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1.
Proc Natl Acad Sci U S A ; 112(44): 13447-54, 2015 Nov 03.
Artigo em Inglês | MEDLINE | ID: mdl-26483459

RESUMO

The activated B-cell-like (ABC) subtype of diffuse large B-cell lymphoma (DLBCL) relies on chronic active B-cell receptor (BCR) signaling. BCR pathway inhibitors induce remissions in a subset of ABC DLBCL patients. BCR microclusters on the surface of ABC cells resemble those generated following antigen engagement of normal B cells. We speculated that binding of lymphoma BCRs to self-antigens initiates and maintains chronic active BCR signaling in ABC DLBCL. To assess whether antigenic engagement of the BCR is required for the ongoing survival of ABC cells, we developed isogenic ABC cells that differed solely with respect to the IgH V region of their BCRs. In competitive assays with wild-type cells, substitution of a heterologous V region impaired the survival of three ABC lines. The viability of one VH4-34(+) ABC line and the ability of its BCR to bind to its own cell surface depended on V region residues that mediate the intrinsic autoreactivity of VH4-34 to self-glycoproteins. The BCR of another ABC line reacted with self-antigens in apoptotic debris, and the survival of a third ABC line was sustained by reactivity of its BCR to an idiotypic epitope in its own V region. Hence, a diverse set of self-antigens is responsible for maintaining the malignant survival of ABC DLBCL cells. IgH V regions used by the BCRs of ABC DLBCL biopsy samples varied in their ability to sustain survival of these ABC lines, suggesting a screening procedure to identify patients who might benefit from BCR pathway inhibition.


Assuntos
Autoantígenos/metabolismo , Linfócitos B/metabolismo , Linfoma Difuso de Grandes Células B/metabolismo , Receptores de Antígenos de Linfócitos B/metabolismo , Sequência de Aminoácidos , Apoptose/genética , Linfócitos B/patologia , Western Blotting , Antígenos CD79/genética , Antígenos CD79/metabolismo , Linhagem Celular Tumoral , Sobrevivência Celular/genética , Citometria de Fluxo , Humanos , Ativação Linfocitária/genética , Linfoma Difuso de Grandes Células B/genética , Linfoma Difuso de Grandes Células B/patologia , Dados de Sequência Molecular , Mutação , Ligação Proteica , Interferência de RNA , Receptores de Antígenos de Linfócitos B/genética , Transdução de Sinais/genética
2.
J Allergy Clin Immunol ; 133(4): 1149-61, 2014 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-24373350

RESUMO

BACKGROUND: Signaling lymphocytic activation molecule (SLAM)-associated protein (SAP) can mediate the function of SLAM molecules, which have been proposed to be involved in the development of autoimmunity in mice. OBJECTIVE: We sought to determine whether the SLAM/SAP pathway regulates the establishment of human B-cell tolerance and what mechanisms of B-cell tolerance could be affected by SAP deficiency. METHODS: We tested the reactivity of antibodies isolated from single B cells from SAP-deficient patients with X-linked lymphoproliferative disease (XLP). The expressions of SAP and SLAM family members were assessed in human bone marrow-developing B cells. We also analyzed regulatory T (Treg) cell function in patients with XLP and healthy control subjects. RESULTS: We found that new emigrant/transitional B cells from patients with XLP were enriched in autoreactive clones, revealing a defective central B-cell tolerance checkpoint in the absence of functional SAP. In agreement with a B cell-intrinsic regulation of central tolerance, we identified SAP expression in a discrete subset of bone marrow immature B cells. SAP colocalized with SLAMF6 only in association with clustered B-cell receptors likely recognizing self-antigens, suggesting that SLAM/SAP regulate B-cell receptor-mediated central tolerance. In addition, patients with XLP displayed defective peripheral B-cell tolerance, which is normally controlled by Treg cells. Treg cells in patients with XLP seem functional, but SAP-deficient T cells were resistant to Treg cell-mediated suppression. Indeed, SAP-deficient T cells were hyperresponsive to T-cell receptor stimulation, which resulted in increased secretion of IL-2, IFN-γ, and TNF-α. CONCLUSIONS: SAP expression is required for the counterselection of developing autoreactive B cells and prevents their T cell-dependent accumulation in the periphery.


Assuntos
Antígenos CD/metabolismo , Linfócitos B/imunologia , Linfócitos B/metabolismo , Tolerância Imunológica , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Receptores de Superfície Celular/metabolismo , Transdução de Sinais , Autoimunidade/genética , Autoimunidade/imunologia , Fator Ativador de Células B/sangue , Expressão Gênica , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/deficiência , Peptídeos e Proteínas de Sinalização Intracelular/genética , Ativação Linfocitária/imunologia , Transtornos Linfoproliferativos/genética , Transtornos Linfoproliferativos/imunologia , Transtornos Linfoproliferativos/metabolismo , Ligação Proteica , Transporte Proteico , Receptores de Antígenos de Linfócitos B/metabolismo , Proteína Associada à Molécula de Sinalização da Ativação Linfocitária , Família de Moléculas de Sinalização da Ativação Linfocitária , Membro 1 da Família de Moléculas de Sinalização da Ativação Linfocitária , Linfócitos T Reguladores/imunologia , Linfócitos T Reguladores/metabolismo
3.
J Allergy Clin Immunol ; 134(6): 1365-1374, 2014 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-25218284

