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1.
Blood ; 115(8): 1554-63, 2010 Feb 25.
Artigo em Inglês | MEDLINE | ID: mdl-20009034

RESUMO

The identification of molecules responsible for apoptotic cell (AC) uptake by dendritic cells (DCs) and induction of T-cell immunity against AC-associated antigens is a challenge in immunology. DCs differentiated in the presence of interferon-alpha (IFN-alpha-conditioned DCs) exhibit a marked phagocytic activity and a special attitude in inducing CD8(+) T-cell response. In this study, we found marked overexpression of the scavenger receptor oxidized low-density lipoprotein receptor 1 (LOX-1) in IFN-alpha-conditioned DCs, which was associated with increased levels of genes belonging to immune response families and high competence in inducing T-cell immunity against antigens derived from allogeneic apoptotic lymphocytes. In particular, the capture of ACs by IFN-alpha DCs led to a substantial subcellular rearrangement of major histocompatibility complex class I and class II molecules, along with enhanced cross-priming of autologous CD8(+) T cells and CD4(+) T-cell activation. Remarkably, AC uptake, CD8(+) T-cell cross-priming, and, to a lesser extent, priming of CD4(+) T lymphocytes were inhibited by a neutralizing antibody to the scavenger receptor LOX-1 protein. These results unravel a novel LOX-1-dependent pathway by which IFN-alpha can, under both physiologic and pathologic conditions, render DCs fully competent for presenting AC-associated antigens for cross-priming CD8(+) effector T cells, concomitantly with CD4(+) T helper cell activation.


Assuntos
Apresentação de Antígeno/imunologia , Apoptose/efeitos dos fármacos , Células Dendríticas/imunologia , Fatores Imunológicos/farmacologia , Interferon-alfa/farmacologia , Receptores Depuradores Classe E/imunologia , Transdução de Sinais/efeitos dos fármacos , Apoptose/imunologia , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD8-Positivos/imunologia , Células Cultivadas , Humanos , Imunidade Celular/efeitos dos fármacos , Fatores Imunológicos/imunologia , Interferon-alfa/imunologia , Ativação Linfocitária/efeitos dos fármacos , Transdução de Sinais/imunologia
2.
J Exp Med ; 196(11): 1415-25, 2002 Dec 02.
Artigo em Inglês | MEDLINE | ID: mdl-12461077

RESUMO

Interferon (IFN) consensus sequence-binding protein (ICSBP) is a transcription factor playing a critical role in the regulation of lineage commitment, especially in myeloid cell differentiation. In this study, we have characterized the phenotype and activation pattern of subsets of dendritic cells (DCs) in ICSBP(-/-) mice. Remarkably, the recently identified mouse IFN-producing cells (mIPCs) were absent in all lymphoid organs from ICSBP(-/-) mice, as revealed by lack of CD11c(low)B220(+)Ly6C(+)CD11b(-) cells. In parallel, CD11c(+) cells isolated from ICSBP(-/-) spleens were unable to produce type I IFNs in response to viral stimulation. ICSBP(-/-) mice also displayed a marked reduction of the DC subset expressing the CD8alpha marker (CD8alpha(+) DCs) in spleen, lymph nodes, and thymus. Moreover, ICSBP(-/-) CD8alpha(+) DCs exhibited a markedly impaired phenotype when compared with WT DCs. They expressed very low levels of costimulatory molecules (intercellular adhesion molecule [ICAM]-1, CD40, CD80, CD86) and of the T cell area-homing chemokine receptor CCR7, whereas they showed higher levels of CCR2 and CCR6, as revealed by reverse transcription PCR. In addition, these cells were unable to undergo full phenotypic activation upon in vitro culture in presence of maturation stimuli such as lipopolysaccharide or poly (I:C), which paralleled with lack of Toll-like receptor (TLR)3 mRNA expression. Finally, cytokine expression pattern was also altered in ICSBP(-/-) DCs, as they did not express interleukin (IL)-12p40 or IL-15, but they displayed detectable IL-4 mRNA levels. On the whole, these results indicate that ICSBP is a crucial factor in the regulation of two possibly linked processes: (a) the development and activity of mIPCs, whose lack in ICSBP(-/-) mice may explain their high susceptibility to virus infections; (b) the generation and activation of CD8alpha(+) DCs, whose impairment in ICSBP(-/-) mice can be responsible for the defective generation of a Th1 type of immune response.


