RESUMO
Antigen presentation to T cells by MHC molecules is essential for adaptive immune responses. To determine the exact position of a gene affecting expression of MHC molecules, we finely mapped a previously defined rat quantitative trait locus regulating MHC class II on microglia in an advanced intercross line. We identified a small interval including the gene MHC class II transactivator (Mhc2ta) and, using a map over six inbred strains combined with gene sequencing and expression analysis, two conserved Mhc2ta haplotypes segregating with MHC class II levels. In humans, a -168A --> G polymorphism in the type III promoter of the MHC class II transactivator (MHC2TA) was associated with increased susceptibility to rheumatoid arthritis, multiple sclerosis and myocardial infarction, as well as lower expression of MHC2TA after stimulation of leukocytes with interferon-gamma. We conclude that polymorphisms in Mhc2ta and MHC2TA result in differential MHC molecule expression and are associated with susceptibility to common complex diseases with inflammatory components.
Assuntos
Artrite Reumatoide/genética , Complexo Principal de Histocompatibilidade , Esclerose Múltipla/genética , Infarto do Miocárdio/genética , Proteínas Nucleares/genética , Transativadores/genética , Animais , Predisposição Genética para Doença , Genótipo , Antígenos de Histocompatibilidade Classe II/imunologia , Polimorfismo de Nucleotídeo Único , Ratos , Medula Espinal/imunologiaRESUMO
EAE, an animal model for multiple sclerosis, is a Th17- and Th1-cell-mediated auto-immune disease, but the mechanisms leading to priming of encephalitogenic T cells in autoimmune neuroinflammation are poorly understood. To investigate the role of dendritic cells (DCs) in the initiation of autoimmune Th17- and Th1-cell responses and EAE, we used mice transgenic for a simian diphtheria toxin receptor (DTR) expressed under the control of the murine CD11c promoter (CD11c-DTR mice o nC57BL/6 background).EAE was induced by immunization with myelin oligodendrocyte glycoprotein (MOG) protein in CFA. DCs were depleted on the day before and 8 days after MOG immunization. The mean clinical EAE score was only mildly reduced in DC-depleted mice when DCs were ablated before EAE induction. The frequency of activated Th cells was not altered, and MOG-induced Th17 or Th1-cell responses were not altered, in the spleens of DC-depleted mice. Similar results were obtained if DCs were ablated the first 10 days after MOG immunization with repeated DC depletions. Unexpectedly, transient depletion of DCs did not affect priming or differentiation of MOG-induced Th17 and Th1-cell responses or the incidence of EAE. Thus, the mechanism of priming of Th cells in EAE remains to be elucidated.
Assuntos
Células Dendríticas/imunologia , Encefalomielite Autoimune Experimental/imunologia , Esclerose Múltipla/imunologia , Células Th1/imunologia , Células Th17/imunologia , Animais , Antígeno CD11c/genética , Células Cultivadas , Toxina Diftérica/administração & dosagem , Feminino , Fator de Crescimento Semelhante a EGF de Ligação à Heparina , Humanos , Peptídeos e Proteínas de Sinalização Intercelular/genética , Ativação Linfocitária/efeitos dos fármacos , Ativação Linfocitária/genética , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Glicoproteína Mielina-Oligodendrócito/imunologia , Regiões Promotoras GenéticasRESUMO
Autoimmune polyendocrine syndrome type 1 (APS-1) is a multiorgan autoimmune disease caused by mutations in the autoimmune regulator (AIRE) gene. Chronic mucocutaneous candidiasis, hypoparathyroidism and adrenal failure are hallmarks of the disease. The critical mechanisms causing chronic mucocutaneous candidiasis in APS-1 patients have not been identified although autoantibodies to cytokines are implicated in the pathogenesis. To investigate whether the Th reactivity to Candida albicans (C. albicans) and other stimuli was altered, we isolated PBMC from APS-1 patients and matched healthy controls. The Th17 pathway was upregulated in response to C. albicans in APS-1 patients, whereas the IL-22 secretion was reduced. Autoantibodies against IL-22, IL-17A and IL-17F were detected in sera from APS-1 patients by immunoprecipitation. In addition, Aire-deficient (Aire(0/0) ) mice were much more susceptible than Aire(+/+) mice to mucosal candidiasis and C. albicans-induced Th17- and Th1-cell responses were increased in Aire(0/0) mice. Thus an excessive IL-17A reactivity towards C. albicans was observed in APS-1 patients and Aire(0/0) mice.
