RESUMO
Type 1 diabetes (T1D) is a chronic multi-factorial disorder characterized by the immune-mediated destruction of insulin-producing pancreatic beta cells. Variations at a large number of genes influence susceptibility to spontaneous autoimmune T1D in non-obese diabetic (NOD) mice, one of the most frequently studied animal models for human disease. The genetic analysis of these mice allowed the identification of many insulin-dependent diabetes (Idd) loci and candidate genes, one of them being Cd101. CD101 is a heavily glycosylated transmembrane molecule which exhibits negative-costimulatory functions and promotes regulatory T (Treg) function. It is abundantly expressed on subsets of lymphoid and myeloid cells, particularly within the gastrointestinal tract. We have recently reported that the genotype-dependent expression of CD101 correlates with a decreased susceptibility to T1D in NOD.B6 Idd10 congenic mice compared to parental NOD controls. Here we show that the knockout of CD101 within the introgressed B6-derived Idd10 region increased T1D frequency to that of the NOD strain. This loss of protection from T1D was paralleled by decreased Gr1-expressing myeloid cells and FoxP3+ Tregs and an enhanced accumulation of CD4-positive over CD8-positive T lymphocytes in pancreatic tissues. As compared to CD101+/+ NOD.B6 Idd10 donors, adoptive T cell transfers from CD101-/- NOD.B6 Idd10 mice increased T1D frequency in lymphopenic NOD scid and NOD.B6 Idd10 scid recipients. Increased T1D frequency correlated with a more rapid expansion of the transferred CD101-/- T cells and a lower proportion of recipient Gr1-expressing myeloid cells in the pancreatic lymph nodes. Fewer of the Gr1+ cells in the recipients receiving CD101-/- T cells expressed CD101 and the cells had lower levels of IL-10 and TGF-ß mRNA. Thus, our results connect the Cd101 haplotype-dependent protection from T1D to an anti-diabetogenic function of CD101-expressing Tregs and Gr1-positive myeloid cells and confirm the identity of Cd101 as Idd10.
Assuntos
Antígenos CD/genética , Antígenos Ly/genética , Diabetes Mellitus Tipo 1/genética , Pâncreas/metabolismo , Animais , Linfócitos T CD4-Positivos/metabolismo , Linfócitos T CD8-Positivos/metabolismo , Diabetes Mellitus Tipo 1/patologia , Regulação da Expressão Gênica/genética , Predisposição Genética para Doença , Haplótipos/genética , Humanos , Linfonodos/metabolismo , Linfonodos/patologia , Camundongos , Camundongos Endogâmicos NOD , Camundongos Knockout , Células Mieloides/imunologia , Células Mieloides/metabolismo , Pâncreas/patologia , Linfócitos T Reguladores/imunologia , Linfócitos T Reguladores/metabolismoRESUMO
By congenic strain mapping using autoimmune NOD.C57BL/6J congenic mice, we demonstrated previously that the type 1 diabetes (T1D) protection associated with the insulin-dependent diabetes (Idd)10 locus on chromosome 3, originally identified by linkage analysis, was in fact due to three closely linked Idd loci: Idd10, Idd18.1, and Idd18.3. In this study, we define two additional Idd loci--Idd18.2 and Idd18.4--within the boundaries of this cluster of disease-associated genes. Idd18.2 is 1.31 Mb and contains 18 genes, including Ptpn22, which encodes a phosphatase that negatively regulates T and B cell signaling. The human ortholog of Ptpn22, PTPN22, is associated with numerous autoimmune diseases, including T1D. We, therefore, assessed Ptpn22 as a candidate for Idd18.2; resequencing of the NOD Ptpn22 allele revealed 183 single nucleotide polymorphisms with the C57BL/6J (B6) allele--6 exonic and 177 intronic. Functional studies showed higher expression of full-length Ptpn22 RNA and protein, and decreased TCR signaling in congenic strains with B6-derived Idd18.2 susceptibility alleles. The 953-kb Idd18.4 locus contains eight genes, including the candidate Cd2. The CD2 pathway is associated with the human autoimmune disease, multiple sclerosis, and mice with NOD-derived susceptibility alleles at Idd18.4 have lower CD2 expression on B cells. Furthermore, we observed that susceptibility alleles at Idd18.2 can mask the protection provided by Idd10/Cd101 or Idd18.1/Vav3 and Idd18.3. In summary, we describe two new T1D loci, Idd18.2 and Idd18.4, candidate genes within each region, and demonstrate the complex nature of genetic interactions underlying the development of T1D in the NOD mouse model.
