RESUMO
Thymic regulatory T cells (tTregs) and induced regulatory T cells (iTregs) suppress murine acute graft-versus-host disease (GVHD). Previously, we demonstrated that the plasmacytoid dendritic cell indoleamine 2,3-dioxygenase (IDO) fosters the in vitro development of human iTregs via tryptophan depletion and kynurenine (Kyn) metabolites. We now show that stimulation of naïve CD4+ T cells in low tryptophan (low Trp) plus Kyn supports human iTreg generation. In vitro, low Trp + Kyn iTregs and tTregs potently suppress T effector cell proliferation equivalently but are phenotypically distinct. Compared with tTregs or T effector cells, bioenergetics profiling reveals that low Trp + Kyn iTregs have increased basal glycolysis and oxidative phosphorylation and use glutaminolysis as an energy source. Low Trp + Kyn iTreg viability was reliant on interleukin (IL)-2 in vitro. Although in vivo IL-2 administration increased low Trp + Kyn iTreg persistence on adoptive transfer into immunodeficient mice given peripheral blood mononuclear cells to induce GVHD, IL-2-supported iTregs did not improve recipient survival. We conclude that low Trp + Kyn create suppressive iTregs that have high metabolic needs that will need to be addressed before clinical translation.
Assuntos
Transplante de Medula Óssea , Doença Enxerto-Hospedeiro/imunologia , Tolerância Imunológica/imunologia , Cinurenina/metabolismo , Linfócitos T Reguladores/imunologia , Triptofano/metabolismo , Animais , Células Cultivadas , Doença Enxerto-Hospedeiro/metabolismo , Doença Enxerto-Hospedeiro/prevenção & controle , Humanos , Técnicas In Vitro , Camundongos , Taxa de SobrevidaRESUMO
Adoptive transfer of thymus-derived natural regulatory T cells (nTregs) effectively suppresses disease in murine models of autoimmunity and graft-versus-host disease (GVHD). TGFß induces Foxp3 expression and suppressive function in stimulated murine CD4+25- T cells, and these induced Treg (iTregs), like nTreg, suppress auto- and allo-reactivity in vivo. However, while TGFß induces Foxp3 expression in stimulated human T cells, the expanded cells lack suppressor cell function. Here we show that Rapamycin (Rapa) enhances TGFß-dependent Foxp3 expression and induces a potent suppressor function in naive (CD4+ 25-45RA+) T cells. Rapa/TGFß iTregs are anergic, express CD25 at levels higher than expanded nTregs and few cells secrete IL-2, IFNγ or IL-17 even after PMA and Ionomycin stimulation in vitro. Unlike other published methods of inducing Treg function, Rapa/TGFß induces suppressive function even in the presence of memory CD4+ T cells. A single apheresis unit of blood yields an average ~240 × 109 (range ~ 70-560 × 109) iTregs from CD4+25- T cells in ≤ 2 weeks of culture. Most importantly, Rapa/TGFß iTregs suppress disease in a xenogeneic model of GVHD. This study opens the door for iTreg cellular therapy for human diseases.
Assuntos
Doença Enxerto-Hospedeiro/prevenção & controle , Linfócitos T Reguladores/imunologia , Fator de Crescimento Transformador beta/fisiologia , Animais , Fatores de Transcrição Forkhead/metabolismo , Humanos , Camundongos , Camundongos Knockout , Sirolimo/farmacologiaRESUMO
Telomerase, a ribonucleoprotein that is capable of synthesizing telomeric repeats, is expressed in germline and malignant cells, and is absent in most normal human somatic cells. The selective expression of telomerase has thus been proposed to be a basis for the immortality of the germline and of malignant cells. In the present study, telomerase activity was analyzed in normal human T lymphocytes. It was found that telomerase is expressed at a high level in thymocyte subpopulations, at an intermediate level in tonsil T lymphocytes, and at a low to undetectable level in peripheral blood T lymphocytes. Moreover, telomerase activity is highly inducible in peripheral T lymphocytes by activation through CD3 with or without CD28 costimulation, or by stimulation with phorbol myristate acetate (PMA)/ionomycin. The induction of telomerase by anti-CD3 plus anti-CD28 (anti-CD3/CD28) stimulation required RNA and protein synthesis, and was blocked by herbimycin A, an inhibitor of S pi protein tyrosine kinases. The immunosuppressive drug cyclosporin A selectively inhibited telomerase induction by PMA/ionomycin and by anti-CD3, but not by anti-CD3/CD28. Although telomerase activity in peripheral T lymphocytes was activation dependent and correlated with cell proliferation, it was not cell cycle phase restricted. These results indicate that the expression of telomerase in normal human T lymphocytes is both developmentally regulated and activation induced. Telomerase may thus play a permissive role in T cell development and in determining the capacity of lymphoid cells for cell division and clonal expansion.