Assuntos
Candida albicans/imunologia , Candidíase/imunologia , Interleucina-17/imunologia , Adolescente , Adulto , Animais , Autoanticorpos/sangue , Autoanticorpos/imunologia , Modelos Animais de Doenças , Feminino , Humanos , Interleucinas/imunologia , Interleucinas/metabolismo , Leucócitos Mononucleares/imunologia , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Mutantes , Pessoa de Meia-Idade , Poliendocrinopatias Autoimunes/imunologia , Células Th17/imunologia , Fatores de Transcrição/genética , Regulação para Cima/imunologia , Adulto Jovem , Proteína AIRE , Interleucina 22RESUMO
Traumatic brain injury (TBI) in the mouse results in the rapid appearance of scattered clusters of cells expressing the chemokine Cxcl10 in cortical and subcortical areas. To extend the observation of this unique pattern, we used neuropathological mouse models using quantitative reverse transcriptase-polymerase chain reaction, gene array analysis, in-situ hybridization and flow cytometry. As for TBI, cell clusters of 150-200 mum expressing Cxcl10 characterize the cerebral cortex of mice carrying a transgene encoding the Swedish mutation of amyloid precursor protein, a model of amyloid Alzheimer pathology. The same pattern was found in experimental autoimmune encephalomyelitis in mice modelling multiple sclerosis. In contrast, mice carrying a SOD1(G93A) mutant mimicking amyotrophic lateral sclerosis pathology lacked such cell clusters in the cerebral cortex, whereas clusters appeared in the brainstem and spinal cord. Mice homozygous for a null mutation of the Cxcl10 gene did not show detectable levels of Cxcl10 transcript after TBI, confirming the quantitative reverse transcriptase-polymerase chain reaction and in-situ hybridization signals. Moreover, unbiased microarray expression analysis showed that Cxcl10 was among 112 transcripts in the neocortex upregulated at least threefold in both TBI and ageing TgSwe mice, many of them involved in inflammation. The identity of the Cxcl10(+) cells remains unclear but flow cytometry showed increased numbers of activated microglia/macrophages as well as myeloid dendritic cells in the TBI and experimental autoimmune encephalomyelitis models. It is concluded that the Cxcl10(+) cells appear in the inflamed central nervous system and may represent a novel population of cells that it may be possible to target pharmacologically in a broad range of neurodegenerative conditions.
Assuntos
Lesões Encefálicas/metabolismo , Lesões Encefálicas/patologia , Quimiocina CXCL10/biossíntese , Doenças Neurodegenerativas/metabolismo , Doenças Neurodegenerativas/patologia , Animais , Separação Celular , Modelos Animais de Doenças , Citometria de Fluxo , Expressão Gênica , Perfilação da Expressão Gênica , Hibridização In Situ , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Análise de Sequência com Séries de Oligonucleotídeos , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
EAE, an animal model for MS, is a Th17 and Th1-cell-mediated autoimmune disease, but the mechanisms leading to priming of encephalitogenic T cells in autoimmune neuroinflammation are poorly understood. To investigate the role of plasmacytoid DC (pDC) in the initiation of autoimmune Th17- and Th1-cell responses and EAE, we depleted pDC with anti-pDC Ag-1 (anti-PDCA1) mAb prior to immunization of C57BL/6 mice with myelin oligodendrocyte glycoprotein (MOG). pDC-depleted mice developed less severe clinical and histopathological signs of EAE than control mice, which demonstrates a promoting role for pDC in the initiation of EAE. The levels of type I IFN were much lower in the sera from anti-PDCA1-treated mice. However, neutralization of type I IFN ameliorated the early phase of EAE but did not alter the severity of disease. Thus, only a minor part of the EAE-promoting effect of pDC appears to be mediated by IFN-alpha/beta secretion. The numbers of MOG-specific Th17 cells, but not Th1 cells, were lower in spleen from anti-PDCA1-treated mice compared with controls. In contrast, pDC depletion a week after MOG immunization resulted in more severe clinical signs of EAE. In conclusion, we demonstrate that pDC promote initiation of MOG-induced Th17-cell responses and EAE.
