Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 26
Filtrar
1.
Nat Immunol ; 16(11): 1153-61, 2015 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-26437244

RESUMO

Central to adaptive immunity is the interaction between the αß T cell receptor (TCR) and peptide presented by the major histocompatibility complex (MHC) molecule. Presumably reflecting TCR-MHC bias and T cell signaling constraints, the TCR universally adopts a canonical polarity atop the MHC. We report the structures of two TCRs, derived from human induced T regulatory (iT(reg)) cells, complexed to an MHC class II molecule presenting a proinsulin-derived peptide. The ternary complexes revealed a 180° polarity reversal compared to all other TCR-peptide-MHC complex structures. Namely, the iT(reg) TCR α-chain and ß-chain are overlaid with the α-chain and ß-chain of MHC class II, respectively. Nevertheless, this TCR interaction elicited a peptide-reactive, MHC-restricted T cell signal. Thus TCRs are not 'hardwired' to interact with MHC molecules in a stereotypic manner to elicit a T cell signal, a finding that fundamentally challenges our understanding of TCR recognition.


Assuntos
Autoantígenos/metabolismo , Complexo Principal de Histocompatibilidade/imunologia , Receptores de Antígenos de Linfócitos T/metabolismo , Imunidade Adaptativa , Apresentação de Antígeno , Autoantígenos/química , Autoantígenos/genética , Células Cultivadas , Antígeno HLA-DR4/química , Antígeno HLA-DR4/genética , Antígeno HLA-DR4/metabolismo , Antígenos de Histocompatibilidade Classe II/química , Antígenos de Histocompatibilidade Classe II/genética , Antígenos de Histocompatibilidade Classe II/metabolismo , Humanos , Complexo Principal de Histocompatibilidade/genética , Modelos Moleculares , Mutagênese Sítio-Dirigida , Proinsulina/química , Proinsulina/genética , Proinsulina/imunologia , Domínios e Motivos de Interação entre Proteínas , Receptores de Antígenos de Linfócitos T/química , Receptores de Antígenos de Linfócitos T/genética , Linfócitos T Reguladores/imunologia
2.
Blood ; 141(11): 1277-1292, 2023 03 16.
Artigo em Inglês | MEDLINE | ID: mdl-36044666

RESUMO

Acute graft-versus-host disease (aGVHD) is an immune cell‒driven, potentially lethal complication of allogeneic hematopoietic stem cell transplantation affecting diverse organs, including the skin, liver, and gastrointestinal (GI) tract. We applied mass cytometry (CyTOF) to dissect circulating myeloid and lymphoid cells in children with severe (grade III-IV) aGVHD treated with immune suppressive drugs alone (first-line therapy) or in combination with mesenchymal stromal cells (MSCs; second-line therapy). These results were compared with CyTOF data generated in children who underwent transplantation with no aGVHD or age-matched healthy control participants. Onset of aGVHD was associated with the appearance of CD11b+CD163+ myeloid cells in the blood and accumulation in the skin and GI tract. Distinct T-cell populations, including TCRγδ+ cells, expressing activation markers and chemokine receptors guiding homing to the skin and GI tract were found in the same blood samples. CXCR3+ T cells released inflammation-promoting factors after overnight stimulation. These results indicate that lymphoid and myeloid compartments are triggered at aGVHD onset. Immunoglobulin M (IgM) presumably class switched, plasmablasts, and 2 distinct CD11b- dendritic cell subsets were other prominent immune populations found early during the course of aGVHD in patients refractory to both first- and second-line (MSC-based) therapy. In these nonresponding patients, effector and regulatory T cells with skin- or gut-homing receptors also remained proportionally high over time, whereas their frequencies declined in therapy responders. Our results underscore the additive value of high-dimensional immune cell profiling for clinical response evaluation, which may assist timely decision-making in the management of severe aGVHD.


Assuntos
Doença Enxerto-Hospedeiro , Transplante de Células-Tronco Hematopoéticas , Transplante de Células-Tronco Mesenquimais , Criança , Humanos , Transplante de Células-Tronco Hematopoéticas/efeitos adversos , Doença Enxerto-Hospedeiro/etiologia , Transplante de Células-Tronco Mesenquimais/métodos , Terapia de Imunossupressão , Doença Aguda
3.
Eur J Immunol ; 51(6): 1377-1389, 2021 06.
Artigo em Inglês | MEDLINE | ID: mdl-33728639

RESUMO

The Ig superfamily protein glycoprotein A33 (GPA33) has been implicated in immune dysregulation, but little is known about its expression in the immune compartment. Here, we comprehensively determined GPA33 expression patterns on human blood leukocyte subsets, using mass and flow cytometry. We found that GPA33 was expressed on fractions of B, dendritic, natural killer and innate lymphoid cells. Most prominent expression was found in the CD4+ T cell compartment. Naïve and CXCR5+ regulatory T cells were GPA33high , and naïve conventional CD4+ T cells expressed intermediate GPA33 levels. The expression pattern of GPA33 identified functional heterogeneity within the CD4+ central memory T cell (Tcm) population. GPA33+ CD4+ Tcm cells were fully undifferentiated, bona fide Tcm cells that lack immediate effector function, whereas GPA33- Tcm cells exhibited rapid effector functions and may represent an early stage of differentiation into effector/effector memory T cells before loss of CD62L. Expression of GPA33 in conventional CD4+ T cells suggests a role in localization and/or preservation of an undifferentiated state. These results form a basis to study the function of GPA33 and show it to be a useful marker to discriminate between different cellular subsets, especially in the CD4+ T cell lineage.


