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1.
J Immunol ; 201(7): 1907-1917, 2018 10 01.
Artigo em Inglês | MEDLINE | ID: mdl-30127089

RESUMO

In both NOD mice and humans, the development of type 1 diabetes (T1D) is dependent in part on autoreactive CD8+ T cells recognizing pancreatic ß cell peptides presented by often quite common MHC class I variants. Studies in NOD mice previously revealed that the common H2-Kd and/or H2-Db class I molecules expressed by this strain aberrantly lose the ability to mediate the thymic deletion of pathogenic CD8+ T cell responses through interactions with T1D susceptibility genes outside the MHC. A gene(s) mapping to proximal chromosome 7 was previously shown to be an important contributor to the failure of the common class I molecules expressed by NOD mice to mediate the normal thymic negative selection of diabetogenic CD8+ T cells. Using an inducible model of thymic negative selection and mRNA transcript analyses, we initially identified an elevated Nfkbid expression variant as a likely NOD-proximal chromosome 7 region gene contributing to impaired thymic deletion of diabetogenic CD8+ T cells. CRISPR/Cas9-mediated genetic attenuation of Nfkbid expression in NOD mice resulted in improved negative selection of autoreactive diabetogenic AI4 and NY8.3 CD8+ T cells. These results indicated that allelic variants of Nfkbid contribute to the efficiency of intrathymic deletion of diabetogenic CD8+ T cells. However, although enhancing thymic deletion of pathogenic CD8+ T cells, ablating Nfkbid expression surprisingly accelerated T1D onset that was associated with numeric decreases in both regulatory T and B lymphocytes in NOD mice.


Assuntos
Linfócitos T CD8-Positivos/imunologia , Cromossomos Humanos Par 7/genética , Diabetes Mellitus Tipo 1/imunologia , Proteínas I-kappa B/genética , Timo/imunologia , Alelos , Animais , Autoantígenos/imunologia , Diferenciação Celular , Células Cultivadas , Deleção Clonal , Modelos Animais de Doenças , Suscetibilidade a Doenças , Humanos , Proteínas I-kappa B/metabolismo , Camundongos , Camundongos Endogâmicos NOD , Polimorfismo Genético
2.
J Immunol ; 200(10): 3353-3363, 2018 05 15.
Artigo em Inglês | MEDLINE | ID: mdl-29632144

RESUMO

Type 1 diabetes (T1D) is characterized by T cell-mediated destruction of the insulin-producing ß cells of the pancreatic islets. Among the loci associated with T1D risk, those most predisposing are found in the MHC region. HLA-B*39:06 is the most predisposing class I MHC allele and is associated with an early age of onset. To establish an NOD mouse model for the study of HLA-B*39:06, we expressed it in the absence of murine class I MHC. HLA-B*39:06 was able to mediate the development of CD8 T cells, support lymphocytic infiltration of the islets, and confer T1D susceptibility. Because reduced thymic insulin expression is associated with impaired immunological tolerance to insulin and increased T1D risk in patients, we incorporated this in our model as well, finding that HLA-B*39:06-transgenic NOD mice with reduced thymic insulin expression have an earlier age of disease onset and a higher overall prevalence as compared with littermates with typical thymic insulin expression. This was despite virtually indistinguishable blood insulin levels, T cell subset percentages, and TCR Vß family usage, confirming that reduced thymic insulin expression does not impact T cell development on a global scale. Rather, it will facilitate the thymic escape of insulin-reactive HLA-B*39:06-restricted T cells, which participate in ß cell destruction. We also found that in mice expressing either HLA-B*39:06 or HLA-A*02:01 in the absence of murine class I MHC, HLA transgene identity alters TCR Vß usage by CD8 T cells, demonstrating that some TCR Vß families have a preference for particular class I MHC alleles.


Assuntos
Diabetes Mellitus Tipo 1/genética , Antígenos HLA-B/genética , Insulina/genética , Timo/metabolismo , Alelos , Animais , Linfócitos T CD8-Positivos/metabolismo , Diabetes Mellitus Tipo 1/metabolismo , Modelos Animais de Doenças , Genes MHC Classe I/genética , Antígeno HLA-A2/genética , Células Secretoras de Insulina/metabolismo , Camundongos , Camundongos Endogâmicos NOD , Camundongos Transgênicos
3.
Comp Med ; 67(4): 335-343, 2017 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-28830580

RESUMO

Segmented filamentous bacterium (SFB) a gram-positive, anaerobic, and intestinal commensal organism directly influences the development of Th17 helper cells in the small intestine of mice. In NOD mice, SFB colonization interferes with the development of type 1 diabetes (T1D), a T-cell-mediated autoimmune disease, suggesting that SFB may influence Th17 cells to inhibit Th1 populations associated with the anti-ß-cell immune response. This effect is a serious concern for investigators who use NOD mice for diabetes research because the expected incidence of disease decreases markedly when they are colonized by SFB. A room housing mice for T1D studies at The Jackson Laboratory was determined by fecal PCR testing to have widespread SFB colonization of multiple NOD strains after a steady decline in the incidence of T1D was noted. Rederivation of all NOD-related mouse strains was not feasible; therefore an alternative treatment using antibiotics to eliminate SFB from colonized mice was undertaken. After antibiotic treatment, soiled bedding from NOD mouse strains housed in SFB-free high-health-status production barrier rooms was used to reintroduce the gastrointestinal microbiota. Over the past 16 mo since treating the mice and disinfecting the mouse room, regular PCR testing has shown that no additional SFB colonization of mice has occurred, and the expected incidence of T1D has been reestablished in the offspring of treated mice.