Assuntos
Autoimunidade/imunologia , Células Dendríticas/imunologia , Encefalomielite Autoimune Experimental/imunologia , Interleucina-17/metabolismo , Ativação Linfocitária/imunologia , Linfócitos T Auxiliares-Indutores/imunologia , Animais , Anticorpos/imunologia , Anticorpos/farmacologia , Anticorpos Monoclonais/imunologia , Anticorpos Monoclonais/farmacologia , Antígenos de Diferenciação de Linfócitos T/genética , Contagem de Células , Sistema Nervoso Central/patologia , Células Dendríticas/citologia , Células Dendríticas/efeitos dos fármacos , Células Dendríticas/metabolismo , Encefalomielite Autoimune Experimental/sangue , Encefalomielite Autoimune Experimental/etiologia , Encefalomielite Autoimune Experimental/patologia , Feminino , Fatores de Transcrição Forkhead/genética , Expressão Gênica/genética , Proteína Coestimuladora de Linfócitos T Induzíveis , Interferon-alfa/sangue , Interferon-alfa/imunologia , Interferon beta/sangue , Interferon beta/imunologia , Interleucina-10/genética , Interleucina-17/genética , Interleucina-23/genética , Interleucina-6/sangue , Interleucina-6/genética , Camundongos , Camundongos Endogâmicos C57BL , Proteínas da Mielina , Glicoproteína Associada a Mielina/imunologia , Glicoproteína Mielina-Oligodendrócito , Polirradiculoneuropatia , Baço/citologia , Baço/metabolismo , Subpopulações de Linfócitos T/citologia , Subpopulações de Linfócitos T/metabolismo , Linfócitos T Auxiliares-Indutores/citologia , Linfócitos T Auxiliares-Indutores/metabolismo , Linfócitos T Reguladores/citologia , Linfócitos T Reguladores/metabolismo , Células Th1/citologia , Células Th1/metabolismo , VacinaçãoRESUMO
Neutralizing autoantibodies to type I, but not type II, interferons (IFNs) are found at high titers in almost every patient with autoimmune polyendocrinopathy candidiasis ectodermal dystrophy (APECED), a disease caused by AIRE gene mutations that lead to defects in thymic T-cell selection. Combining genome-wide expression array with real time RT-PCR assays, we here demonstrate that antibodies against IFN-alpha cause highly significant down-regulation of interferon-stimulated gene expression in cells from APECED patients' blood by blocking their highly dilute endogenous IFNs. This down-regulation was lost progressively as these APECED cells matured in cultures without neutralizing autoantibodies. Most interestingly, a rare APECED patient with autoantibodies to IFN-omega but not IFN-alpha showed a marked increase in expression of the same interferon-stimulated genes. We also report unexpected increases in serum CXCL10 levels in APECED. Our results argue that the breakdown of tolerance to IFNs in AIRE deficiency is associated with impaired responses to them in thymus, and highlight APECED as another autoimmune disease with associated dysregulation of IFN activity.
Assuntos
Autoanticorpos/imunologia , Regulação para Baixo/genética , Interferons/imunologia , Fatores de Transcrição/deficiência , Adolescente , Adulto , Células Sanguíneas/metabolismo , Estudos de Casos e Controles , Linhagem Celular , Quimiocina CXCL10/sangue , Células Dendríticas/imunologia , Feminino , Humanos , Interferon Tipo I/imunologia , Masculino , Pessoa de Meia-Idade , Modelos Imunológicos , Monócitos/imunologia , Testes de Neutralização , Análise de Sequência com Séries de Oligonucleotídeos , Fosforilação , Poliendocrinopatias Autoimunes/sangue , Poliendocrinopatias Autoimunes/genética , Fator de Transcrição STAT1/metabolismo , Fatores de Transcrição/imunologia , Proteína AIRERESUMO
Multiple sclerosis (MS) is a chronic, inflammatory, demyelinating disease of the CNS, most frequently starting with a series of bouts, each followed by complete remission and then a secondary, progressive phase during which the neurological deficit increases steadily. The underlying molecular mechanisms responsible for disease progression are still unclear. Herein, we demonstrate that high mobility group box chromosomal protein 1 (HMGB1), a DNA-binding protein with proinflammatory properties, is evident in active lesions of MS and experimental autoimmune encephalomyelitis (EAE) and that HMGB1 levels correlate with active inflammation. Furthermore, the expression of the innate HMGB1 receptors--receptor for advanced glycation end products, TLR2, and TLR4--was also highly increased in MS and rodent EAE. Additionally, in vitro activation of rodent CNS-derived microglia and bone marrow-derived macrophages demonstrated that microglia were equally as capable as macrophages of translocating HMGB1 following LPS/IFN-gamma stimulation. Significant expression of HMGB1 and its receptors on accumulating activated macrophages and resident microglia may thus provide a positive feedback loop that amplifies the inflammatory response during MS and EAE pathogenesis.
