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1.
J Immunol ; 194(4): 1514-22, 2015 Feb 15.
Artigo em Inglês | MEDLINE | ID: mdl-25582858

RESUMO

A human La/Sjögren's syndrome-B (hLa)-specific TCR/hLa neo-self-Ag double-transgenic (Tg) mouse model was developed and used to investigate cellular tolerance and autoimmunity to the ubiquitous RNA-binding La Ag often targeted in systemic lupus erythematosus and Sjögren's syndrome. Extensive thymic clonal deletion of CD4(+) T cells occurred in H-2(k/k) double-Tg mice presenting high levels of the I-E(k)-restricted hLa T cell epitope. In contrast, deletion was less extensive in H-2(k/b) double-Tg mice presenting lower levels of the epitope, and some surviving thymocytes were positively selected as thymic regulatory T cells (tTreg). These mice remained serologically tolerant to hLa and healthy. H-2(k/b) double-Tg mice deficient of all endogenous Tcra genes, a deficiency known to impair Treg development and function, produced IgG anti-hLa autoantibodies and displayed defective tTreg development. These autoimmune mice had interstitial lung disease characterized by lymphocytic aggregates containing Tg T cells with an activated, effector memory phenotype. Salivary gland infiltrates were notably absent. Thus, expression of nuclear hLa Ag induces thymic clonal deletion and tTreg selection, and lymphocytic infiltration of the lung is a consequence of La-specific CD4(+) T cell autoimmunity.


Assuntos
Autoantígenos/imunologia , Autoimunidade/imunologia , Doenças Pulmonares Intersticiais/imunologia , Ribonucleoproteínas/imunologia , Linfócitos T Reguladores/imunologia , Animais , Apresentação de Antígeno/imunologia , Autoanticorpos/imunologia , Ensaio de Imunoadsorção Enzimática , Epitopos de Linfócito T/imunologia , Citometria de Fluxo , Genes Codificadores da Cadeia alfa de Receptores de Linfócitos T/imunologia , Humanos , Tolerância Imunológica/imunologia , Imuno-Histoquímica , Ativação Linfocitária/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Reação em Cadeia da Polimerase , Receptores de Antígenos de Linfócitos T , Síndrome de Sjogren/complicações , Síndrome de Sjogren/imunologia , Timo/citologia , Timo/imunologia , Antígeno SS-B
2.
J Immunol ; 184(6): 3063-71, 2010 Mar 15.
Artigo em Inglês | MEDLINE | ID: mdl-20139278

RESUMO

Recombination of germline TCR alpha and beta genes generates polypeptide receptors for MHC peptide. Ag exposure during long-term herpes simplex infections may shape the T cell repertoire over time. We investigated the CD8 T cell response to HSV-2 in chronically infected individuals by sequencing the hypervariable regions encoding TCR alpha and beta polypeptides from T cell clones recognizing virion protein 22 aa 49-57, an immunodominant epitope. The most commonly detected TCRBV gene segment, found in four of five subjects and in 12 of 50 independently derived T cell clones, was TCRBV12-4. Nineteen to seventy-two percent of tetramer-binding cells in PBMCs were stained ex vivo with a TCRBV12 mAb. Three alpha-chain and three beta-chain public TCR sequences were shared between individuals. Public heterodimers were also detected. Promiscuous pairing of a specific TCRVA1-1 sequence with several different TCRB polypeptides was observed, implying a dominant structural role for the TCRA chain for these clonotypes. Functional avidity for cytotoxicity and IFN-gamma release was relatively invariant, except for one subject with both high avidity and unique TCR sequences and lower HSV-2 shedding. These data indicate that the CD8 response to a dominant alpha-herpesvirus epitope converges on preferred TCR sequences with relatively constant functional avidity.


Assuntos
Epitopos de Linfócito T/imunologia , Genes Codificadores da Cadeia alfa de Receptores de Linfócitos T/imunologia , Genes Codificadores da Cadeia beta de Receptores de Linfócitos T/imunologia , Herpesvirus Humano 2/imunologia , Linfócitos T Citotóxicos/imunologia , Linfócitos T Citotóxicos/virologia , Sequência de Aminoácidos , Células Apresentadoras de Antígenos/imunologia , Células Apresentadoras de Antígenos/metabolismo , Células Apresentadoras de Antígenos/virologia , Linfócitos T CD8-Positivos/imunologia , Linfócitos T CD8-Positivos/metabolismo , Linfócitos T CD8-Positivos/virologia , Linhagem Celular Transformada , Células Clonais , Testes Imunológicos de Citotoxicidade/métodos , Epitopos de Linfócito T/biossíntese , Epitopos de Linfócito T/genética , Antígenos HLA-B/biossíntese , Antígenos HLA-B/genética , Antígenos HLA-B/imunologia , Antígeno HLA-B7 , Humanos , Região Variável de Imunoglobulina/biossíntese , Região Variável de Imunoglobulina/genética , Interferon gama/metabolismo , Dados de Sequência Molecular , Linfócitos T Citotóxicos/metabolismo , Latência Viral/imunologia
3.
Am J Transplant ; 9(4): 709-18, 2009 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-19344462

RESUMO

The relative contributions of the direct and indirect pathways in alloimmune responses have not been fully elucidated. We report a novel murine TCR transgenic system that can simultaneously track the CD4-direct (CD4-d), CD4-indirect (CD4-i) and CD8-direct (CD8-d) pathways after transplantation. Using this system, we have observed a profoundly greater proliferation of CD4-i T cells relative to CD4-d and CD8-d T cells after transplantation. Furthermore, a much larger proportion of CD4-i T cells attain an effector phenotype. We also analyzed endogenous, wild-type T cells using enzyme-linked immunospot analysis. In naïve mice, T cells with indirect reactivity were undetectable, but T cells with direct reactivity were abundant. However, 10 days after skin or heterotopic heart transplantation, CD4-i T cells comprised approximately 10% of the CD4+ response. Consistent with increased priming of the CD4-i pathway, we observed that the CD4-i T cells were further enriched in the effector cells migrating to the allograft and in memory-like T cells persisting after rejection. Thus, priming of the CD4-i pathway is favored after transplantation, allowing a rare population to rapidly become a major component of the CD4+ T-cell response in acute allograft rejection. The generalizability of this observation to other models remains to be determined.