Assuntos
Biomarcadores/metabolismo , Linfócitos T CD4-Positivos/imunologia , Leucócitos Mononucleares/imunologia , Glicoproteínas de Membrana/metabolismo , Subpopulações de Linfócitos T/imunologia , Linfócitos T Reguladores/imunologia , Diferenciação Celular , Linhagem da Célula , Separação Celular , Citometria de Fluxo , Células HEK293 , Humanos , Imunidade Inata , Memória Imunológica , Glicoproteínas de Membrana/genética , Receptores CXCR5/metabolismo
4.
Cytotherapy ; 23(3): 242-255, 2021 03.
Artigo em Inglês | MEDLINE | ID: mdl-33461863

RESUMO

Autologous, antigen-specific, tolerogenic dendritic cells (tolDCs) are presently assessed to reverse and possibly cure autoimmune diseases such as type 1 diabetes (T1D). Good Manufacturing Practice production and clinical implementation of such cell therapies critically depend on their stability and reproducible production from healthy donors and, more importantly, patient-derived monocytes. Here the authors demonstrate that tolDCs (modulated using 1,25-dihydroxyvitamin D3 and dexamethasone) displayed similar features, including protein, transcriptome and epigenome profiles, between two international clinical centers and between T1D and healthy donors, validating reproducible production. In addition, neither phenotype nor function of tolDCs was affected by repeated stimulation with inflammatory stimuli, underscoring their stability as semi-mature DCs. Furthermore, tolDCs exhibited differential DNA methylation profiles compared with inflammatory mature DCs (mDCs), and this was already largely established prior to maturation, indicating that tolDCs are locked into an immature state. Finally, approximately 80% of differentially expressed known T1D risk genes displayed a corresponding differential DNA methylome in tolDCs versus mDCs and metabolic and immune pathway genes were also differentially methylated and expressed. In summary, tolDCs are reproducible and stable clinical cell products unaffected by the T1D status of donors. The observed stable, semi-mature phenotype and function of tolDCs are exemplified by epigenetic modifications representative of immature-stage cells. Together, the authors' data provide a strong basis for the production and clinical implementation of tolDCs in the treatment of autoimmune diseases such as T1D.


Assuntos
Calcitriol , Diabetes Mellitus Tipo 1 , Calcitriol/farmacologia , Células Dendríticas , Epigênese Genética , Humanos , Tolerância Imunológica
5.
Diabetologia ; 63(2): 351-361, 2020 02.
Artigo em Inglês | MEDLINE | ID: mdl-31754749

RESUMO

AIMS/HYPOTHESIS: Heterogeneity in individuals with type 1 diabetes has become more generally appreciated, but has not yet been extensively and systematically characterised. Here, we aimed to characterise type 1 diabetes heterogeneity by creating immunological, genetic and clinical profiles for individuals with juvenile-onset type 1 diabetes in a cross-sectional study. METHODS: Participants were HLA-genotyped to determine HLA-DR-DQ risk, and SNP-genotyped to generate a non-HLA genetic risk score (GRS) based on 93 type 1 diabetes-associated SNP variants outside the MHC region. Islet autoimmunity was assessed as T cell proliferation upon stimulation with the beta cell antigens GAD65, islet antigen-2 (IA-2), preproinsulin (PPI) and defective ribosomal product of the insulin gene (INS-DRIP). Clinical parameters were collected retrospectively. RESULTS: Of 80 individuals, 67 had proliferation responses to one or more islet antigens, with vast differences in the extent of proliferation. Based on the multitude and amplitude of the proliferation responses, individuals were clustered into non-, intermediate and high responders. High responders could not be characterised entirely by enrichment for the highest risk HLA-DR3-DQ2/DR4-DQ8 genotype. However, high responders did have a significantly higher non-HLA GRS. Clinically, high T cell responses to beta cell antigens did not reflect in worsened glycaemic control, increased complications, development of associated autoimmunity or younger age at disease onset. The number of beta cell antigens that an individual responded to increased with disease duration, pointing to chronic islet autoimmunity and epitope spreading. CONCLUSIONS/INTERPRETATION: Collectively, these data provide new insights into type 1 diabetes disease heterogeneity and highlight the importance of stratifying patients on the basis of their genetic and autoimmune signatures for immunotherapy and personalised disease management.


Assuntos
Autoimunidade/fisiologia , Diabetes Mellitus Tipo 1/imunologia , Diabetes Mellitus Tipo 1/metabolismo , Adolescente , Adulto , Autoimunidade/genética , Proliferação de Células/genética , Proliferação de Células/fisiologia , Criança , Pré-Escolar , Estudos Transversais , Diabetes Mellitus Tipo 1/genética , Feminino , Genótipo , Antígenos HLA-DQ/metabolismo , Antígenos HLA-DR/metabolismo , Humanos , Insulina/metabolismo , Masculino , Análise de Componente Principal , Precursores de Proteínas/metabolismo , Estudos Retrospectivos , Linfócitos T/metabolismo , Adulto Jovem
6.
J Autoimmun ; 107: 102361, 2020 02.
Artigo em Inglês | MEDLINE | ID: mdl-31776056