Assuntos
Proteína HMGB1/genética , Esclerose Múltipla/genética , Animais , Células da Medula Óssea/citologia , Células da Medula Óssea/fisiologia , Modelos Animais de Doenças , Encefalomielite Autoimune Experimental/genética , Encefalomielite Autoimune Experimental/patologia , Feminino , Humanos , Hibridização In Situ , Macrófagos/citologia , Microglia/citologia , RNA Mensageiro/genética , Ratos , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Medula Espinal/patologiaRESUMO
Expression of T cell immunoglobulin- and mucin-domain-containing molecules (TIMs) can be used as T helper (Th) differentiation markers in the human and mouse. We examined the expression of TIM-1 and -3 mRNAs in rat MBP(63-88)-induced experimental autoimmune encephalomyelitis (EAE). TIM-3 expression was upregulated in the spinal cord during EAE and following antigen restimulation of the encephalitogenic TCRBV8S2+ population. Interestingly, TIM-3 expression was also detected by in situ hybridization in resident cells of the nervous system. TIM-1 was expressed in B cells but not in resident CNS cells and TIM-1 mRNA levels in spinal cord were unchanged throughout the course of EAE. These results support the notion that TIM-3 can also be used as a Th1 differentiation marker in the rat. However, expression of TIM-1 and -3 is not restricted solely to T cells and the presence of TIM-3 in resident CNS cells may indicate a role for this molecule in the interaction between the nervous and immune systems.
Assuntos
Encefalomielite Autoimune Experimental/metabolismo , Regulação da Expressão Gênica/fisiologia , Sistema Imunitário/metabolismo , Proteínas de Membrana/metabolismo , Sistema Nervoso/metabolismo , Receptores Virais/metabolismo , Animais , Células Cultivadas , Encefalomielite Autoimune Experimental/induzido quimicamente , Feminino , Citometria de Fluxo/métodos , Receptor Celular 1 do Vírus da Hepatite A , Receptor Celular 2 do Vírus da Hepatite A , Hibridização In Situ/métodos , Interferon gama/genética , Interferon gama/metabolismo , Interleucina-10/genética , Interleucina-10/metabolismo , Proteínas de Membrana/genética , Proteína Básica da Mielina , Sistema Nervoso/patologia , Fragmentos de Peptídeos , RNA Mensageiro/biossíntese , Ratos , Ratos Endogâmicos Lew , Receptores Virais/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa/métodosRESUMO
DNA vaccines encoding encephalitogenic peptides protect against subsequent development of rat experimental autoimmune encephalomyelitis (EAE) through unknown mechanisms. We investigated immune cell phenotypes at different time points after DNA vaccination with vaccine encoding myelin oligodendrocyte glycoprotein peptide 91-108 and subsequent induction of EAE. In protected rats, we observed (i) no alterations in antigen-specific Th2 or Th3 responses, (ii) reduced MHC II expression on splenocytes early after EAE induction, (iii) antigen-specific upregulation of IFNbeta upon recall stimulation and (iv) reduced IL-12Rbeta2 on lymphocytes. We suggest that the underlying mechanism of DNA vaccination is associated with immunomodulation exerted by induced IFNbeta.