Assuntos
Regulação da Expressão Gênica no Desenvolvimento , Ativação Linfocitária , Linfócitos T/enzimologia , Linfócitos T/imunologia , Telomerase/biossíntese , Antígenos CD/imunologia , Antígenos CD28/imunologia , Complexo CD3/imunologia , Criança , Pré-Escolar , Citometria de Fluxo , Humanos , Lactente , Ionomicina/farmacologia , Tonsila Palatina/imunologia , Subpopulações de Linfócitos T/imunologia , Linfócitos T/efeitos dos fármacos , Acetato de Tetradecanoilforbol/farmacologia , Timo/imunologiaRESUMO
Toxic shock syndrome (TSS) is a multi system disorder resulting from superantigen-mediated cytokine production. Nearly 90% of the clinical cases of TSS arise due to an exotoxin, toxic shock syndrome toxin-1 (TSST-1), elaborated by toxigenic strains of Staphylococcus aureus. It is clearly established that besides antigen-specific signals a variety of costimulatory signals are required for full T cell activation. However, the nature and potential redundancy of costimulatory signals are incompletely understood, particularly with regards to superantigen-mediated T cell activation in vivo. Here we report that CD28-deficient mice (CD28-/-) are completely resistant to TSST-1-induced lethal TSS while CD28 (+/-) littermate mice were partially resistant to TSST-1. The mechanism for the resistance of the CD28 (-/-) mice was a complete abrogation of TNF-alpha accumulation in the serum and a nearly complete (90%) impairment of IFN-gamma secretion in response to TSST-1 injection. In contrast, the serum level of IL-2 was only moderately influenced by the variation of CD28 expression. CD28 (-/-) mice retained sensitivity to TNF-alpha as demonstrated by equivalent lethality after cytokine injection. These findings establish an essential requirement for CD28 costimulatory signals in TSST-1-induced TSS. The hierarchy of TSST-1 resistance among CD28 wild-type (CD28+/+), CD28 heterozygous (CD28+/-), and CD28-/- mice suggests a gene-dose effect, implying that the levels of T cell surface CD28 expression critically regulate superantigen-mediated costimulation. Finally, as these results demonstrate the primary and non-redundant role of CD28 receptors in the initiation of the in vivo cytokine cascade, they suggest therapeutic approaches for superantigen-mediated immunopathology.
Assuntos
Toxinas Bacterianas , Antígenos CD28/genética , Choque Séptico/imunologia , Linfócitos T/imunologia , Animais , Cruzamentos Genéticos , Enterotoxinas/imunologia , Feminino , Heterozigoto , Imunidade Inata/genética , Interferon gama/biossíntese , Interferon gama/fisiologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Choque Séptico/genética , Choque Séptico/prevenção & controle , Staphylococcus aureus , Superantígenos/imunologia , Fator de Necrose Tumoral alfa/biossíntese , Fator de Necrose Tumoral alfa/fisiologiaRESUMO
CD45 is a transmembrane tyrosine phosphatase implicated in T cell antigen receptor (TCR)-mediated activation. In T cell variants expressing progressively lower levels of CD45 (from normal to undetectable), CD45 expression was inversely related to spontaneous tyrosine phosphorylation of multiple proteins, including the TCR zeta chain, and was directly correlated with TCR-driven phosphoinositide hydrolysis. The Ca2+ response in these cells was altered in an unexpected fashion. Unlike wild-type cells, stimulated CD45- cell populations did not manifest an early increase in intracellular Ca2+, but did exhibit a delayed and gradual increase in mean intracellular Ca2+. Computer-aided fluorescence imaging of individual cells revealed that CD45- cells experienced late Ca2+ oscillations that were not blocked by removal of extracellular Ca2+. CD45 revertants had the signaling properties of wild-type cells. Thus, CD45 has a profound influence on both TCR-mediated signaling and phosphotyrosine homeostasis, and its loss reveals a novel role for this tyrosine phosphatase in Ca2+ regulation.