Assuntos
Linfócitos T CD4-Positivos/imunologia , Transplante de Coração/imunologia , Transplante de Pele/imunologia , Linfócitos T/imunologia , Transplante Homólogo/imunologia , Transferência Adotiva , Animais , Linfócitos T CD8-Positivos/imunologia , Ensaio de Imunoadsorção Enzimática , Citometria de Fluxo , Genes Codificadores da Cadeia alfa de Receptores de Linfócitos T/imunologia , Memória Imunológica , Interferon gama/imunologia , Interleucina-2/imunologia , Linfócitos/imunologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Antígenos Thy-1/imunologia
4.
Int Immunol ; 21(2): 179-85, 2009 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-19106232

RESUMO

Cells bearing invariant Valpha19-Jalpha33 TCR alpha chains are believed to participate in the regulation of inflammatory autoimmune diseases. In this study, the potential to produce immunoregulatory cytokines by these cells was characterized in order to find the mechanism underlying their immunoregulatory functions. Serum levels of IL-4, IL-10, transforming growth factor-beta, IFN-gamma and IL-17 increased in mice over-expressing an invariant Valpha19-Jalpha33 TCR alpha transgene (Valpha19 Tg) in response to anti-CD3 antibody injection. NK1.1(+) Valpha19 Tg(+), but not NK1.1(-) Valpha19 Tg(+) cells, promptly produced immunoregulatory IL-4, IFN-gamma and IL-17 upon invariant TCR engagement with immobilized anti-CD3 antibody in culture. The activation of Valpha19 Tg(+) cells then triggered the production of pro-inflammatory cytokines by bystander cells. Interestingly, the ratio of T(h)2 cytokines such as IL-4, IL-5 and IL-10, but not pro-inflammatory IL-17, to IFN-gamma was increased when the intensity of the stimulation to invariant TCR was attenuated. Collectively, these findings suggest that invariant Valpha19 TCR(+) cells have the potential to participate in the regulation of inflammatory autoimmunity by producing T(h)2-biased cytokines in certain circumstances.


Assuntos
Citocinas/metabolismo , Genes Codificadores da Cadeia alfa de Receptores de Linfócitos T/imunologia , Fígado/metabolismo , Receptores de Antígenos de Linfócitos T/metabolismo , Células Th2/metabolismo , Animais , Anticorpos Monoclonais , Antígenos Ly , Complexo CD3/imunologia , Células Cultivadas , Citocinas/genética , Regulação da Expressão Gênica/imunologia , Genes Codificadores da Cadeia alfa de Receptores de Linfócitos T/genética , Fígado/citologia , Ativação Linfocitária , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Subfamília B de Receptores Semelhantes a Lectina de Células NK , Ligação Proteica/imunologia , Estabilidade Proteica , Receptores de Antígenos de Linfócitos T/genética , Receptores de Antígenos de Linfócitos T alfa-beta/genética , Receptores de Antígenos de Linfócitos T alfa-beta/imunologia , Transdução de Sinais/imunologia , Especificidade da Espécie , Células Th2/citologia , Células Th2/imunologia
5.
Int Immunol ; 20(12): 1517-25, 2008 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-18927318

RESUMO

The anti-tumor response of human invariant NKT (NKT) cells is well established. A novel T cell subset, mucosal-associated invariant T (MAIT) cells, possesses similar regulatory properties to NKT cells in autoimmune models and disease. Here, we examined the clonality of four T cell subsets expressing invariant alphaTCR, including Valpha7.2-Jalpha33 of MAIT cells, in 19 kidney and brain tumors. The MAIT clonotype was identified and co-expressed with NKT clonotype in half of the tumors. In contrast, two other invariant T cell clonotypes (Valpha4 and Valpha19) were not present in tumors. Such tumors also expressed Vbeta2 and Vbeta13, the restricted TCRbeta chain of MAIT cells and the antigen-presenting molecule MR1. A high percentage of infiltrating T cells was CD8+ and expressed HLA-DR suggesting activation. Although the MAIT alphaTCR was identified in both peripheral CD56+ and CD56- subsets, infiltrating lymphocytes were CD56 negative. The clonal presence of MAIT cells in tumors correlated with the expression of pro-inflammatory cytokines but no IL-4, IL-5 and IL-10, suggesting that a pro-inflammatory subset of human MAIT cells may exist. Our data imply that a CD56- subset of MAIT cells may participate in tumor immune responses similarly to NKT cells.