RESUMO

BACKGROUND: Dedicator of cytokinesis 8 (DOCK8) deficiency is typified by recurrent infections, increased serum IgE levels, eosinophilia, and a high incidence of allergic and autoimmune manifestations. OBJECTIVE: We sought to determine the role of DOCK8 in the establishment and maintenance of human B-cell tolerance. METHODS: Autoantibodies were measured in the plasma of DOCK8-deficient patients. The antibody-coding genes from new emigrant/transitional and mature naive B cells were cloned and assessed for their ability to bind self-antigens. Regulatory T (Treg) cells in the blood were analyzed by means of flow cytometry, and their function was tested by examining their capacity to inhibit the proliferation of CD4(+)CD25(-) effector T cells. RESULTS: DOCK8-deficient patients had increased levels of autoantibodies in their plasma. We determined that central B-cell tolerance did not require DOCK8, as evidenced by the normally low frequency of polyreactive new emigrant/transitional B cells in DOCK8-deficient patients. In contrast, autoreactive B cells were enriched in the mature naive B-cell compartment, revealing a defective peripheral B-cell tolerance checkpoint. In addition, we found that Treg cells were decreased and exhibited impaired suppressive activity in DOCK8-deficient patients. CONCLUSIONS: Our data support a critical role for DOCK8 in Treg cell homeostasis and function and the enforcement of peripheral B-cell tolerance.


Assuntos
Linfócitos B/imunologia , Fatores de Troca do Nucleotídeo Guanina/deficiência , Fatores de Troca do Nucleotídeo Guanina/imunologia , Síndromes de Imunodeficiência/imunologia , Linfócitos T Reguladores/imunologia , Adolescente , Autoanticorpos/sangue , Criança , Pré-Escolar , Feminino , Humanos , Tolerância Imunológica/imunologia , Lactente , Contagem de Linfócitos , Masculino
4.
Front Immunol ; 14: 1151748, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-37795090

RESUMO

Background: Immune cell expression profiling from patient samples is critical for the successful development of immuno-oncology agents and is useful to understand mechanism-of-action, to identify exploratory biomarkers predictive of response, and to guide treatment selection and combination therapy strategies. LAG-3 is an inhibitory immune checkpoint that can suppress antitumor T-cell responses and targeting LAG-3, in combination with PD-1, is a rational approach to enhance antitumor immunity that has recently demonstrated clinical success. Here, we sought to identify human immune cell subsets that express LAG-3 and its ligands, to characterize the marker expression profile of these subsets, and to investigate the potential relationship between LAG-3 expressing subsets and clinical outcomes to immuno-oncology therapies. Methods: Comprehensive high-parameter immunophenotyping was performed using mass and flow cytometry of tumor-infiltrating lymphocytes (TILs) and peripheral blood mononuclear cells (PBMCs) from two independent cohorts of samples from patients with various solid tumor types. Profiling of circulating immune cells by single cell RNA-seq was conducted on samples from a clinical trial cohort of melanoma patients treated with immunotherapy. Results: LAG-3 was most highly expressed by subsets of tumor-infiltrating CD8 T central memory (TCM) and effector memory (TEM) cells and was frequently co-expressed with PD-1. We determined that these PD-1+ LAG-3+ CD8 memory T cells exhibited a unique marker profile, with greater expression of activation (CD69, HLA-DR), inhibitory (TIM-3, TIGIT, CTLA-4) and stimulatory (4-1BB, ICOS) markers compared to cells that expressed only PD-1 or LAG-3, or that were negative for both checkpoints. In contrast to tumors, LAG-3 expression was more limited in circulating immune cells from healthy donors and solid tumor patients. Additionally, we found abundant expression of the LAG-3 ligands MHC-II and galectin-3 in diverse immune cell types, whereas FGL1 and LSECtin were minimally expressed by immune cells in the tumor microenvironment (TME). Lastly, we found an inverse relationship between baseline and on-treatment levels of circulating LAG3 transcript-expressing CD8 memory T cells and response to combination PD-1 and CTLA-4 blockade in a clinical trial cohort of melanoma patients profiled by scRNAseq. Conclusions: These results provide insights into the nature of LAG-3- and ligand-expressing immune cells within the TME, and suggest a biological basis for informing mechanistic hypotheses, treatment selection strategies, and combination immunotherapy approaches to support continued development of dual PD-1 and LAG-3 blockade.


