex functional network of diverse cells and soluble molecules orchestrating innate and adaptive immunity. Biological information, to run these intricate interactions, is not only stored in protein sequences but also in the structure of the glycan part of the glycoconjugates. The spatially accessible carbohydrate structures that contribute to the cell's glycome are decoded by versatile recognition systems in order to maintain the immune homeostasis of an organism. Microbial carbohydrate structures are recognized by pathogen associated molecular pattern (PAMP) receptors of innate immunity including C-type lectins such as MBL, the tandem-repeat-type macrophage mannose receptor, DC-SIGN or dectin-1 of dendritic cells, certain TLRS or the TCR of NKT cells. Natural autoantibodies, a long known effector branch of this network-based operation, are effective to home in on non-self and self-glycosylation also. The recirculating pool of mammalian immune cells is recruited to inflammatory sites by a reaction pathway involving the self-carbohydrate-binding selectins as initial recognition step. Galectins, further key sensors reading the high-density sugar code, exert regulatory functions on activated T cells, among other activities. Autoimmune diseases are being associated with defined changes of glycosylation. This correlation deserves to be thoroughly studied on the levels of structural mimicry and dysregulation as well as effector molecules to devise innovative anti-inflammatory strategies. This review briefly summarizes data on sensor systems for carbohydrate epitopes and implications for autoimmunity.
Assuntos
Autoimunidade , Carboidratos/química , Carboidratos/imunologia , Epitopos/química , Epitopos/imunologia , Animais , Humanos , Transdução de Sinais/imunologia , Biologia de SistemasRESUMO
The gastrointestinal tract constitutes the largest surface of the body and thus has developed multitude mechanisms to either prevent pathogen entry or to efficiently eliminate invading pathogens. At the same time, the gastrointestinal system has to avoid unwanted immune responses against self and harmless nonself antigens, such as nutrients and commensal microbiota. Therefore, it is somewhat not unexpected that the gastrointestinal mucosa serves as the largest repository of immune cells throughout the body, harboring both potent pro- as well as anti-inflammatory properties. One additional key element of this regulatory machinery is created by trillions of symbiotic commensal bacteria in the gut. The microbiota not only simply contribute to the breakdown of nutrients, but are essential in limiting the expansion of pathogens, directing the development of the intestinal immune system, and establishing mucosal tolerance by fostering the induction of regulatory T cells (Tregs). In this review, we will discuss our current understanding about the microenvironmental factors fostering the de novo generation of Tregs within the gastrointestinal immune system, focusing on unique properties of antigen-presenting cells, tolerogenic cytokines, commensal-derived metabolites and the contribution of lymph node stromal cells.
Assuntos
Microambiente Celular , Trato Gastrointestinal/imunologia , Linfonodos/imunologia , Linfócitos T Reguladores/imunologia , Animais , Fatores de Transcrição Forkhead , Humanos , Sistema ImunitárioRESUMO
Recently, a transglutaminase 3 knockout (TGM3/KO) mouse was generated that showed impaired hair development, but no gross defects in the epidermal barrier, although increased fragility of isolated corneocytes was demonstrated. Here we investigated the functionality of skin barrier in vivo by percutaneous sensitization to FITC in TGM3/KO (n=64) and C57BL/6 wild-type (WT) mice (n=36). Cutaneous inflammation was evaluated by mouse ear swelling test (MEST), histology, serum IgE levels, and by flow cytometry from draining lymph nodes. Inflammation-induced significant MEST difference (P<0.0001) was detected between KO and WT mice and was supported also by histopathology. A significant increase of CD4+ CD25+-activated T cells (P<0.01) and elevated serum IgE levels (P<0.05) in KO mice indicated more the development of FITC sensitization than an irritative reaction. Propionibacter acnes-induced intracutaneous inflammation showed no difference (P=0.2254) between the reactivity of WT and KO immune system. As in vivo tracer, FITC penetration from skin surface followed by two-photon microscopy demonstrated a more invasive percutaneous penetration in KO mice. The clinically uninvolved skin in TGM3/KO mice showed impaired barrier function and higher susceptibility to FITC sensitization indicating that TGM3 has a significant contribution to the functionally intact cutaneous barrier.
