RESUMO
The ability to generate large numbers of distinct types of human dendritic cells (DCs) in vitro is critical for accelerating our understanding of DC biology and harnessing them clinically. We developed a DC differentiation method from human CD34+ precursors leading to high yields of plasmacytoid DCs (pDCs) and both types of conventional DCs (cDC1s and cDC2s). The identity of the cells generated in vitro and their strong homology to their blood counterparts were demonstrated by phenotypic, functional, and single-cell RNA-sequencing analyses. This culture system revealed a critical role of Notch signaling and GM-CSF for promoting cDC1 generation. Moreover, we discovered a pre-terminal differentiation state for each DC type, characterized by high expression of cell-cycle genes and lack of XCR1 in the case of cDC1. Our culture system will greatly facilitate the simultaneous and comprehensive study of primary, otherwise rare human DC types, including their mutual interactions.
Assuntos
Linhagem da Célula/imunologia , Células Dendríticas/imunologia , Fator Estimulador de Colônias de Granulócitos e Macrófagos/genética , Peptídeos e Proteínas de Sinalização Intercelular/genética , Proteínas de Membrana/genética , Receptor Notch1/genética , Antígenos CD34/genética , Antígenos CD34/imunologia , Proteínas de Ligação ao Cálcio , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/imunologia , Diferenciação Celular/efeitos dos fármacos , Células Dendríticas/citologia , Células Dendríticas/efeitos dos fármacos , Expressão Gênica , Perfilação da Expressão Gênica , Fator Estimulador de Colônias de Granulócitos e Macrófagos/imunologia , Humanos , Imidazóis/farmacologia , Imunofenotipagem , Peptídeos e Proteínas de Sinalização Intercelular/imunologia , Lipopolissacarídeos/farmacologia , Proteínas de Membrana/imunologia , Poli I-C/farmacologia , Cultura Primária de Células , Receptor Notch1/imunologia , Receptores Acoplados a Proteínas G/deficiência , Receptores Acoplados a Proteínas G/genética , Receptores Acoplados a Proteínas G/imunologia , Transdução de Sinais , Análise de Célula ÚnicaRESUMO
Bovine neonatal pancytopenia (BNP) is a recently described haemorrhagic disease of calves characterised by thrombocytopenia, leucopenia and bone marrow depletion. Feeding colostrum from cows that have previously produced a BNP affected calf has been shown to induce the disease in some calves, leading to the hypothesis that alloantibodies in colostrum from dams of affected calves mediate destruction of blood and bone marrow cells in the recipient calves. The aims of the current experimental study were first to confirm the role of colostrum-derived antibody in mediating the disease and second to investigate the haematopoietic cell lineages and maturation stages depleted by the causative antibodies. Clinical, haematological and pathological changes were examined in 5 calves given a standardised pool of colostrum from known BNP dams, and 5 control calves given an equivalent pool of colostrum from non-BNP dams. All calves fed challenge colostrum showed progressive depletion of bone marrow haematopoietic cells and haematological changes consistent with the development of BNP. Administration of a standardised dose of the same colostrum pool to each calf resulted in a consistent response within the groups, allowing detailed interpretation of the cellular changes not previously described. Analyses of blood and serial bone marrow changes revealed evidence of differential effects on different blood cell lineages. Peripheral blood cell depletion was confined to leucocytes and platelets, while bone marrow damage occurred to the primitive precursors and lineage committed cells of the thrombocyte, lymphocyte and monocyte lineages, but only to the more primitive precursors in the neutrophil, erythrocyte and eosinophil lineages. Such differences between lineages may reflect cell type-dependent differences in levels of expression or conformational nature of the target antigens.