Assuntos
Ampicilina/farmacologia , Antibacterianos/farmacologia , Diabetes Mellitus Tipo 1/microbiologia , Microbioma Gastrointestinal/efeitos dos fármacos , Bactérias Gram-Positivas/efeitos dos fármacos , Infecções por Bactérias Gram-Positivas/veterinária , Intestinos/efeitos dos fármacos , Criação de Animais Domésticos/métodos , Animais , Descontaminação/métodos , Diabetes Mellitus Tipo 1/genética , Diabetes Mellitus Tipo 1/imunologia , Modelos Animais de Doenças , Monitoramento Ambiental/métodos , Fezes/microbiologia , Predisposição Genética para Doença , Bactérias Gram-Positivas/classificação , Bactérias Gram-Positivas/imunologia , Infecções por Bactérias Gram-Positivas/tratamento farmacológico , Infecções por Bactérias Gram-Positivas/imunologia , Infecções por Bactérias Gram-Positivas/microbiologia , Interações Hospedeiro-Patógeno , Intestinos/imunologia , Intestinos/microbiologia , Camundongos Endogâmicos NOD , Fenótipo , Células Th1/imunologia , Células Th1/microbiologia , Células Th17/imunologia , Células Th17/microbiologia , Fatores de Tempo
4.
J Immunol ; 198(11): 4255-4267, 2017 06 01.
Artigo em Inglês | MEDLINE | ID: mdl-28461573

RESUMO

B lymphocytes play a key role in type 1 diabetes (T1D) development by serving as a subset of APCs preferentially supporting the expansion of autoreactive pathogenic T cells. As a result of their pathogenic importance, B lymphocyte-targeted therapies have received considerable interest as potential T1D interventions. Unfortunately, the B lymphocyte-directed T1D interventions tested to date failed to halt ß cell demise. IgG autoantibodies marking humans at future risk for T1D indicate that B lymphocytes producing them have undergone the affinity-maturation processes of class switch recombination and, possibly, somatic hypermutation. This study found that CRISPR/Cas9-mediated ablation of the activation-induced cytidine deaminase gene required for class switch recombination/somatic hypermutation induction inhibits T1D development in the NOD mouse model. The activation-induced cytidine deaminase protein induces genome-wide DNA breaks that, if not repaired through RAD51-mediated homologous recombination, result in B lymphocyte death. Treatment with the RAD51 inhibitor 4,4'-diisothiocyanatostilbene-2, 2'-disulfonic acid also strongly inhibited T1D development in NOD mice. The genetic and small molecule-targeting approaches expanded CD73+ B lymphocytes that exert regulatory activity suppressing diabetogenic T cell responses. Hence, an initial CRISPR/Cas9-mediated genetic modification approach has identified the AID/RAD51 axis as a target for a potentially clinically translatable pharmacological approach that can block T1D development by converting B lymphocytes to a disease-inhibitory CD73+ regulatory state.


Assuntos
Linfócitos B Reguladores/imunologia , Proteínas de Transporte/antagonistas & inibidores , Citidina Desaminase/antagonistas & inibidores , Diabetes Mellitus Tipo 1/imunologia , Diabetes Mellitus Tipo 1/prevenção & controle , Ativação Linfocitária , Proteínas Nucleares/antagonistas & inibidores , Ácido 4,4'-Di-Isotiocianoestilbeno-2,2'-Dissulfônico/farmacologia , 5'-Nucleotidase/imunologia , Animais , Autoanticorpos/imunologia , Sistemas CRISPR-Cas , Proteínas de Transporte/genética , Proteínas de Transporte/metabolismo , Citidina Desaminase/genética , Citidina Desaminase/metabolismo , Diabetes Mellitus Experimental , Switching de Imunoglobulina , Camundongos , Camundongos Endogâmicos NOD , Proteínas Nucleares/deficiência , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Hipermutação Somática de Imunoglobulina
5.
Diabetes ; 66(3): 710-721, 2017 03.
Artigo em Inglês | MEDLINE | ID: mdl-27920091

RESUMO

Type 1 diabetes development in the NOD mouse model is widely reported to be dependent on high-level production by autoreactive CD4+ and CD8+ T cells of interferon-γ (IFN-γ), generally considered a proinflammatory cytokine. However, IFN-γ can also participate in tolerance-induction pathways, indicating it is not solely proinflammatory. This study addresses how IFN-γ can suppress activation of diabetogenic CD8+ T cells. CD8+ T cells transgenically expressing the diabetogenic AI4 T-cell receptor adoptively transferred disease to otherwise unmanipulated NOD.IFN-γnull , but not standard NOD, mice. AI4 T cells only underwent vigorous intrasplenic proliferation in NOD.IFN-γnull recipients. Disease-protective IFN-γ could be derived from any lymphocyte source and suppressed diabetogenic CD8+ T-cell responses both directly and through an intermediary nonlymphoid cell population. Suppression was not dependent on regulatory T cells, but was associated with increased inhibitory STAT1 to STAT4 expression levels in pathogenic AI4 T cells. Importantly, IFN-γ exposure during activation reduced the cytotoxicity of human-origin type 1 diabetes-relevant autoreactive CD8+ T cells. Collectively, these results indicate that rather than marking the most proinflammatory lymphocytes in diabetes development, IFN-γ production could represent an attempted limitation of pathogenic CD8+ T-cell activation. Thus, great care should be taken when designing possible diabetic intervention approaches modulating IFN-γ production.