Assuntos
Antígenos CD/metabolismo , Cálcio/metabolismo , Antígenos de Histocompatibilidade/metabolismo , Proteínas Tirosina Fosfatases/metabolismo , Receptores de Antígenos de Linfócitos T/metabolismo , Tirosina/análogos & derivados , Animais , Citometria de Fluxo , Homeostase , Antígenos Comuns de Leucócito , Camundongos , Camundongos Endogâmicos A , Fosforilação , Fosfotirosina , Tirosina/metabolismoRESUMO
Lymphoproliferative diseases are characterized by massive accumulation of CD4(-)CD8(-)B220(+) (double-negative [DN]) T cells in peripheral organs. Although evidence indicates these cells are derived from mature autoreactive alpha/beta T cells, the significance of coreceptor downregulation is not known. In this study, we examined the role CD4 coreceptor plays in the survival of repeatedly stimulated T cells. CD4(+/+) and CD4(-/-) T cells from AND T cell receptor (TCR) transgenic mice exhibited similar phenotypes after antigenic stimulation, but the CD4(-/-) T cells survived in much larger numbers than the CD4(+/+) cells upon primary and secondary major histocompatibility complex (MHC)/peptide stimulation. Enhanced survival of CD4(-/-) T cells was due to decreased apoptosis rather than enhanced proliferation. Similarly, circumvention of the CD4/MHC interaction by using a surrogate TCR ligand that does not engage CD4 led to significant enhancement of CD4(+/+) cells than when stimulated with MHC/peptide. Finally, we generated DN B220(+) T cells using an in vitro model system and showed they were more tolerant to chronic stimulation than CD4(+/+) cells. Together, these results indicate that coreceptor engagement controls expansion of normal T cells. In the absence of coreceptor, T cells survive chronic stimulation and express B220 as seen in autoimmune lymphoproliferative diseases.
Assuntos
Autoimunidade/imunologia , Ativação Linfocitária/imunologia , Receptores de Antígenos de Linfócitos T alfa-beta , Receptores Imunológicos/imunologia , Subpopulações de Linfócitos T/imunologia , Animais , Apoptose , Complexo CD3/imunologia , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD8-Positivos/imunologia , Sobrevivência Celular , Antígenos Comuns de Leucócito/imunologia , Transtornos Linfoproliferativos/etiologia , Camundongos , Camundongos Transgênicos , Subpopulações de Linfócitos T/citologiaRESUMO
To address the possible role of replicative senescence in human immunodeficiency virus (HIV) infection, telomere length, telomerase activity, and in vitro replicative capacity were assessed in peripheral blood T cells from HIV+ and HIV- donors. Genetic and age-specific effects on these parameters were controlled by studying HIV-discordant pairs of monozygotic twins. Telomere terminal restriction fragment (TRF) lengths from CD4+ T cells of HIV+ donors were significantly greater than those from HIV- twins. In contrast, telomere lengths in CD8+ T cells from HIV+ donors were shorter than in HIV- donors. The in vitro replicative capacity of CD4+ cells from HIV+ donors was equivalent to that of HIV- donors in response to stimulation through T cell receptor CD3 and CD28. Little or no telomerase activity was detected in freshly isolated CD4+ or CD8+ lymphocytes from HIV+ or HIV- donors, but was induced by in vitro stimulation of both HIV+ and HIV- donor cells. These results suggest that HIV infection is associated with alterations in the population dynamics of both CD4+ and CD8+ T cells, but fail to provide evidence for clonal exhaustion or replicative senescence as a mechanism underlying the decline in CD4+ T cells of HIV-infected donors.