Assuntos
Encefalomielite Autoimune Experimental/prevenção & controle , Regulação da Expressão Gênica , Interferon beta/biossíntese , Vacinas de DNA/administração & dosagem , Animais , Antígenos de Superfície/genética , Antígenos de Superfície/metabolismo , Células Cultivadas , Sistema Nervoso Central/citologia , Citocinas/genética , Citocinas/metabolismo , Modelos Animais de Doenças , Encefalomielite Autoimune Experimental/imunologia , Citometria de Fluxo/métodos , Regulação da Expressão Gênica/imunologia , Receptor Celular 2 do Vírus da Hepatite A , Humanos , Interferon beta/genética , Lectinas Tipo C/genética , Lectinas Tipo C/metabolismo , Lipopolissacarídeos/farmacologia , Linfócitos/imunologia , Linfócitos/metabolismo , Complexo Principal de Histocompatibilidade/genética , Camundongos , Proteínas da Mielina , Glicoproteína Associada a Mielina/imunologia , Glicoproteína Mielina-Oligodendrócito , Subfamília B de Receptores Semelhantes a Lectina de Células NK , Fragmentos de Peptídeos/imunologia , RNA Mensageiro/biossíntese , Ratos , Ratos Endogâmicos Lew , Receptores de Interleucina/genética , Receptores de Interleucina/metabolismo , Receptores de Interleucina-12 , Receptores Virais/genética , Receptores Virais/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa/métodos , Alinhamento de Sequência , Linfócitos T/efeitos dos fármacos , Linfócitos T/imunologia , Fatores de Tempo , Vacinação , Vacinas de DNA/imunologiaRESUMO
AIMS/HYPOTHESIS: Diabetes mellitus is one of the most common endocrine disorders in dogs and is commonly proposed to be of autoimmune origin. Although the clinical presentation of human type 1 diabetes (T1D) and canine diabetes are similar, the aetiologies may differ. The aim of this study was to investigate if autoimmune aetiology resembling human T1D is as prevalent in dogs as previously reported. METHODS: Sera from 121 diabetic dogs representing 40 different breeds were tested for islet cell antibodies (ICA) and GAD65 autoantibodies (GADA) and compared with sera from 133 healthy dogs. ICA was detected by indirect immunofluorescence using both canine and human frozen sections. GADA was detected by in vitro transcription and translation (ITT) of human and canine GAD65, followed by immune precipitation. Sections of pancreata from five diabetic dogs and two control dogs were examined histopathologically including immunostaining for insulin, glucagon, somatostatin and pancreas polypeptide. RESULTS: None of the canine sera analysed tested positive for ICA on sections of frozen canine or human ICA pancreas. However, serum from one diabetic dog was weakly positive in the canine GADA assay and serum from one healthy dog was weakly positive in the human GADA assay. Histopathology showed marked degenerative changes in endocrine islets, including vacuolisation and variable loss of immune-staining for insulin. No sign of inflammation was noted. CONCLUSIONS/INTERPRETATIONS: Contrary to previous observations, based on results from tests for humoral autoreactivity towards islet proteins using four different assays, and histopathological examinations, we do not find any support for an islet autoimmune aetiology in canine diabetes mellitus.
Assuntos
Diabetes Mellitus/veterinária , Doenças do Cão/imunologia , Ilhotas Pancreáticas/imunologia , Animais , Autoanticorpos/metabolismo , Diabetes Mellitus/sangue , Diabetes Mellitus/imunologia , Doenças do Cão/sangue , Cães , Imunofluorescência , Humanos , Ilhotas Pancreáticas/metabolismoRESUMO
BACKGROUND: Gene variants known to contribute to Autoimmune Addison's disease (AAD) susceptibility include those at the MHC, MICA, CIITA, CTLA4, PTPN22, CYP27B1, NLRP-1 and CD274 loci. The majority of the genetic component to disease susceptibility has yet to be accounted for. AIM: To investigate the role of 19 candidate genes in AAD susceptibility in six European case-control cohorts. METHODS: A sequential association study design was employed with genotyping using Sequenom iPlex technology. In phase one, 85 SNPs in 19 genes were genotyped in UK and Norwegian AAD cohorts (691 AAD, 715 controls). In phase two, 21 SNPs in 11 genes were genotyped in German, Swedish, Italian and Polish cohorts (1264 AAD, 1221 controls). In phase three, to explore association of GATA3 polymorphisms with AAD and to determine if this association extended to other autoimmune conditions, 15 SNPs in GATA3 were studied in UK and Norwegian AAD cohorts, 1195 type 1 diabetes patients from Norway, 650 rheumatoid arthritis patients from New Zealand and in 283 UK Graves' disease patients. Meta-analysis was used to compare genotype frequencies between the participating centres, allowing for heterogeneity. RESULTS: We report significant association with alleles of two STAT4 markers in AAD cohorts (rs4274624: Pâ=â0.00016; rs10931481: Pâ=â0.0007). In addition, nominal association of AAD with alleles at GATA3 was found in 3 patient cohorts and supported by meta-analysis. Association of AAD with CYP27B1 alleles was also confirmed, which replicates previous published data. Finally, nominal association was found at SNPs in both the NF-κB1 and IL23A genes in the UK and Italian cohorts respectively. CONCLUSIONS: Variants in the STAT4 gene, previously associated with other autoimmune conditions, confer susceptibility to AAD. Additionally, we report association of GATA3 variants with AAD: this adds to the recent report of association of GATA3 variants with rheumatoid arthritis.