Assuntos
Neoplasias Encefálicas/metabolismo , Carcinoma de Células Renais/metabolismo , Glioblastoma/metabolismo , Neoplasias Renais/metabolismo , Linfócitos do Interstício Tumoral/metabolismo , Neoplasias Meníngeas/metabolismo , Meningioma/metabolismo , Células T Matadoras Naturais/metabolismo , Subpopulações de Linfócitos T/metabolismo , Neoplasias Encefálicas/genética , Neoplasias Encefálicas/patologia , Antígeno CD56/biossíntese , Antígeno CD56/genética , Carcinoma de Células Renais/genética , Carcinoma de Células Renais/patologia , Separação Celular , Citocinas/metabolismo , Citometria de Fluxo , Expressão Gênica/imunologia , Genes Codificadores da Cadeia alfa de Receptores de Linfócitos T/imunologia , Glioblastoma/genética , Glioblastoma/patologia , Antígenos HLA-DR/biossíntese , Antígenos HLA-DR/genética , Humanos , Imunidade nas Mucosas , Imuno-Histoquímica , Neoplasias Renais/genética , Neoplasias Renais/patologia , Ativação Linfocitária , Neoplasias Meníngeas/genética , Neoplasias Meníngeas/patologia , Meningioma/genética , Meningioma/patologia , Polimorfismo Conformacional de Fita Simples
6.
Int Immunol ; 20(7): 869-79, 2008 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-18487227

RESUMO

CTLs are thought to be major effectors for clearing viruses in acute infections including hepatitis B virus (HBV). Persistent HBV infection is characterized by a lack of or a weak CTL response to HBV, which is thought to reflect tolerance to HBV antigens. In the present study, we found that alpha-galactosylceramide (alpha-GalCer), a ligand for Valpha14-positive NKT cells, strongly enhanced the induction and proliferation of HBV-specific CTLs by HBsAg. In HBsAg transgenic mice, which are thought to be tolerant to HBV-encoded antigens, administration of HBsAg or alpha-GalCer alone failed to induce HBsAg-specific CTLs, but they were induced by co-administration of both compounds. Furthermore, by limiting dilution analysis, we confirmed the existence of HBsAg-specific CTL precursors in the HBsAg transgenic mice immunized with HBsAg and alpha-GalCer. A blocking experiment using antibodies to cytokines and CD40 ligand showed that IL-2 and CD40-CD40L interaction mediate the enhancement of CTL induction caused by alpha-GalCer through NKT cell activation. Our results may open up a new method for clearing the virus from patients with persistent HBV infection.


Assuntos
Genes Codificadores da Cadeia alfa de Receptores de Linfócitos T/imunologia , Vírus da Hepatite B/imunologia , Hepatite B/imunologia , Tolerância Imunológica , Células T Matadoras Naturais/imunologia , Animais , Antígenos Virais/administração & dosagem , Antígenos Virais/imunologia , Antígenos Virais/metabolismo , Linhagem Celular Tumoral , Células Cultivadas , Galactosilceramidas/administração & dosagem , Galactosilceramidas/imunologia , Galactosilceramidas/metabolismo , Tolerância Imunológica/imunologia , Ativação Linfocitária , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Knockout , Camundongos Transgênicos , Especificidade do Receptor de Antígeno de Linfócitos T , Subpopulações de Linfócitos T/metabolismo , Linfócitos T Citotóxicos/imunologia
7.
J Immunol ; 179(1): 449-54, 2007 Jul 01.
Artigo em Inglês | MEDLINE | ID: mdl-17579065

RESUMO

The TCR delta- and alpha-chain genes lie in a single complex locus, the TCRalpha/delta locus. TCRdelta-chain genes are assembled in CD4(-)CD8(-) (double negative (DN)) thymocytes and TCRalpha-chain genes are assembled in CD4(+)CD8(+) (double positive) thymocytes due, in part, to the developmental stage-specific activities of the TCRdelta and TCRalpha enhancers (Edelta and Ealpha), respectively. Edelta functions with TCRdelta promoters to mediate TCRdelta-chain gene assembly in DN thymocytes. However, Edelta is unable to function with TCRalpha promoters such as the TEA promoter to drive TCRalpha-chain gene assembly in these cells. This is important, because the premature assembly of TCRalpha-chain genes in DN thymocytes would disrupt alphabeta and gammadelta T cell development. The basis for TEA inactivity in DN thymocytes is unclear, because Edelta can activate the Vdelta5 gene segment promoter that lies only 4 kb upstream of TEA promoter. In this study, we use gene targeting to construct a modified TCRalpha/delta locus (TCRalpha/delta(5DeltaT)) in which the TEA promoter lies in the same location as the Vdelta5 gene segment on the wild-type TCRalpha/delta allele. Remarkably, the TEA promoter on this allele exhibits normal developmental stage-specific regulation, being active in double positive thymocytes but not in DN thymocytes as is the case with the Vdelta5 promoter. Thus, the inactivity of the TEA promoter in DN thymocytes is due primarily to intrinsic developmental stage-specific features of the promoter itself and not to its location relative to other cis-acting elements in the locus, such as Edelta.


Assuntos
Diferenciação Celular/genética , Diferenciação Celular/imunologia , Elementos Facilitadores Genéticos/imunologia , Rearranjo Gênico da Cadeia alfa dos Receptores de Antígenos dos Linfócitos T , Genes Codificadores da Cadeia alfa de Receptores de Linfócitos T/imunologia , Regiões Promotoras Genéticas/imunologia , Processamento de Proteína Pós-Traducional/imunologia , Receptores de Antígenos de Linfócitos T alfa-beta/genética , Animais , Rearranjo Gênico da Cadeia delta dos Receptores de Antígenos dos Linfócitos T , Marcação de Genes , Genes Codificadores da Cadeia delta de Receptores de Linfócitos T , Marcadores Genéticos/imunologia , Região de Junção de Imunoglobulinas/genética , Região de Junção de Imunoglobulinas/metabolismo , Região Variável de Imunoglobulina/genética , Região Variável de Imunoglobulina/metabolismo , Camundongos , Camundongos Knockout , Processamento de Proteína Pós-Traducional/genética , Receptores de Antígenos de Linfócitos T alfa-beta/biossíntese , Receptores de Antígenos de Linfócitos T alfa-beta/deficiência , Receptores de Antígenos de Linfócitos T gama-delta/biossíntese , Receptores de Antígenos de Linfócitos T gama-delta/genética , Receptores de Antígenos de Linfócitos T gama-delta/metabolismo , Subpopulações de Linfócitos T/citologia , Subpopulações de Linfócitos T/imunologia , Subpopulações de Linfócitos T/metabolismo , Transcrição Gênica/imunologia
8.
Nat Immunol ; 8(4): 378-87, 2007 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-17334367