Assuntos
Antígenos CD2/genética , Cromossomos de Mamíferos/genética , Diabetes Mellitus Tipo 1/genética , Predisposição Genética para Doença , Polimorfismo de Nucleotídeo Único , Proteína Tirosina Fosfatase não Receptora Tipo 22/genética , Alelos , Animais , Linfócitos B/imunologia , Linfócitos B/patologia , Antígenos CD2/imunologia , Cromossomos de Mamíferos/imunologia , Diabetes Mellitus Tipo 1/imunologia , Regulação da Expressão Gênica/imunologia , Loci Gênicos/imunologia , Humanos , Camundongos , Camundongos Endogâmicos NOD , Camundongos Transgênicos , Dados de Sequência Molecular , Proteína Tirosina Fosfatase não Receptora Tipo 22/imunologia , Transdução de Sinais/genética , Transdução de Sinais/imunologia , Linfócitos T/imunologia , Linfócitos T/patologiaRESUMO
Environmental and genetic factors define the susceptibility of an individual to autoimmune disease. Although common genetic pathways affect general immunological tolerance mechanisms in autoimmunity, the effects of such genes could vary under distinct immune challenges within different tissues. In this study, we demonstrate this by observing that autoimmune type 1 diabetes-protective haplotypes at the insulin-dependent diabetes susceptibility region 10 (Idd10) introgressed from chromosome 3 of C57BL/6 (B6) and A/J mice onto the NOD background increase the severity of autoimmune primary biliary cirrhosis induced by infection with Novosphingobium aromaticivorans, a ubiquitous alphaproteobacterium, when compared with mice having the NOD and NOD.CAST Idd10 type 1 diabetes-susceptible haplotypes. Substantially increased liver pathology in mice having the B6 and A/J Idd10 haplotypes correlates with reduced expression of CD101 on dendritic cells, macrophages, and granulocytes following infection, delayed clearance of N. aromaticivorans, and the promotion of overzealous IFN-γ- and IL-17-dominated T cell responses essential for the adoptive transfer of liver lesions. CD101-knockout mice generated on the B6 background also exhibit substantially more severe N. aromaticivorans-induced liver disease correlating with increased IFN-γ and IL-17 responses compared with wild-type mice. These data strongly support the hypothesis that allelic variation of the Cd101 gene, located in the Idd10 region, alters the severity of liver autoimmunity induced by N. aromaticivorans.
Assuntos
Antígenos CD/genética , Predisposição Genética para Doença/genética , Infecções por Bactérias Gram-Negativas/imunologia , Hepatite Autoimune/imunologia , Cirrose Hepática Biliar/imunologia , Sphingomonadaceae/imunologia , Animais , Antígenos CD/imunologia , Feminino , Infecções por Bactérias Gram-Negativas/genética , Infecções por Bactérias Gram-Negativas/patologia , Hepatite Autoimune/genética , Hepatite Autoimune/microbiologia , Cirrose Hepática Biliar/genética , Cirrose Hepática Biliar/patologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos NOD , Camundongos Knockout , Camundongos Transgênicos , Índice de Gravidade de DoençaRESUMO
We have previously proposed that sequence variation of the CD101 gene between NOD and C57BL/6 mice accounts for the protection from type 1 diabetes (T1D) provided by the insulin-dependent diabetes susceptibility region 10 (Idd10), a <1 Mb region on mouse chromosome 3. In this study, we provide further support for the hypothesis that Cd101 is Idd10 using haplotype and expression analyses of novel Idd10 congenic strains coupled to the development of a CD101 knockout mouse. Susceptibility to T1D was correlated with genotype-dependent CD101 expression on multiple cell subsets, including Foxp3(+) regulatory CD4(+) T cells, CD11c(+) dendritic cells, and Gr1(+) myeloid cells. The correlation of CD101 expression on immune cells from four independent Idd10 haplotypes with the development of T1D supports the identity of Cd101 as Idd10. Because CD101 has been associated with regulatory T and Ag presentation cell functions, our results provide a further link between immune regulation and susceptibility to T1D.