Assuntos
Doença de Addison/genética , Alelos , Doenças Autoimunes/genética , Fator de Transcrição GATA3/genética , Estudos de Associação Genética , Predisposição Genética para Doença , Fator de Transcrição STAT4/genética , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Estudos de Casos e Controles , Criança , Pré-Escolar , Estudos de Coortes , Feminino , Heterogeneidade Genética , Técnicas de Genotipagem , Humanos , Lactente , Recém-Nascido , Masculino , Pessoa de Meia-Idade , Noruega , Reino Unido , População Branca , Adulto JovemRESUMO
Experimental autoimmune encephalomyelitis (EAE) is a Th17-mediated autoimmune disease and an animal model for multiple sclerosis (MS). Complete Freund's adjuvant (CFA) contains pathogen-associated molecular patterns (PAMPs) that bind toll-like receptors (TLRs), and is necessary to induce EAE. Upstream TLR signals modify innate and adaptive immune responses in EAE. In detail, the common TLR adaptor molecule MyD88 is necessary for induction of EAE, and mediates activation of peripheral myeloid dendritic cells (mDCs) and differentiation of autoimmune Th17 cells. The stimulatory TLRs have not yet been identified for Th17 cells. TLR4 down regulates disease severity in EAE and Th17 cell responses, but promotes Th1 cell responses, which may inhibit the differentiation of Th17 cells. Moreover, treatment with a TLR4 ligand tolerizes mice and prevents EAE. TLR9 down regulates disease severity in myelin oligodendrocyte glycoprotein (MOG)-induced EAE, whereas it promotes disease in MOG(35-55)-induced EAE. Thus MyD88, TLR4 and TLR9 modify the disease process in EAE. Both endogenous and CFA-derived TLR ligands are implicated to modulate the disease process.
Assuntos
Encefalomielite Autoimune Experimental/imunologia , Esclerose Múltipla/imunologia , Fator 88 de Diferenciação Mieloide/imunologia , Receptores Toll-Like/imunologia , Animais , Diferenciação Celular , Adjuvante de Freund , Glicoproteínas/imunologia , Humanos , Fatores Imunológicos , Interleucina-17/imunologia , Camundongos , Proteínas da Mielina , Glicoproteína Associada a Mielina/imunologia , Glicoproteína Mielina-Oligodendrócito , Fragmentos de Peptídeos/imunologia , Subpopulações de Linfócitos T/imunologia , Subpopulações de Linfócitos T/patologia , Células Th1/imunologia , Células Th1/patologiaRESUMO
Innate immune mechanisms essential for priming encephalitogenic T cells in autoimmune neuroinflammation are poorly understood. Experimental autoimmune encephalomyelitis (EAE) is a IL-17-producing Th (Th17) cell-mediated autoimmune disease and an animal model of multiple sclerosis. To investigate how upstream TLR signals influence autoimmune T cell responses, we studied the role of individual TLR and MyD88, the common TLR adaptor molecule, in the initiation of innate and adaptive immune responses in EAE. Wild type (WT) C57BL/6, TLR-deficient and MyD88-deficient mice were immunized with myelin oligodendrocyte glycoprotein (MOG) in CFA. MyD88(-/-) mice were completely EAE resistant. Purified splenic myeloid DC (mDC) from MyD88(-/-) mice expressed much less IL-6 and IL-23, and serum and T cell IL-17 were absent. TLR4(-/-) and TLR9(-/-) mice surprisingly exhibited more severe EAE symptoms than WT mice. IL-6 and IL-23 expression by mDC and Th17 responses were higher in TLR4(-/-) mice, suggesting a regulatory role of TLR4 in priming Th17 cells. IL-6 expression by splenocytes was higher in TLR9(-/-) mice. Our data suggest that MyD88 mediates the induction of mDC IL-6 and IL-23 responses after MOG immunization, which in turn drives IL-17-producing encephalitogenic Th17 cell activation. Importantly, we demonstrate that TLR4 and TLR9 regulate disease severity in MOG-induced EAE.
