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1.
Nat Commun ; 12(1): 5074, 2021 08 20.
Artigo em Inglês | MEDLINE | ID: mdl-34417463

RESUMO

ß cells may participate and contribute to their own demise during Type 1 diabetes (T1D). Here we report a role of their expression of Tet2 in regulating immune killing. Tet2 is induced in murine and human ß cells with inflammation but its expression is reduced in surviving ß cells. Tet2-KO mice that receive WT bone marrow transplants develop insulitis but not diabetes and islet infiltrates do not eliminate ß cells even though immune cells from the mice can transfer diabetes to NOD/scid recipients. Tet2-KO recipients are protected from transfer of disease by diabetogenic immune cells.Tet2-KO ß cells show reduced expression of IFNγ-induced inflammatory genes that are needed to activate diabetogenic T cells. Here we show that Tet2 regulates pathologic interactions between ß cells and immune cells and controls damaging inflammatory pathways. Our data suggests that eliminating TET2 in ß cells may reduce activating pathologic immune cells and killing of ß cells.


Assuntos
Proteínas de Ligação a DNA/metabolismo , Diabetes Mellitus Tipo 1/patologia , Inflamação/patologia , Células Secretoras de Insulina/metabolismo , Células Secretoras de Insulina/patologia , Proteínas Proto-Oncogênicas/metabolismo , Animais , Sequência de Bases , Citotoxicidade Imunológica , Diabetes Mellitus Tipo 1/genética , Diabetes Mellitus Tipo 1/imunologia , Progressão da Doença , Feminino , Humanos , Imunidade , Inflamação/genética , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos NOD , Linfócitos T/imunologia , Transcrição Genética
2.
Mol Metab ; 27S: S104-S113, 2019 09.
Artigo em Inglês | MEDLINE | ID: mdl-31500821

RESUMO

BACKGROUND: The extended and clinically silent progression of Type 1 diabetes (T1D) creates a challenge for clinical interventions and for understanding the mechanisms that underlie its pathogenesis. Over the course of the development of Type 1 diabetes, studies in animal models and of human tissues have identified adaptive changes in ß cells that may affect their immunogenicity and susceptibility to killing. Loss of ß cells has traditionally been identified by impairment in function but environmental factors may affect these measurements. SCOPE OF REVIEW: In this review we will highlight features of ß cell responses to cell death, particularly in the setting of inflammation, and focus on methods of detecting ß cell death in vivo. MAJOR CONCLUSIONS: We developed an assay to measure ß cell death in vivo by detecting cell free DNA with epigenetic modifications of the INS gene that are found in ß cells. This assay has robust technical performance and identifies killing in individuals at very high risk for disease, but its ability to identify ß cell killing in at-risk relatives is limited by the short half-life of the cell free DNA and the need for repeated sampling over an extended course. We present results from the Diabetes Prevention Trial-1 using this assay. In addition, recent studies have identified cellular adaptations in some ß cells that may avoid killing but impair metabolic function. Cells with these characteristics may aggravate the autoimmune response but also may represent a potentially recoverable source of functional ß cells.


Assuntos
Diabetes Mellitus Tipo 1/metabolismo , Inflamação/metabolismo , Células Secretoras de Insulina/metabolismo , Animais , Humanos
3.
Curr Opin Immunol ; 49: 44-50, 2017 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-28992525

RESUMO

Studies with immunologics have shown that the natural history of Type 1 diabetes can be modified. These studies have targeted key mediators of the disease and recent analyses, together with studies in preclinical models have identified mechanisms that may be involved in the clinical effects. Several issues remain including specificity of the interventions, adverse effects of the treatments, and duration of their effects. Future studies are likely to include more specific approaches with agents such as cell therapies with selected immune regulatory subsets, antigen specific therapies, and combinations of agents with complementary mechanisms of activity.


Assuntos
Autoantígenos/uso terapêutico , Produtos Biológicos/uso terapêutico , Diabetes Mellitus Tipo 1/terapia , Imunoterapia/métodos , Células Secretoras de Insulina/patologia , Animais , Autoantígenos/imunologia , Diabetes Mellitus Tipo 1/imunologia , Modelos Animais de Doenças , Humanos , Tolerância Imunológica , Células Secretoras de Insulina/efeitos dos fármacos , Terapia de Alvo Molecular
4.
Cell Metab ; 25(3): 727-738, 2017 03 07.
Artigo em Inglês | MEDLINE | ID: mdl-28190773

RESUMO

Type 1 diabetes (T1D) is a chronic autoimmune disease that involves immune-mediated destruction of ß cells. How ß cells respond to immune attack is unknown. We identified a population of ß cells during the progression of T1D in non-obese diabetic (NOD) mice that survives immune attack. This population develops from normal ß cells confronted with islet infiltrates. Pathways involving cell movement, growth and proliferation, immune responses, and cell death and survival are activated in these cells. There is reduced expression of ß cell identity genes and diabetes antigens and increased immune inhibitory markers and stemness genes. This new subpopulation is resistant to killing when diabetes is precipitated with cyclophosphamide. Human ß cells show similar changes when cultured with immune cells. These changes may account for the chronicity of the disease and the long-term survival of ß cells in some patients.


Assuntos
Diabetes Mellitus Tipo 1/imunologia , Diabetes Mellitus Tipo 1/patologia , Progressão da Doença , Células Secretoras de Insulina/imunologia , Células Secretoras de Insulina/patologia , Animais , Anticorpos Monoclonais/farmacologia , Complexo CD3/metabolismo , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/imunologia , Células Cultivadas , Feminino , Humanos , Imunoterapia , Células Secretoras de Insulina/efeitos dos fármacos , Linfócitos/efeitos dos fármacos , Linfócitos/patologia , Camundongos Endogâmicos NOD , Reação em Cadeia da Polimerase em Tempo Real , Análise de Sequência de RNA , Transcrição Genética/efeitos dos fármacos , Transcriptoma/efeitos dos fármacos , Transcriptoma/genética
5.
J Biol Chem ; 291(21): 11230-40, 2016 May 20.
Artigo em Inglês | MEDLINE | ID: mdl-26994137

RESUMO

Type 1 diabetes mellitus is caused by the killing of insulin-producing ß cells by CD8+T cells. The disease progression, which is chronic, does not follow a course like responses to conventional antigens such as viruses, but accelerates as glucose tolerance deteriorates. To identify the unique features of the autoimmune effectors that may explain this behavior, we analyzed diabetogenic CD8+ T cells that recognize a peptide from the diabetes antigen IGRP (NRP-V7-reactive) in prediabetic NOD mice and compared them to others that shared their phenotype (CD44(+)CD62L(lo)PD-1(+)CXCR3(+)) but negative for diabetes antigen tetramers and to LCMV (lymphocytic choriomeningitis)-reactive CD8+ T cells. There was an increase in the frequency of the NRP-V7-reactive cells coinciding with the time of glucose intolerance. The T cells persisted in hyperglycemic NOD mice maintained with an insulin pellet despite destruction of ß cells. We compared gene expression in the three groups of cells compared with the other two subsets of cells, and the NRP-V7-reactive cells exhibited gene expression of memory precursor effector cells. They had reduced cellular proliferation and were less dependent on oxidative phosphorylation. When prediabetic NOD mice were treated with 2-deoxyglucose to block aerobic glycolysis, there was a reduction in the diabetes antigen versus other cells of similar phenotype and loss of lymphoid cells infiltrating the islets. In addition, treatment of NOD mice with 2-deoxyglucose resulted in improved ß cell granularity. These findings identify a link between metabolic disturbances and autoreactive T cells that promotes development of autoimmune diabetes.


Assuntos
Linfócitos T CD8-Positivos/imunologia , Diabetes Mellitus Tipo 1/genética , Animais , Insulina , Ilhotas Pancreáticas/imunologia , Camundongos Endogâmicos NOD , Camundongos Transgênicos , Estado Pré-Diabético/imunologia
6.
J Autoimmun ; 71: 51-8, 2016 07.
Artigo em Inglês | MEDLINE | ID: mdl-27017348

RESUMO

Type 1 diabetes (T1D) is an autoimmune disorder characterized by the destruction of insulin-producing pancreatic ß cells. Immune modulators have achieved some success in modifying the course of disease progression in T1D. However, there are parallel declines in C-peptide levels in treated and control groups after initial responses. In this review, we discuss mechanisms of ß cell death in T1D that involve necrosis and apoptosis. New technologies are being developed to enable visualization of insulitis and ß cell mass involving positron emission transmission that identifies ß cell ligands and magnetic resonance imaging that can identify vascular leakage. Molecular signatures that identify ß cell derived insulin DNA that is released from dying cells have been described and applied to clinical settings. We also consider changes in ß cells that occur during disease progression including the induction of DNA methyltransferases that may affect the function and differentiation of ß cells. Our findings from newer data suggest that the model of chronic long standing ß cell killing should be reconsidered. These studies indicate that the pathophysiology is accelerated in the peridiagnosis period and manifest by increased rates of ß cell killing and insulin secretory impairments over a shorter period than previously thought. Finally, we consider cellular explanations to account for the ongoing loss of insulin production despite continued immune therapy that may identify potential targets for treatment. The progressive decline in ß cell function raises the question as to whether ß cell failure that is independent of immune attack may be involved.


Assuntos
Diabetes Mellitus Tipo 1/etiologia , Diabetes Mellitus Tipo 1/metabolismo , Células Secretoras de Insulina/imunologia , Células Secretoras de Insulina/metabolismo , Animais , Autoimunidade , Biomarcadores , Morte Celular/genética , Morte Celular/imunologia , Sobrevivência Celular/genética , Sobrevivência Celular/imunologia , Diabetes Mellitus Tipo 1/patologia , Diabetes Mellitus Tipo 1/terapia , Predisposição Genética para Doença , Humanos , Imunoterapia/efeitos adversos , Imunoterapia/métodos , Células Secretoras de Insulina/patologia , Necrose , Estresse Fisiológico
7.
Diabetologia ; 59(5): 1021-9, 2016 May.
Artigo em Inglês | MEDLINE | ID: mdl-26910463

RESUMO

AIMS/HYPOTHESIS: Type 1 diabetes is caused by the immunological destruction of pancreatic beta cells. Preclinical and clinical data indicate that there are changes in beta cell function at different stages of the disease, but the fate of beta cells has not been closely studied. We studied how immune factors affect the function and epigenetics of beta cells during disease progression and identified possible triggers of these changes. METHODS: We studied FACS sorted beta cells and infiltrating lymphocytes from NOD mouse and human islets. Gene expression was measured by quantitative real-time RT-PCR (qRT-PCR) and methylation of the insulin genes was investigated by high-throughput and Sanger sequencing. To understand the role of DNA methyltransferases, Dnmt3a was knocked down with small interfering RNA (siRNA). The effects of cytokines on methylation and expression of the insulin gene were studied in humans and mice. RESULTS: During disease progression in NOD mice, there was an inverse relationship between the proportion of infiltrating lymphocytes and the beta cell mass. In beta cells, methylation marks in the Ins1 and Ins2 genes changed over time. Insulin gene expression appears to be most closely regulated by the methylation of Ins1 exon 2 and Ins2 exon 1. Cytokine transcription increased with age in NOD mice, and these cytokines could induce methylation marks in the insulin DNA by inducing methyltransferases. Similar changes were induced by cytokines in human beta cells in vitro. CONCLUSIONS/INTERPRETATION: Epigenetic modification of DNA by methylation in response to immunological stressors may be a mechanism that affects insulin gene expression during the progression of type 1 diabetes.


Assuntos
Metilação de DNA/genética , DNA/genética , Células Secretoras de Insulina/metabolismo , Insulina/genética , Adulto , Animais , Citocinas , Diabetes Mellitus Tipo 1 , Feminino , Humanos , Camundongos , Camundongos Endogâmicos NOD
8.
Methods Mol Biol ; 1323: 151-8, 2016.
Artigo em Inglês | MEDLINE | ID: mdl-26294406

RESUMO

Reconstituted thymus organ culture is based on fetal thymus organ culture (FTOC). Purified thymocyte populations, from genetically modified mice or even from other species, are cultured in vitro with thymic lobes depleted of their endogenous thymocytes (by 2'-deoxyguanosine treatment) to form a new thymus. This potent and timesaving method is distinct from FTOC, which assesses development of unmodified thymic lobes, and reaggregate thymic organ culture, in which epithelial cells are separately purified before being aggregated with thymocytes.


Assuntos
Técnicas de Cultura de Órgãos , Timo , Animais , Técnicas de Cultura de Células , Separação Celular , Feminino , Feto , Vetores Genéticos/genética , Camundongos , Células Precursoras de Linfócitos B/citologia , Células Precursoras de Linfócitos B/metabolismo , Gravidez , Retroviridae/genética , Timócitos/citologia , Timócitos/metabolismo , Transdução Genética
9.
J Immunol ; 189(3): 1380-90, 2012 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-22730529

RESUMO

Intrathymic CD4/CD8 differentiation is a process that establishes the mutually exclusive expression profiles of the CD4 and CD8 T cell lineage. The RUNX3-mediated silencing of CD4 in CD8 lineage cells has been well documented; however, it is unclear how CD8 is silenced during CD4 lineage differentiation. In this study, we report that, by directly binding the CD8 locus, ThPOK works as a negative regulator that mediates the deacetylation of Cd8 genes and repositions the CD8 alleles close to heterochromatin during the development of the CD4 lineage. The ectopic expression of ThPOK resulted in increased recruitment of histone deacetylases at Cd8 loci; the enhanced deacetylation of Cd8 genes eventually led to impaired Cd8 transcription. In the absence of ThPOK, the enhanced acetylation and transcription of Cd8 genes were observed. The results of these studies showed that Cd8 loci are the direct targets of ThPOK, and, more importantly, they provide new insights into CD8 silencing during CD4 lineage commitment.


Assuntos
Linfócitos T CD4-Positivos/imunologia , Antígenos CD8/genética , Diferenciação Celular/genética , Diferenciação Celular/imunologia , Proteínas de Ligação a DNA/fisiologia , Inativação Gênica/imunologia , Fatores de Transcrição/fisiologia , Acetilação , Animais , Linfócitos T CD4-Positivos/citologia , Linfócitos T CD4-Positivos/metabolismo , Antígenos CD8/metabolismo , Linhagem Celular , Proteínas de Ligação a DNA/deficiência , Regulação para Baixo/genética , Regulação para Baixo/imunologia , Epigênese Genética/imunologia , Células HEK293 , Humanos , Camundongos , Camundongos Knockout , Camundongos Transgênicos , Supressão Genética/imunologia , Fatores de Transcrição/deficiência
10.
J Immunol ; 185(7): 3960-9, 2010 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-20810990

RESUMO

The lineage-specifying factor Th-inducing POK (ThPOK) directs the intrathymic differentiation of CD4 T cells. Although the regulation of ThPOK at the transcription level has been extensively studied, specific posttranslational modifications regulating the activity of ThPOK have not been addressed. In this paper, we show that ThPOK is an unstable protein that is more readily degraded in CD8 T cells compared with CD4 T cells. Among the various proteins that bind ThPOK, acetyltransferase p300 specifically promotes the acetylation of ThPOK at K210, K216, and K339, outcompeting ubiquitination, thereby stabilizing the protein. In CD4 T cells, attenuation of p300-mediated acetylation promotes the degradation of ThPOK. In contrast, mutation of lysines 210, 216, and 339 to arginines stabilizes ThPOK and enhances its ability to suppress the expression of CD8 molecule and cytotoxic effectors in CD8 T cells. Our results reveal an essential role of p300-mediated acetylation in regulating the stability of ThPOK and suggest that such regulation may play a part in CD4/CD8 lineage differentiation.


Assuntos
Linfócitos T CD4-Positivos/citologia , Linfócitos T CD8-Positivos/citologia , Linhagem da Célula/imunologia , Proteínas de Ligação a DNA/metabolismo , Fatores de Transcrição/metabolismo , Fatores de Transcrição de p300-CBP/metabolismo , Acetilação , Animais , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD4-Positivos/metabolismo , Linfócitos T CD8-Positivos/imunologia , Linfócitos T CD8-Positivos/metabolismo , Diferenciação Celular/imunologia , Linhagem Celular , Separação Celular , Proteínas de Ligação a DNA/imunologia , Citometria de Fluxo , Humanos , Immunoblotting , Imunoprecipitação , Células Jurkat , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Fatores de Transcrição/imunologia , Fatores de Transcrição de p300-CBP/imunologia
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