Assuntos
Doenças dos Bovinos/imunologia , Colostro/imunologia , Isoanticorpos/administração & dosagem , Isoanticorpos/efeitos adversos , Pancitopenia/veterinária , Animais , Animais Recém-Nascidos , Células Sanguíneas/imunologia , Células Sanguíneas/patologia , Medula Óssea/imunologia , Medula Óssea/patologia , Bovinos , Doenças dos Bovinos/sangue , Doenças dos Bovinos/patologia , Linhagem da Célula/imunologia , Feminino , Genes MHC da Classe II , Células-Tronco Hematopoéticas/imunologia , Células-Tronco Hematopoéticas/patologia , Modelos Imunológicos , Pancitopenia/imunologia , Pancitopenia/patologia , GravidezRESUMO
Eicosanoids have been implicated in the physiological regulation of hematopoiesis with pleiotropic effects on hematopoietic stem cells and various classes of lineage restricted progenitor cells. Herein we review the effects of eicosanoids on hematopoiesis, focusing on new findings implicating prostaglandin E(2) in enhancing hematopoietic stem cell engraftment by enhancing stem cell homing, survival and self-renewal. We also describe a role for cannabinoids in hematopoiesis. Lastly, we discuss the yin and yang of various eicosanoids in modulating hematopoietic stem and progenitor cell functions and summarize potential strategies to take advantage of these effects for therapeutic benefit for hematopoietic stem cell transplantation.
Assuntos
Canabinoides/imunologia , Dinoprostona/imunologia , Hematopoese/imunologia , Células-Tronco Hematopoéticas/fisiologia , Transdução de Sinais/imunologia , Animais , Apoptose/genética , Apoptose/imunologia , Canabinoides/genética , Canabinoides/metabolismo , Ciclo Celular/genética , Ciclo Celular/imunologia , Linhagem da Célula/genética , Linhagem da Célula/imunologia , Movimento Celular/genética , Movimento Celular/imunologia , Quimiocina CXCL12/genética , Quimiocina CXCL12/imunologia , Quimiocina CXCL12/metabolismo , Dinoprostona/genética , Dinoprostona/metabolismo , Regulação da Expressão Gênica/imunologia , Sobrevivência de Enxerto/genética , Sobrevivência de Enxerto/imunologia , Hematopoese/genética , Transplante de Células-Tronco Hematopoéticas , Humanos , Camundongos , Receptores CXCR4/genética , Receptores CXCR4/imunologia , Receptores CXCR4/metabolismo , Transdução de Sinais/genética , Yin-YangRESUMO
Although it has been established that maternal leukocytes traffic from colostrum into the neonatal circulation, the effects of these cells on neonatal immunity are only beginning to be understood. This study examined the effects of maternal colostral leukocytes on development and maturation of neonatal antigen presenting cells. At birth, groups of neonatal calves received whole or cell-free colostrum (CFC) from their respective dams. Peripheral blood samples were obtained over the first 4 weeks of life, and expression of surface markers associated with cellular activation and physiological stress were monitored on monocyte lineage cells. Calves receiving cell-free colostrum at birth expressed elevated levels of CD11a, CD11c, and CD14, compared to calves receiving whole colostrum (C). Calves receiving cell-free colostrum had an elevated number of monocytes in the peripheral blood during the first 2 weeks of life, however, these cells expressed lower levels of expression of CD25 and MHC class I compared to calves receiving whole colostrum. The most significant differences in marker expression occurred within the first 7 days of life.
Assuntos
Bovinos/imunologia , Colostro/imunologia , Leucócitos Mononucleares/imunologia , Animais , Animais Recém-Nascidos , Apresentação de Antígeno , Antígeno CD11a/biossíntese , Antígeno CD11a/sangue , Antígeno CD11c/biossíntese , Antígeno CD11c/sangue , Bovinos/sangue , Linhagem da Célula/imunologia , Colostro/citologia , Ensaio de Imunoadsorção Enzimática/veterinária , Feminino , Citometria de Fluxo/veterinária , Antígenos de Histocompatibilidade Classe I/imunologia , Antígenos de Histocompatibilidade Classe II/imunologia , Imunoglobulina G/sangue , Imunofenotipagem/veterinária , Receptores de Lipopolissacarídeos/biossíntese , Receptores de Lipopolissacarídeos/sangue , Óxido Nítrico/biossíntese , Óxido Nítrico/sangueRESUMO
Adoptive T-cell immunotherapy may provide complementary therapy for childhood B-cell precursor acute lymphoblastic leukemia (BCP-ALL). In this study, we have analyzed the functional characteristics of anti-BCP-ALL effector T cells generated by co-culturing T lymphocytes and dendritic cells (DC) from allogeneic human stem cell transplantation (HSCT) donors. After 21-day co-culture with DC pulsed with CD40L+ apoptotic BCP-ALL blasts, T cells presented with both effector and central memory phenotype, and showed high and specific cytotoxic activity against leukemic cells (average lysis = 77%), mostly mediated by CD8+ T cells. Noticeably, growth of CD4 T cells was maintained (45% of total cells), which actively produced Th1 cytokines (IFN-gamma, TNF-alpha, IL-2), but not IL-4, IL-5 and IL-10. Anti-BCP-ALL T cells expressed CD49d and CXCR4 (implicated in the recruitment to bone marrow), and CD62L and CCR7 (involved in the migration to lymphoid organs). In accordance with this profile, T cells significantly migrated in response to the chemokines CXCL12 and CCL19. In conclusion, stimulation of T cells with CD40L+BCP-ALL cells-loaded DC not only elicited the generation of potent and specific anti-leukemic cytotoxic effectors, but also the differentiation of specific and functional Th-1 CD4 lymphocytes. These effectors are fully equipped to reach leukemia-infiltrated tissues and have characteristics to support and orchestrate the anti-tumor immune-response.