RESUMO

Induction of antigen-specific regulatory T cells (Tregs) in vivo is the holy grail of current immune-regulating therapies in autoimmune diseases, such as type 1 diabetes. Tolerogenic dendritic cells (tolDCs) generated from monocytes by a combined treatment with vitamin D and dexamethasone (marked by CD52hi and CD86lo expression) induce antigen-specific Tregs. We evaluated the phenotypes of these Tregs using high-dimensional mass cytometry to identify a surface-based T cell signature of tolerogenic modulation. Naïve CD4+ T cells were stimulated with tolDCs or mature inflammatory DCs pulsed with proinsulin peptide, after which the suppressive capacity, cytokine production and phenotype of stimulated T cells were analysed. TolDCs induced suppressive T cell lines that were dominated by a naïve phenotype (CD45RA+CCR7+). These naïve T cells, however, did not show suppressive capacity, but were arrested in their naïve status. T cell cultures stimulated by tolDC further contained memory-like (CD45RA-CCR7-) T cells expressing regulatory markers Lag-3, CD161 and ICOS. T cells expressing CD25lo or CD25hi were most prominent and suppressed CD4+ proliferation, while CD25hi Tregs also effectively supressed effector CD8+ T cells. We conclude that tolDCs induce antigen-specific Tregs with various phenotypes. This extends our earlier findings pointing to a functionally diverse pool of antigen-induced and specific Tregs and provides the basis for immune-monitoring in clinical trials with tolDC.


Assuntos
Autoimunidade , Células Dendríticas/imunologia , Células Dendríticas/metabolismo , Tolerância Imunológica , Peptídeos/imunologia , Proinsulina/imunologia , Linfócitos T Reguladores/imunologia , Linfócitos T Reguladores/metabolismo , Animais , Biomarcadores , Citocinas/metabolismo , Diabetes Mellitus Tipo 1/imunologia , Diabetes Mellitus Tipo 1/metabolismo , Humanos , Imunofenotipagem , Monócitos/imunologia , Monócitos/metabolismo
7.
J Immunol ; 196(8): 3253-63, 2016 Apr 15.
Artigo em Inglês | MEDLINE | ID: mdl-26944932

RESUMO

Identifying T cell epitopes of islet autoantigens is important for understanding type 1 diabetes (T1D) immunopathogenesis and to design immune monitoring and intervention strategies in relationship to disease progression. Naturally processed T cell epitopes have been discovered by elution from HLA-DR4 of pulsed B lymphocytes. The designated professional APC directing immune responses is the dendritic cell (DC). To identify naturally processed epitopes, monocyte-derived DC were pulsed with preproinsulin (PPI), glutamic acid decarboxylase (65-kDa isoform; GAD65), and insulinoma-associated Ag-2 (IA-2), and peptides were eluted of HLA-DR3 and -DR4, which are associated with highest risk for T1D development. Proteome analysis confirmed uptake and processing of islet Ags by DC. PPI peptides generated by DC differed from those processed by B lymphocytes; PPI signal-sequence peptides were eluted from HLA-DR4 and -DR3/4 that proved completely identical to a primary target epitope of diabetogenic HLA-A2-restricted CD8 T cells. HLA-DR4 binding was confirmed. GAD65 peptides, eluted from HLA-DR3 and -DR4, encompassed two core regions overlapping the two most immunodominant and frequently studied CD4 T cell targets. GAD65 peptides bound to HLA-DR3. Strikingly, the IA-2 ligandome of HLA-DR was exclusively generated from the extracellular part of IA-2, whereas most previous immune studies have focused on intracellular IA-2 epitopes. The newly identified IA-2 peptides bound to HLA-DR3 and -DR4. Differential T cell responses were detected against the newly identified IA-2 epitopes in blood from T1D patients. The core regions to which DC may draw attention from autoreactive T cells are largely distinct and more restricted than are those of B cells. GAD65 peptides presented by DC focus on highly immunogenic T cell targets, whereas HLA-DR-binding peptides derived from IA-2 are distinct from the target regions of IA-2 autoantibodies.


Assuntos
Autoimunidade/imunologia , Células Dendríticas/imunologia , Diabetes Mellitus Tipo 1/imunologia , Antígeno HLA-DR3/imunologia , Antígeno HLA-DR4/imunologia , Ilhotas Pancreáticas/imunologia , Autoantígenos/imunologia , Linfócitos B/imunologia , Linfócitos T CD8-Positivos/imunologia , Células Cultivadas , Epitopos de Linfócito T/imunologia , Glutamato Descarboxilase/metabolismo , Humanos , Insulina/metabolismo , Ativação Linfocitária/imunologia , Ligação Proteica/imunologia , Precursores de Proteínas/metabolismo , Proteínas Tirosina Fosfatases Classe 8 Semelhantes a Receptores/metabolismo
8.
Eur J Immunol ; 43(1): 85-92, 2013 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-23042025

RESUMO

Type 1 diabetes is a T-cell-mediated autoimmune disease in which autoreactive CD8(+) T cells destroy the insulin-producing pancreatic beta cells. Vitamin D3 and dexamethasone-modulated dendritic cells (Combi-DCs) loaded with islet antigens inducing islet-specific regulatory CD4(+) T cells may offer a tissue-specific intervention therapy. The effect of Combi-DCs on CD8(+) T cells, however, remains unknown. To investigate the interaction of CD8(+) T cells with Combi-DCs presenting epitopes on HLA class I, naive, and memory CD8(+) T cells were co-cultured with DCs and proliferation and function of peptide-specific T cells were analyzed. Antigen-loaded Combi-DCs were unable to prime naïve CD8(+) T cells to proliferate, although a proportion of T cells converted to a memory phenotype. Moreover, expansion of CD8(+) T cells that had been primed by mature monocyte-derived DCs (moDCs) was curtailed by Combi-DCs in co-cultures. Combi-DCs expanded memory T cells once, but CD8(+) T-cell numbers collapsed by subsequent re-stimulation with Combi-DCs. Our data point that (re)activation of CD8(+) T cells by antigen-pulsed Combi-DCs does not promote, but rather deteriorates, CD8(+) T-cell immunity. Yet, Combi-DCs pulsed with CD8(+) T-cell epitopes also act as targets of cytotoxicity, which is undesirable for survival of Combi-DCs infused into patients in therapeutic immune intervention strategies.