Assuntos
Encefalomielite Autoimune Experimental/imunologia , Encefalomielite Autoimune Experimental/metabolismo , Receptor 4 Toll-Like/fisiologia , Receptor Toll-Like 9/fisiologia , Animais , Células Cultivadas , Feminino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Proteínas da Mielina , Glicoproteína Associada a Mielina/imunologia , Glicoproteína Mielina-Oligodendrócito , Fator 88 de Diferenciação Mieloide/deficiência , Fator 88 de Diferenciação Mieloide/genética , Fator 88 de Diferenciação Mieloide/fisiologia , Índice de Gravidade de Doença , Receptor 4 Toll-Like/deficiência , Receptor 4 Toll-Like/genética , Receptor Toll-Like 9/deficiência , Receptor Toll-Like 9/genéticaRESUMO
Presentation of Ag bound to MHC class II (MHC II) molecules to CD4+ T cells is a key event in adaptive immune responses. Genetic differences in MHC II expression in the rat CNS were recently positioned to allelic variability in the CIITA gene (Mhc2ta), located within the Vra4 locus on rat chromosome 10. In this study, we have examined reciprocal Vra4-congenic strains on the DA and PVGav1 backgrounds, respectively. After experimental nerve injury the strain-specific MHC II expression on microglia was reversed in the congenic strains. Similar findings were obtained after intraparenchymal injection of IFN-gamma in the brain. Expression of MHC class II was also lower on B cells and dendritic cells from the DA.PVGav1-Vra4- congenic strain compared with DA rats after in vitro stimulation with IFN-gamma. We next explored whether Vra4 may affect the outcome of experimental autoimmune disease. In experimental autoimmune encephalomyelitis induced by immunization with myelin oligodendrocyte glycoprotein, DA.PVGav1-Vra4 rats displayed a lower disease incidence and milder disease course compared with DA, whereas both PVGav1 and PVGav1.DA-Vra4 rats were completely protected. These results demonstrate that naturally occurring allelic differences in Mhc2ta have profound effects on the quantity of MHC II expression in the CNS and on immune cells and that this genetic variability also modulates susceptibility to autoimmune neuroinflammation.
Assuntos
Alelos , Encefalomielite Autoimune Experimental/genética , Encefalomielite Autoimune Experimental/imunologia , Predisposição Genética para Doença , Antígenos de Histocompatibilidade Classe II/genética , Proteínas Nucleares/genética , Transativadores/genética , Animais , Animais Congênicos , Células Cultivadas , Regulação para Baixo/genética , Regulação para Baixo/imunologia , Encefalomielite Autoimune Experimental/patologia , Feminino , Marcadores Genéticos , Antígenos de Histocompatibilidade Classe II/biossíntese , Masculino , Inflamação Neurogênica/genética , Inflamação Neurogênica/imunologia , Proteínas Nucleares/biossíntese , Ratos , Ratos Endogâmicos , Rizotomia , Raízes Nervosas Espinhais/patologia , Raízes Nervosas Espinhais/cirurgia , Transativadores/biossínteseRESUMO
BACKGROUND: We have previously shown that vaccination with DNA encoding the encephalitogenic peptide myelin oligodendrocyte glycoprotein (MOG)(91-108) (pMOG) suppresses MOG(91-108)-induced rat Experimental Autoimmune Encephalomyelitis (EAE), a model for human Multiple Sclerosis (MS). The suppressive effect of pMOG is dependent on inclusion of CpG DNA in the plasmid backbone and is associated with early induction of Interferon (IFN)-beta. PRINCIPAL FINDINGS: In this study we examined the mechanisms underlying pMOG-induced protection. We found that in the DNA vaccinated cohort proinflammatory Interleukin (IL)-17 and IL-21 responses were dramatically reduced compared to in the control group, but that the expression of Foxp3 and Tumor Growth Factor (TGF)-beta1, which are associated with regulatory T cells, was not enhanced. Moreover, genes associated with Type I IFNs were upregulated. To delineate the role of IFN-beta in the protective mechanism we employed short interfering RNA (siRNA) to IFN-beta in the DNA vaccine. SiRNA to IFN-beta completely abrogated the protective effects of the vaccine, demonstrating that a local early elaboration of IFN-beta is important for EAE protection. IL-17 responses comparable to those in control rats developed in rats injected with the IFN-beta-silencing DNA vaccine. CONCLUSIONS: We herein demonstrate that DNA vaccination protects from proinflammatory Th17 cell responses during induction of EAE. The mechanism involves IFN-beta as IL-17 responses are rescued by silencing of IFN-beta during DNA vaccination.