Assuntos
Antígenos CD8/análise , Células Dendríticas/fisiologia , Interferon Tipo I/biossíntese , Proteínas Repressoras/fisiologia , Animais , Antígenos CD/análise , Antígeno B7-1/análise , Antígeno B7-2 , Antígeno CD11a , Antígenos CD40/análise , Citocinas/biossíntese , Células Dendríticas/imunologia , Molécula 1 de Adesão Intercelular/análise , Fatores Reguladores de Interferon , Glicoproteínas de Membrana/análise , Camundongos , Camundongos Endogâmicos C57BL , Receptores de Quimiocinas/análise
3.
J Immunol ; 181(3): 1673-82, 2008 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-18641303

RESUMO

Regulatory T (Treg) cells are critical in inducing and maintaining tolerance. Despite progress in understanding the basis of immune tolerance, mechanisms and molecules involved in the generation of Treg cells remain poorly understood. IFN regulatory factor (IRF)-1 is a pleiotropic transcription factor implicated in the regulation of various immune processes. In this study, we report that IRF-1 negatively regulates CD4(+)CD25(+) Treg cell development and function by specifically repressing Foxp3 expression. IRF-1-deficient (IRF-1(-/-)) mice showed a selective and marked increase of highly activated and differentiated CD4(+)CD25(+)Foxp3(+) Treg cells in thymus and in all peripheral lymphoid organs. Furthermore, IRF-1(-/-) CD4(+)CD25(-) T cells showed extremely high bent to differentiate into CD4(+)CD25(+)Foxp3(+) Treg cells, whereas restoring IRF-1 expression in IRF-1(-/-) CD4(+)CD25(-) T cells impaired their differentiation into CD25(+)Foxp3(+) cells. Functionally, both isolated and TGF-beta-induced CD4(+)CD25(+) Treg cells from IRF-1(-/-) mice exhibited more increased suppressive activity than wild-type Treg cells. Such phenotype and functional characteristics were explained at a mechanistic level by the finding that IRF-1 binds a highly conserved IRF consensus element sequence (IRF-E) in the foxp3 gene promoter in vivo and negatively regulates its transcriptional activity. We conclude that IRF-1 is a key negative regulator of CD4(+)CD25(+) Treg cells through direct repression of Foxp3 expression.


Assuntos
Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD4-Positivos/metabolismo , Diferenciação Celular/imunologia , Fatores de Transcrição Forkhead/metabolismo , Fator Regulador 1 de Interferon/metabolismo , Subunidade alfa de Receptor de Interleucina-2/imunologia , Animais , Linfócitos T CD4-Positivos/citologia , Células Cultivadas , Sequência Consenso , Regulação para Baixo , Fatores de Transcrição Forkhead/genética , Humanos , Fator Regulador 1 de Interferon/deficiência , Fator Regulador 1 de Interferon/genética , Camundongos , Camundongos Knockout , Regiões Promotoras Genéticas/genética , Ligação Proteica , Transcrição Gênica/genética
4.
PLoS One ; 8(8): e72833, 2013.
Artigo em Inglês | MEDLINE | ID: mdl-23977359

RESUMO

Type I interferon (IFN-I) have emerged as crucial mediators of cellular signals controlling DC differentiation and function. Human DC differentiated from monocytes in the presence of IFN-α (IFN-α DC) show a partially mature phenotype and a special capability of stimulating CD4+ T cell and cross-priming CD8+ T cells. Likewise, plasmacytoid DC (pDC) are blood DC highly specialized in the production of IFN-α in response to viruses and other danger signals, whose functional features may be shaped by IFN-I. Here, we investigated the molecular mechanisms stimulated by IFN-α in driving human monocyte-derived DC differentiation and performed parallel studies on peripheral unstimulated and IFN-α-treated pDC. A specific miRNA signature was induced in IFN-α DC and selected miRNAs, among which miR-23a and miR-125b, proved to be negatively associated with up-modulation of Blimp-1 occurring during IFN-α-driven DC differentiation. Of note, monocyte-derived IFN-α DC and in vitro IFN-α-treated pDC shared a restricted pattern of miRNAs regulating Blimp-1 expression as well as some similar phenotypic, molecular and functional hallmarks, supporting the existence of a potential relationship between these DC populations. On the whole, these data uncover a new role of Blimp-1 in human DC differentiation driven by IFN-α and identify Blimp-1 as an IFN-α-mediated key regulator potentially accounting for shared functional features between IFN-α DC and pDC.