RESUMO

Reversible contraction of immunoglobulin loci juxtaposes the variable (V) genes next to the (diversity)-joining-constant ((D)JC) gene domain, thus facilitating V-(D)J recombination. Here we show that the T cell receptor beta (Tcrb) and T cell receptor alphadelta (Tcra-Tcrd) loci also underwent long-range interactions by looping in double-negative and double-positive thymocytes, respectively. Contraction of the Tcrb and Tcra loci occurred in rearranging thymocytes and was reversed at the next developmental stage. Decontraction of the Tcrb locus probably prevented further V(beta)-DJ(beta) rearrangements in double-positive thymocytes by separating the V(beta) genes from the DJC(beta) domain. In most double-negative cells, one Tcrb allele was recruited to pericentromeric heterochromatin. Such allelic positioning may facilitate asynchronous V(beta)-DJ(beta) recombination. Hence, pericentromeric recruitment and locus 'decontraction' seem to contribute to the initiation and maintenance of allelic exclusion at the Tcrb locus.


Assuntos
Rearranjo Gênico do Linfócito T/imunologia , Genes Codificadores da Cadeia alfa de Receptores de Linfócitos T/imunologia , Genes Codificadores da Cadeia beta de Receptores de Linfócitos T/imunologia , Genes Codificadores da Cadeia delta de Receptores de Linfócitos T/imunologia , Linfócitos T/imunologia , Animais , Diferenciação Celular , Proteínas de Ligação a DNA/imunologia , Regiões Constantes de Imunoglobulina/imunologia , Região Variável de Imunoglobulina/imunologia , Hibridização in Situ Fluorescente , Linfopoese/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Camundongos Transgênicos , Microscopia Confocal , Fator de Transcrição PAX5/imunologia
9.
Int Immunol ; 18(6): 911-20, 2006 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-16641111

RESUMO

Expression and signalling through the pre-TCR and the TCRalphabeta resemble two critical checkpoints during T cell development. We investigated to which extent a pre-TCR can functionally replace mature TCRalpha chains during T cell development. For this purpose, transgenic mice were generated expressing the pre-TCRalpha (pTalpha) under the transcriptional control of TCRbeta regulatory elements. We report here on the interesting finding that constitutive pTalpha expression allows complete T cell maturation. The pre-TCR complex permits a subset of beta-selected thymocytes to mature in the absence of TCRalpha into peripheral T cells (betaT cells) comprising up to 10% of all lymphocytes. Lymphopenia-driven proliferation of these betaT cells is similar to that of conventional alphabetaT cells. Furthermore, betaT cells proliferated and acquired effector function upon stimulation with allogeneic MHC.


Assuntos
Diferenciação Celular/imunologia , Regulação da Expressão Gênica/imunologia , Genes Codificadores da Cadeia alfa de Receptores de Linfócitos T/imunologia , Genes Codificadores da Cadeia beta de Receptores de Linfócitos T/imunologia , Receptores de Antígenos de Linfócitos T alfa-beta/imunologia , Linfócitos T/imunologia , Animais , Diferenciação Celular/genética , Proliferação de Células , Regulação da Expressão Gênica/genética , Genes Codificadores da Cadeia alfa de Receptores de Linfócitos T/genética , Genes Codificadores da Cadeia beta de Receptores de Linfócitos T/genética , Linfopenia/genética , Linfopenia/imunologia , Complexo Principal de Histocompatibilidade/genética , Complexo Principal de Histocompatibilidade/imunologia , Camundongos , Camundongos Knockout , Receptores de Antígenos de Linfócitos T alfa-beta/genética , Elementos de Resposta/genética , Elementos de Resposta/imunologia , Transdução de Sinais/genética , Transdução de Sinais/imunologia
10.
J Exp Med ; 203(3): 661-73, 2006 Mar 20.
Artigo em Inglês | MEDLINE | ID: mdl-16505140

RESUMO

Little is known regarding the basis for selection of the semi-invariant alphabeta T cell receptor (TCR) expressed by natural killer T (NKT) cells or how this mediates recognition of CD1d-glycolipid complexes. We have determined the structures of two human NKT TCRs that differ in their CDR3beta composition and length. Both TCRs contain a conserved, positively charged pocket at the ligand interface that is lined by residues from the invariant TCR alpha- and semi-invariant beta-chains. The cavity is centrally located and ideally suited to interact with the exposed glycosyl head group of glycolipid antigens. Sequences common to mouse and human invariant NKT TCRs reveal a contiguous conserved "hot spot" that provides a basis for the reactivity of NKT cells across species. Structural and functional data suggest that the CDR3beta loop provides a plasticity mechanism that accommodates recognition of a variety of glycolipid antigens presented by CD1d. We propose a model of NKT TCR-CD1d-glycolipid interaction in which the invariant CDR3alpha loop is predicted to play a major role in determining the inherent bias toward CD1d. The findings define a structural basis for the selection of the semi-invariant alphabeta TCR and the unique antigen specificity of NKT cells.