Assuntos
Dermatite de Contato/imunologia , Dermatite de Contato/microbiologia , Infecções por Bactérias Gram-Positivas/imunologia , Propionibacterium acnes/imunologia , Transglutaminases/imunologia , Animais , Dermatite de Contato/etiologia , Edema/imunologia , Edema/metabolismo , Feminino , Citometria de Fluxo , Fluoresceína-5-Isotiocianato/toxicidade , Infecções por Bactérias Gram-Positivas/metabolismo , Imunoglobulina E/imunologia , Linfonodos/imunologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Propionibacterium acnes/metabolismo , Pele/imunologia , Pele/metabolismo , Pele/microbiologia , Transglutaminases/genéticaRESUMO
Since the 1970s, numerous reports have described elevated hexosaminidase activities in rheumatoid arthritis. However, due to the overlapping substrate specificities of different hexosaminidases, identification of the exact enzyme(s) responsible for the elevated activity remains incomplete. In this work we tested if the recently described enzyme, hexosaminidase D was expressed in human arthritic joints, and could contribute to the elevated hexosaminidase activity in rheumatoid arthritis. Thermostable ß-d-N-acetyl-galactosaminidase (hexosaminidase D) activities were determined in synovial fluid samples, synovial membranes, synovial fibroblast cell strains and synovial fibroblast-derived extracellular vesicles of patients with rheumatoid arthritis and osteoarthritis using chromogenic substrates. Expression of the HEXDC gene was detected both in steady state and in TGF-ß treated synovial fibroblasts by real time PCR. Strikingly, hexosaminidase D accounted for approximately 50% of the total ß-N-acetyl-galactosaminidase activity in synovial membranes and synovial fibroblasts, and it was responsible for the vast majority of the ß-d-N-acetyl-galactosaminidase activity in synovial fluid samples. TGF-ß downregulated the expression of hexosaminidase D in synovial fibroblasts dose-dependently. Of note, significant activity of hexosaminidase D was also found in association with extracellular vesicles released by synovial fibroblasts. This first study that describes the expression and disease relevance of the HEXDC gene in humans demonstrates the expression of this novel enzyme within the joints, and suggests that its activity may significantly contribute to the overall local exoglycosidase activity.
Assuntos
Artrite Reumatoide/enzimologia , Regulação Enzimológica da Expressão Gênica , Líquido Sinovial/enzimologia , Membrana Sinovial/enzimologia , beta-N-Acetil-Hexosaminidases/biossíntese , Adulto , Idoso , Artrite Reumatoide/imunologia , Artrite Reumatoide/patologia , Linhagem Celular , Feminino , Fibroblastos/enzimologia , Fibroblastos/imunologia , Fibroblastos/patologia , Humanos , Masculino , Pessoa de Meia-Idade , Líquido Sinovial/imunologia , Membrana Sinovial/imunologia , Membrana Sinovial/patologia , beta-N-Acetil-Hexosaminidases/imunologiaRESUMO
The P70-84 peptide (also called 5/4E8 epitope) of the human cartilage proteoglycan (PG) aggrecan is the dominant/arthritogenic epitope in both humans and arthritis-prone BALB/c mice (PG-induced arthritis, PGIA). An elevated T cell reactivity was demonstrated to a citrullinated version of the P70-84 epitope in most of the patients with rheumatoid arthritis (RA). The goal of this study was to understand better how a T cell epitope, if citrullinated, may affect antigenicity/arthritogenicity in PGIA, a murine model of RA. T cell reactivity to differentially citrullinated versions of either the human PG aggrecan P70-84 peptide or the corresponding mouse sequence was assessed in peptide or aggrecan-immunized and arthritic BALB/c mice as well as in T cell receptor transgenic mice specific for peptide P70-84 sequence. Peripheral T cell responses were induced by priming BALB/c mice with either the human wild-type or its citrullinated versions. Unexpectedly, priming with the citrullinated self-peptide induced a higher T cell response compared to the wild-type sequence (p<0.001), and the citrullination of the human peptide abolished T cell reactivity in PGIA. Our data suggest that T cells reactive to the citrullinated P70-84 peptide escaped thymic selection and are present in the peripheral T cell repertoire. Results of this study provide evidence that citrullination of an immunodominant T cell epitope may substantially alter, either increase or abolish, T cell recognition at the periphery in an experimental model of arthritis.