Assuntos
Encefalomielite Autoimune Experimental/imunologia , Encefalomielite Autoimune Experimental/prevenção & controle , Linfócitos T Auxiliares-Indutores/imunologia , Vacinas de DNA/imunologia , Animais , Células Cultivadas , Feminino , Imunossupressores/administração & dosagem , Imunossupressores/imunologia , Interferon beta/imunologia , Interleucina-17/genética , Interleucina-17/imunologia , Interleucinas/imunologia , Proteínas da Mielina , Glicoproteína Associada a Mielina/imunologia , Glicoproteína Mielina-Oligodendrócito , RNA Interferente Pequeno/metabolismo , Ratos , Ratos Endogâmicos Lew , Transfecção , Vacinas de DNA/administração & dosagemRESUMO
Regeneration of cells in the central nervous system is a process that might be affected during neurological disease and trauma. Because nitric oxide (NO) and its derivatives are powerful mediators in the inflammatory cascade, we have investigated the effects of pathophysiological concentrations of NO on neurogenesis, gliogenesis, and the expression of proneural genes in primary adult neural stem cell cultures. After exposure to NO, neurogenesis was downregulated, and this corresponded to decreased expression of the proneural gene neurogenin-2 and beta-III-tubulin. The decreased ability to generate neurons was also found to be transmitted to the progeny of the cells. NO exposure was instead beneficial for astroglial differentiation, which was confirmed by increased activation of the Janus tyrosine kinase/signal transducer and activator of transcription transduction pathway. Our findings reveal a new role for NO during neuroinflammatory conditions, whereby its proastroglial fate-determining effect on neural stem cells might directly influence the neuroregenerative process.
Assuntos
Astrócitos/citologia , Diferenciação Celular/efeitos dos fármacos , Linhagem da Célula/efeitos dos fármacos , Neurônios/citologia , Óxido Nítrico/farmacologia , Células-Tronco/citologia , Animais , Astrócitos/efeitos dos fármacos , Morte Celular/efeitos dos fármacos , Células Cultivadas , Regulação para Baixo/efeitos dos fármacos , Feminino , Expressão Gênica/efeitos dos fármacos , Proteína Glial Fibrilar Ácida/metabolismo , Neurônios/efeitos dos fármacos , Óxido Nítrico/análise , Óxido Nítrico/metabolismo , Compostos Nitrosos/farmacologia , Oxigênio/metabolismo , Ratos , Células-Tronco/efeitos dos fármacosRESUMO
Vaccination with DNA encoding a myelin basic protein peptide suppresses Lewis rat experimental autoimmune encephalomyelitis (EAE) induced with the same peptide. Additional myelin proteins, such as myelin oligodendrocyte glycoprotein (MOG), may be important in multiple sclerosis. Here we demonstrate that DNA vaccination also suppresses MOG peptide-induced EAE. MOG(91-108) is encephalitogenic in DA rats and MHC-congenic LEW.1AV1 (RT1(av1)) and LEW.1N (RT1(n)) rats. We examined the effects of DNA vaccines encoding MOG(91-108) in tandem, with or without targeting of the hybrid gene product to IgG. In all investigated rat strains DNA vaccination suppressed clinical signs of EAE. There was no requirement for targeting the gene product to IgG, but T1-promoting CpG DNA motifs in the plasmid backbone of the construct were necessary for efficient DNA vaccination, similar to the case in DNA vaccination in myelin basic protein-induced EAE. We failed to detect any effects on ex vivo MOG-peptide-induced IFN-gamma, TNF-alpha, IL-6, IL-4, IL-10, and brain-derived neurotropic factor expression in splenocytes or CNS-derived lymphocytes. In CNS-derived lymphocytes, Fas ligand expression was down-regulated in DNA-vaccinated rats compared with controls. However, MOG-specific IgG2b responses were enhanced after DNA vaccination. The enhanced IgG2b responses together with the requirement for CpG DNA motifs in the vaccine suggest a protective mechanism involving induction of a T1-biased immune response.