Assuntos
Infecções por HIV/imunologia , Subpopulações de Linfócitos T/imunologia , Telomerase/análise , Telômero/ultraestrutura , Gêmeos Monozigóticos , Adulto , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD8-Positivos/imunologia , Células Cultivadas , Senescência Celular , Infecções por HIV/genética , Infecções por HIV/patologia , Humanos , Ativação Linfocitária , Pessoa de Meia-Idade , Subpopulações de Linfócitos T/citologiaRESUMO
CD4 T cells activated in vitro by anti-CD3/28-coated beads are resistant to infection by CC chemokine receptor 5 (CCR5)-dependent HIV-1 isolates. In vivo, antigen-presenting cells (APCs) activate CD4 T cells in part by signaling through the T cell receptor and CD28, yet cells stimulated in this manner are susceptible to HIV-1 infection. We show that cytotoxic T lymphocyte antigen 4 (CTLA-4) engagement counteracts the CD28 antiviral effects, and that the ratio of CTLA-4 to CD28 engagement determines the susceptibility of HIV-1 infection. Furthermore, unopposed CTLA-4 signaling provided by CD28 blockade promotes vigorous HIV-1 replication, despite minimal T cell proliferation. Finally, CTLA-4 antibodies decrease the susceptibility of antigen-activated CD4 T cells to HIV, suggesting a potential approach to prevent or limit viral spread in HIV-1-infected individuals.
Assuntos
Antígenos de Diferenciação/imunologia , HIV-1/imunologia , Imunoconjugados , Linfócitos T Citotóxicos/imunologia , Linfócitos T Citotóxicos/virologia , Abatacepte , Antígenos CD , Antígenos CD28/imunologia , Antígeno CTLA-4 , Células Cultivadas , Quimiocinas CC/biossíntese , Regulação para Baixo/imunologia , HIV-1/fisiologia , Humanos , Fito-Hemaglutininas/farmacologia , Receptores CCR5/biossíntese , Linfócitos T Citotóxicos/citologia , Linfócitos T Citotóxicos/efeitos dos fármacosRESUMO
Surface immunoglobulin (sIg)-mediated stimulation of B lymphocytes induces a tyrosine kinase-dependent sequence of events leading to rapid and large elevations in intracellular ionized calcium ([Ca2+]i). These early biochemical events do not necessarily lead to proliferation of B cells, however, and conversely, the absence of or inhibition of these events does not necessarily prevent cellular proliferation. We now show by digital image analysis of single B cells that conditions which lead to B cell proliferation are associated with low-level but persistent sustained or cyclic elevations in [Ca2+]i. In marked contrast, early and nonsustained elevations in [Ca2+]i are induced in B cells by stimuli that lead to G1 transition but fail to progress to DNA synthesis. Thus, when B cells were stimulated with mitogenic and nonmitogenic anti-IgD antibodies, both of which induce entry of cells into G1 and early calcium transients of comparable magnitude, persistent low-level calcium elevations were only detected in cells stimulated with the mitogenic antibody. Furthermore, persistent calcium elevations were also seen when B cells were stimulated with a multivalent dextran-anti-Ig conjugate which induced very high levels of B cell proliferation in the absence of detectable phosphatidylinositol 4,5-biphosphate hydrolysis or elevations in [Ca2+]i as detected by flow cytometry. Finally, B cells from X-linked B cell-defective mice, which do not proliferate in response to anti-Ig antibody, show marked and early increases in [Ca2+]i, but do not show persistent calcium elevations. These data suggest that the rapid and large increases of [Ca2+]i seen in lymphocytes within seconds after antigen receptor ligation may be associated with entry in G1, whereas low-level but persistent elevations may be the hallmark of a cell destined to synthesize DNA.
Assuntos
Linfócitos B/metabolismo , Cálcio/metabolismo , DNA/biossíntese , Ativação Linfocitária , Receptores de Antígenos de Linfócitos B/fisiologia , Animais , Anticorpos Anti-Idiotípicos/imunologia , Linfócitos B/imunologia , Camundongos , Camundongos Endogâmicos DBARESUMO
CD28 is an adhesion receptor expressed as a 44-kD dimer on the surface of a major subset of human T cells. The CD28 receptor regulates the production of multiple lymphokines, including interleukin 2 (IL-2), by activation of a signal transduction pathway that is poorly understood. Here we show that ligation of CD28 by a monoclonal antibody (mAb) or by a natural ligand, B7/BB1, induces protein tyrosine phosphorylation that is distinct from T cell receptor (TCR)-induced tyrosine phosphorylation. CD28-induced protein tyrosine phosphorylation was greatly enhanced in cells that had been preactivated by ligation of the TCR, or by pretreatment with phorbol esters. Rapid and prolonged tyrosine phosphorylation of a single substrate, pp100, was induced in T cells after interaction with B7/BB1 presented on transfected Chinese hamster ovary (CHO) cells. Anti-B7 mAb inhibited B7/BB1 receptor-induced tyrosine phosphorylation, indicating that B7-CD28 interaction was required. CD28-induced tyrosine phosphorylation was independent of the TCR because it occurred in a variant of the Jurkat T cell line that does not express the TCR. Herbimycin A, a protein tyrosine kinase inhibitor, could prevent CD28-induced tyrosine phosphorylation and CD28-induced IL-2 production in normal T cells. The simultaneous crosslinking of CD28 and CD45, a tyrosine phosphatase, could prevent tyrosine phosphorylation of pp100. These results suggest that specific tyrosine phosphorylation, particularly of pp100, occurs directly as a result of CD28 ligand binding and is involved in transducing the signal delivered through CD28 by accessory cells that express the B7/BB1 receptor. Thus, this particular form of signal transduction may be relevant to lymphokine production and, potentially may provide a means to study the induction of self-tolerance, given the putative role of the costimulatory signal in the induction of T cell activation or anergy.
Assuntos
Antígenos CD/metabolismo , Antígenos de Diferenciação de Linfócitos T/metabolismo , Receptores de Superfície Celular/metabolismo , Linfócitos T/metabolismo , Anticorpos Monoclonais/imunologia , Antígenos CD/fisiologia , Benzoquinonas , Antígenos CD28 , Antígenos de Histocompatibilidade/fisiologia , Humanos , Técnicas In Vitro , Interleucina-2/farmacologia , Lactamas Macrocíclicas , Antígenos Comuns de Leucócito , Peso Molecular , Fosfoproteínas/metabolismo , Fosforilação , Fosfotirosina , Quinonas/farmacologia , Agregação de Receptores , Rifabutina/análogos & derivados , Transdução de Sinais , Acetato de Tetradecanoilforbol/farmacologia , Fatores de Tempo , Células Tumorais Cultivadas , Tirosina/análogos & derivados , Tirosina/metabolismoRESUMO
CD8+ T lymphocytes confer significant but ultimately insufficient protection against HIV infection. Here we report that activated neonatal CD8+ T cells can be productively infected in vitro by macrophage-tropic (M-tropic) HIV-1 isolates, which are responsible for disease transmission, whereas they are resistant to T cell-tropic (T-tropic) HIV strains. Physiological activation of CD8-alpha/beta+ CD4- T cell receptor-alpha/beta+ neonatal T cells, including activation by allogeneic dendritic cells, induces the accumulation of CD4 messenger RNA and the expression of CD4 Ag on the cell surface. The large majority of anti-CD3/B7.1-activated cord blood CD8+ T cells coexpress CD4, the primary HIV receptor, as well as CCR5 and CXCR4, the coreceptors used by M- and T-tropic HIV-1 strains, respectively, to enter target cells. These findings are relevant to the rapid progression of neonatal HIV infection. Infection of primary HIV-specific CD8+ T cells may compromise their survival and thus significantly contribute to the failure of the immune system to control the infection. Furthermore, these results indicate a previously unsuspected level of plasticity in the neonatal immune system in the regulation of CD4 expression by costimulation.
Assuntos
Linfócitos T CD8-Positivos/imunologia , HIV-1/patogenicidade , Antígenos CD4/imunologia , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD4-Positivos/virologia , Linfócitos T CD8-Positivos/virologia , Sobrevivência Celular/imunologia , Citometria de Fluxo , Infecções por HIV/imunologia , Infecções por HIV/metabolismo , Humanos , Recém-Nascido , Macrófagos/metabolismo , RNA Mensageiro/metabolismo , Receptores de Antígenos de Linfócitos T/metabolismo , Receptores de HIV/metabolismoRESUMO
Signals generated through CD28-B7 and CD40 ligand (CD40L)-CD40 interactions have been shown to be crucial for the induction of long-term allograft survivability. We have recently demonstrated that humanized anti-CD40L (hu5C8) prevents rejection of mismatched renal allografts in primates. To investigate potential mechanisms of CD40L-induced allograft acceptance, we coimmobilized hu5C8 with suboptimal amounts of anti-CD3 to stimulate CD4(+) T cells. We now report that anti-CD3/CD40L costimulation results in CD28-independent activation and subsequent deletion of resting T cells. Coligation of CD3 and CD40L increased expression of CD69, CD25, and CD54 on CD4(+) T cells. We also found that costimulation with anti-CD3/CD40L resulted in enhanced production of interleukin (IL)-10, interferon gamma, and tumor necrosis factor alpha but not IL-2 or IL-6. Interestingly, after several days, anti-CD3/CD40L-mediated activation was followed by apoptosis in a significant population of cells. Consistent with that observation, anti-CD3/CD40L did not enhance the antiapoptotic proteins Bcl-2 and Bcl-xL. Further, the addition of CD28 at 24 h failed to rescue those cells induced to die after costimulation with anti-CD3/CD40L. Together, these data suggest that the graft-sparing effect of hu5C8 in vivo may result in part from early and direct effects on CD4(+) T cells, including a vigorous induction of immunomodulatory cytokines and/or apoptosis of allograft-specific T cells.
Assuntos
Apoptose , Linfócitos T CD4-Positivos/imunologia , Antígenos CD40/fisiologia , Citocinas/biossíntese , Glicoproteínas de Membrana/fisiologia , Antígenos CD/análise , Antígenos CD/fisiologia , Linfócitos T CD4-Positivos/citologia , Linfócitos T CD4-Positivos/efeitos dos fármacos , Antígenos CD40/genética , Ligante de CD40 , Células Cultivadas , Citocinas/genética , Regulação da Expressão Gênica , Humanos , Imunoglobulina G/farmacologia , Interferon gama/biossíntese , Interferon gama/genética , Interleucinas/biossíntese , Interleucinas/genética , Ativação Linfocitária , Complexo Principal de Histocompatibilidade , Glicoproteínas de Membrana/imunologia , Reação em Cadeia da Polimerase , Proteínas Recombinantes/metabolismo , Transdução de Sinais , Transfecção , Células Tumorais CultivadasRESUMO
We have been studying the re-activation of tumor-associated antigen (TAA)-specific CD8(+) T cells in sentinel lymph nodes (SLN) of melanoma patients upon intradermal administration of the CpG-B oligodeoxynucleotide PF-3512676. To facilitate functional testing of T cells from small SLN samples, high-efficiency polyclonal T cell expansion is required. In this study, SLN cells were expanded via classic methodologies with plate- or bead-bound anti-CD3/CD28 antibodies and with the K562/CD32/4-1BBL artificial APC system (K32/4-1BBL aAPC) and analyzed for responsiveness to common recall or TAA-derived peptides. K32/4-1BBL-expanded T cell populations contained significantly more effector/memory CD8(+) T cells. Moreover, recall and melanoma antigen-specific CD8(+) T cells were more frequently detected in K32/4-1BBL-expanded samples as compared with anti-CD3/CD28-expanded samples. We conclude that K32/4-1BBL aAPC are superior to anti-CD3/CD28 antibodies for the expansion of in vivo-primed specific CD8(+) T cells and that their use facilitates the sensitive monitoring of functional anti-tumor T cell immunity in SLN.