Assuntos
Melanoma , Receptor de Morte Celular Programada 1 , Humanos , Antígeno CTLA-4 , Receptor de Morte Celular Programada 1/metabolismo , Leucócitos Mononucleares , Imunofenotipagem , Ligantes , Microambiente Tumoral , Fibrinogênio/uso terapêutico
5.
Blood ; 115(24): 5026-36, 2010 Jun 17.
Artigo em Inglês | MEDLINE | ID: mdl-20231422

RESUMO

Complement receptor 2-negative (CR2/CD21(-)) B cells have been found enriched in patients with autoimmune diseases and in common variable immunodeficiency (CVID) patients who are prone to autoimmunity. However, the physiology of CD21(-/lo) B cells remains poorly characterized. We found that some rheumatoid arthritis (RA) patients also display an increased frequency of CD21(-/lo) B cells in their blood. A majority of CD21(-/lo) B cells from RA and CVID patients expressed germline autoreactive antibodies, which recognized nuclear and cytoplasmic structures. In addition, these B cells were unable to induce calcium flux, become activated, or proliferate in response to B-cell receptor and/or CD40 triggering, suggesting that these autoreactive B cells may be anergic. Moreover, gene array analyses of CD21(-/lo) B cells revealed molecules specifically expressed in these B cells and that are likely to induce their unresponsive stage. Thus, CD21(-/lo) B cells contain mostly autoreactive unresponsive clones, which express a specific set of molecules that may represent new biomarkers to identify anergic B cells in humans.


Assuntos
Doenças Autoimunes/imunologia , Linfócitos B/imunologia , Biomarcadores , Anergia Clonal/imunologia , Receptores de Complemento 3d/genética , Receptores de Complemento 3d/imunologia , Adulto , Antígenos CD/imunologia , Antígenos de Diferenciação de Linfócitos T/imunologia , Apoptose/imunologia , Autoanticorpos/imunologia , Doenças Autoimunes/genética , Linfócitos B/citologia , Antígenos CD40/imunologia , Divisão Celular/imunologia , Anergia Clonal/genética , Feminino , Perfilação da Expressão Gênica , Humanos , Imunofenotipagem , Ligante Coestimulador de Linfócitos T Induzíveis , Subunidade alfa de Receptor de Interleucina-2/imunologia , Lectinas Tipo C/imunologia , Ativação Linfocitária/imunologia , Masculino , Receptores de IgE/imunologia , Proteína Transmembrana Ativadora e Interagente do CAML/imunologia , Adulto Jovem
6.
Arthritis Rheum ; 63(5): 1237-45, 2011 May.
Artigo em Inglês | MEDLINE | ID: mdl-21538313

RESUMO

OBJECTIVE: Rheumatoid arthritis (RA) patients who have never received treatment for RA have been found to have defective early B cell tolerance checkpoints, resulting in impaired removal of developing autoreactive B cells. However, it is unclear whether these defects in B cell tolerance checkpoints are a primary aspect of the disease or are the result of ongoing inflammatory processes in these patients. The aim of this study was to assess the impact of standard immunosuppressive treatments, methotrexate and anti-tumor necrosis factor α (anti-TNFα) agents, on early B cell tolerance checkpoints in RA patients. METHODS: Blood samples were obtained from RA patients before and after treatment with methotrexate and/or anti-TNFα agents. B cells were tested pre- and posttherapy for reactivity of recombinant antibodies cloned from single B cells, which allowed us to determine the evolution of the frequency of autoreactive clones in the mature naive B cell compartment in RA patients before and after treatment. B cells from healthy donors were used as controls. RESULTS: Posttreatment frequencies of autoreactive mature naive B cells were elevated in the blood of RA patients. Nevertheless, the frequencies after treatment remained similar to those observed in the same patients before treatment. CONCLUSION: Despite the achievement of clinical improvement in RA patients following treatment with methotrexate and/or anti-TNFα agents, these therapies did not correct the accumulation of peripheral autoreactive mature naive B cells in these patients, suggesting that inflammation is not responsible for the defective early B cell tolerance checkpoints in RA.


Assuntos
Artrite Reumatoide/imunologia , Autoanticorpos/imunologia , Linfócitos B/imunologia , Tolerância Imunológica/imunologia , Inflamação/imunologia , Anticorpos Monoclonais/uso terapêutico , Antirreumáticos/uso terapêutico , Artrite Reumatoide/tratamento farmacológico , Ensaio de Imunoadsorção Enzimática , Feminino , Citometria de Fluxo , Imunofluorescência , Humanos , Inflamação/tratamento farmacológico , Masculino , Metotrexato/uso terapêutico , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Resultado do Tratamento , Fator de Necrose Tumoral alfa/antagonistas & inibidores
7.
J Immunol ; 184(12): 7022-9, 2010 Jun 15.
Artigo em Inglês | MEDLINE | ID: mdl-20488791