Assuntos
Diabetes Mellitus Tipo 1/imunologia , Interferon gama/imunologia , Ativação Linfocitária/imunologia , RNA Mensageiro/metabolismo , Linfócitos T Reguladores/imunologia , Transferência Adotiva , Animais , Linfócitos T CD8-Positivos/efeitos dos fármacos , Linfócitos T CD8-Positivos/imunologia , Células Cultivadas , Feminino , Humanos , Interferon gama/genética , Interferon gama/farmacologia , Ativação Linfocitária/efeitos dos fármacos , Camundongos , Camundongos Endogâmicos NOD , Camundongos Transgênicos , Receptores de Antígenos de Linfócitos T/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Fator de Transcrição STAT1/metabolismo , Fator de Transcrição STAT4/metabolismo , Baço/citologia , Linfócitos T Reguladores/efeitos dos fármacos
6.
Diabetes ; 65(7): 1977-1987, 2016 07.
Artigo em Inglês | MEDLINE | ID: mdl-26961115

RESUMO

While the autoimmune destruction of pancreatic ß-cells underlying type 1 diabetes (1D) development is ultimately mediated by T-cells in NOD mice and also likely humans, B-lymphocytes play an additional key pathogenic role. It appears expression of plasma membrane bound immunoglobulin (Ig) molecules that efficiently capture ß-cell antigens allows autoreactive B-lymphocytes bypassing normal tolerance induction processes to be the subset of antigen presenting cells most efficiently activating diabetogenic T-cells. NOD mice transgenically expressing Ig molecules recognizing antigens that are (insulin) or not (hen egg lysozyme; HEL) expressed by ß-cells have proven useful in dissecting the developmental basis of diabetogenic B-lymphocytes. However, these transgenic Ig specificities were originally selected for their ability to recognize insulin or HEL as foreign, rather than autoantigens. Thus, we generated and characterized NOD mice transgenically expressing an Ig molecule representative of a large proportion of naturally occurring islet-infiltrating B-lymphocytes in NOD mice recognizing the neuronal antigen peripherin. Transgenic peripherin autoreactive B-lymphocytes infiltrate NOD pancreatic islets, acquire an activated proliferative phenotype, and potently support accelerated T1D development. These results support the concept of neuronal autoimmunity as a pathogenic feature of T1D, and targeting such responses could ultimately provide an effective disease intervention approach.

7.
J Immunol ; 192(7): 3080-90, 2014 Apr 01.
Artigo em Inglês | MEDLINE | ID: mdl-24610011

RESUMO

Autoreactive B cells are essential for the pathogenesis of type 1 diabetes. The genesis and dynamics of autoreactive B cells remain unknown. In this study, we analyzed the immune response in the NOD mouse model to the neuronal protein peripherin (PRPH), a target Ag of islet-infiltrating B cells. PRPH autoreactive B cells recognized a single linear epitope of this protein, in contrast to the multiple epitope recognition commonly observed during autoreactive B cell responses. Autoantibodies to this epitope were also detected in the disease-resistant NOR and C57BL/6 strains. To specifically detect the accumulation of these B cells, we developed a novel approach, octameric peptide display, to follow the dynamics and localization of anti-PRPH B cells during disease progression. Before extended insulitis was established, anti-PRPH B cells preferentially accumulated in the peritoneum. Anti-PRPH B cells were likewise detected in C57BL/6 mice, albeit at lower frequencies. As disease unfolded in NOD mice, anti-PRPH B cells invaded the islets and increased in number at the peritoneum of diabetic but not prediabetic mice. Isotype-switched B cells were only detected in the peritoneum. Anti-PRPH B cells represent a heterogeneous population composed of both B1 and B2 subsets. In the spleen, anti-PRPH B cell were predominantly in the follicular subset. Therefore, anti-PRPH B cells represent a heterogeneous population that is generated early in life but proliferates as diabetes is established. These findings on the temporal and spatial progression of autoreactive B cells should be relevant for our understanding of B cell function in diabetes pathogenesis.


Assuntos
Linfócitos B/imunologia , Diabetes Mellitus Tipo 1/imunologia , Ilhotas Pancreáticas/imunologia , Periferinas/imunologia , Sequência de Aminoácidos , Animais , Autoanticorpos/imunologia , Autoanticorpos/metabolismo , Linfócitos B/metabolismo , Linfócitos B/patologia , Western Blotting , Linhagem Celular Tumoral , Diabetes Mellitus Tipo 1/metabolismo , Diabetes Mellitus Tipo 1/patologia , Progressão da Doença , Mapeamento de Epitopos/métodos , Epitopos de Linfócito B/imunologia , Epitopos de Linfócito B/metabolismo , Feminino , Ilhotas Pancreáticas/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos NOD , Microscopia de Fluorescência , Dados de Sequência Molecular , Periferinas/genética , Periferinas/metabolismo , Peritônio/imunologia , Peritônio/metabolismo , Isoformas de Proteínas/imunologia , Baço/imunologia , Baço/metabolismo
8.
PLoS One ; 7(1): e30217, 2012.
Artigo em Inglês | MEDLINE | ID: mdl-22272310

RESUMO

Glycine is the major inhibitory neurotransmitter in the spinal cord and some brain regions. The presynaptic glycine transporter, GlyT2, is required for sustained glycinergic transmission through presynaptic reuptake and recycling of glycine. Mutations in SLC6A5, encoding GlyT2, cause hereditary hyperekplexia in humans, and similar phenotypes in knock-out mice, and variants are associated with schizophrenia. We identified a spontaneous mutation in mouse Slc6a5, caused by a MusD retrotransposon insertion. The GlyT2 protein is undetectable in homozygous mutants, indicating a null allele. Homozygous mutant mice are normal at birth, but develop handling-induced spasms at five days of age, and only survive for two weeks, but allow the study of early activity-regulated developmental processes. At the neuromuscular junction, synapse elimination and the switch from embryonic to adult acetylcholine receptor subunits are hastened, consistent with a presumed increase in motor neuron activity, and transcription of acetylcholine receptors is elevated. Heterozygous mice, which show no reduction in lifespan but nonetheless have reduced levels of GlyT2, have a normal thermal sensitivity with the hot-plate test, but differences in repetitive grooming and decreased sleep time with home-cage monitoring. Open-field and elevated plus-maze tests did not detect anxiety-like behaviors; however, the latter showed a hyperactivity phenotype. Importantly, grooming and hyperactivity are observed in mouse schizophrenia models. Thus, mutations in Slc6a5 show changes in neuromuscular junction development as homozygotes, and behavioral phenotypes as heterozygotes, indicating their usefulness for studies related to glycinergic dysfunction.