Assuntos
Apresentação de Antígeno/imunologia , Antígenos CD1/imunologia , Glicolipídeos/imunologia , Células Matadoras Naturais/imunologia , Receptores de Antígenos de Linfócitos T alfa-beta/imunologia , Linfócitos T/imunologia , Animais , Apresentação de Antígeno/genética , Antígenos CD1/genética , Genes Codificadores da Cadeia alfa de Receptores de Linfócitos T/genética , Genes Codificadores da Cadeia alfa de Receptores de Linfócitos T/imunologia , Genes Codificadores da Cadeia beta de Receptores de Linfócitos T/genética , Genes Codificadores da Cadeia beta de Receptores de Linfócitos T/imunologia , Humanos , Camundongos , Ligação Proteica/genética , Ligação Proteica/imunologia , Estrutura Quaternária de Proteína , Estrutura Terciária de Proteína/fisiologia , Especificidade da Espécie , Homologia Estrutural de Proteína , Relação Estrutura-Atividade
11.
Mol Immunol ; 43(3): 246-54, 2006 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-16199261

RESUMO

The T cell receptor alpha chain repertoire and the possible influence of the alpha chain on the development and phenotype of naturally arising mouse CD4+CD25+ T cells have not been extensively analysed. We used all available Valpha-specific monoclonal antibodies and a sensitive multiplex genomic DNA PCR assay to study the Valpha repertoire of CD4+CD25+ T cells in normal mice. To address whether CD4+CD25+ T cells express two TCR alpha chains, we have carried out four-colour flow cytometry using combinations of the available anti-Valpha reagents in mice where one allele of the TCRA locus had been inactivated. Results indicate that the Valpha repertoire of CD4+CD25+ T cells is as diverse as their CD25- partners. In addition, CD4+CD25+ T cells develop normally in Tcralpha+/- mice and we show for the first time that despite expressing only one TCRalpha chain, they retain their characteristic CD4(low), CD3(low), TCRbeta(low), CD5(high), CD45RB(low) and cytoplasmic CD152(high) phenotype.


Assuntos
Linfócitos T CD4-Positivos/imunologia , Diferenciação Celular/imunologia , Rearranjo Gênico da Cadeia alfa dos Receptores de Antígenos dos Linfócitos T/imunologia , Genes Codificadores da Cadeia alfa de Receptores de Linfócitos T/imunologia , Receptores de Interleucina-2/imunologia , Animais , Antígenos CD/imunologia , Citometria de Fluxo , Regulação da Expressão Gênica/imunologia , Camundongos , Camundongos Endogâmicos BALB C , Locos de Características Quantitativas/imunologia
12.
Transplantation ; 80(5): 643-9, 2005 Sep 15.
Artigo em Inglês | MEDLINE | ID: mdl-16177639

RESUMO

BACKGROUND: Ischemia reperfusion injury leading to acute renal failure (ARF) and delayed graft function is an important problem in organ transplantation. CD4+ T cells, essential for transplant rejection, may mediate ischemic ARF. We have demonstrated that the caspase-1 mediated production of IL-18 is pathogenic in ischemic ARF in mice. A potential source of IL-18 in ischemic ARF is the CD4+ T cell. We therefore examined the effect CD4+ T cell depletion on the development of ischemic ARF and the activation of IL-18. METHODS: Functional and histological correlates were examined in two groups of mice with ischemic ARF: 1) CD4 T-cell depleted with the antibody GK1.5, and 2) T-cell receptor alpha-chain deficient (TCRalpha -/-) mice. TCRalpha -/- mice lack the alpha chain of the T-cell receptor and therefore lack functional CD4+ and CD8+ T cells. RESULTS: Flow cytometry of lymph nodes and immunohistochemistry of kidneys demonstrated complete depletion of CD4+ T cells in mice with ischemic ARF treated with GK 1.5. CD4+ T-cell depletion did not confer functional (serum creatinine, BUN and FITC-labeled inulin clearance) or histological protection against ischemic ARF. Likewise, TCRalpha -/- mice were not protected against ischemic ARF. Renal caspase-1 activity and IL-18 protein were similar in CD4+ T-cell depleted and wild-type postischemic reperfusion. CONCLUSIONS: Ischemic ARF can occur in the absence of classical T-cell function. The evaluation of other inflammatory mediators (e.g., macrophages or NK cells) as a source of IL-18 and mediator of ischemic ARF warrants further investigation.


Assuntos
Injúria Renal Aguda/prevenção & controle , Linfócitos T CD4-Positivos/imunologia , Função Retardada do Enxerto/prevenção & controle , Transplante de Rim , Depleção Linfocítica , Injúria Renal Aguda/imunologia , Injúria Renal Aguda/terapia , Animais , Anticorpos Monoclonais/farmacologia , Linfócitos T CD4-Positivos/metabolismo , Caspase 1/metabolismo , Função Retardada do Enxerto/imunologia , Função Retardada do Enxerto/terapia , Genes Codificadores da Cadeia alfa de Receptores de Linfócitos T/imunologia , Interleucina-18/metabolismo , Isquemia/imunologia , Isquemia/prevenção & controle , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Mutantes
13.
Int Immunol ; 17(9): 1167-78, 2005 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-16091384