Assuntos
Antígenos CD/genética , Diabetes Mellitus Tipo 1/genética , Diabetes Mellitus Tipo 1/imunologia , Animais , Antígenos CD/biossíntese , Doenças Autoimunes/genética , Doenças Autoimunes/imunologia , Células CHO , Cricetinae , Cricetulus , Modelos Animais de Doenças , Predisposição Genética para Doença , Genótipo , Haplótipos , Camundongos , Camundongos Congênicos , Camundongos Endogâmicos A , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos NOD , Camundongos Knockout , Dados de Sequência MolecularRESUMO
We have used the public sequencing and annotation of the mouse genome to delimit the previously resolved type 1 diabetes (T1D) insulin-dependent diabetes (Idd)18 interval to a region on chromosome 3 that includes the immunologically relevant candidate gene, Vav3. To test the candidacy of Vav3, we developed a novel congenic strain that enabled the resolution of Idd18 to a 604-kb interval, designated Idd18.1, which contains only two annotated genes: the complete sequence of Vav3 and the last exon of the gene encoding NETRIN G1, Ntng1. Targeted sequencing of Idd18.1 in the NOD mouse strain revealed that allelic variation between NOD and C57BL/6J (B6) occurs in noncoding regions with 138 single nucleotide polymorphisms concentrated in the introns between exons 20 and 27 and immediately after the 3' untranslated region. We observed differential expression of VAV3 RNA transcripts in thymocytes when comparing congenic mouse strains with B6 or NOD alleles at Idd18.1. The T1D protection associated with B6 alleles of Idd18.1/Vav3 requires the presence of B6 protective alleles at Idd3, which are correlated with increased IL-2 production and regulatory T cell function. In the absence of B6 protective alleles at Idd3, we detected a second T1D protective B6 locus, Idd18.3, which is closely linked to, but distinct from, Idd18.1. Therefore, genetic mapping, sequencing, and gene expression evidence indicate that alteration of VAV3 expression is an etiological factor in the development of autoimmune beta-cell destruction in NOD mice. This study also demonstrates that a congenic strain mapping approach can isolate closely linked susceptibility genes.
Assuntos
Diabetes Mellitus Tipo 1/genética , Diabetes Mellitus Tipo 1/imunologia , Loci Gênicos/imunologia , Predisposição Genética para Doença , Insulina/fisiologia , Camundongos Congênicos , Mapeamento Físico do Cromossomo/métodos , Proteínas Proto-Oncogênicas c-vav/genética , Alelos , Animais , Cruzamentos Genéticos , Diabetes Mellitus Tipo 1/patologia , Modelos Animais de Doenças , Éxons/genética , Feminino , Regulação da Expressão Gênica/imunologia , Insulina/genética , Células Secretoras de Insulina/imunologia , Células Secretoras de Insulina/metabolismo , Células Secretoras de Insulina/patologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos NOD , Proteínas do Tecido Nervoso/genética , Netrinas , Proteínas Proto-Oncogênicas c-vav/biossínteseRESUMO
Recommendations on limitation of summer sunlight exposure to prevent skin cancer may conflict with requirements to protect bone health through adequate vitamin D levels, the principal source being UVB in summer sunlight. We determined whether sufficient (> or =20 ng ml(-1)) and proposed optimal (> or =32 ng ml(-1)) 25(OH)D levels are attained by following UK guidance advising casual short exposures to UVB in summer sunlight, and performed the study under known conditions to enhance the specificity of future recommendations. During wintertime, when ambient UVB is negligible, 120 white Caucasians, aged 20-60 years, from Greater Manchester, UK (53.5 degrees N) received a simulated summer's sunlight exposures, specifically 1.3 standard erythemal dose, three times weekly for 6 weeks, while wearing T-shirt and shorts. The baseline winter data predict that 5% (confidence interval (CI): 2.7-8.6) of Greater Manchester white Caucasians have deficient (<5 ng ml(-1)) 25(OH)D, 62.5% (CI: 55.2-69.4) have insufficient, and only 2.9% (CI: 1.4-5.6) have proposed optimal levels. After the simulated summer exposures, 90 (CI: 84.9-93.7) and 26.2% (CI: 20.1-33.2) reached 20 and 32 ng ml(-1) 25(OH)D, respectively. Assuming midday UVB levels, sufficient but suboptimal vitamin D status is attained after a summer's short (13 minutes) sunlight exposures to 35% skin surface area; these findings will assist future public health guidance on vitamin D acquisition.
Assuntos
Estações do Ano , Neoplasias Cutâneas/prevenção & controle , Pele/metabolismo , Pele/efeitos da radiação , Luz Solar , Adulto , Osso e Ossos/metabolismo , Relação Dose-Resposta à Radiação , Exposição Ambiental , Feminino , Guias como Assunto , Humanos , Masculino , Pessoa de Meia-Idade , Fatores de Tempo , Raios Ultravioleta , Reino Unido , Vitamina D/biossíntese , Vitamina D/sangue , População Branca , Adulto JovemRESUMO
The identification of causative genes for the autoimmune disease type 1 diabetes (T1D) in humans and candidate genes in the NOD mouse has made significant progress in recent years. In addition to sharing structural aspects of the MHC class II molecules that confer susceptibility or resistance to T1D, genes and pathways contributing to autoimmune pathogenesis are held in common by the two species. There are data demonstrating a similar need to establish central tolerance to insulin. Gene variants for the interacting molecules IL2 and CD25, members of a pathway that is essential for immune homeostasis, are present in mice and humans, respectively. Variation of two molecules that negatively regulate T cells, CTLA-4 and the tyrosine phosphatase LYP/PEP, are associated with susceptibility to human and NOD T1D. These observations underscore the value of the NOD mouse model for mechanistic studies on human T1D-associated molecular and cellular pathways.