Assuntos
Proteínas da Membrana Plasmática de Transporte de Glicina/genética , Mutagênese Insercional , Junção Neuromuscular/fisiologia , Desempenho Psicomotor/fisiologia , Retroelementos/genética , Animais , Ansiedade , Western Blotting , Mapeamento Cromossômico , Cromossomos de Mamíferos/genética , Feminino , Estudos de Associação Genética , Proteínas da Membrana Plasmática de Transporte de Glicina/metabolismo , Asseio Animal , Humanos , Masculino , Aprendizagem em Labirinto , Camundongos , Camundongos Endogâmicos DBA , Camundongos Endogâmicos NOD , Camundongos Knockout , Camundongos SCID , Atividade Motora , Mutação , Inibição Neural/genética , Inibição Neural/fisiologia , Junção Neuromuscular/genética , Medula Espinal/metabolismo , Medula Espinal/fisiopatologia
9.
Diabetes ; 60(11): 2914-21, 2011 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-21926271

RESUMO

OBJECTIVE: Consistent with studies in NOD mice, early clinical trials addressing whether depletion of B cells by the Rituximab CD20-specific antibody provides an effective means for type 1 diabetes reversal have produced promising results. However, to improve therapeutic efficacy, additional B-cell-depleting agents, as well as attempts seeking diabetes prevention, are being considered. RESEARCH DESIGN AND METHODS: Autoantibodies, including those against insulin (IAAs), are used to identify at-risk subjects for inclusion in diabetes prevention trials. Therefore, we tested the ability of anti-CD20 to prevent diabetes in NOD mice when administered either before or after IAA onset. RESULTS: The murine CD20-specific 18B12 antibody that like Rituximab, depletes the follicular (FO) but not marginal zone subset of B cells, efficiently inhibited diabetes development in NOD mice in a likely regulatory T-cell-dependent manner only when treatment was initiated before IAA detection. One implication of these results is that the FO subset of B cells preferentially contributes to early diabetes initiation events. However, most important, the inefficient ability of anti-CD20 treatment to exert late-stage diabetes prevention was found to be attributable to downregulation of CD20 expression upon B cell entry into pancreatic islets. CONCLUSIONS: These findings provide important guidance for designing strategies targeting B cells as a potential means of diabetes intervention.


Assuntos
Anticorpos Monoclonais Murinos/uso terapêutico , Antígenos CD20/metabolismo , Linfócitos B/efeitos dos fármacos , Diabetes Mellitus Tipo 1/metabolismo , Hipoglicemiantes/uso terapêutico , Ilhotas Pancreáticas/efeitos dos fármacos , Ilhotas Pancreáticas/metabolismo , Animais , Antígenos CD20/química , Autoanticorpos/análise , Linfócitos B/imunologia , Linfócitos B/metabolismo , Diabetes Mellitus Tipo 1/sangue , Diabetes Mellitus Tipo 1/imunologia , Diabetes Mellitus Tipo 1/prevenção & controle , Progressão da Doença , Feminino , Ilhotas Pancreáticas/imunologia , Depleção Linfocítica , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCID , Terapia de Alvo Molecular , Plasmócitos/efeitos dos fármacos , Plasmócitos/imunologia , Plasmócitos/metabolismo , Estado Pré-Diabético/sangue , Estado Pré-Diabético/imunologia , Estado Pré-Diabético/metabolismo , Rituximab
10.
J Proteome Res ; 9(3): 1203-8, 2010 Mar 05.
Artigo em Inglês | MEDLINE | ID: mdl-20113007

RESUMO

Type 1 diabetes (T1D) is an autoimmune disorder that results from the destruction of insulin-producing beta-cells in the islets of Langerhans. To date, autoimmune T-cell response and antibody reactivity to more than 20 autoantigens have been linked to this disease. Some studies have described the intermediate filament protein peripherin (PRPH) as an autoantigen associated with T1D in non-obese diabetic (NOD) mice. We evaluated immune reactivity of mouse and rabbit sera and human plasma to a 58 kDa protein expressed in RIN-m5F rat insulinoma cells. The protein was isolated using 2-DE and identified by mass spectrometry as PRPH. Antibodies from healthy humans and T1D patients, CD-1 mice, C57BL/6 mice, NOR (non-obese diabetes resistant) mice, and NOD mice reacted with PRPH on Western blots. However, antibody response to PRPH was stronger in NOD than non-autoimmune prone C57BL/6 mice. We conclude that immune reactivity to PRPH is not exclusively associated with NOD mice or human patients with T1D. Furthermore, the frequent occurrence of PRPH-reactive antibodies in mouse and human blood suggests that binding may be non-specific or could reflect the presence of natural autoantibodies against PRPH. These findings point to the need for a re-evaluation of PRPH as a T1D autoantigen in NOD mice and raise the question of the physiological relevance of such widespread immune reactivity against this peripheral nervous system protein.


Assuntos
Autoanticorpos/sangue , Proteínas de Filamentos Intermediários/imunologia , Glicoproteínas de Membrana/imunologia , Proteínas do Tecido Nervoso/imunologia , Adulto , Animais , Autoanticorpos/metabolismo , Linhagem Celular Tumoral , Diabetes Mellitus Tipo 1/sangue , Diabetes Mellitus Tipo 1/imunologia , Diabetes Mellitus Tipo 1/metabolismo , Eletroforese em Gel Bidimensional , Feminino , Humanos , Insulinoma/imunologia , Insulinoma/metabolismo , Proteínas de Filamentos Intermediários/química , Proteínas de Filamentos Intermediários/metabolismo , Masculino , Glicoproteínas de Membrana/química , Glicoproteínas de Membrana/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos NOD , Proteínas do Tecido Nervoso/química , Proteínas do Tecido Nervoso/metabolismo , Neuroblastoma/imunologia , Neuroblastoma/metabolismo , Periferinas , Coelhos , Ratos , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz , Frações Subcelulares/química , Frações Subcelulares/metabolismo
11.
Diabetes ; 57(12): 3273-80, 2008 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-18776136