Assuntos
Linfócitos T CD8-Positivos/imunologia , Células Dendríticas/imunologia , Células Dendríticas/transplante , Diabetes Mellitus Tipo 1/terapia , Depleção Linfocítica , Subpopulações de Linfócitos T/imunologia , Diferenciação Celular , Proliferação de Células , Células Cultivadas , Colecalciferol/imunologia , Deleção Clonal , Técnicas de Cocultura , Citotoxicidade Imunológica , Dexametasona/imunologia , Diabetes Mellitus Tipo 1/imunologia , Epitopos de Linfócito T/imunologia , Epitopos de Linfócito T/metabolismo , Antígenos HLA/metabolismo , Humanos , Tolerância Imunológica , Memória Imunológica , Ativação Linfocitária
9.
J Immunol ; 187(12): 6357-64, 2011 Dec 15.
Artigo em Inglês | MEDLINE | ID: mdl-22084438

RESUMO

Infectious tolerance is a term generally assigned to the process through which regulatory T cells (Tregs) transfer immunoregulatory properties to other T cells. In this study, we demonstrated that a similar process applies to human dendritic cells (DCs), albeit through a different mechanism. We induced and cloned proinsulin-specific Tregs using tolerogenic DCs and investigated mechanisms by which induced Ag-specific regulatory T cells (iaTregs) endorse the suppressive effects. iaTregs expressed FOXP3, programmed death-1, and membrane-bound TGF-ß and upregulated IL-10 and CTLA-4 after stimulation with the cognate Ag. The iaTregs suppressed effector T cells only when both encountered the cognate Ags on the same APCs (linked suppression). This occurred independently of IL-10, TGF-ß, programmed death-1, or CTLA-4. Instead, iaTregs used a granzyme B-mediated mechanism to kill B cells and monocytes, whereas proinflammatory DCs that resisted being killed were induced to upregulate the inhibitory receptors B7 (family) homolog 3 and ICOS ligand. These re-educated mature monocyte-derived dendritic cells (mDCs) suppressed effector T cells and induced IL-10-producing cells from the naive T cell pool. Our data indicated that human tolerogenic DCs confer infectious tolerance by inducing Ag-specific Tregs, which, in turn, re-educate proinflammatory mature DCs into DCs with regulatory properties.


Assuntos
Células Dendríticas/imunologia , Células Dendríticas/patologia , Epitopos de Linfócito T/imunologia , Tolerância Imunológica , Ativação Linfocitária/imunologia , Linfócitos T Reguladores/imunologia , Diferenciação Celular/imunologia , Células Cultivadas , Colecalciferol/fisiologia , Células Clonais , Técnicas de Cocultura , Células Dendríticas/metabolismo , Fatores de Transcrição Forkhead/biossíntese , Cadeias HLA-DRB1/fisiologia , Humanos , Inflamação/imunologia , Inflamação/metabolismo , Inflamação/patologia , Mediadores da Inflamação/fisiologia , Subunidade alfa de Receptor de Interleucina-2/biossíntese , Monócitos/citologia , Monócitos/imunologia , Monócitos/metabolismo , Receptor de Morte Celular Programada 1/biossíntese , Proinsulina/biossíntese , Proinsulina/imunologia , Linfócitos T Reguladores/metabolismo , Linfócitos T Reguladores/patologia
10.
Nat Med ; 12(4): 417-24, 2006 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-16550190

RESUMO

Defects in major histocompatibility complex (MHC) class I-restricted antigen presentation are frequently observed in human cancers and result in escape of tumors from cytotoxic T lymphocyte (CTL) immune surveillance in mice. Here, we show the existence of a unique category of CTLs that can prevent this escape. The CTLs target an alternative repertoire of peptide epitopes that emerge in MHC class I at the surface of cells with impaired function of transporter associated with antigen processing (TAP), tapasin or the proteasome. These peptides, although derived from self antigens such as the commonly expressed Lass5 protein (also known as Trh4), are not presented by normal cells. This explains why they act as immunogenic neoantigens. The newly discovered epitopes can be exploited for immune intervention against processing-deficient tumors through adoptive T-cell transfer or peptide vaccination.


Assuntos
Marcação de Genes , Variação Genética , Linfócitos T Citotóxicos/imunologia , Evasão Tumoral , Animais , Apresentação de Antígeno , Antiporters/deficiência , Antiporters/genética , Antiporters/fisiologia , Linfócitos T CD8-Positivos/imunologia , Linhagem Celular Transformada , Linhagem Celular Tumoral , Transformação Celular Neoplásica , Transformação Celular Viral , Células Clonais , Testes Imunológicos de Citotoxicidade , Epitopos , Genes MHC Classe I , Antígenos de Histocompatibilidade Classe I/imunologia , Imunoglobulinas/deficiência , Imunoglobulinas/genética , Imunoglobulinas/fisiologia , Vigilância Imunológica , Imunoterapia , Imunoterapia Adotiva , Proteínas de Membrana Transportadoras , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos , Camundongos Knockout , Dados de Sequência Molecular , Vacinas Sintéticas/uso terapêutico
11.
J Immunol ; 185(3): 1412-8, 2010 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-20574005