RESUMO

The suppressor of cytokine signaling (SOCS) 1 is a negative regulator in multiple cytokine-related aspects to maintain immunological homeostasis. Here, we studied a role of SOCS1 on dendritic cell (DC) maturation in the mice lacking either TCRalpha chain or CD28 in SOCS1-deficient background, and found that the SOCS1 could restore acute phase of inflammatory response in SOCS1-deficient mice. The CD11c+ CD8- DC population in freshly isolated splenic DCs from normal mice highly expressed SOCS1. However, in SOCS1-deficient environment, the proportion of CD8alpha+ DCs (CD8 DCs) noticeably increased without affecting the cell number of conventional and plasmacytoid DC populations. This population revealed the CD11cdull CD8alpha+ CD11b- CD45RA- B220- phenotype, which is a minor population in normal mice. Localization of the abnormal CD8 DCs in splenic microenvironments was mainly restricted to deep within red pulp. The CD8 DCs secrete a large amount of IFN-gamma, IL-12 and B lymphocyte stimulator/B cell activation factor of the tumor necrosis factor family in response to LPS and CpG stimulation. This is responsible for the development of DC-mediated systemic autoimmunity in the old age of SOCS1-deficient mice. Moreover, the CD8 DC subsets expressed more indoleamine 2,3-dioxygenase and IL-10, and hence inhibit the allogeneic proliferative T cell response and antigen-induced Th1 responses. Therefore, SOCS1 expression during DC maturation plays a role in surveillance in controlling the aberrant expansion of abnormal DC subset to maintain homeostasis of immune system.


Assuntos
Antígenos CD28/imunologia , Antígenos CD8/imunologia , Proteínas de Transporte/imunologia , Proliferação de Células , Células Dendríticas/imunologia , Genes Codificadores da Cadeia alfa de Receptores de Linfócitos T/imunologia , Proteínas Repressoras/imunologia , Proteínas Supressoras da Sinalização de Citocina/imunologia , Animais , Antígenos CD/imunologia , Antígenos CD28/genética , Diferenciação Celular/genética , Diferenciação Celular/imunologia , Citocinas/imunologia , Células Dendríticas/citologia , Genes Codificadores da Cadeia alfa de Receptores de Linfócitos T/genética , Camundongos , Camundongos Knockout , Plasmócitos/citologia , Plasmócitos/imunologia , Baço/citologia , Baço/imunologia , Proteína 1 Supressora da Sinalização de Citocina , Proteínas Supressoras da Sinalização de Citocina/deficiência , Células Th1/citologia , Células Th1/imunologia
14.
J Exp Med ; 202(4): 467-72, 2005 Aug 15.
Artigo em Inglês | MEDLINE | ID: mdl-16087716

RESUMO

Murine Tcrd and Tcra gene segments reside in a single genetic locus and undergo recombination in CD4- CD8- (double negative [DN]) and CD4+ CD8+ (double positive [DP]) thymocytes, respectively. TcraTcrd locus variable gene segments are subject to complex regulation. Only a small subset of approximately 100 variable gene segments contributes substantially to the adult TCRdelta repertoire. Moreover, although most contribute to the TCRalpha repertoire, variable gene segments that are Jalpha proximal are preferentially used during primary Tcra recombination. We investigate the role of local chromatin accessibility in determining the developmental pattern of TcraTcrd locus variable gene segment recombination. We find variable gene segments to be heterogeneous with respect to acetylation of histones H3 and H4. Those that dominate the adult TCRdelta repertoire are hyperacetylated in DN thymocytes, independent of their position in the locus. Moreover, proximal variable gene segments show dramatic increases in histone acetylation and germline transcription in DP thymocytes, a result of super long-distance regulation by the Tcra enhancer. Our results imply that differences in chromatin accessibility contribute to biases in TcraTcrd locus variable gene segment recombination in DN and DP thymocytes and extend the distance over which the Tcra enhancer can regulate chromatin structure to a remarkable 525 kb.


Assuntos
Montagem e Desmontagem da Cromatina/genética , Cromatina/genética , Rearranjo Gênico da Cadeia alfa dos Receptores de Antígenos dos Linfócitos T/genética , Rearranjo Gênico da Cadeia delta dos Receptores de Antígenos dos Linfócitos T/genética , Genes Codificadores da Cadeia alfa de Receptores de Linfócitos T/genética , Genes Codificadores da Cadeia delta de Receptores de Linfócitos T/genética , Acetilação , Animais , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD8-Positivos/imunologia , Cromatina/imunologia , Montagem e Desmontagem da Cromatina/imunologia , Elementos Facilitadores Genéticos/genética , Elementos Facilitadores Genéticos/imunologia , Regulação da Expressão Gênica/genética , Regulação da Expressão Gênica/imunologia , Rearranjo Gênico da Cadeia alfa dos Receptores de Antígenos dos Linfócitos T/imunologia , Rearranjo Gênico da Cadeia delta dos Receptores de Antígenos dos Linfócitos T/imunologia , Genes Codificadores da Cadeia alfa de Receptores de Linfócitos T/imunologia , Genes Codificadores da Cadeia delta de Receptores de Linfócitos T/imunologia , Histonas/imunologia , Camundongos , Camundongos Knockout , Locos de Características Quantitativas/genética , Locos de Características Quantitativas/imunologia , Recombinação Genética/genética , Recombinação Genética/imunologia , Timo/citologia , Timo/imunologia , Transcrição Gênica/genética , Transcrição Gênica/imunologia
15.
Immunol Res ; 30(2): 231-40, 2004.
Artigo em Inglês | MEDLINE | ID: mdl-15477663

RESUMO

Owing to ordered, stage-specific T-cell receptor (TCR) gene rearrangements and cell division during T-cell development, small cohorts of "half-sibling" T cells sharing an ancestral TCR VDJbeta rearrangement but expressing different TCR alpha-locus rearrangements may be selected into the mature T-cell repertoire. We wondered whether different alphabetaTCRs expressed by T cells from the same ancestral VDJbeta cohort might be capable of recognizing the same foreign peptide-major histocompatibility complex complex (pMHC). By a combined flow cytometric and single-cell polymerase chain reaction (PCR) approach to analyze TCRs selected by the previously defined foreign antigen, pCW3170-179/H-2Kd, we were able to identify cohorts of half-sibling antigen-specific CD8 T cells after their expansion in immunized mice. We amplified residual DJbeta rearrangements as clonal markers to confirm that the shared VDJbeta sequences represent ancestral rearrangements rather than identical but independent ones. An intriguing explanation of our findings would be that only a very limited repertoire of TCR alpha-chains is selected to pair with a given TCR beta-chain during T-cell development.