RESUMO

OBJECTIVE: Although the H2(g7) major histocompatibility complex (MHC) provides the primary pathogenic component, the development of T-cell-mediated autoimmune type 1 diabetes in NOD mice also requires contributions from other susceptibility (Idd) genes. Despite sharing the H2(g7) MHC, the closely NOD-related NOR strain remains type 1 diabetes resistant because of contributions of protective Idd5.2, Idd9/11, and Idd13 region alleles. To aid their eventual identification, we evaluated cell types in which non-MHC Idd resistance genes in NOR mice exert disease-protective effects. RESEARCH DESIGN AND METHODS: Adoptive transfer and bone marrow chimerism approaches tested the diabetogenic activity of CD4 and CD8 T-cells from NOR mice and NOD stocks congenic for NOR-derived Idd resistance loci. Tetramer staining and mimotope stimulation tested the frequency and proliferative capacity of CD4 BDC2.5-like cells. Regulatory T-cells (Tregs) were identified by Foxp3 staining and functionally assessed by in vitro suppression assays. RESULTS: NOR CD4 T-cells were less diabetogenic than those from NOD mice. The failure of NOR CD4 T-cells to induce type 1 diabetes was not due to decreased proliferative capacity of BDC2.5 clonotypic-like cells. The frequency and function of Tregs in NOD and NOR mice were also equivalent. However, bone marrow chimerism experiments demonstrated that intrinsic factors inhibited the pathogenic activity of NOR CD4 T-cells. The NOR Idd9/11 resistance region on chromosome 4 was found to diminish the diabetogenic activity of CD4 but not CD8 T-cells. CONCLUSIONS: In conclusion, we demonstrated that a gene(s) within the Idd9/11 region regulates the diabetogenic activity of CD4 T-cells.


Assuntos
Linfócitos T CD4-Positivos/imunologia , Diabetes Mellitus Tipo 1/genética , Diabetes Mellitus Tipo 1/imunologia , Complexo Principal de Histocompatibilidade , Camundongos Endogâmicos NOD/genética , Animais , Células Apresentadoras de Antígenos/imunologia , Células Apresentadoras de Antígenos/patologia , Linfócitos T CD4-Positivos/patologia , Antígenos CD8/genética , Antígenos CD8/imunologia , Linfócitos T CD8-Positivos/imunologia , Linfócitos T CD8-Positivos/patologia , Mapeamento Cromossômico , Diabetes Mellitus Tipo 1/patologia , Predisposição Genética para Doença , Camundongos , Camundongos Endogâmicos C57BL , Camundongos SCID , Receptores de Antígenos de Linfócitos T alfa-beta/genética , Receptores de Antígenos de Linfócitos T alfa-beta/imunologia , Subpopulações de Linfócitos T/imunologia , Subpopulações de Linfócitos T/patologia , Linfócitos T Reguladores/imunologia , Linfócitos T Reguladores/patologia
12.
J Immunol ; 180(5): 3250-9, 2008 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-18292549

RESUMO

When expressed in NOD, but not C57BL/6 (B6) genetic background mice, the common class I variants encoded by the H2g7 MHC haplotype aberrantly lose the ability to mediate the thymic deletion of autoreactive CD8+ T cells contributing to type 1 diabetes (T1D). This indicated some subset of the T1D susceptibility (Idd) genes located outside the MHC of NOD mice interactively impair the negative selection of diabetogenic CD8+ T cells. In this study, using both linkage and congenic strain analyses, we demonstrate contributions from a polymorphic gene(s) in the previously described Idd7 locus on the proximal portion of Chromosome 7 predominantly, but not exclusively, determines the extent to which H2g7 class I molecules can mediate the thymic deletion of diabetogenic CD8+ T cells as illustrated using the AI4 TCR transgenic system. The polymorphic Idd7 region gene(s) appears to control events that respectively result in high vs low expression of the AI4 clonotypic TCR alpha-chain on developing thymocytes in B6.H2g7 and NOD background mice. This expression difference likely lowers levels of the clonotypic AI4 TCR in NOD, but not B6.H2g7 thymocytes, below the threshold presumably necessary to induce a signaling response sufficient to trigger negative selection upon Ag engagement. These findings provide further insight to how susceptibility genes, both within and outside the MHC, may interact to elicit autoreactive T cell responses mediating T1D development in both NOD mice and human patients.


Assuntos
Linfócitos T CD8-Positivos/imunologia , Deleção Clonal/genética , Diabetes Mellitus Tipo 1/genética , Diabetes Mellitus Tipo 1/imunologia , Predisposição Genética para Doença , Receptores de Antígenos de Linfócitos T alfa-beta/biossíntese , Receptores de Antígenos de Linfócitos T alfa-beta/genética , Timo/imunologia , Animais , Linfócitos T CD8-Positivos/metabolismo , Linfócitos T CD8-Positivos/patologia , Diferenciação Celular/genética , Diferenciação Celular/imunologia , Mapeamento Cromossômico , Deleção Clonal/imunologia , Células Clonais , Diabetes Mellitus Tipo 1/metabolismo , Marcadores Genéticos , Camundongos , Camundongos Congênicos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos NOD , Camundongos Transgênicos , Locos de Características Quantitativas/imunologia , Receptores de Antígenos de Linfócitos T alfa-beta/metabolismo , Timo/metabolismo , Timo/patologia
13.
Diabetes ; 56(2): 424-30, 2007 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-17259387