Assuntos
Transferência Adotiva/métodos , Ligante de CD40/metabolismo , Células Dendríticas/imunologia , Leucemia-Linfoma Linfoblástico de Células Precursoras/imunologia , Leucemia-Linfoma Linfoblástico de Células Precursoras/terapia , Células Th1/imunologia , Células Apresentadoras de Antígenos/citologia , Células Apresentadoras de Antígenos/imunologia , Linfócitos B/citologia , Linfócitos B/imunologia , Células da Medula Óssea/citologia , Comunicação Celular/imunologia , Linhagem da Célula/imunologia , Movimento Celular/imunologia , Células Cultivadas , Criança , Técnicas de Cocultura , Células Dendríticas/citologia , Humanos , Memória Imunológica , Imunofenotipagem , Células Th1/citologia , Células Th1/metabolismoRESUMO
Astragali Radix (AR), is a popular herbal medicine used to treat allergic diseases in Korea, Japan and China. Our study examined the effect of an AR ethanol extract on both in vitro and in vivo murine CD4 T cells' differentiation into Th1 and Th2 subsets. CD4 T cells from Balb/c mice were activated with anti-CD3/anti-CD28 mAb in the presence of AR for 2 d. AR treated cells showed an elevated level of IL-4 but a reduced level of IFN-gamma secretion. In addition, in vitro Th1/Th2 polarization experiments revealed that AR enhanced the levels of IL-4 in Th2 cells but reduced the levels of IFN-gamma in Th1 cells. To elucidate the effects of AR in Th1/Th2 lineage development during the in vivo condition, AR was administrated orally to BALB/c mice. The results demonstrated that AR administration significantly increased IL-4 production in both the serum and supernatant of splenocyte culture, while IFN-gamma secretion was diminished upon in vivo activation with anti-CD3 antibody. Our data clearly indicates that AR selectively alters Th1/Th2 cytokine secretion patterns and provides the pharmacological basis for AR's clinical applications.
Assuntos
Astrágalo , Fatores Imunológicos/farmacologia , Células Th1/citologia , Células Th2/citologia , Animais , Diferenciação Celular/efeitos dos fármacos , Diferenciação Celular/imunologia , Linhagem da Célula/efeitos dos fármacos , Linhagem da Célula/imunologia , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/imunologia , Células Cultivadas , Relação Dose-Resposta a Droga , Feminino , Fatores Imunológicos/isolamento & purificação , Camundongos , Camundongos Endogâmicos BALB C , Extratos Vegetais/isolamento & purificação , Extratos Vegetais/farmacologia , Raízes de Plantas , Células Th1/efeitos dos fármacos , Células Th1/imunologia , Células Th2/efeitos dos fármacos , Células Th2/imunologiaRESUMO
The present study was designed to evaluate the effect of So-Cheong-Ryong-Tang (SCRT, also called Sho-Seiryu-To or Xiao-Qing-Long-Tang) on helper T cell development by monitoring Th1/Th2-specific cytokine secretion patterns in artificially induced Th1 or Th2 polarized conditions. The results demonstrated that Th2 cells were dramatically underpopulated in the Th2-driven condition triggered by SCRT treatment, while the Th1 cells were not altered in the Th1-skewed condition. Furthermore, under Th2-skewed conditions the levels of interleukin-4 were considerably decreased with SCRT treatment. The expression of GATA-3, a transcription factor that plays a pivotal role in Th2 lineage programming, did not change with SCRT treatment, while the expression of another Th2 transcription factor, c-Maf, was dramatically suppressed. These data suggest that SCRT modulates Th2 development by suppressing c-Maf expression. This study implies that the SCRT effect on CD4(+) T cells is a key pharmacologic point of effect for treating IgE-mediated allergic asthma. These results also suggest that SCRT might be a useful agent for the correction of Th2-dominant pathologic disorders.