Assuntos
Linfócitos T CD8-Positivos/imunologia , Linhagem da Célula/genética , Rearranjo Gênico da Cadeia beta dos Receptores de Antígenos dos Linfócitos T/genética , Genes Codificadores da Cadeia beta de Receptores de Linfócitos T/genética , Antígenos HLA-C/imunologia , Peptídeos/imunologia , Animais , Linhagem da Célula/imunologia , Rearranjo Gênico da Cadeia alfa dos Receptores de Antígenos dos Linfócitos T/genética , Rearranjo Gênico da Cadeia alfa dos Receptores de Antígenos dos Linfócitos T/imunologia , Rearranjo Gênico da Cadeia beta dos Receptores de Antígenos dos Linfócitos T/imunologia , Genes Codificadores da Cadeia alfa de Receptores de Linfócitos T/genética , Genes Codificadores da Cadeia alfa de Receptores de Linfócitos T/imunologia , Genes Codificadores da Cadeia beta de Receptores de Linfócitos T/imunologia , Antígenos HLA-C/genética , Humanos , Ligantes , Camundongos , Camundongos Endogâmicos DBA
16.
Int Immunol ; 16(8): 1069-80, 2004 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-15197172

RESUMO

Early thymocyte development was compared in normal, recombinase-activating gene 2-inactivated (RAG-2 KO) and pre-T cell receptor alpha-inactivated (pre-Talpha KO) mice, mutants representing either a complete (RAG-2 KO) or partial (pre-Talpha KO) block in progenitor development. Using three colour analysis with antibodies to CD117, CD44 and CD25, cell numbers in each progenitor subset were quantified, demonstrating an accumulation of cells prior to the block. Progenitor number was influenced both by the nature of the genetic block and thymus size, as shown in the enlarged thymus of a transgenic mouse line. By four colour staining for CD3, CD117, CD44 and CD25 and deliberately not gating out CD3(-) cells, a novel aspect of gamma delta T cell development in pre-Talpha KO mice was identified. 5-bromodeoxyuridine labelling and subsequent four colour staining for BrdU, CD117, CD44 and CD25 showed firstly that DN1 cells were cycling, secondly that the developmental block in pre-Talpha KO mice corresponded to a decrease in DN4 cell proliferation, and thirdly provided a novel 'snapshot' of T cell receptor beta-selected cells transiting the DN3 to DN4 compartment. Taken together, these results emphasise the need for a more detailed qualitative and quantitative analysis of the progenitor compartment in the thymus.


Assuntos
Diferenciação Celular/imunologia , Proliferação de Células , Proteínas de Ligação a DNA/imunologia , Genes Codificadores da Cadeia alfa de Receptores de Linfócitos T/imunologia , Linfócitos T/imunologia , Timo/imunologia , Animais , Antígenos CD/imunologia , Diferenciação Celular/genética , Proteínas de Ligação a DNA/genética , Genes Codificadores da Cadeia alfa de Receptores de Linfócitos T/genética , Genes Codificadores da Cadeia beta de Receptores de Linfócitos T/genética , Genes Codificadores da Cadeia beta de Receptores de Linfócitos T/imunologia , Camundongos , Camundongos Knockout , Timo/citologia
17.
J Immunol ; 170(3): 1209-17, 2003 Feb 01.
Artigo em Inglês | MEDLINE | ID: mdl-12538678

RESUMO

The adoptive transfer of tumor-specific T cells expanded in vitro can be of significant therapeutic value in select cancer patients. This strategy is limited though, as it is often difficult, if not impossible, to obtain T cells of clinical value. The transfer of TCR genes to mature T cells to generate tumor-reactive T cells provides a potential mechanism to overcome these limitations. To evaluate the feasibility of such an approach and the quality of the resulting T cells, we generated replication-deficient retroviral vectors using the well-characterized OT-1 TCR genes. After transducing murine T cells, we were able to expand large numbers of Ag-specific T cells that were functionally active against tumor cells expressing the relevant Ag. Furthermore, we found that T cells expressing retrovirally encoded TCR had avidity that was similar to that of the parental clone. This maintenance of avidity was despite variable expression of the retrovirally encoded TCR and the presence of potentially competing endogenous TCRs. These results suggest that the inherent qualities of the TCR, as dictated by the coding sequence, are the most critical parameters in the generation of high-avidity T cells.