RESUMO

TNF is a pleiotropic cytokine with differential effects on immune cells and diseases. Anti-TNF therapy was shown to be effective in rheumatoid arthritis but proved inefficient or even detrimental in other autoimmune diseases. We studied the role of TNF in the induction of Ag-specific regulatory T cells (Tregs) by tolerogenic vitamin D3-modulated human dendritic cells (VD3-DCs), which previously were shown to release high amounts of soluble TNF (sTNF) upon maturation with LPS. First, production of TNF by modulated VD3-DCs was analyzed upon maturation with LPS or CD40L with respect to both secreted (cleaved) TNF (sTNF) and expression of the membrane-bound (uncleaved) form of TNF (mTNF). Next, TNF antagonists were tested for their effect on induction of Ag-specific Tregs by modulated DCs and the subsequent functionality of these Tregs. VD3-DCs expressed greater amounts of mTNF than did control DCs (nontreated DCs), independent of the maturation protocol. Inhibition of TNF with anti-TNF Ab (blocking both sTNF and mTNF) during the priming of Tregs with VD3-DCs prevented generation of Tregs and their suppression of proliferation of CD4(+) T cells. In contrast, sTNF receptor II (sTNFRII), mainly blocking sTNF, did not change the suppressive capacity of Tregs. Blocking of TNFRII by anti-CD120b Ab during Treg induction similarly abrogated their subsequent suppressive function. These data point to a specific role for mTNF on VD3-DCs in the induction of Ag-specific Tregs. Interaction between mTNF and TNFRII instructs the induction of suppressive Tregs by VD3-DCs. Anti-TNF therapy may therefore act adversely in different patients or disease pathways.


Assuntos
Células Dendríticas/imunologia , Células Dendríticas/metabolismo , Epitopos de Linfócito T/imunologia , Tolerância Imunológica , Ativação Linfocitária/imunologia , Linfócitos T Reguladores/citologia , Linfócitos T Reguladores/imunologia , Fator de Necrose Tumoral alfa/fisiologia , Diferenciação Celular/imunologia , Membrana Celular/imunologia , Membrana Celular/metabolismo , Colecalciferol/farmacologia , Humanos , Ligação Proteica/imunologia , Isoformas de Proteínas/antagonistas & inibidores , Isoformas de Proteínas/metabolismo , Isoformas de Proteínas/fisiologia , Linfócitos T Reguladores/metabolismo , Fator de Necrose Tumoral alfa/antagonistas & inibidores , Fator de Necrose Tumoral alfa/metabolismo
12.
Front Immunol ; 13: 1054968, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-36505460

RESUMO

Introduction: Restoration of immune tolerance may halt progression of autoimmune diseases. Tolerogenic dendritic cells (tolDC) inhibit antigen-specific proinflammatory T-cells, generate antigen-specific regulatory T-cells and promote IL-10 production in-vitro, providing an appealing immunotherapy to intervene in autoimmune disease progression. Methods: A placebo-controlled, dose escalation phase 1 clinical trial in nine adult patients with long-standing type 1 diabetes (T1D) demonstrated the safety and feasibility of two (prime-boost) vaccinations with tolDC pulsed with a proinsulin peptide. Immunoregulatory effects were monitored by antigen-specific T-cell assays and flow and mass cytometry. Results: The tolDC vaccine induced a profound and durable decline in pre-existing autoimmune responses to the vaccine peptide up to 3 years after therapy and temporary decline in CD4 and CD8+ T-cell responses to other islet autoantigens. While major leukocyte subsets remained stable, ICOS+CCR4+TIGIT+ Tregs and CD103+ tissue-resident and CCR6+ effector memory CD4+ T-cells increased in response to the first tolDC injection, the latter declining thereafter below baseline levels. Discussion: Our data identify immune correlates of mechanistic efficacy of intradermally injected tolDC reducing proinsulin autoimmunity in T1D.


Assuntos
Doenças Autoimunes , Diabetes Mellitus Tipo 1 , Adulto , Humanos , Células Dendríticas , Diabetes Mellitus Tipo 1/terapia , Tolerância Imunológica , Proinsulina
13.
J Autoimmun ; 37(3): 151-9, 2011 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-21636247

RESUMO

Autoreactive cytotoxic CD8 T-cells (CTLs) play a key pathogenic role in the destruction of insulin-producing beta-cells resulting in type 1 diabetes. However, knowledge regarding their targets is limited, restricting the ability to monitor the course of the disease and immune interventions. In a multi-step discovery process to identify novel CTL epitopes in human preproinsulin (PPI), PPI was digested with purified human proteasomes, and resulting COOH-fragments aligned with algorithm-predicted HLA-binding peptides to yield nine potential HLA-A1, -A2, -A3 or -B7-restricted candidates. An UV-exchange method allowed the generation of a repertoire of multimers including low-affinity HLA-binding peptides. These were labeled with quantum dot-fluorochromes and encoded in a combinatorial fashion, allowing parallel and sensitive detection of specific, low-avidity T-cells. Significantly increased frequencies of T-cells against four novel PPI epitopes (PPI(4-13)/B7, PPI(29-38)/A2, PPI(76-84)/A3 and PPI(79-88)/A3) were detected in stored blood of patients with recent onset diabetes but not in controls. Changes in frequencies of circulating CD8 T-cells against these novel epitopes were detected in blood of islet graft recipients at different time points after transplantation, which correlated with clinical outcome. In conclusion, our novel strategy involving a sensitive multiplex detection technology and requiring minimal volumes of stored blood represents a major improvement in the direct ex-vivo characterization and enumeration of immune cells in the pathogenesis of type 1 diabetes.