RESUMO

The H2(g7) (K(d), A(g7), E(null), and D(b)) major histocompatibility complex (MHC) is the primary genetic contributor to type 1 diabetes in NOD mice. NOD stocks congenically expressing other MHC haplotypes such as H2(nb1) (K(b), A(nb1), E(k), and D(b)) in a heterozygous state are type 1 diabetes resistant. Hematopoietically derived antigen-presenting cells (APCs) expressing H2(nb1) MHC molecules delete or inactivate autoreactive diabetogenic T-cells. Thus, provided a relatively benign preconditioning protocol is ultimately developed, hematopoietic chimerization by APCs expressing dominantly protective MHC molecules could conceivably provide a means for type 1 diabetes prevention in humans. Before hematopoietic chimerization can be considered for type 1 diabetes prevention, it must be determined what subtype(s) of APCs (B-cells, macrophages, and/or dendritic cells) expressing protective MHC molecules most efficiently inhibit disease, as well as the engraftment level they must achieve to accomplish this. These issues were addressed through analyses of NOD background bone marrow chimeras in which H2(nb1) molecules were selectively expressed on variable proportions of different APC subtypes. While a modest B-cell effect was observed, the strongest type 1 diabetes protection resulted from at least 50% of dendritic cells and macrophages expressing H2(nb1) molecules. At this engraftment level, H2(nb1)-expressing dendritic cells and macrophages mediated virtually complete deletion of a highly pathogenic CD8 T-cell population.


Assuntos
Células Apresentadoras de Antígenos/imunologia , Diabetes Mellitus Tipo 1/prevenção & controle , Células Secretoras de Insulina/imunologia , Complexo Principal de Histocompatibilidade/genética , Animais , Células Apresentadoras de Antígenos/classificação , Linfócitos B/metabolismo , Medula Óssea/imunologia , Antígenos CD8/genética , Quimera , Células Dendríticas/imunologia , Diabetes Mellitus Tipo 1/genética , Diabetes Mellitus Tipo 1/imunologia , Modelos Animais de Doenças , Suscetibilidade a Doenças , Feminino , Haplótipos , Imunidade Inata , Macrófagos/imunologia , Complexo Principal de Histocompatibilidade/fisiologia , Camundongos , Camundongos Endogâmicos NOD
14.
J Immunol ; 177(10): 6675-84, 2006 Nov 15.
Artigo em Inglês | MEDLINE | ID: mdl-17082580

RESUMO

In both humans and NOD mice, particular combinations of MHC genes provide the primary risk factor for development of the autoreactive T cell responses causing type 1 diabetes (T1D). Conversely, other MHC variants can confer dominant T1D resistance, and previous studies in NOD mice have shown their expression on hemopoietically derived APC is sufficient to induce disease protection. Although allogeneic hemopoietic chimerization can clearly provide a means for blocking T1D development, its clinical use for this purpose has been obviated by a requirement to precondition the host with what would be a lethal irradiation dose if bone marrow engraftment is not successful. There have been reports in which T1D-protective allogeneic hemopoietic chimerization was established in NOD mice that were preconditioned by protocols not including a lethal dose of irradiation. In most of these studies, virtually all the hemopoietic cells in the NOD recipients eventually converted to donor type. We now report that a concern about such full allogeneic chimeras is that they are severely immunocompromised potentially because their T cells are positively selected in the thymus by MHC molecules differing from those expressed by the APC available in the periphery to activate T cell effector functions. However, this undesirable side effect of generalized immunosuppression is obviated by a new protocol that establishes without a lethal preconditioning component, a stable state of mixed allogeneic hemopoietic chimerism sufficient to inhibit T1D development and also induce donor-specific tolerance in NOD recipients.


Assuntos
Diabetes Mellitus Tipo 1/imunologia , Diabetes Mellitus Tipo 1/prevenção & controle , Transplante de Células-Tronco Hematopoéticas , Quimera por Radiação/imunologia , Tolerância ao Transplante/imunologia , Animais , Linhagem Celular , Cricetinae , Células Dendríticas/imunologia , Diabetes Mellitus Tipo 1/genética , Diabetes Mellitus Tipo 1/mortalidade , Feminino , Transplante de Células-Tronco Hematopoéticas/métodos , Hemocianinas/administração & dosagem , Hemocianinas/imunologia , Depleção Linfocítica , Macrófagos/imunologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos NOD , Camundongos SCID , Camundongos Transgênicos , Transplante de Pele/imunologia , Transplante de Pele/métodos , Timo/imunologia , Timo/patologia , Tolerância ao Transplante/genética
15.
J Immunol ; 177(10): 7033-41, 2006 Nov 15.
Artigo em Inglês | MEDLINE | ID: mdl-17082619

RESUMO

Autoreactive T cells clearly mediate the pancreatic beta cell destruction causing type 1 diabetes (T1D). However, studies in NOD mice indicate that B cells also contribute to pathogenesis because their ablation by introduction of an Igmunull mutation elicits T1D resistance. T1D susceptibility is restored in NOD.Igmunull mice that are irradiated and reconstituted with syngeneic bone marrow plus NOD B cells, but not syngeneic bone marrow alone. Thus, we hypothesized some non-MHC T1D susceptibility (Idd) genes contribute to disease by allowing development of pathogenic B cells. Supporting this hypothesis was the finding that unlike those from NOD donors, engraftment with B cells from H2g7 MHC-matched, but T1D-resistant, nonobese-resistant (NOR) mice failed to restore full disease susceptibility in NOD.Igmunull recipients. T1D resistance in NOR mice is mainly encoded within the Idd13, Idd5.2, and Idd9/11 loci. B cells from NOD congenic stocks containing Idd9/11 or Idd5.1/5.2-resistance loci, respectively, derived from the NOR or C57BL/10 strains were characterized by suppressed diabetogenic activity. Immature autoreactive B cells in NOD mice have an impaired ability to be rendered anergic upon Ag engagement. Interestingly, both Idd5.1/5.2 and Idd9/11-resistance loci were found to normalize this B cell tolerogenic process, which may represent a mechanism contributing to the inhibition of T1D.