Assuntos
Citotoxicidade Imunológica/genética , Receptores de Antígenos de Linfócitos T alfa-beta/genética , Subpopulações de Linfócitos T/imunologia , Subpopulações de Linfócitos T/metabolismo , Transdução Genética/métodos , Animais , Diferenciação Celular/genética , Diferenciação Celular/imunologia , Células Cultivadas , Proteínas do Ovo/genética , Proteínas do Ovo/imunologia , Genes Codificadores da Cadeia alfa de Receptores de Linfócitos T/imunologia , Genes Codificadores da Cadeia beta de Receptores de Linfócitos T/imunologia , Vetores Genéticos/síntese química , Vetores Genéticos/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Vírus da Leucemia Murina de Moloney/genética , Vírus da Leucemia Murina de Moloney/imunologia , Ovalbumina/genética , Ovalbumina/imunologia , Fragmentos de Peptídeos , Receptores de Antígenos de Linfócitos T alfa-beta/biossíntese , Subpopulações de Linfócitos T/virologia , Células Tumorais Cultivadas
19.
J Immunol ; 168(7): 3259-65, 2002 Apr 01.
Artigo em Inglês | MEDLINE | ID: mdl-11907080

RESUMO

Immunization with superantigen in vivo induces transient activation of superantigen-specific T cells, followed by a superantigen-nonresponsive state. In this study, using a TCR alpha knock-in mouse in which the knock-in alpha-chain can be replaced with endogenous alpha-chain through secondary rearrangement, we show that immunization of superantigen changes the TCR alpha-chain expression on peripheral superantigen-specific T cells, induces expression of recombination-activating genes, and generates DNA double-strand breaks at the TCR alpha-chain locus. These results suggest that viral superantigens are capable of inducing peripheral TCR revision. Our findings thus provide a new perspective on pathogen-immune system interaction.


Assuntos
Rearranjo Gênico da Cadeia alfa dos Receptores de Antígenos dos Linfócitos T/imunologia , Genes Codificadores da Cadeia alfa de Receptores de Linfócitos T/imunologia , Tolerância Imunológica/genética , Ativação Linfocitária/genética , Superantígenos/imunologia , Linfócitos T/imunologia , Linfócitos T/metabolismo , Animais , Antígenos/farmacologia , Diferenciação Celular/genética , Diferenciação Celular/imunologia , Columbidae , Grupo dos Citocromos c/imunologia , Grupo dos Citocromos c/farmacologia , Regulação para Baixo/genética , Regulação para Baixo/imunologia , Imunização , Injeções Intravenosas , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos CBA , Camundongos Knockout , Camundongos Transgênicos , Receptores de Antígenos de Linfócitos T alfa-beta/antagonistas & inibidores , Receptores de Antígenos de Linfócitos T alfa-beta/biossíntese , Receptores de Antígenos de Linfócitos T alfa-beta/genética , Baço/citologia , Baço/imunologia , Superantígenos/administração & dosagem , Superantígenos/farmacologia , Linfócitos T/citologia
20.
J Immunol ; 166(3): 1755-62, 2001 Feb 01.
Artigo em Inglês | MEDLINE | ID: mdl-11160221

RESUMO

The NK1.1(+)TCRalphabeta(int) CD4(+), or double negative T cells (NK T cells) consist of a mixture of CD1d-restricted and CD1d-unrestricted cells. The relationships between CD4(+)NK1.1(+) T cells and conventional T cells are not understood. To compare their respective TCR repertoires, NK1.1(+)TCRalphabeta(int), CD4(+) T cells have been sorted out of the thymus, liver, spleen, and bone marrow of C57BL/6 mice. Molecular analysis showed that thymus and liver used predominantly the Valpha14-Jalpha281 and Vbeta 2, 7, and 8 segments. These cells are CD1d restricted and obey the original definition of NK T cells. The complementarity-determining region 3 (CDR3) sequences of the TCR Vbeta8.2-Jbeta2.5 chain of liver and thymus CD4(+) NK T cells were determined and compared with those of the same rearrangements of conventional CD4(+) T cells. No amino acid sequence or usage characteristic of NK T cells could be evidenced: the Vbeta8.2-Jbeta2.5 diversity regions being primarily the same in NK T and in T cells. No clonal expansion of the beta-chains was observed in thymus and liver CD1d-restricted CD4(+)NK T cells, suggesting the absence of acute or chronic Ag-driven stimulation. Molecular analysis of the TCR used by Valpha14-Jalpha281 transgenic mice on a Calpha(-/-) background showed that the alpha-chain can associate with beta-chains using any Vbeta segment, except in NK T cells in which it paired predominately with Vbeta 2, 7, and 8(+) beta-chains. The structure of the TCR of NK T cells thus reflects the affinity for the CD1d molecule rather than a structural constraint leading to the association of the invariant alpha-chain with a distinctive subset of Vbeta segment.


Assuntos
Antígenos CD4/biossíntese , Células Matadoras Naturais/metabolismo , Proteínas , Receptores de Antígenos de Linfócitos T alfa-beta/imunologia , Receptores de Antígenos de Linfócitos T alfa-beta/metabolismo , Subpopulações de Linfócitos T/metabolismo , Animais , Antígenos/biossíntese , Antígenos Ly , Antígenos de Superfície , Células da Medula Óssea/imunologia , Células da Medula Óssea/metabolismo , Separação Celular , Células Clonais , Regulação da Expressão Gênica/imunologia , Rearranjo Gênico da Cadeia beta dos Receptores de Antígenos dos Linfócitos T/imunologia , Genes Codificadores da Cadeia alfa de Receptores de Linfócitos T/imunologia , Células Matadoras Naturais/citologia , Células Matadoras Naturais/imunologia , Lectinas Tipo C , Fígado/citologia , Fígado/imunologia , Fígado/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Subfamília B de Receptores Semelhantes a Lectina de Células NK , Especificidade de Órgãos/imunologia , Biossíntese de Proteínas , Receptores de Antígenos de Linfócitos T alfa-beta/biossíntese , Receptores de Antígenos de Linfócitos T alfa-beta/genética , Baço/citologia , Baço/imunologia , Baço/metabolismo , Subpopulações de Linfócitos T/citologia , Subpopulações de Linfócitos T/imunologia , Timo/citologia , Timo/imunologia , Timo/metabolismo
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