Assuntos
Autoimunidade , Linfócitos T CD8-Positivos/metabolismo , Técnicas de Química Combinatória , Diabetes Mellitus Tipo 1/imunologia , Células Secretoras de Insulina/metabolismo , Insulina/química , Peptídeos/química , Precursores de Proteínas/química , Algoritmos , Sequência de Aminoácidos , Linfócitos T CD8-Positivos/imunologia , Linfócitos T CD8-Positivos/patologia , Diabetes Mellitus Tipo 1/diagnóstico , Diabetes Mellitus Tipo 1/metabolismo , Diabetes Mellitus Tipo 1/patologia , Epitopos , Epitopos de Linfócito T/química , Epitopos de Linfócito T/imunologia , Epitopos de Linfócito T/metabolismo , Rejeição de Enxerto/diagnóstico , Rejeição de Enxerto/imunologia , Rejeição de Enxerto/metabolismo , Antígenos HLA-A/química , Antígenos HLA-A/imunologia , Antígenos HLA-A/metabolismo , Antígeno HLA-B7/química , Antígeno HLA-B7/imunologia , Antígeno HLA-B7/metabolismo , Humanos , Insulina/imunologia , Insulina/metabolismo , Células Secretoras de Insulina/imunologia , Células Secretoras de Insulina/patologia , Células Secretoras de Insulina/transplante , Transplante das Ilhotas Pancreáticas/imunologia , Complexo Principal de Histocompatibilidade , Dados de Sequência Molecular , Peptídeos/análise , Peptídeos/imunologia , Peptídeos/metabolismo , Complexo de Endopeptidases do Proteassoma/metabolismo , Ligação Proteica , Precursores de Proteínas/imunologia , Precursores de Proteínas/metabolismo , Pontos Quânticos
14.
J Immunol ; 183(5): 2984-94, 2009 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-19648269

RESUMO

Tolerogenic dendritic cells (TDC) offer a promising therapeutic potential to ameliorate autoimmune diseases. Reported to inhibit adaptive immune responses, little is known about their innate immunity receptor repertoire. In this study, we compared three types of human TDC (IL-10-DC, dexamethasone (DX)-DC, and 1,25(OH)(2)D(3)-DC) by their TLR expression and response to a set of TLR ligands. TDC are endowed with the same TLR set as standard monocyte-derived dendritic cells but respond differentially to the TLR stimuli Pam3CSK4, polyinosinic-polycytidylic acid, LPS, and flagellin. TDC expressed low or no IL-12-related cytokines and remarkably elevated IL-10 levels. Interestingly, only TDC up-regulated the expression of TLR2 upon stimulation. This boosted the tolerogenic potential of these cells, because IL-10 production was up-regulated in TLR2-stimulated, LPS-primed DX-DC, whereas IL-12 and TNF-alpha secretion remained low. When comparing the TDC subsets, DX-DC and 1,25(OH)(2)D(3)-DC up-regulated TLR2 irrespective of the TLR triggered, whereas in IL-10-DC this effect was only mediated by LPS. Likewise, DX-DC and 1,25(OH)(2)D(3)-DC exhibited impaired ability to mature, reduced allostimulatory properties, and hampered capacity to induce Th1 differentiation. Therefore, both DX-DC and 1,25(OH)(2)D(3)-DC display the strongest tolerogenic and anti-inflammatory features and might be most suitable tools for the treatment of autoimmune diseases.


Assuntos
Células Dendríticas/imunologia , Células Dendríticas/metabolismo , Tolerância Imunológica , Mediadores da Inflamação/antagonistas & inibidores , Receptor 2 Toll-Like/biossíntese , Receptor 2 Toll-Like/sangue , Regulação para Cima/imunologia , Diferenciação Celular/imunologia , Células Cultivadas , Células Dendríticas/classificação , Regulação para Baixo/imunologia , Retroalimentação Fisiológica/imunologia , Flagelina/antagonistas & inibidores , Flagelina/metabolismo , Humanos , Mediadores da Inflamação/sangue , Mediadores da Inflamação/fisiologia , Ligantes , Lipopeptídeos/antagonistas & inibidores , Lipopeptídeos/metabolismo , Lipopolissacarídeos/antagonistas & inibidores , Lipopolissacarídeos/metabolismo , Monócitos/citologia , Monócitos/imunologia , Monócitos/metabolismo , Poli I-C/antagonistas & inibidores , Poli I-C/metabolismo , Receptor 2 Toll-Like/agonistas
15.
Eur J Immunol ; 39(11): 3147-59, 2009 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-19688742

RESUMO

Specific therapy with modulated DC may restore immunological tolerance, thereby obviating the need for chronic immunosuppression in transplantation or autoimmunity. In this study we compared the tolerizing capacity of dexamethasone (Dex)- and 1 alpha,25-dihydroxyvitamin D3 (VD3)-modulated DC. Treatment of monocytes with either VD3 or Dex resulted in DC with stable, semi-mature phenotypes compared with standard DC, with intermediate levels of co-stimulatory and MHC class II molecules, which remained unaltered after subsequent pro-inflammatory stimulation. IL-12p70 secretion was lost by VD3- and Dex-DC, whereas IL-10 secretion was unaffected. VD3-DC distinctly produced large amounts of TNF-alpha. Both VD3- and Dex-DC possessed the capacity to convert CD4 T cells into IL-10-secreting Treg potently suppressing the proliferation of responder T cells. However, only Treg induced by VD3-DC exhibited antigen specificity. VD3-, but not Dex-, DC expressed significant high levels of PD-L1 (programmed death-1 ligand), upon activation. Blockade of PD-L1 during priming redirected T cells to produce IFN-gamma instead of IL-10 and abolished acquisition of regulatory capacity. Our findings demonstrate that both VD3- and Dex-DC possess durable but differential tolerogenic features, acting via different mechanisms. Both are potentially useful to specifically down-regulate unwanted immune responses and induce immune tolerance. These modulated DC appear suitable as adjuvant in antigen-specific clinical vaccination intervention strategies.