Assuntos
Medicamentos de Ervas Chinesas/farmacologia , Inibidores do Crescimento/farmacologia , Imunossupressores/farmacologia , Medicina Kampo , Células Th2/citologia , Células Th2/efeitos dos fármacos , Animais , Linhagem da Célula/efeitos dos fármacos , Linhagem da Célula/imunologia , Medicamentos de Ervas Chinesas/isolamento & purificação , Feminino , Coreia (Geográfico) , Camundongos , Camundongos Endogâmicos BALB CRESUMO
For decades immunologists have relied heavily on the mouse model for their experimental designs. With the realization of the important role innate immunity plays in orchestrating immune responses, invertebrates such as worms and flies have been added to the repertoire. Here, we discuss the advent of the zebrafish as a powerful vertebrate model organism that promises to positively impact immunologic research.
Assuntos
Imunidade Inata , Peixe-Zebra/imunologia , Animais , Linhagem da Célula/imunologia , Modelos Animais de Doenças , Avaliação Pré-Clínica de Medicamentos , Expressão Gênica , CamundongosRESUMO
Interleukin 18 induces both T helper 1 and T helper 2 cytokines, proinflammatory cytokines, chemokines, and IgE and IgG1 production. A role of interleukin 18 in inflammatory cutaneous reactions is still unclear, however. Here we generated keratin 5/interleukin 18 transgenic mice overexpressing mature murine interleukin 18 in the skin using a human keratin 5 promoter. In the contact hypersensitivity model, trinitrochlorobenzene elicited a stronger ear swelling in keratin 5/interleukin 18 transgenic mice compared with control littermate wild-type or immunoglobulin/interleukin 18 transgenic mice in which mature interleukin 18 was expressed by B and T cells under the control of the immunoglobulin promoter. Application of an irritant, croton oil, induced stronger and more sustained ear swelling in keratin 5/interleukin 18 transgenic mice than in immunoglobulin/interleukin 18 transgenic or wild-type mice. Repetitive topical application (weekly for six consecutive weeks) of trinitrochlorobenzene to their ears also elicited a stronger cutaneous inflammation in keratin 5/interleukin 18 transgenic mice than seen in immunoglobulin/interleukin 18 transgenic or wild-type mice. After these six trinitrochlorobenzene applications, the expression of interferon-gamma, interleukin-4, and CCL20 mRNA in the ear tissue was increased and dermal changes, such as acanthosis and eosinophilic, neutrophilic, and mast cell infiltration, were greater in keratin 5/interleukin 18 transgenic mice than in wild-type mice. Furthermore, the repetitive application elicited a significant increase in serum IgE levels and the number of B cells in the draining lymph node in keratin 5/interleukin 18 transgenic mice. These results suggest that overexpression of interleukin 18 in the skin aggravates allergic and nonallergic cutaneous inflammation, which is accompanied by high expression of T helper 1 and T helper 2 cytokines and chemokines in the skin.
Assuntos
Dermatite Alérgica de Contato/imunologia , Dermatite Alérgica de Contato/fisiopatologia , Interleucina-18/genética , Interleucina-18/imunologia , Pele/imunologia , Animais , Linhagem da Célula/imunologia , Quimiocinas/genética , Óleo de Cróton , Citocinas/genética , Dermatite Alérgica de Contato/patologia , Orelha Externa , Feminino , Expressão Gênica/imunologia , Irritantes , Queratina-15 , Queratina-5 , Queratinócitos/patologia , Queratinócitos/fisiologia , Queratinas/genética , Linfonodos/citologia , Linfonodos/imunologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Cloreto de Picrila , Regiões Promotoras Genéticas , RNA Mensageiro/análise , Pele/patologiaRESUMO
The proximal promoter of lck directs gene expression exclusively in T cells. To investigate the developmental regulation of the lck proximal promoter activity and its relationship to T cell lineage commitment, a green fluorescence protein (GFP) transgenic (Tg) mouse in which the GFP expression is under the control of the proximal promoter of lck was created. In the adult GFP-Tg mice, >90% of CD4(+)CD8(+) and CD4(+)CD8(-) thymocytes, and the majority of CD4(-)CD8(+) and CD4(-)CD8(-) [double-negative (DN)] thymocytes were highly positive for GFP. Slightly lower but substantial levels of expression of GFP was also observed in mature splenic T cells. No GFP(+) cells was detected in non-T lineage subsets, including mature and immature B cells, CD5(+) B cells, and NK cells, indicating a preserved tissue specificity of the promoter. The earliest GFP(+) cells detected were found in the CD44(+)CD25(-) DN thymocyte subpopulation. The developmental potential of GFP(-) and GFP(+) cells in the CD44(+)CD25(-) DN fraction was examined using in vitro culture systems. The generation of substantial numbers of alphabeta and gammadelta T cells as well as NK cells was demonstrated from both GFP(-) and GFP(+) cells. However, no development of B cells or dendritic cells was detected from GFP(+) CD44(+)CD25(-) DN thymocytes. These results suggest that the progenitors expressing lck proximal promoter activity in the CD44(+)CD25(-) DN thymocyte subset have lost most of the progenitor potential for the B and dendritic cell lineage. Thus, progression of T cell lineage restriction in the earliest thymic population can be visualized by lck proximal promoter activity, suggesting a potential role of Lck in the T cell lineage commitment.
Assuntos
Proteína Tirosina Quinase p56(lck) Linfócito-Específica/biossíntese , Proteína Tirosina Quinase p56(lck) Linfócito-Específica/genética , Regiões Promotoras Genéticas/imunologia , Subpopulações de Linfócitos T/enzimologia , Subpopulações de Linfócitos T/imunologia , Timo/citologia , Timo/enzimologia , Animais , Linfócitos B/citologia , Diferenciação Celular/genética , Diferenciação Celular/imunologia , Linhagem da Célula/genética , Linhagem da Célula/imunologia , Células Cultivadas , Células Dendríticas/citologia , Regulação da Expressão Gênica/imunologia , Proteínas de Fluorescência Verde , Receptores de Hialuronatos/biossíntese , Células Matadoras Naturais/citologia , Células Matadoras Naturais/metabolismo , Proteínas Luminescentes/biossíntese , Proteínas Luminescentes/genética , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Microscopia Confocal , Receptores de Antígenos de Linfócitos T alfa-beta/biossíntese , Receptores de Antígenos de Linfócitos T gama-delta/biossíntese , Receptores de Interleucina-2/biossíntese , Cifozoários , Baço/imunologia , Baço/metabolismo , Subpopulações de Linfócitos T/citologia , Subpopulações de Linfócitos T/metabolismo , Timo/crescimento & desenvolvimento , Timo/imunologiaRESUMO
Virchow-Robin's perivascular spaces lie between the basement membrane around pericytes and the basement membrane at the surface of the glia limitans of the brain vessels. They are directly connected to the subpial space and harbour a population of cells distinct from pericytes, perivascular microglia and other cells within perivascular spaces (e.g. T cells and mast cells) in their ability to quickly phagocytose particles from the cerebrospinal fluid (CSF). Morphology, function, and cell surface proteins of these perivascular cells suggest an origin from the monocyte/macrophage lineage. It is currently unclear to what extent these brain perivascular cells represent a resident population of histiocytes or undergo continuous supplementation from blood monocytes. Using transplants of green-fluorescent-protein (GFP)-transfected bone marrow cells, we therefore investigated the replacement of perivascular cells by blood-borne macrophages in adult mice. GFP-positive cells in the perivascular spaces were found as early as 2 weeks post transplantation. The substitution of host perivascular cells by donor-derived macrophages was then evaluated using immunocytochemistry and intraventricular injection of hydrophilic rhodamine-fluorescent tracers. Such tracers diffuse along perivascular spaces and are subsequently phagocytosed by perivascular cells leading to stable phagocytosis-dependent labelling. Thus, the population of newly immigrated macrophages could be related to the total number of perivascular macrophages. This approach revealed a continuous increase of donor-derived perivascular cells. At 14 weeks post transplantation, all perivascular cells were donor-derived. These data show that brain perivascular cells are a population of migratory macrophages and not resident histiocytes.