Assuntos
Subpopulações de Linfócitos B/imunologia , Subpopulações de Linfócitos B/patologia , Diabetes Mellitus Tipo 1/genética , Diabetes Mellitus Tipo 1/imunologia , Predisposição Genética para Doença , Animais , Subpopulações de Linfócitos B/metabolismo , Diferenciação Celular/genética , Diferenciação Celular/imunologia , Anergia Clonal/genética , Diabetes Mellitus Tipo 1/patologia , Feminino , Deleção de Genes , Marcadores Genéticos , Imunidade Inata/genética , Ativação Linfocitária/genética , Camundongos , Camundongos Congênicos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos NOD , Camundongos Transgênicos
16.
J Immunol ; 177(5): 2939-47, 2006 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-16920929

RESUMO

T cell-mediated autoimmune type-1 diabetes (T1D) in NOD mice partly results from this strain's numerical and functional defects in invariant NK T (iNKT) cells. T1D is inhibited in NOD mice treated with the iNKT cell superagonist alpha-galactosylceramide through a process involving enhanced accumulation of immunotolerogenic dendritic cells in pancreatic lymph nodes. Conversely, T1D is accelerated in NOD mice lacking CD38 molecules that play a role in dendritic cell migration to inflamed tissues. Unlike in standard NOD mice, alpha-galactosylceramide pretreatment did not protect the CD38-deficient stock from T1D induced by an adoptively transferred pancreatic beta cell-autoreactive CD8 T cell clone (AI4). We found that in the absence of CD38, ADP-ribosyltransferase 2 preferentially activates apoptotic deletion of peripheral iNKT cells, especially the CD4+ subset. Therefore, this study documents a previously unrecognized role for CD38 in maintaining survival of an iNKT cell subset that preferentially contributes to the maintenance of immunological tolerance.


Assuntos
ADP-Ribosil Ciclase 1/imunologia , Linfócitos T CD4-Positivos/citologia , Linfócitos T CD4-Positivos/imunologia , ADP Ribose Transferases/metabolismo , ADP-Ribosil Ciclase 1/deficiência , ADP-Ribosil Ciclase 1/genética , ADP-Ribosil Ciclase 1/metabolismo , Animais , Sobrevivência Celular , Células Cultivadas , Diabetes Mellitus/imunologia , Diabetes Mellitus/metabolismo , Ativação Linfocitária/imunologia , Camundongos , Camundongos Endogâmicos NOD
17.
J Immunol ; 174(3): 1196-204, 2005 Feb 01.
Artigo em Inglês | MEDLINE | ID: mdl-15661873

RESUMO

NKT cell activation by alpha-galactosylceramide (alpha-GalCer) inhibits autoimmune diabetes in NOD mice, in part by inducing recruitment to pancreatic lymph nodes (PLNs) of mature dendritic cells (DCs) with disease-protective effects. However, how activated NKT cells promote DC maturation, and what downstream effect this has on diabetogenic T cells was unknown. Activated NKT cells were found to produce a soluble factor(s) inducing DC maturation. Initially, there was a preferential accumulation of mature DCs in the PLNs of alpha-GalCer-treated NOD mice, followed by a substantial increase in T cells. Adoptive transfer of a diabetogenic CD8 T cell population (AI4) induced a high rate of disease (75%) in PBS-treated NOD recipients, but not in those pretreated with alpha-GalCer (8%). Significantly, more AI4 T cells accumulated in PLNs of alpha-GalCer than PBS-treated recipients, while no differences were found in mesenteric lymph nodes from each group. Compared with those in mesenteric lymph nodes, AI4 T cells entering PLNs underwent greater levels of apoptosis, and the survivors became functionally anergic. NKT cell activation enhanced this process. Hence, activated NKT cells elicit diabetes protection in NOD mice by producing a soluble factor(s) that induces DC maturation and accumulation in PLNs, where they subsequently recruit and tolerize pathogenic T cells.


Assuntos
Movimento Celular/imunologia , Células Dendríticas/imunologia , Diabetes Mellitus Tipo 1/prevenção & controle , Tolerância Imunológica , Células Matadoras Naturais/imunologia , Linfonodos/imunologia , Ativação Linfocitária , Pâncreas/imunologia , Subpopulações de Linfócitos T/imunologia , Adjuvantes Imunológicos/metabolismo , Adjuvantes Imunológicos/fisiologia , Animais , Subpopulações de Linfócitos B/citologia , Linfócitos T CD8-Positivos/citologia , Agregação Celular/imunologia , Diferenciação Celular/genética , Diferenciação Celular/imunologia , Movimento Celular/genética , Proliferação de Células , Células Cultivadas , Células Clonais , Células Dendríticas/citologia , Diabetes Mellitus Tipo 1/imunologia , Feminino , Galactosilceramidas/administração & dosagem , Galactosilceramidas/farmacologia , Galactosilceramidas/uso terapêutico , Tolerância Imunológica/genética , Células Matadoras Naturais/citologia , Células Matadoras Naturais/metabolismo , Linfonodos/citologia , Ativação Linfocitária/genética , Masculino , Camundongos , Camundongos Endogâmicos NOD , Camundongos Knockout , Pâncreas/citologia , Solubilidade , Subpopulações de Linfócitos T/citologia , Subpopulações de Linfócitos T/metabolismo
18.
J Immunol ; 173(6): 3791-800, 2004 Sep 15.
Artigo em Inglês | MEDLINE | ID: mdl-15356126

RESUMO

For unknown reasons, the common MHC class I variants encoded by the H2g7 haplotype (Kd, Db) aberrantly elicit autoreactive CD8 T cell responses essential to type 1 diabetes development when expressed in NOD mice, but not other strains. In this study, we show that interactive non-MHC genes allow a NOD-derived diabetogenic CD8 T cell clonotype (AI4) to be negatively selected at far greater efficiency in C57BL/6 mice congenically expressing H2g7 (B6.H2g7). However, the few AI4 T cells escaping negative selection in B6.H2g7 mice are exported from the thymus more efficiently, and are more functionally aggressive than those of NOD origin. This provides mechanistic insight to previous findings that resistant mouse strains carry some genes conferring greater diabetes susceptibility than the corresponding NOD allele. In the B6.H2g7 stock, non-MHC gene-controlled elevations in TCR expression are associated with both enhanced negative selection of diabetogenic CD8 T cells and increased aggressiveness of those escaping this process. An implication of this finding is that the same phenotype, in this case relatively high TCR expression levels, could have double-edged sword effects, contributing to type 1 diabetes resistance at one level of T cell development, but at another actually promoting pathogenesis.


Assuntos
Linfócitos T CD8-Positivos/imunologia , Linfócitos T CD8-Positivos/patologia , Morte Celular/genética , Morte Celular/imunologia , Diabetes Mellitus Tipo 1/genética , Diabetes Mellitus Tipo 1/imunologia , Predisposição Genética para Doença , Antígenos H-2/genética , Animais , Antígenos de Diferenciação de Linfócitos T/fisiologia , Apoptose/imunologia , Diferenciação Celular/imunologia , Membrana Celular/imunologia , Membrana Celular/metabolismo , Movimento Celular/genética , Movimento Celular/imunologia , Deleção Clonal/genética , Deleção Clonal/imunologia , Células Clonais , Diabetes Mellitus Tipo 1/patologia , Feminino , Antígenos H-2/fisiologia , Homeostase/genética , Homeostase/imunologia , Tolerância Imunológica/genética , Linfopenia/genética , Linfopenia/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos DBA , Camundongos Endogâmicos NOD , Camundongos SCID , Camundongos Transgênicos , Receptores de Antígenos de Linfócitos T/biossíntese , Timo/imunologia , Timo/patologia
19.
J Immunol ; 172(8): 5086-94, 2004 Apr 15.
Artigo em Inglês | MEDLINE | ID: mdl-15067092

RESUMO

One mechanism whereby B cells contribute to type 1 diabetes in nonobese diabetic (NOD) mice is as a subset of APCs that preferentially presents MHC class II-bound pancreatic beta cell Ags to autoreactive CD4 T cells. This results from their ability to use cell surface Ig to specifically capture beta cell Ags. Hence, we postulated a diabetogenic role for defects in the tolerance mechanisms normally blocking the maturation and/or activation of B cells expressing autoreactive Ig receptors. We compared B cell tolerance mechanisms in NOD mice with nonautoimmune strains by using the IgHEL and Ig3-83 transgenic systems, in which the majority of B cells recognize one defined Ag. NOD- and nonautoimmune-prone mice did not differ in ability to delete or receptor edit B cells recognizing membrane-bound self Ags. However, in contrast to the nonautoimmune-prone background, B cells recognizing soluble self Ags in NOD mice did not undergo partial deletion and were also not efficiently anergized. The defective induction of B cell tolerance to soluble autoantigens is most likely responsible for the generation of diabetogenic APC in NOD mice.


Assuntos
Células Apresentadoras de Antígenos/citologia , Células Apresentadoras de Antígenos/imunologia , Subpopulações de Linfócitos B/imunologia , Subpopulações de Linfócitos B/patologia , Diabetes Mellitus Tipo 1/imunologia , Diabetes Mellitus Tipo 1/patologia , Animais , Apresentação do Antígeno/genética , Células Apresentadoras de Antígenos/metabolismo , Autoantígenos/metabolismo , Subpopulações de Linfócitos B/metabolismo , Diferenciação Celular/genética , Diferenciação Celular/imunologia , Anergia Clonal/genética , Deleção Clonal/genética , Diabetes Mellitus Tipo 1/genética , Feminino , Proteínas de Membrana/imunologia , Proteínas de Membrana/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos NOD , Camundongos Transgênicos , Edição de RNA/genética , Receptores de Antígenos de Linfócitos T/genética , Solubilidade
20.
Diabetes ; 52(5): 1119-27, 2003 May.
Artigo em Inglês | MEDLINE | ID: mdl-12716741

RESUMO

Genes in the early region 3 (E3) of the adenovirus genome allow the virus to evade host immune responses by interfering with major histocompatibility (MHC) class I-mediated antigen presentation and tumor necrosis factor-alpha (TNF-alpha)- or Fas-induced apoptosis of infected cells. Autoimmune type 1 diabetes (T1D) is inhibited in NOD mice transgenically expressing all E3 genes under control of a rat insulin promoter (RIPE3/NOD). For dissecting the protective mechanisms afforded by various E3 genes, they were subdivided into RIP-driven transgene constructs. Strong T1D protection mediated at the beta-cell level characterized DL704/NOD mice lacking the E3 gp19K gene suppressing MHC class I expression but retaining the 10.4K, 14.5K, and 14.7K genes inhibiting Fas- or TNF-alpha-induced apoptosis and TNF-alpha-induced NF-kB activation. Much weaker protection characterized DL309/NOD mice expressing the gp19K but not the 10.4K, 14.5K, and 14.7K genes. While RIPE3/NOD splenocytes had an unexpected decrease in ability to adoptively transfer T1D, splenocytes from both the DL704 and DL309 stocks efficiently did so. These findings indicate that all E3 genes must be expressed to inhibit the diabetogenic potential of NOD immune cells. They also demonstrate that the antiapoptotic E3 genes most effectively protect pancreatic beta-cells from diabetogenic immune responses.


Assuntos
Adenoviridae/genética , Diabetes Mellitus Tipo 1/prevenção & controle , Diabetes Mellitus Tipo 1/virologia , Genoma Viral , Animais , Células da Medula Óssea/patologia , Diabetes Mellitus Tipo 1/genética , Diabetes Mellitus Tipo 1/imunologia , Feminino , Insulina/genética , Complexo Principal de Histocompatibilidade , Masculino , Camundongos , Camundongos Endogâmicos NOD , Peso Molecular , Regiões Promotoras Genéticas , Ratos , Proteínas Virais/genética
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