RESUMO

Alpha-defensins (or Cryptdins [Crps]) are a group of antimicrobial peptides produced as a component of Paneth cell (PC) secretory granules in the small intestine. In vivo ligation of TLR9 by synthetic agonists leads to PC degranulation, although the mechanism by which this occurs remains uncertain. In this report, we investigated TLR9-dependent mechanisms, triggered by the parasite Toxoplasma gondii, inducing Crp release in the lumen. Oral challenge of C57BL/6J (B6) wild-type (WT) mice with T. gondii induced TLR9 mRNA upregulation associated with a marked increase of type I IFN mRNA expression. PC secretory granules were released, and Crp-3/-5 mRNA expression by purified epithelial cells was increased following oral challenge of B6 WT mice. Although PCs failed to degranulate in infected B6 TLR9-/- mice, i.p. injection of mouse IFN-beta alone led to Crp-3/-5 mRNA upregulation in B6 WT and TLR9-/- mice. In addition, modulation of Crp mRNA expression in response to T. gondii infection was abrogated in B6 IFNAR-/- mice, which lack a functional type I IFN receptor. Taken together, these data demonstrate that T. gondii induces Crp-3/-5 production and release by PCs via a TLR9-dependent production of type I IFNs. Crps have a limited direct effect against T. gondii but may indirectly affect the early control of T. gondii invasiveness by promoting the initiation of a protective Th1 response against the parasite.


Assuntos
Celulas de Paneth/metabolismo , Receptor Toll-Like 9/imunologia , Toxoplasmose Animal/imunologia , Toxoplasmose/imunologia , alfa-Defensinas/metabolismo , Animais , Degranulação Celular/imunologia , Feminino , Expressão Gênica , Imunidade nas Mucosas/imunologia , Immunoblotting , Intestino Delgado/imunologia , Intestino Delgado/microbiologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Celulas de Paneth/imunologia , RNA Mensageiro/análise , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Toxoplasma , Toxoplasmose/metabolismo , alfa-Defensinas/imunologia
8.
Oncol Rep ; 47(2)2022 02.
Artigo em Inglês | MEDLINE | ID: mdl-34958112

RESUMO

Hepatocellular carcinoma (HCC) is an immunogenic malignancy, which exhibits low responsiveness to programmed cell death protein­1 (PD­1)/programmed death ligand­1 (PD­L1) antibodies. Therefore, the identification of novel immunotherapeutic targets to treat HCC is imperative. Systematic characterization of the HCC tumor microenvironment (TME) can provide novel insight into additional therapeutic approaches. In the present study, the RNA­sequencing (RNA­seq) data of 360 patients with HCC were integrated from The Cancer Genome Atlas to assess the expression of membrane spanning 4­domains A1 (MS4A1; encoding CD20) in tumors and normal liver tissues. Immunofluorescence and multiplex tissue fluorescence analyses were performed and combined with flow cytometry staining to measure CD20/CD19 expression at the protein level. In addition, published single cell RNA­seq data of CD45+ cells were derived from 16 treatment­naïve patients from Beijing Shijitan Hospital with HCC to illustrate the characteristics of CD19+ B cells. The results indicated that the HCC TME included nuclear receptor subfamily 4 group A member 2+ (NR4A2) B cells. Patients with HCC and high density of intratumoral B cells demonstrated compromised antitumor immunity manifested by low percentages of cytotoxic CD8+ T cells and high density of regulatory T cells. Furthermore, PD­L1 expression was significantly correlated with the B cell signature marker CD20. The present study indicated that tumor­infiltrating B cells may play a negative role in antitumor immunity and serve as a promising target for HCC immunotherapy, alone or in combination with anti­PD­L1/PD­1 antibodies.


Assuntos
Linfócitos B/imunologia , Carcinoma Hepatocelular/tratamento farmacológico , Carcinoma Hepatocelular/imunologia , Imunoterapia/métodos , Neoplasias Hepáticas/tratamento farmacológico , Neoplasias Hepáticas/imunologia , Microambiente Tumoral/imunologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Linfócitos B/efeitos dos fármacos , China , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Microambiente Tumoral/efeitos dos fármacos
9.
J Clin Invest ; 132(2)2022 01 18.
Artigo em Inglês | MEDLINE | ID: mdl-34813502

RESUMO

Although negative selection of developing B cells in the periphery is well described, yet poorly understood, evidence of naive B cell positive selection remains elusive. Using 2 humanized mouse models, we demonstrate that there was strong skewing of the expressed immunoglobulin repertoire upon transit into the peripheral naive B cell pool. This positive selection of expanded naive B cells in humanized mice resembled that observed in healthy human donors and was independent of autologous thymic tissue. In contrast, negative selection of autoreactive B cells required thymus-derived Tregs and MHC class II-restricted self-antigen presentation by B cells. Indeed, both defective MHC class II expression on B cells of patients with rare bare lymphocyte syndrome and prevention of self-antigen presentation via HLA-DM inhibition in humanized mice resulted in the production of autoreactive naive B cells. These latter observations suggest that Tregs repressed autoreactive naive B cells continuously produced by the bone marrow. Thus, a model emerged, in which both positive and negative selection shaped the human naive B cell repertoire and that each process was mediated by fundamentally different molecular and cellular mechanisms.