Assuntos
Células Dendríticas/citologia , Células Dendríticas/metabolismo , Interferon-alfa/farmacologia , MicroRNAs/genética , Monócitos/citologia , Proteínas Repressoras/genética , Diferenciação Celular/efeitos dos fármacos , Diferenciação Celular/genética , Células Dendríticas/efeitos dos fármacos , Perfilação da Expressão Gênica , Células HeLa , Humanos , Fenótipo , Fator 1 de Ligação ao Domínio I Regulador Positivo , Proteínas Repressoras/metabolismo
5.
Neoplasia ; 14(9): 855-67, 2012 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-23019417

RESUMO

Currently approved combination regimens available for the treatment of metastatic tumors, such as breast cancer, have been shown to increase response rates, often at the cost of a substantial increase in toxicity. An ideal combination strategy may consist of agents with different mechanisms of action leading to complementary antitumor activities and safety profiles. In the present study, we investigated the effects of the epigenetic modulator apicidin in combination with the cytotoxic agent docetaxel in tumor breast cell lines characterized by different grades of invasiveness. We report that combined treatment of apicidin and docetaxel, at low toxicity doses, stimulates in metastatic breast cancer cells the expression of CTCF-like protein and other cancer antigens, thus potentially favoring an antitumor immune response. In addition, apicidin and docetaxel co-treatment specifically stimulates apoptosis, characterized by an increased Bax/Bcl-2 ratio and caspase-8 activation. Importantly, following combined exposure to these agents, metastatic cells were also found to induce signals of immunogenic apoptosis such as cell surface expression of calreticulin and release of considerable amounts of high-mobility group box 1 protein, thus potentially promoting the translation of induced cell death into antitumor immune response. Altogether, our results indicate that the combined use of apicidin and docetaxel, at a low toxicity profile, may represent a potential innovative strategy able to activate complementary antitumor pathways in metastatic breast cancer cells, associated with a potential control of metastatic growth and possible induction of antitumor immunity.


Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Neoplasias da Mama/genética , Proteínas de Ligação a DNA/genética , Proteína HMGB1/genética , Peptídeos Cíclicos/farmacologia , Taxoides/farmacologia , Apoptose/efeitos dos fármacos , Apoptose/imunologia , Neoplasias da Mama/imunologia , Neoplasias da Mama/patologia , Proteínas de Ligação a DNA/imunologia , Células Dendríticas/efeitos dos fármacos , Células Dendríticas/imunologia , Docetaxel , Sinergismo Farmacológico , Feminino , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Proteína HMGB1/imunologia , Humanos , Imunomodulação/efeitos dos fármacos , Metástase Neoplásica , Peptídeos Cíclicos/administração & dosagem , Taxoides/administração & dosagem
6.
Viral Immunol ; 25(2): 117-30, 2012 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-22413916

RESUMO

Despite the fact that murine cells are not permissive for human immunodeficiency virus type 1 (HIV-1) infection, several investigators have constructed transgenic (Tg) mice to model HIV-1-induced diseases to overcome this restriction. The generation of Tg mice expressing selected HIV-1 genes revealed that Nef harbors a major disease determinant. HIV-1 Nef protein is a molecular adapter able to interact with several cellular partners, interfering with cellular functions. The phenotype of Nef Tg mice was extensively characterized regarding in vivo development of AIDS-like disease and the effects of Nef expression in T lymphocytes, but the functions eventually corrupted by Nef in monocytes and macrophages were less studied. Nef treatment of human monocyte-derived macrophages induces the internalization of the protein and modulates the production and secretion of different chemokines and cytokines by activating specific intracellular signaling pathways (i.e., NF-κB, MAPK, and IRF3). Therefore we set up an in vitro murine macrophage-based model using stabilized cell lines and primary peritoneal macrophages, and treated them with recombinant myristoylated Nef(SF2) (recNef). Like human cells, murine macrophages responded to Nef treatment, activating IKK-α and IKK-ß, JNK, and p38 MAP kinases. Activation of the NF-κB pathway is mandatory for the synthesis and release of a pool of cytokines and chemokines, including IFN-ß, that induce tyrosine phosphorylation of the signal transducer and activator of transcription (STAT)-1, STAT-2, and STAT-3, in an autocrine and paracrine manner, confirming that murine macrophages respond to Nef similarly to human ones. These data extend the results previously obtained in human primary macrophages, allowing the use of murine cells in culture to study signaling events modulated by Nef in myeloid-derived cells. In particular, it may be feasible to use macrophages derived from mice knocked out in specific signaling intermediates to obtain greater insight into the mechanism of Nef-induced effects.


Assuntos
HIV-1/patogenicidade , Ativação de Macrófagos , Macrófagos/efeitos dos fármacos , Macrófagos/imunologia , Transdução de Sinais , Fatores de Virulência/imunologia , Produtos do Gene nef do Vírus da Imunodeficiência Humana/imunologia , Animais , Células Cultivadas , Citocinas/metabolismo , HIV-1/imunologia , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Transgênicos
7.
Immunity ; 27(2): 228-39, 2007 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-17702615

RESUMO

Dendritic cells (DCs) produce type I interferons (IFNs) in greater amounts than other cells, but the mechanisms remain elusive. Here we studied the role of a transcription factor, IRF8, in DC induction of type I IFNs. Upon newcastle disease virus (NDV) infection, bone marrow-derived plasmacytoid and conventional DCs induced IFN transcripts, exhibiting two-phase kinetics. The second, amplifying phase represented an IFN feedback response that accounted for much of IFN protein production. Induction of second phase transcription required IRF8. Mouse cytomegalovirus (MCMV) and Toll-like receptor-mediated IFN induction in DCs also required IRF8. Chromatin immunoprecipitation analysis showed that IRF7, IRF8, and RNA polymerase II were recruited to the IFN promoters upon stimulation. Moreover, sustained RNA polymerase II recruitment to the promoters critically depended on IRF8. Together, these data indicate that IRF8 magnifies the second phase of IFN transcription in DCs by prolonging binding of basic transcription machinery to the IFN promoters, thereby playing a role in innate immunity.


Assuntos
Células Dendríticas/imunologia , Regulação da Expressão Gênica , Fatores Reguladores de Interferon/metabolismo , Interferon Tipo I/genética , Animais , Imunoprecipitação da Cromatina , Retroalimentação Fisiológica , Fibroblastos/imunologia , Fator Regulador 7 de Interferon/metabolismo , Fatores Reguladores de Interferon/genética , Camundongos , Camundongos Mutantes , Regiões Promotoras Genéticas , RNA Polimerase II/metabolismo
8.
Blood ; 108(2): 609-17, 2006 Jul 15.
Artigo em Inglês | MEDLINE | ID: mdl-16569763

RESUMO

Interferon consensus sequence-binding protein (ICSBP)/interferon regulatory factor 8 (IRF-8) is a transcription factor that plays critical roles in the differentiation of defined dendritic-cell (DC) populations and in the immune response to many pathogens. In this study, we show that splenic DCs (s-DCs) from ICSBP(-/-) mice are markedly defective in their ability to capture and to present exogenous antigens (Ags) to naive CD4(+) T lymphocytes. We found that CD8alpha(+) DCs and, to a lesser extent, CD8alpha(-) DCs from ICSBP(-/-) mice are impaired at internalizing Ags, either through a receptor-mediated pathway or by macropinocytosis, in spite of having a more immature phenotype than their wild-type (WT) counterparts. These features reflected a greatly impaired ability of ICSBP(-/-) s-DCs to present injected soluble ovalbumin (OVA) to OVA-specific CD4(+) T cells in vivo. Conversely, bone marrow (BM)-derived DCs from ICSBP(-/-) mice, in keeping with their immature phenotype, exhibited higher endocytic activity than WT cells. However, Ag-loaded ICSBP(-/-) BM-DCs were defective in priming Ag-specific CD4(+) T lymphocytes and failed to induce a contact hypersensitivity (CHS) response when injected into competent WT hosts. Together, these results indicate that, throughout the developmental program of DCs, ICSBP differentially controls Ag uptake and MHC class II (MHC-II) presentation affecting both functions only in differentiated peripheral DCs.


Assuntos
Apresentação de Antígeno , Linfócitos T CD4-Positivos/imunologia , Células Dendríticas/citologia , Fatores Reguladores de Interferon/fisiologia , Animais , Células Cultivadas , Células Dendríticas/imunologia , Antígenos de Histocompatibilidade Classe II/imunologia , Fatores Reguladores de Interferon/deficiência , Fatores Reguladores de Interferon/imunologia , Camundongos , Camundongos Knockout , Pinocitose , Baço/citologia
9.
Blood ; 103(3): 980-7, 2004 Feb 01.
Artigo em Inglês | MEDLINE | ID: mdl-14525781

RESUMO

Chronic myelogenous leukemia (CML) is a malignant myeloproliferative disease arising from the clonal expansion of a stem cell expressing the bcr/abl oncogene. CML patients frequently respond to treatment with interferon-alpha (IFN-alpha), even though the mechanisms of the response remain unclear. In the present study, we evaluated the role of IFN-alpha in differentiation and activity of monocyte-derived dendritic cells (DCs) from CML patients as well as in modulation of the cell response to lipopolysaccharide (LPS). Treatment of CML monocytes with IFN-alpha and granulocyte-macrophage colony-stimulating factor (GM-CSF) resulted in the rapid generation of activated DCs (CML-IFN-DCs) expressing interleukin-15 (IL-15) and the antiapoptotic bcl-2 gene. These cells were fully competent to induce IFN-gamma production by cocultured autologous T lymphocytes and expansion of CD8(+) T cells. LPS treatment of CML-IFN-DCs, but not of immature DCs generated in the presence of IL-4/GM-CSF, induced the generation of CD8(+) T cells reactive against autologous leukemic CD34(+) cells. Altogether, these results suggest that (1) the generation of highly active monocyte-derived DCs could be important for the induction of an antitumor response in IFN-treated CML patients and (2) IFN-alpha can represent a valuable cytokine for the rapid generation of active monocyte-derived DCs to be utilized for vaccination strategies of CML patients.


Assuntos
Células Dendríticas/efeitos dos fármacos , Interferon Tipo I/farmacologia , Leucemia Mielogênica Crônica BCR-ABL Positiva/tratamento farmacológico , Leucemia Mielogênica Crônica BCR-ABL Positiva/patologia , Monócitos/efeitos dos fármacos , Adulto , Idoso , Sequência de Bases , Linfócitos T CD8-Positivos/imunologia , Diferenciação Celular/efeitos dos fármacos , Quimiocina CXCL10 , Quimiocinas CXC/genética , Técnicas de Cocultura , Células Dendríticas/imunologia , Células Dendríticas/patologia , Feminino , Expressão Gênica/efeitos dos fármacos , Fator Estimulador de Colônias de Granulócitos e Macrófagos/farmacologia , Humanos , Interferon gama/biossíntese , Interleucina-15/genética , Leucemia Mielogênica Crônica BCR-ABL Positiva/imunologia , Lipopolissacarídeos/farmacologia , Masculino , Pessoa de Meia-Idade , Monócitos/imunologia , Monócitos/patologia , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Proteínas Recombinantes
10.
Blood ; 103(6): 2221-8, 2004 Mar 15.
Artigo em Inglês | MEDLINE | ID: mdl-14615368

RESUMO

Interferon consensus sequence-binding protein (ICSBP) is a transcription factor belonging to the interferon regulatory factor (IRF) family, recently shown to play a critical role in dendritic cell (DC) differentiation. Here, we analyzed the role of ICSBP in the development and trafficking of epidermal Langerhans cells (LCs) and dermal DCs and the implications for initiation of a competent immune response. ICSBP-/- mice exhibited a reduced frequency of LCs and a delayed mobility of DCs from skin that reflected a slower turnover rate in lymph nodes during steady-state conditions. Even under inflammatory changes, ICSBP-/- DCs displayed reduced mobility from skin to lymph nodes and, as a consequence, failed to induce a contact hypersensitivity (CHS) response, suggesting that these DCs were unable to initiate a competent antigen (Ag)-specific T-cell-mediated immunity. Moreover, bone marrow (BM)-derived DCs from ICSBP-/- mice exhibited an immature phenotype and a severe reduction of interleukin 12 (IL-12) expression. These BM DCs also showed a marked defect in their migratory response to macrophage inflammatory protein 3 alpha (MIP-3 alpha), MIP-3 beta, and the CC chemokine CCL21/6Ckine, which was paralleled by an impaired expression of the CC chemokine receptors, CCR6 and CCR7. Together, these results indicate that ICSBP is critically required for the development and trafficking of skin DCs, thus playing a critical role in the DC-mediated initiation of T-cell immunity.


Assuntos
Movimento Celular/imunologia , Dermatite/imunologia , Células de Langerhans/fisiologia , Proteínas Repressoras/genética , Proteínas Repressoras/metabolismo , Animais , Comunicação Celular/imunologia , Quimiotaxia/imunologia , Derme/citologia , Derme/imunologia , Fatores Reguladores de Interferon , Células de Langerhans/citologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Mutantes , Linfócitos T/citologia , Linfócitos T/imunologia
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