Assuntos
Linfócitos T CD8-Positivos/imunologia , Memória Imunológica/imunologia , Linfonodos/imunologia , Ativação Linfocitária/imunologia , Melanoma/imunologia , Biópsia de Linfonodo Sentinela , Membro 9 da Superfamília de Receptores de Fatores de Necrose Tumoral/metabolismo , Ligante 4-1BB/genética , Anticorpos Monoclonais/imunologia , Células Apresentadoras de Antígenos/imunologia , Antígenos CD/metabolismo , Linfócitos T CD8-Positivos/citologia , Linfócitos T CD8-Positivos/efeitos dos fármacos , Linfócitos T CD8-Positivos/metabolismo , Contagem de Células , Proliferação de Células , Epitopos de Linfócito T/imunologia , Humanos , Interferon gama/metabolismo , Interleucinas/metabolismo , Células K562 , Linfonodos/efeitos dos fármacos , Linfonodos/cirurgia , Ativação Linfocitária/efeitos dos fármacos , Proteína 1 de Membrana Associada ao Lisossomo/metabolismo , Antígenos Específicos de Melanoma/imunologia , Oligodesoxirribonucleotídeos/administração & dosagem , Oligodesoxirribonucleotídeos/farmacologia , Receptores de IgG/genética , Subpopulações de Linfócitos T/citologia , Subpopulações de Linfócitos T/efeitos dos fármacos , Subpopulações de Linfócitos T/imunologia , Subpopulações de Linfócitos T/metabolismo , Transfecção , Fator de Necrose Tumoral alfa/metabolismoRESUMO
The binding of antigen or monoclonal antibody to the T cell receptor for antigen or the closely associated CD3 complex causes increases in the concentration of intracellular ionized calcium and subsequent cell proliferation. By measuring second messenger production in primary cultures of human immunodeficiency virus (HIV-1)--infected T cells stimulated with monoclonal antibodies specific for either CD3 or CD2, a specific impairment of membrane signaling was revealed. The HIV-1--infected T cells were unable to mobilize Ca2+ after stimulation with anti-CD3, whereas CD2-induced calcium mobilization remained intact. Furthermore, the HIV-1--infected cells proliferated poorly after CD3 stimulation, although the cells retained normal DNA synthesis in response to interleukin-2 stimulation. These results show that the signals initiated by CD2 and CD3 can be regulated independently within the same T cell; uncoupling of signal transduction after antigen-specific stimulation provides a biochemical mechanism to explain, in part, the profound immunodeficiency of patients with HIV-1 infection.
Assuntos
Síndrome da Imunodeficiência Adquirida/fisiopatologia , Antígenos de Diferenciação de Linfócitos T/fisiologia , Receptores de Antígenos de Linfócitos T/fisiologia , Linfócitos T/fisiologia , Anticorpos Monoclonais/imunologia , Antígenos de Diferenciação/fisiologia , Antígenos CD2 , Complexo CD3 , Cálcio/fisiologia , HIV , Humanos , Receptores Imunológicos/fisiologia , Linfócitos T/microbiologia , Fatores de TempoRESUMO
Selection of the T cell receptor (TCR) repertoire in the thymus probably involves TCR-mediated signals transduced in developing thymocytes after interaction with thymic stromal cells bearing self ligands. TCR-transduced signals should have identifiable consequences that would distinguish thymocytes whose TCRs have been engaged by self ligands from those whose TCRs have not. Among thymocytes expressing a transgenic TCR of defined specificity, a large number had elevated intracellular calcium concentrations but only when resident in a negatively selecting thymus in which their self ligand was expressed. Thus, developing thymocytes are stimulated by endogenous ligands in vivo to mobilize intracellular calcium, and increased intracellular calcium concentrations may reflect the consequences of intrathymic signaling associated with thymic negative selection.
Assuntos
Cálcio/metabolismo , Receptores de Antígenos de Linfócitos T/fisiologia , Transdução de Sinais , Linfócitos T/fisiologia , Animais , Antígenos CD4/análise , Antígenos CD8/análise , Feminino , Masculino , Camundongos , Camundongos Transgênicos , Fenótipo , Receptores de Antígenos de Linfócitos T/genética , Caracteres Sexuais , Baço/imunologia , Subpopulações de Linfócitos T/imunologia , Linfócitos T/imunologia , Timo/imunologiaRESUMO
Quiescent T cells can be induced to express many genes by mitogen or antigen stimulation. The messenger RNAs of some of these genes undergo relatively rapid degradation compared to messenger RNAs from constitutively expressed genes. A T cell activation pathway that specifically regulates the stability of messenger RNAs for the lymphokines interleukin-2, interferon-gamma, tumor necrosis factor-alpha, and granulocyte-macrophage colony-stimulating factor is induced by stimulation of the CD28 surface molecule. This pathway does not directly affect the steady-state messenger RNA level, transcription, or messenger RNA half-life of other T cell activation genes, including c-myc, c-fos, IL-2 receptor, and the 4F2HC surface antigen. These data show that stimuli received at the cell surface can alter gene expression by inducing specific changes in messenger RNA degradation.