Assuntos
Apresentação de Antígeno , Linfócitos B/imunologia , Antígenos de Histocompatibilidade Classe II/imunologia , Imunodeficiência Combinada Severa/imunologia , Linfócitos T Reguladores/imunologia , Animais , Feminino , Humanos , Masculino , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCID
10.
Front Immunol ; 12: 681941, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34295332

RESUMO

Background: While all salivary glands (SGs) can be involved in primary Sjögren's syndrome (pSS), their respective role in pathogenesis remains unclear. Our objective was to assess immunopathway activation in paired parotid and labial gland tissue from biopsy-positive and biopsy-negative pSS and non-SS sicca patients. Methods: Paraffin-embedded, paired parotid and labial salivary gland tissue and peripheral blood mononuclear cells were obtained from 39 pSS and 20 non-SS sicca patients. RNA was extracted, complementary DNA libraries were prepared and sequenced. For analysis of differentially expressed genes (DEGs), patients were subdivided based on fulfillment of ACR-EULAR criteria and histopathology. Results: With principal component analysis, only biopsy-positive pSS could be separated from non-SS sicca patients based on SG gene expression. When comparing the transcriptome of biopsy-positive pSS and biopsy-negative non-SS sicca patients, 1235 and 624 DEGs (FDR<0.05, log2FC<-1 or >1) were identified for parotid and labial glands, respectively. The number of DEGs between biopsy-negative pSS and non-SS sicca patients was scarce. Overall, transcript expression levels correlated strongly between parotid and labial glands (R2 = 0.86, p-value<0.0001). Gene signatures present in both glands of biopsy-positive pSS patients included IFN-α signaling, IL-12/IL-18 signaling, CD3/CD28 T-cell activation, CD40 signaling in B-cells, DN2 B-cells, and FcRL4+ B-cells. Signature scores varied considerably amongst pSS patients. Conclusion: Transcriptomes of paired major and minor SGs in pSS were overall comparable, although significant inter-individual heterogeneity in immunopathway activation existed. The SG transcriptome of biopsy-negative pSS was indistinguishable from non-SS sicca patients. Different patterns of SG immunopathway activation in pSS argue for personalized treatment approaches.


Assuntos
Glândulas Salivares Menores/metabolismo , Glândulas Salivares/metabolismo , Síndrome de Sjogren/etiologia , Transcriptoma , Autoimunidade , Linfócitos B/imunologia , Linfócitos B/metabolismo , Biomarcadores , Biópsia , Suscetibilidade a Doenças , Feminino , Perfilação da Expressão Gênica/métodos , Regulação da Expressão Gênica , Humanos , Ativação Linfocitária/genética , Ativação Linfocitária/imunologia , Masculino , Modelos Biológicos , Glândulas Salivares/patologia , Glândulas Salivares Menores/patologia , Síndrome de Sjogren/diagnóstico , Síndrome de Sjogren/metabolismo
11.
Bioanalysis ; 12(18): 1311-1324, 2020 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-32945691

RESUMO

Background: S1PR1, a G protein-coupled receptor (GPCR) protein, is a therapeutic target for treatment of autoimmune diseases. As a potential biomarker for drug effect and patient stratification, it is of great significance to measure it in biological samples. However, due to the hydrophobic nature of S1PR1 and the difficulties in extraction and solubilization, as well as low expression levels, quantitative determination of S1PR1 remains challenging. Results: In this work, a peptide immunoaffinity LC-MS/MS method was developed to quantify S1PR1 in biopsy-sized colon samples with an LLOQ of 7.81 pM. Conclusion: Peptide immunoaffinity LC-MS/MS based strategy has achieved the desired sensitivity for low abundance S1PR1, and the same strategy could be applied to quantify S1PR1 in multiple species and other GPCR proteins.


Assuntos
Cromatografia Líquida/métodos , Colo/imunologia , Peptídeos/química , Receptores de Esfingosina-1-Fosfato/imunologia , Espectrometria de Massas em Tandem/métodos , Biópsia , Humanos
12.
Science ; 369(6501): 320-325, 2020 07 17.
Artigo em Inglês | MEDLINE | ID: mdl-32675374

RESUMO

Restricted V(D)J recombination during fetal development was postulated to limit antibody repertoire breadth and prevent autoimmunity. However, newborn serum contains abundant autoantibodies, suggesting that B cell tolerance during gestation is not yet fully established. To investigate this apparent paradox, we evaluated the reactivities of more than 450 antibodies cloned from single B cells from human fetal liver, bone marrow, and spleen. We found that incomplete B cell tolerance in early human fetal life favored the accumulation of polyreactive B cells that bound both apoptotic cells and commensal bacteria from healthy adults. Thus, the restricted fetal preimmune repertoire contains potentially beneficial self-reactive innate-like B cell specificities that may facilitate the removal of apoptotic cells during development and shape gut microbiota assembly after birth.