Assuntos
Anti-Inflamatórios/farmacologia , Antígeno B7-1/imunologia , Colecalciferol/farmacologia , Células Dendríticas/efeitos dos fármacos , Dexametasona/farmacologia , Glicoproteínas de Membrana/imunologia , Peptídeos/imunologia , Linfócitos T Reguladores/imunologia , Antígeno B7-H1 , Diferenciação Celular/efeitos dos fármacos , Diferenciação Celular/imunologia , Separação Celular , Quimiotaxia de Leucócito/imunologia , Citocinas/biossíntese , Citocinas/imunologia , Células Dendríticas/citologia , Células Dendríticas/imunologia , Citometria de Fluxo , Humanos , Tolerância Imunológica/efeitos dos fármacos , Tolerância Imunológica/imunologia , Imunomodulação/imunologia , Ativação Linfocitária/efeitos dos fármacos , Ativação Linfocitária/imunologia , Monócitos/efeitos dos fármacos , Monócitos/imunologia , Reação em Cadeia da Polimerase
16.
Diabetes ; 69(4): 670-680, 2020 04.
Artigo em Inglês | MEDLINE | ID: mdl-31896552

RESUMO

The signal peptide of preproinsulin is a major source for HLA class I autoantigen epitopes implicated in CD8 T cell (CTL)-mediated ß-cell destruction in type 1 diabetes (T1D). Among them, the 10-mer epitope located at the C-terminal end of the signal peptide was found to be the most prevalent in patients with recent-onset T1D. While the combined action of signal peptide peptidase and endoplasmic reticulum (ER) aminopeptidase 1 (ERAP1) is required for processing of the signal peptide, the mechanisms controlling signal peptide trimming and the contribution of the T1D inflammatory milieu on these mechanisms are unknown. Here, we show in human ß-cells that ER stress regulates ERAP1 gene expression at posttranscriptional level via the IRE1α/miR-17-5p axis and demonstrate that inhibition of the IRE1α activity impairs processing of preproinsulin signal peptide antigen and its recognition by specific autoreactive CTLs during inflammation. These results underscore the impact of ER stress in the increased visibility of ß-cells to the immune system and position the IRE1α/miR-17 pathway as a central component in ß-cell destruction processes and as a potential target for the treatment of autoimmune T1D.


Assuntos
Aminopeptidases/metabolismo , Linfócitos T CD8-Positivos/metabolismo , Estresse do Retículo Endoplasmático/fisiologia , Endorribonucleases/metabolismo , Células Secretoras de Insulina/metabolismo , Insulina/metabolismo , Antígenos de Histocompatibilidade Menor/metabolismo , Precursores de Proteínas/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , Aminopeptidases/genética , Linfócitos T CD8-Positivos/efeitos dos fármacos , Linfócitos T CD8-Positivos/imunologia , Regulação para Baixo/efeitos dos fármacos , Estresse do Retículo Endoplasmático/efeitos dos fármacos , Endorribonucleases/genética , Células HEK293 , Humanos , Inflamação/genética , Inflamação/metabolismo , Células Secretoras de Insulina/efeitos dos fármacos , Células Secretoras de Insulina/imunologia , Interferon gama/farmacologia , Interleucina-1beta/farmacologia , MicroRNAs/genética , MicroRNAs/metabolismo , Antígenos de Histocompatibilidade Menor/genética , Proteínas Serina-Treonina Quinases/genética , Regulação para Cima
17.
Front Immunol ; 11: 571300, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-33193353

RESUMO

During healthy pregnancy, a balanced microenvironment at the maternal-fetal interface with coordinated interaction between various immune cells is necessary to maintain immunological tolerance. While specific decidual immune cell subsets have been investigated, a system-wide unbiased approach is lacking. Here, mass cytometry was applied for data-driven, in-depth immune profiling of the total leukocyte population isolated from first, second, and third trimester decidua, as well as maternal peripheral blood at time of delivery. The maternal-fetal interface showed a unique composition of immune cells, different from peripheral blood, with significant differences between early and term pregnancy samples. Profiling revealed substantial heterogeneity in the decidual lymphoid and myeloid cell lineages that shape gestational-specific immune networks and putative differentiation trajectories over time during gestation. Uncovering the overall complexity at the maternal-fetal interface throughout pregnancy resulted in a human atlas that may serve as a foundation upon which comprehension of the immune microenvironment and alterations thereof in pregnancy complications can be built.