Assuntos
Ativação Linfocitária , Linfocinas/genética , RNA Mensageiro/metabolismo , Linfócitos T/imunologia , Antígenos de Diferenciação de Linfócitos T/imunologia , Antígenos CD28 , Complexo CD3 , Fatores Estimuladores de Colônias/genética , Estabilidade de Medicamentos , Regulação da Expressão Gênica , Fator Estimulador de Colônias de Granulócitos e Macrófagos , Substâncias de Crescimento/genética , Interferon gama/genética , Interleucina-2/genética , RNA Mensageiro/genética , Receptores de Antígenos de Linfócitos T/imunologia , Transcrição Gênica , Fator de Necrose Tumoral alfa/genéticaRESUMO
The CD45 protein is a transmembrane tyrosine phosphatase that is required for normal T cell receptor (TCR)-mediated signaling. A chimeric complementary DNA encoding the intracellular enzymatically active portion of murine CD45 preceded by a short amino-terminal sequence from p60c-src was transfected into CD45- T cells. Expression of this chimeric protein corrected most of the TCR signaling abnormalities observed in the absence of CD45, including TCR-mediated enhancement of tyrosine kinase activity and Ca2+ flux. Thus, the enzymatically active intracellular portion of CD45 is sufficient to allow TCR transmembrane signaling.
Assuntos
Antígenos Comuns de Leucócito/metabolismo , Receptores de Antígenos de Linfócitos T/metabolismo , Transdução de Sinais , Animais , Sequência de Bases , Complexo CD3/imunologia , Cálcio/metabolismo , Membrana Celular/metabolismo , Antígenos Comuns de Leucócito/genética , Proteínas de Membrana/metabolismo , Camundongos , Dados de Sequência Molecular , Fosforilação , Proteínas Tirosina Quinases/metabolismo , Proteínas Recombinantes de Fusão/metabolismo , Células Tumorais Cultivadas , Tirosina/metabolismoRESUMO
Most immature CD4+CD8+ thymocytes express only a small number of T cell receptor (TCR) molecules on their surface, and the TCR molecules they do express are only marginally capable of transducing intracellular signals. TCR expression and function was not intrinsically low in immature CD4+CD8+ thymocytes, but was found to be actively inhibited by CD4-mediated signals. Indeed, release of CD4+CD8+ thymocytes from CD4-mediated signals resulted in significant increases in both TCR expression and signaling function. These results suggest that, in CD4+CD8+ cells developing in the thymus, increased TCR expression and function requires release from CD4-mediated inhibition.
Assuntos
Antígenos de Diferenciação de Linfócitos T/imunologia , Antígenos CD4/imunologia , Receptores de Antígenos de Linfócitos T/fisiologia , Linfócitos T/imunologia , Animais , Anticorpos Monoclonais/imunologia , Antígenos CD8 , Membrana Celular/imunologia , Células Cultivadas , Antígenos de Histocompatibilidade Classe II/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Receptores de Antígenos de Linfócitos T/biossínteseRESUMO
Activation of CD4(+) T lymphocytes from human immunodeficiency virus-type 1 (HIV-1)-infected donors with immobilized antibodies to CD3 and CD28 induces a virus-resistant state. This effect is specific for macrophage-tropic HIV-1. Transcripts encoding CXCR4/Fusin, the fusion cofactor used by T cell line-tropic isolates, were abundant in CD3/CD28-stimulated cells, but transcripts encoding CCR5, the fusion cofactor used by macrophage-tropic viruses, were not detectable. Thus, CD3/CD28 costimulation induces an HIV-1-resistant phenotype similar to that seen in some highly exposed and HIV-uninfected individuals.