Assuntos
Anticorpos/imunologia , Linfócitos B/imunologia , Feto/imunologia , Autoanticorpos/imunologia , Autoantígenos/imunologia , Autoimunidade , Bactérias/imunologia , Feminino , Humanos , Imunidade Inata , Especificidade de Órgãos , Gravidez , Recombinação V(D)J
13.
Front Immunol ; 9: 2728, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30534127

RESUMO

Checkpoint inhibitors target the inhibitory receptors expressed by tumor-infiltrating T cells in order to reinvigorate an anti-tumor immune response. Therefore, understanding T cell composition and phenotype in human tumors is crucial. We analyzed by flow cytometry tumor-infiltrating lymphocytes (TILs) from two independent cohorts of patients with different cancer types, including RCC, lung, and colon cancer. In healthy donors, peripheral T cells are usually either CD4+ or CD8+ with a small percentage of CD4+ CD8+ DP cells (<5%). Compared to several other cancer types, including lung, and colorectal cancers, TILs from about a third of RCC patients showed an increased proportion of DP CD4+CD8+ T cells (>5%, reaching 30-50% of T cells in some patients). These DP T cells have an effector memory phenotype and express CD38, 4-1BB, and HLA-DR, suggesting antigen-driven expansion. In fact, TCR sequencing analysis revealed a high degree of clonality in DP T cells. Additionally, there were high levels of PD-1 and TIM-3 expression on DP T cells, which correlated with higher expression of PD-1 and TIM-3 in conventional single positive CD8 T cells from the same patients. These results suggest that DP T cells could be dysfunctional tumor-specific T cells with the potential to be reactivated by checkpoint inhibitors.


Assuntos
Antígenos de Diferenciação/imunologia , Antígenos CD4/imunologia , Antígenos CD8/imunologia , Carcinoma de Células Renais/imunologia , Neoplasias Renais/imunologia , Linfócitos do Interstício Tumoral/imunologia , Proteínas de Neoplasias/imunologia , Linfócitos T/imunologia , Idoso , Idoso de 80 Anos ou mais , Carcinoma de Células Renais/patologia , Feminino , Regulação Neoplásica da Expressão Gênica/imunologia , Humanos , Neoplasias Renais/patologia , Linfócitos do Interstício Tumoral/patologia , Masculino , Pessoa de Meia-Idade , Linfócitos T/patologia
14.
Front Immunol ; 9: 1613, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30123214

RESUMO

Agents targeting the PD1-PDL1 axis have transformed cancer therapy. Factors that influence clinical response to PD1-PDL1 inhibitors include tumor mutational burden, immune infiltration of the tumor, and local PDL1 expression. To identify peripheral correlates of the anti-tumor immune response in the absence of checkpoint blockade, we performed a retrospective study of circulating T cell subpopulations and matched tumor gene expression in melanoma and non-small cell lung cancer (NSCLC) patients. Notably, both melanoma and NSCLC patients whose tumors exhibited increased inflammatory gene transcripts presented high CD4+ and CD8+ central memory T cell (CM) to effector T cell (Eff) ratios in blood. Consequently, we evaluated CM/Eff T cell ratios in a second cohort of NSCLC. The data showed that high CM/Eff T cell ratios correlated with increased tumor PDL1 expression. Furthermore, of the 22 patients within this NSCLC cohort who received nivolumab, those with high CM/Eff T cell ratios, had longer progression-free survival (PFS) (median survival: 91 vs. 215 days). These findings show that by providing a window into the state of the immune system, peripheral T cell subpopulations inform about the state of the anti-tumor immune response and identify potential blood biomarkers of clinical response to checkpoint inhibitors in melanoma and NSCLC.


Assuntos
Carcinoma Pulmonar de Células não Pequenas/imunologia , Neoplasias Pulmonares/imunologia , Melanoma/imunologia , Receptor de Morte Celular Programada 1/antagonistas & inibidores , Subpopulações de Linfócitos T/imunologia , Idoso , Antineoplásicos Imunológicos/uso terapêutico , Antígeno B7-H1/imunologia , Antígeno B7-H1/metabolismo , Biomarcadores Tumorais , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Humanos , Neoplasias Pulmonares/tratamento farmacológico , Masculino , Pessoa de Meia-Idade , Nivolumabe/uso terapêutico , Receptor de Morte Celular Programada 1/imunologia , Receptor de Morte Celular Programada 1/metabolismo , Intervalo Livre de Progressão , Subpopulações de Linfócitos T/metabolismo
15.
Sci Immunol ; 1(1)2016.
Artigo em Inglês | MEDLINE | ID: mdl-27917411

RESUMO

The 1858T protein tyrosine phosphatase nonreceptor type 22 (PTPN22 T) allele is one of the main risk factors associated with many autoimmune diseases and correlates with a defective removal of developing autoreactive B cells in humans. To determine whether inhibiting PTPN22 favors the elimination of autoreactive B cells, we first demonstrated that the PTPN22 T allele interfered with the establishment of central B cell tolerance using NOD-scid-common γ chain knockout (NSG) mice engrafted with human hematopoietic stem cells expressing this allele. In contrast, the inhibition of either PTPN22 enzymatic activity or its expression by RNA interference restored defective central B cell tolerance in this model. Thus, PTPN22 blockade may represent a therapeutic strategy for the prevention or treatment of autoimmunity.

16.
JCI Insight ; 1(9): e87310, 2016 06 16.
Artigo em Inglês | MEDLINE | ID: mdl-27699274

RESUMO

Systemic lupus erythematosus (SLE) is a complex systemic autoimmune disease driven by both innate and adaptive immune cells. African Americans tend to present with more severe disease at an earlier age compared with patients of European ancestry. In order to better understand the immunological differences between African American and European American patients, we analyzed the frequencies of B cell subsets and the expression of B cell activation markers from a total of 68 SLE patients and 69 normal healthy volunteers. We found that B cells expressing the activation markers CD86, CD80, PD1, and CD40L, as well as CD19+CD27-IgD- double-negative B cells, were enriched in African American patients vs. patients of European ancestry. In addition to increased expression of CD40L, surface levels of CD40 on B cells were lower, suggesting the engagement of the CD40 pathway. In vitro experiments confirmed that CD40L expressed by B cells could lead to CD40 activation and internalization on adjacent B cells. To conclude, these results indicate that, compared with European American patients, African American SLE patients present with a particularly active B cell component, possibly via the activation of the CD40/CD40L pathway. These data may help guide the development of novel therapies.


Assuntos
Linfócitos B/citologia , Lúpus Eritematoso Sistêmico/etnologia , Negro ou Afro-Americano , Antígenos de Superfície/análise , Antígeno B7-2/análise , Antígenos CD40/análise , Ligante de CD40/análise , Humanos , Fenótipo
17.
J Clin Invest ; 126(11): 4289-4302, 2016 11 01.
Artigo em Inglês | MEDLINE | ID: mdl-27701145

RESUMO

Patients with mutations in AICDA, which encodes activation-induced cytidine deaminase (AID), display an impaired peripheral B cell tolerance. AID mediates class-switch recombination (CSR) and somatic hypermutation (SHM) in B cells, but the mechanism by which AID prevents the accumulation of autoreactive B cells in blood is unclear. Here, we analyzed B cell tolerance in AID-deficient patients, patients with autosomal dominant AID mutations (AD-AID), asymptomatic AICDA heterozygotes (AID+/-), and patients with uracil N-glycosylase (UNG) deficiency, which impairs CSR but not SHM. The low frequency of autoreactive mature naive B cells in UNG-deficient patients resembled that of healthy subjects, revealing that impaired CSR does not interfere with the peripheral B cell tolerance checkpoint. In contrast, we observed decreased frequencies of SHM in memory B cells from AD-AID patients and AID+/- subjects, who were unable to prevent the accumulation of autoreactive mature naive B cells. In addition, the individuals with AICDA mutations, but not UNG-deficient patients, displayed Tregs with defective suppressive capacity that correlated with increases in circulating T follicular helper cells and enhanced cytokine production. We conclude that SHM, but not CSR, regulates peripheral B cell tolerance through the production of mutated antibodies that clear antigens and prevent sustained interleukin secretions that interfere with Treg function.


Assuntos
Linfócitos B/imunologia , Pontos de Checagem do Ciclo Celular/imunologia , Citidina Desaminase/deficiência , Tolerância Imunológica , Memória Imunológica , Mutação , Hipermutação Somática de Imunoglobulina/imunologia , Linfócitos B/patologia , Pontos de Checagem do Ciclo Celular/genética , Citidina Desaminase/imunologia , Feminino , Humanos , Masculino , Linfócitos T Reguladores/imunologia , Linfócitos T Reguladores/patologia
18.
J Clin Invest ; 123(10): 4283-93, 2013 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-24051380

RESUMO

Common variable immune deficiency (CVID) is an assorted group of primary diseases that clinically manifest with antibody deficiency, infection susceptibility, and autoimmunity. Heterozygous mutations in the gene encoding the tumor necrosis factor receptor superfamily member TACI are associated with CVID and autoimmune manifestations, whereas two mutated alleles prevent autoimmunity. To assess how the number of TACI mutations affects B cell activation and tolerance checkpoints, we analyzed healthy individuals and CVID patients carrying one or two TACI mutations. We found that TACI interacts with the cleaved, mature forms of TLR7 and TLR9 and plays an important role during B cell activation and the central removal of autoreactive B cells in healthy donors and CVID patients. However, only subjects with a single TACI mutation displayed a breached immune tolerance and secreted antinuclear antibodies (ANAs). These antibodies were associated with the presence of circulating B cell lymphoma 6-expressing T follicular helper (Tfh) cells, likely stimulating autoreactive B cells. Thus, TACI mutations may favor CVID by altering B cell activation with coincident impairment of central B cell tolerance, whereas residual B cell responsiveness in patients with one, but not two, TACI mutations enables autoimmune complications.


Assuntos
Linfócitos B/imunologia , Imunodeficiência de Variável Comum/genética , Ativação Linfocitária , Proteína Transmembrana Ativadora e Interagente do CAML/genética , Adulto , Anticorpos Antinucleares/metabolismo , Autoanticorpos/metabolismo , Fator Ativador de Células B/sangue , Linfócitos B/metabolismo , Estudos de Casos e Controles , Proliferação de Células , Células Cultivadas , Tolerância Central , Imunodeficiência de Variável Comum/sangue , Imunodeficiência de Variável Comum/imunologia , Proteínas de Ligação a DNA/metabolismo , Dosagem de Genes , Heterozigoto , Humanos , Masculino , Pessoa de Meia-Idade , Mutação , Tolerância Periférica , Proteínas Proto-Oncogênicas c-bcl-6 , Linfócitos T Auxiliares-Indutores/imunologia , Linfócitos T Auxiliares-Indutores/metabolismo , Receptor 7 Toll-Like/metabolismo , Receptor Toll-Like 9/metabolismo
19.
J Clin Invest ; 121(9): 3635-44, 2011 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-21804190

RESUMO

Protein tyrosine phosphatase nonreceptor type 22 (PTPN22) gene polymorphisms are associated with many autoimmune diseases. The major risk allele encodes an R620W amino acid change that alters B cell receptor (BCR) signaling involved in the regulation of central B cell tolerance. To assess whether this PTPN22 risk allele affects the removal of developing autoreactive B cells, we tested by ELISA the reactivity of recombinant antibodies isolated from single B cells from asymptomatic healthy individuals carrying one or two PTPN22 risk allele(s) encoding the PTPN22 R620W variant. We found that new emigrant/transitional and mature naive B cells from carriers of this PTPN22 risk allele contained high frequencies of autoreactive clones compared with those from non-carriers, revealing defective central and peripheral B cell tolerance checkpoints. Hence, a single PTPN22 risk allele has a dominant effect on altering autoreactive B cell counterselection before any onset of autoimmunity. In addition, gene array experiments analyzing mature naive B cells displaying PTPN22 risk allele(s) revealed that the association strength of PTPN22 for autoimmunity may be due not only to the impaired removal of autoreactive B cells but also to the upregulation of genes such as CD40, TRAF1, and IRF5, which encode proteins that promote B cell activation and have been identified as susceptibility genes associated with autoimmune diseases. These data demonstrate that early B cell tolerance defects in autoimmunity can result from specific polymorphisms and precede the onset of disease.


Assuntos
Alelos , Autoimunidade/imunologia , Linfócitos B/imunologia , Isoenzimas/genética , Proteína Tirosina Fosfatase não Receptora Tipo 22/genética , Autoanticorpos/imunologia , Doenças Autoimunes/genética , Doenças Autoimunes/imunologia , Linfócitos B/citologia , Predisposição Genética para Doença , Humanos , Isoenzimas/metabolismo , Polimorfismo de Nucleotídeo Único , Proteína Tirosina Fosfatase não Receptora Tipo 22/metabolismo , Fatores de Risco , Tolerância a Antígenos Próprios/genética , Tolerância a Antígenos Próprios/imunologia
20.
J Immunol ; 179(7): 4857-66, 2007 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-17878385

RESUMO

Aside from being the precursors of the Ab-secreting cells, B cells are engaged in other immune functions such as Ag presentation to T cells or cytokine production. These functions may contribute to the pathogenic role of B cells in a wide range of autoimmune diseases. We demonstrate that B cells acquire the capacity to amplify IFN-gamma production by CD4 and CD8 T cells during the course of the Th1 inflammatory response to Toxoplasma gondii infection. Using the two following different strategies, we observed that B cells from T. gondii-infected mice, but not from naive mice, induce higher IFN-gamma expression by splenic host T cells: 1) reconstitution of B cell-deficient mice with B cells expressing an alloantigen different from the recipients, and 2) adoptive transfer of B and T cells into RAG-/- mice. In vitro assays allowing the physical separation of T and B cells demonstrate that Ag-primed B cells enhance IFN-gamma production by T cells in a contact-dependent fashion. Using an OVA-transgenic strain of T. gondii and OVA-specific CD4 T cells, we observed that the proinflammatory effect of B cells is neither Ag specific nor requires MHCII expression. However, TNF-alpha expressed on the surface of B cells appears to mediate in part the up-regulation of IFN-gamma by the effector T cells.


Assuntos
Linfócitos B/metabolismo , Interferon gama/biossíntese , Linfócitos T/metabolismo , Fator de Necrose Tumoral alfa/metabolismo , Animais , Apresentação de Antígeno/imunologia , Linfócitos B/imunologia , Quimiocina CCL3 , Quimiocina CCL4 , Quimiocinas CC/metabolismo , Feminino , Antígenos de Histocompatibilidade Classe II/imunologia , Inflamação/imunologia , Ativação Linfocitária/imunologia , Proteínas Inflamatórias de Macrófagos/metabolismo , Camundongos , Camundongos Knockout , Baço/imunologia , Baço/metabolismo , Linfócitos T/imunologia , Fator de Necrose Tumoral alfa/deficiência , Fator de Necrose Tumoral alfa/genética
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