Assuntos
Decídua/imunologia , Leucócitos/imunologia , Linfócitos/imunologia , Perinatologia/métodos , Placenta/imunologia , Complicações na Gravidez/imunologia , Gravidez , Adulto , Células Cultivadas , Microambiente Celular , Feminino , Citometria de Fluxo , Humanos , Tolerância Imunológica , Imunofenotipagem
18.
Sci Transl Med ; 12(524)2020 01 01.
Artigo em Inglês | MEDLINE | ID: mdl-31894102

RESUMO

Helminth infections induce strong type 2 and regulatory responses, but the degree of heterogeneity of such cells is not well characterized. Using mass cytometry, we profiled these cells in Europeans and Indonesians not exposed to helminths and in Indonesians residing in rural areas infected with soil-transmitted helminths. To assign immune alteration to helminth infection, the profiling was performed before and 1 year after deworming. Very distinct signatures were found in Europeans and Indonesians, showing expanded frequencies of T helper 2 cells, particularly CD161+ cells and ILC2s in helminth-infected Indonesians, which was confirmed functionally through analysis of cytokine-producing cells. Besides ILC2s and CD4+ T cells, CD8+ T cells and γδ T cells in Indonesians produced type 2 cytokines. Regulatory T cells were also expanded in Indonesians, but only those expressing CTLA-4, and some coexpressed CD38, HLA-DR, ICOS, or CD161. CD11c+ B cells were found to be the main IL-10 producers among B cells in Indonesians, a subset that was almost absent in Europeans. A number of the distinct immune profiles were driven by helminths as the profiles reverted after clearance of helminth infections. Moreover, Indonesians with no helminth infections residing in an urban area showed immune profiles that resembled Europeans rather than rural Indonesians, which excludes a major role for ethnicity. Detailed insight into the human type 2 and regulatory networks could provide opportunities to target these cells for more precise interventions.


Assuntos
Helmintíase/imunologia , Helmintos/fisiologia , Linfócitos T Reguladores/imunologia , Células Th2/imunologia , Animais , Anti-Helmínticos/farmacologia , Anti-Helmínticos/uso terapêutico , Europa (Continente) , Helmintíase/tratamento farmacológico , Humanos , Indonésia , Interleucina-10/metabolismo , Subfamília B de Receptores Semelhantes a Lectina de Células NK/metabolismo , População Rural
19.
Cancer Res ; 67(18): 8450-5, 2007 Sep 15.
Artigo em Inglês | MEDLINE | ID: mdl-17875682

RESUMO

A large proportion of human cancers show deficiencies in the MHC class I antigen-processing machinery. Such defects render tumors resistant to immune eradication by tumoricidal CTLs. We recently identified a unique population of CTL that selectively targets tumor immune-escape variants through recognition of MHC-presented peptides, termed TEIPP (T cell epitopes associated with impaired peptide processing), expressed on cells lacking functional TAP-peptide transporters. Previously, we showed that vaccination with TEIPP peptides mediates protection against TAP-deficient tumors. Here, we further explored the concept of TEIPP-targeted therapy using a dendritic cell (DC)-based cellular vaccine. Impairment of TAP function in DC induced the presentation of endogenous TEIPP antigens by MHC class I molecules, and immunization with these DCs protected mice against the outgrowth of TAP-deficient lymphomas and fibrosarcomas. Immune analysis of vaccinated mice revealed strong TEIPP-specific CTL responses, and a crucial role for CD8(+) cells in tumor resistance. Finally, we show that TEIPP antigens could be successfully induced in wild-type DC by introducing the viral TAP inhibitor UL49.5. Our results imply that immune intervention strategies with TAP-inhibited DC could be developed for the treatment of antigen processing-deficient cancers in humans.


Assuntos
Vacinas Anticâncer/imunologia , Células Dendríticas/imunologia , Imunoterapia Adotiva/métodos , Neoplasias Experimentais/imunologia , Oligopeptídeos/imunologia , Linfócitos T Citotóxicos/imunologia , Animais , Apresentação de Antígeno , Antígenos de Neoplasias/imunologia , Fibrossarcoma/imunologia , Fibrossarcoma/prevenção & controle , Humanos , Células Matadoras Naturais/imunologia , Linfoma/imunologia , Linfoma/prevenção & controle , Camundongos , Neoplasias Experimentais/prevenção & controle , Oligopeptídeos/antagonistas & inibidores
20.
PLoS One ; 13(8): e0200818, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30089176

RESUMO

Auto-reactive CD8 T-cells play an important role in the destruction of pancreatic ß-cells resulting in type 1 diabetes (T1D). However, the phenotype of these auto-reactive cytolytic CD8 T-cells has not yet been extensively described. We used high-dimensional mass cytometry to phenotype autoantigen- (pre-proinsulin), neoantigen- (insulin-DRIP) and virus- (cytomegalovirus) reactive CD8 T-cells in peripheral blood mononuclear cells (PBMCs) of T1D patients. A panel of 33 monoclonal antibodies was designed to further characterise these cells at the single-cell level. HLA-A2 class I tetramers were used for the detection of antigen-specific CD8 T-cells. Using a novel Hierarchical Stochastic Neighbor Embedding (HSNE) tool (implemented in Cytosplore), we identified 42 clusters within the CD8 T-cell compartment of three T1D patients and revealed profound heterogeneity between individuals, as each patient displayed a distinct cluster distribution. Single-cell analysis of pre-proinsulin, insulin-DRIP and cytomegalovirus-specific CD8 T-cells showed that the detected specificities were heterogeneous between and within patients. These findings emphasize the challenge to define the obscure nature of auto-reactive CD8 T-cells.


Assuntos
Linfócitos T CD8-Positivos/citologia , Diabetes Mellitus Tipo 1/imunologia , Adulto , Autoantígenos/imunologia , Biomarcadores/sangue , Linfócitos T CD8-Positivos/imunologia , Linfócitos T CD8-Positivos/fisiologia , Feminino , Antígeno HLA-A2/imunologia , Humanos , Células Secretoras de Insulina/imunologia , Leucócitos Mononucleares/imunologia , Masculino , Fenótipo , Análise de Célula Única/métodos
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA