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This corrects the article DOI: 10.1038/mi.2017.61.
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Atopic dermatitis (AD) often precedes asthma and food allergy, indicating that epicutaneous sensitization to allergens may be important in the induction of allergic responses at other barrier surfaces. Thymic stromal lymphopoietin (TSLP) and interleukin (IL)-33 are two cytokines that may drive type 2 responses in the skin; both are potential targets in the treatment of allergic diseases. We tested the functional role of IL-33 and the interplay between IL-33 and TSLP in mouse models of atopic march and gastrointestinal (GI) allergy. IL-33-driven allergic disease occurred in a TSLP-independent manner. In contrast, mice lacking IL-33 signaling were protected from onset of allergic diarrhea in TSLP-driven disease. Epithelial-derived IL-33 was important in this model, as specific loss of IL-33 expression in the epithelium attenuated cutaneous inflammation. Notably, the development of diarrhea following sensitization with TLSP plus antigen was ameliorated even when IL-33 was blocked after sensitization. Thus, IL-33 has an important role during early cutaneous inflammation and during challenge. These data reveal critical roles for IL-33 in the "atopic march" that leads from AD to GI allergy.
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Dermatitis Atópica/inmunología , Hipersensibilidad a los Alimentos/inmunología , Tracto Gastrointestinal/inmunología , Interleucina-33/metabolismo , Piel/inmunología , Animales , Anticuerpos Neutralizantes/administración & dosificación , Células Cultivadas , Citocinas/metabolismo , Modelos Animales de Enfermedad , Humanos , Inmunoglobulinas/genética , Interleucina-33/inmunología , Ratones , Ratones Noqueados , Receptores de Citocinas/genética , Células Th2/inmunología , Linfopoyetina del Estroma TímicoRESUMEN
Airway epithelial cells are among the first to encounter inhaled allergens and can initiate allergic responses by producing pro-Th2 innate cytokines. In this study, we investigated the role of epithelial-derived cytokines in sensitization to a clinically relevant allergen, cockroach allergen (CRA). Among the epithelial-derived cytokines, granulocyte macrophage colony-stimulating factor (GM-CSF) had a central role in the initiation of Th2 allergic responses to CRA. We show that initial exposure to CRA directly activated airway epithelial cells through a TLR4-MyD88-dependent pathway and MyD88 signaling in epithelial cells induced upregulation of GM-CSF during sensitization. Epithelial-derived GM-CSF was required for allergic sensitization and selectively restored Th2 responses in the absence of MyD88. Thus, we demonstrate that epithelial-derived GM-CSF is a critical early signal during allergic sensitization to CRA.
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Factor Estimulante de Colonias de Granulocitos y Macrófagos/metabolismo , Hipersensibilidad/inmunología , Pulmón/inmunología , Mucosa Respiratoria/inmunología , Células Th2/inmunología , Alérgenos/inmunología , Animales , Células Cultivadas , Cucarachas/inmunología , Inmunización , Proteínas de Insectos/inmunología , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Factor 88 de Diferenciación Mieloide/genética , Factor 88 de Diferenciación Mieloide/metabolismo , Transducción de Señal , Receptor Toll-Like 4/genética , Receptor Toll-Like 4/metabolismoRESUMEN
Thymic stromal lymphopoietin (TSLP) is an interleukin-7 (IL-7)-like cytokine involved in T helper 2 type immune responses. The primary target of TSLP is myeloid dendritic cells (DCs), however, little is known about the mechanism by which TSLP elicits respiratory IgA immune responses upon mucosal immunization. Here, we found that the levels of TSLP and TSLPR were upregulated in the mucosal DCs of mice nasally immunized with pneumococcal surface protein A (PspA) plus cholera toxin (CT) compared with those immunized with PspA alone. PspA-specific IgA responses, but not IgG Ab responses were significantly reduced in both serum and mucosal secretions of TSLPR knockout mice compared with wild-type mice after nasal immunization with PspA plus CT. Furthermore, CD11c+ mucosal DCs isolated from TSLPR knockout mice nasally immunized with PspA plus CT were less activated and exhibited markedly reduced expression of IgA-enhancing cytokines (e.g., APRIL, BAFF, and IL-6) compared with those from equivalently immunized wild-type mice. Finally, exogenous TSLP promoted production of IgAs in an in vitro DC-B cell co-culture system as exhibited by enhanced IL-6 production. These results suggest that TSLP-TSLPR signaling is pivotal in the induction of nasal respiratory immunity against pathogenic pneumococcal infection.
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Linfocitos B/inmunología , Proteínas Bacterianas/inmunología , Toxina del Cólera/inmunología , Citocinas/metabolismo , Células Dendríticas/inmunología , Inmunoglobulinas/metabolismo , Receptores de Citocinas/metabolismo , Mucosa Respiratoria/patología , Administración Intranasal , Animales , Anticuerpos Antibacterianos/metabolismo , Antígeno CD11c/metabolismo , Células Cultivadas , Técnicas de Cocultivo , Inmunidad Humoral , Inmunización , Inmunoglobulina A/metabolismo , Inmunoglobulinas/genética , Ratones , Ratones Endogámicos BALB C , Ratones Noqueados , Receptores de Citocinas/genética , Linfopoyetina del Estroma TímicoRESUMEN
Individuals with one atopic disease are far more likely to develop a second. Approximately half of all atopic dermatitis (AD) patients subsequently develop asthma, particularly those with severe AD. This association, suggesting a role for AD as an entry point for subsequent allergic disease, is a phenomenon known as the "atopic march." Although the underlying cause of the atopic march remains unknown, recent evidence suggests a role for the cytokine thymic stromal lymphopoietin (TSLP). We have established a mouse model to determine whether TSLP plays a role in this phenomenon, and in this study show that mice exposed to the antigen ovalbumin (OVA) in the skin in the presence of TSLP develop severe airway inflammation when later challenged with the same antigen in the lung. Interestingly, neither TSLP production in the lung nor circulating TSLP is required to aggravate the asthma that was induced upon subsequent antigen challenge. However, CD4 T cells are required in the challenge phase of the response, as was challenge with the sensitizing antigen, demonstrating that the response was antigen specific. This study, which provides a clean mouse model to study human atopic march, indicates that skin-derived TSLP may represent an important factor that triggers progression from AD to asthma.
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Asma/inmunología , Linfocitos T CD4-Positivos/inmunología , Citocinas/inmunología , Dermatitis Atópica/inmunología , Neumonía/inmunología , Alérgenos/administración & dosificación , Alérgenos/inmunología , Animales , Asma/complicaciones , Células Cultivadas , Citocinas/administración & dosificación , Dermatitis Atópica/complicaciones , Modelos Animales de Enfermedad , Humanos , Inmunización , Inyecciones Intradérmicas , Pulmón/inmunología , Ratones , Ratones Endogámicos BALB C , Ratones Transgénicos , Ovalbúmina/administración & dosificación , Ovalbúmina/inmunología , Piel/inmunología , Linfopoyetina del Estroma TímicoRESUMEN
Exposure to allergens first occurs at body surfaces in direct contact with the environment such as the skin, airways, and gastrointestinal tract, and compelling evidence suggests that allergic inflammatory responses are profoundly influenced by the products of epithelial cells located at these sites. One such product is thymic stromal lymphopoietin (TSLP), which is capable of affecting multiple cell lineages involved in allergic reactions. In this review we discuss recent work that has provided insight into the role TSLP plays in both aberrant and protective allergic inflammatory responses, as well as regulation, associations with disease, sources, and functions of this important cytokine.
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Citocinas/fisiología , Hipersensibilidad , Animales , Citocinas/genética , Humanos , Hipersensibilidad/fisiopatología , Células Asesinas Naturales/inmunología , Modelos Biológicos , Especificidad de la Especie , Linfopoyetina del Estroma TímicoRESUMEN
Long-term survival after lung transplantation is limited by acute and chronic graft rejection. Induction of immune tolerance by first establishing mixed hematopoietic chimerism (MC) is a promising strategy to improve outcomes. In a preclinical canine model, stable MC was established in recipients after reduced-intensity conditioning and hematopoietic cell transplantation from a DLA-identical donor. Delayed lung transplantation was performed from the stem cell donor without pharmacological immunosuppression. Lung graft survival without loss of function was prolonged in chimeric (n = 5) vs. nonchimeric (n = 7) recipients (p < or = 0.05, Fisher's test). There were histological changes consistent with low-grade rejection in 3/5 of the lung grafts in chimeric recipients at > or =1 year. Chimeric recipients after lung transplantation had a normal immune response to a T-dependent antigen. Compared to normal dogs, there were significant increases of CD4+INFgamma+, CD4+IL-4+ and CD8+ INFgamma+ T-cell subsets in the blood (p < 0.0001 for each of the three T-cell subsets). Markers for regulatory T-cell subsets including foxP3, IL10 and TGFbeta were also increased in CD3+ T cells from the blood and peripheral tissues of chimeric recipients after lung transplantation. Establishing MC is immunomodulatory and observed changes were consistent with activation of both the effector and regulatory immune response.
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Trasplante de Pulmón/inmunología , Animales , Perros , Citometría de Flujo , Rechazo de Injerto/inmunología , Rechazo de Injerto/patología , Supervivencia de Injerto/inmunología , Supervivencia de Injerto/fisiología , Hematopoyesis , Trasplante de Células Madre Hematopoyéticas , Inmunosupresores/uso terapéutico , Trasplante de Pulmón/fisiología , Modelos Animales , Pruebas de Función Respiratoria , Subgrupos de Linfocitos T/inmunología , Quimera por Trasplante , Trasplante HomólogoRESUMEN
INTRODUCTION: Cardiopulmonary bypass (CPB) induces an inflammatory response believed to contribute to postoperative morbidity. We hypothesized that the magnitude of the inflammatory response following CPB would be associated with adverse clinical outcomes. METHODS: Twenty-nine patients had plasma TNF, IL-6, IL-8, elastase, histamine, complement C5a, and complement C3a measured by ELISA before, during, and after cardiac operations employing CPB. Inflammatory mediator levels were analyzed with respect to outcomes. RESULTS: Mediator levels peaked at 4 h post-CPB and either returned to baseline or substantially decreased by 24 h. Patients with peak mediator levels above the median for the group as a whole were classified as 'hyper-responders'; those with levels below the median were classified as 'normal responders'. While IL-8, C3a, and IL-6 levels were independently associated with adverse outcomes, TNF, histamine, and C5a levels were not. Elastase levels trended towards adverse outcomes. IL-8 'hyper-responders' experienced significantly greater postoperative weight gain and had higher IL-8 levels at 24 h (p<0.05), with trends towards renal impairment and protracted supplemental oxygen requirements. C3a 'hyper-responders' strongly trended towards increased bleeding, delayed extubation, greater postoperative weight gain, and decreased levels of independent functioning at discharge (p < or = 0.10). IL-6 'hyper-responders' experienced significantly more postoperative bleeding, delayed extubation, and higher IL-6 levels at 24 h compared to 'normal responders' (p < 0.05). They strongly trended towards greater postoperative weight gain and decreased levels of independent functioning at discharge (p < or = 0.10). CONCLUSIONS: Patients who have an exaggerated inflammatory response to CPB tend to bleed more, require more respiratory support, demonstrate greater capillary leak via weight gain, and display a decline in independent functioning relative to normal responders. Thus, it appears that the magnitude of the inflammatory response to CPB adversely influences clinical outcomes.
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Puente Cardiopulmonar/efectos adversos , Inflamación/etiología , Inflamación/patología , Complicaciones Posoperatorias/patología , Anciano , Biomarcadores/sangre , Complemento C3a/metabolismo , Ensayo de Inmunoadsorción Enzimática , Femenino , Cardiopatías/etiología , Cardiopatías/patología , Humanos , Mediadores de Inflamación/sangre , Interleucina-6/sangre , Interleucina-8/sangre , Enfermedades Renales/etiología , Enfermedades Renales/patología , Enfermedades Pulmonares/etiología , Enfermedades Pulmonares/patología , Masculino , Persona de Mediana Edad , Elastasa Pancreática/sangre , Resultado del TratamientoRESUMEN
In the absence of the recently identified putative transcription factor scurfin, mice develop a lymphoproliferative disorder resulting in death by 3 wk of age from a pathology that resembles TGF-beta or CTLA-4 knockout mice. In this report, we characterize mice that overexpress the scurfin protein and demonstrate that these animals have a dramatically depressed immune system. Mice transgenic for the Foxp3 gene (which encodes the scurfin protein) have fewer T cells than their littermate controls, and those T cells that remain have poor proliferative and cytolytic responses and make little IL-2 after stimulation through the TCR. Although thymic development appears normal in these mice, peripheral lymphoid organs, particularly lymph nodes, are relatively acellular. In a separate transgenic line, forced expression of the gene specifically in the thymus can alter thymic development; however, this does not appear to affect peripheral T cells and is unable to prevent disease in mice lacking a functional Foxp3 gene, indicating that the scurfin protein acts on peripheral T cells. The data indicate a critical role for the Foxp3 gene product in the function of the immune system, with both the number and functionality of peripheral T cells under the aegis of the scurfin protein.
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Proteínas de Unión al ADN/biosíntesis , Proteínas de Unión al ADN/genética , Activación de Linfocitos/genética , Subgrupos de Linfocitos T/inmunología , Subgrupos de Linfocitos T/patología , Animales , Linfocitos T CD4-Positivos/inmunología , Linfocitos T CD4-Positivos/metabolismo , Linfocitos T CD4-Positivos/patología , Linfocitos T CD8-positivos/inmunología , Linfocitos T CD8-positivos/metabolismo , Linfocitos T CD8-positivos/patología , Células Cultivadas , Proteínas de Unión al ADN/fisiología , Factores de Transcripción Forkhead , Regulación de la Expresión Génica/inmunología , Histocitoquímica , Inmunofenotipificación , Activación de Linfocitos/inmunología , Recuento de Linfocitos , Prueba de Cultivo Mixto de Linfocitos , Linfopenia/genética , Linfopenia/inmunología , Linfopenia/patología , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Ratones Mutantes , Ratones Transgénicos , Subgrupos de Linfocitos T/metabolismo , Timo/inmunología , Timo/metabolismo , Timo/patología , Transgenes/inmunologíaRESUMEN
We have recently identified and cloned Foxp3, the gene defective in mice with the scurfy mutation. The immune dysregulation documented in these mice and in humans with mutations in the orthologous gene indicates that the foxp3 gene product, scurfin, is involved in the regulation of T cell activation and differentiation. The autoimmune state observed in these patients with the immune dysregulation polyendocrinopathy, enteropathy, X-linked syndrome, or X-linked autoimmunity-allergic dysregulation syndrome also points to a critical role for scurfin in the regulation of T cell homeostasis. FOXP3 encodes a novel member of the forkhead family of transcription factors. Here we demonstrate that this structural domain is required for nuclear localization and DNA binding. Scurfin, transiently expressed in heterologous cells, represses transcription of a reporter containing a multimeric forkhead binding site. Upon overexpression in CD4 T cells, scurfin attenuates activation-induced cytokine production and proliferation. We have identified FKH binding sequences adjacent to critical NFAT regulatory sites in the promoters of several cytokine genes whose expression is sensitive to changes in SFN abundance. Our findings indicate that the ability of scurfin to bind DNA, and presumably repress transcription, plays a paramount role in determining the amplitude of the response of CD4 T cells to activation.
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Linfocitos T CD4-Positivos/efectos de los fármacos , Proteínas de Unión al ADN/farmacología , Activación de Linfocitos/efectos de los fármacos , Proteínas Nucleares , Transcripción Genética/efectos de los fármacos , Animales , Linfocitos T CD4-Positivos/fisiología , Células COS , Células Cultivadas , Citocinas/biosíntesis , Citocinas/metabolismo , ADN/efectos de los fármacos , ADN/metabolismo , Proteínas de Unión al ADN/genética , Proteínas de Unión al ADN/fisiología , Factores de Transcripción Forkhead , Silenciador del Gen/efectos de los fármacos , Silenciador del Gen/fisiología , Humanos , Activación de Linfocitos/fisiología , Mutación , Factores de Transcripción NFATC , Factores de Transcripción/fisiología , Transcripción Genética/fisiología , TransfecciónRESUMEN
Scurfy (sf) is an X-linked recessive mouse mutant resulting in lethality in hemizygous males 16-25 days after birth, and is characterized by overproliferation of CD4+CD8- T lymphocytes, extensive multiorgan infiltration and elevation of numerous cytokines. Similar to animals that lack expression of either Ctla-4 or Tgf-beta, the pathology observed in sf mice seems to result from an inability to properly regulate CD4+CD8- T-cell activity. Here we identify the gene defective in sf mice by combining high-resolution genetic and physical mapping with large-scale sequence analysis. The protein encoded by this gene (designated Foxp3) is a new member of the forkhead/winged-helix family of transcriptional regulators and is highly conserved in humans. In sf mice, a frameshift mutation results in a product lacking the forkhead domain. Genetic complementation demonstrates that the protein product of Foxp3, scurfin, is essential for normal immune homeostasis.
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Proteínas de Unión al ADN/química , Proteínas de Unión al ADN/metabolismo , Genes Esenciales/genética , Trastornos Linfoproliferativos/genética , Mutación/genética , Secuencias de Aminoácidos , Secuencia de Aminoácidos , Animales , Clonación Molecular , Secuencia Conservada , Análisis Mutacional de ADN , Proteínas de Unión al ADN/genética , Femenino , Factores de Transcripción Forkhead , Perfilación de la Expresión Génica , Genes Recesivos/genética , Prueba de Complementación Genética , Humanos , Ganglios Linfáticos/inmunología , Ganglios Linfáticos/patología , Recuento de Linfocitos , Trastornos Linfoproliferativos/inmunología , Trastornos Linfoproliferativos/patología , Masculino , Ratones , Ratones Mutantes , Ratones Transgénicos , Datos de Secuencia Molecular , Fenotipo , Mapeo Físico de Cromosoma , Estructura Terciaria de Proteína , ARN Mensajero/análisis , ARN Mensajero/genética , Alineación de SecuenciaRESUMEN
The cellular receptor for murine thymic stromal lymphopoietin (TSLP) was detected in a variety of murine, but not human myelomonocytic cell lines by radioligand binding. cDNA clones encoding the receptor were isolated from a murine T helper cell cDNA library. TSLP receptor (TSLPR) is a member of the hematopoietin receptor family. Transfection of TSLPR cDNA resulted in only low affinity binding. Cotransfection of the interleukin 7 (IL-7)Ralpha chain cDNA resulted in conversion to high affinity binding. TSLP did not activate cells from IL-7Ralpha(-/)- mice, but did activate cells from gammac(-/)- mice. Thus, the functional TSLPR requires the IL-7Ralpha chain, but not the gammac chain for signaling.
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Hematopoyesis/efectos de los fármacos , Linfocitos/efectos de los fármacos , Receptores de Citocinas/fisiología , Receptores de Interleucina-7/fisiología , Secuencia de Aminoácidos , Animales , Secuencia de Bases , Línea Celular , Mapeo Cromosómico , Clonación Molecular , Citocinas/farmacología , Humanos , Interleucina-7/farmacología , Linfocitos/fisiología , Masculino , Ratones , Datos de Secuencia Molecular , ARN Mensajero/análisis , Receptores de Citocinas/química , Receptores de Citocinas/genética , Receptores de Interleucina-7/química , Proteínas Recombinantes/química , Transducción de SeñalRESUMEN
Signaling by type I cytokines involves the formation of receptor homodimers, heterodimers or higher order receptor oligomers. Here we report the cloning of a type I cytokine receptor subunit that is most closely related to the common cytokine receptor gamma chain (gamma c). Binding and crosslinking experiments demonstrate that this protein is the receptor for a recently described interleukin 7 (IL-7)-like factor, thymic stromal lymphopoietin (TSLP). Binding of TSLP to the thymic stromal lymphopoietin receptor (TSLPR) is increased markedly in the presence of the IL-7 receptor alpha chain (IL-7R alpha). IL-7R alpha-expressing but not parental 32D cells proliferate in the presence of exogenous TSLP. Moreover, a combination of IL-7R alpha and TSLPR is required for TSLP-dependent activation of a STAT5-dependent reporter construct. Thus it is shown that IL-7R alpha is a component of both the IL-7 and TSLP receptors, which helps to explain why deletion of the gene that encodes IL-7R alpha affects the lymphoid system more severely than deletion of the gene encoding IL-7 does. Cloning of TSLPR should facilitate an understanding of TSLP function and its signaling mechanism.
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Interleucina-7/inmunología , Receptores de Interleucina-7/genética , Receptores de Interleucina-7/inmunología , Transducción de Señal/inmunología , Timo/inmunología , Secuencia de Aminoácidos , Clonación Molecular , Datos de Secuencia Molecular , Transducción de Señal/genéticaRESUMEN
Thymic stromal lymphopoietin (TSLP) is a newly identified cytokine that uniquely promotes B lymphopoiesis to the B220+/IgM+ immature B cell stage. In addition, TSLP shares many biological properties with the related cytokine IL-7. This can be explained by the finding that the receptor complexes for TSLP and IL-7 both contain the IL-7R alpha-chain; IL-7Ralpha is paired with the common gamma-chain (gammac) in the IL-7 receptor complex and the unique TSLP-R chain in the TSLP receptor complex. Although TSLP and IL-7 both induce tyrosine phosphorylation of the transcription factor Stat5, only IL-7-mediated signal transduction could be associated with activation of Janus family kinases (Jaks). Because Stat5 phosphorylation following cytokine stimulation is generally mediated by Jaks, the lack of Jak activation after TSLP treatment suggested the possibility that tyrosine-phosphorylated Stat5 may be nonfunctional. Herein, we demonstrate that TSLP induces a functional Stat5 transcription factor in that TSLP stimulation results in Stat5-DNA complex formation and transcription of the Stat5-responsive gene CIS. We also show that the TSLP receptor complex is functionally reconstituted using TSLP-R and IL-7Ralpha and that TSLP-mediated signal transduction requires Stat5. Moreover, TSLP-mediated signaling is inhibited by suppressor of cytokine signaling (SOCS)-1 and a kinase-deficient version of Tec but not by kinase-deficient forms of Jak1 and Jak2.
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Linfocitos B/inmunología , Citocinas/metabolismo , Proteínas de Unión al ADN/metabolismo , Proteínas Inmediatas-Precoces/metabolismo , Proteínas de la Leche , Transactivadores/metabolismo , Subgrupos de Linfocitos B/inmunología , Oncostatina M , Péptidos , Unión Proteica , Proteínas Tirosina Quinasas/metabolismo , Receptores de Citocinas/metabolismo , Factor de Transcripción STAT5 , Transducción de Señal , Proteínas Supresoras de la Señalización de Citocinas , Proteínas Supresoras de Tumor , Linfopoyetina del Estroma TímicoRESUMEN
Mice hemizygous (Xsf/Y) for the X-linked mutation scurfy (sf) develop a severe and rapidly fatal lymphoproliferative disease mediated by CD4+CD8- T lymphocytes. We have undertaken phenotypic and functional studies to more accurately identify the immunologic pathway(s) affected by this important mutation. Flow cytometric analyses of lymphoid cell populations reveal that scurfy syndrome is characterized by changes in several phenotypic parameters, including an increase in Mac-1+ cells and a decrease in B220+ cells, changes that may result from the production of extremely high levels of the cytokine granulocyte-macrophage CSF by scurfy T cells. Scurfy T cells also exhibit strong up-regulation of cell surface Ags indicative of in vivo activation, including CD69, CD25, CD80, and CD86. Both scurfy and normal T cells are responsive to two distinct signals provided by the TCR and by ligation of CD28; scurfy cells, however, are hyperresponsive to TCR ligation and exhibit a decreased requirement for costimulation through CD28 relative to normal controls. This hypersensitivity may result, in part, from increased costimulation through B7-1 and B7-2, whose expression is up-regulated on scurfy T cells. Although the specific defect leading to this hyperactivation has not been identified, we also demonstrate that scurfy T cells are less sensitive than normal controls to inhibitors of tyrosine kinases such as genistein and herbimycin A, and the immunosuppressant cyclosporin A. One interpretation of our data would suggest that the scurfy mutation results in a defect, which interferes with the normal down-regulation of T cell activation.
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Inmunoconjugados , Trastornos Linfoproliferativos/genética , Linfocitos T/inmunología , Abatacept , Animales , Antígenos CD , Antígenos de Diferenciación/análisis , Antígeno B7-1/análisis , Benzoquinonas , Antígeno CTLA-4 , Ciclosporina/farmacología , Femenino , Genisteína/farmacología , Factor Estimulante de Colonias de Granulocitos y Macrófagos/biosíntesis , Lactamas Macrocíclicas , Antígenos Comunes de Leucocito/análisis , Activación de Linfocitos , Trastornos Linfoproliferativos/inmunología , Masculino , Ratones , Ratones Endogámicos C3H , Ratones Mutantes , Proteínas Nucleares/análisis , Quinonas/farmacología , Receptores de Antígenos de Linfocitos T/fisiología , Rifabutina/análogos & derivados , Factores de Transcripción/análisisRESUMEN
A novel cytokine from a thymic stromal cell line (thymic stromal lymphopoietin (TSLP)) promotes the development of B220+/IgM+ immature B cells when added to fetal liver cultures, long term bone marrow cultures, or bone marrow cells plated in semisolid medium. Because the activities of TSLP overlap with those of IL-7 in some in vitro assays, we compared the signaling mechanisms employed by TSLP and IL-7. Proliferation of a factor-dependent pre-B cell line (NAG8/7) in response to either TSLP or IL-7 was inhibited by anti-IL-7R alpha mAbs, suggesting that the functional TSLP receptor complex uses IL-7R alpha. In contrast, three different Abs to the common cytokine receptor gamma-chain had no effect on the response of these cells to TSLP, indicating that the functional TSLP receptor complex does not use the common cytokine receptor gamma-chain. Both cytokines induced activation of Stat5, but only IL-7 induced activation of the Janus family kinases Jak1 and Jak3. In fact, TSLP failed to activate any of the four known Janus family kinases, suggesting that Stat5 phosphorylation is mediated by a novel mechanism. Taken together, these data support the idea that TSLP can make unique contributions to B lymphopoiesis and indicate that it does so by mechanisms distinct from IL-7.
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Subgrupos de Linfocitos B/citología , Subgrupos de Linfocitos B/inmunología , Citocinas/fisiología , Inmunoglobulina M/biosíntesis , Interleucina-7/fisiología , Proteínas de la Leche , Proteínas Proto-Oncogénicas , Transducción de Señal/inmunología , Timo/metabolismo , Animales , Subgrupos de Linfocitos B/metabolismo , Células de la Médula Ósea/citología , Diferenciación Celular/inmunología , Línea Celular , Células Cultivadas , Ensayo de Unidades Formadoras de Colonias , Proteínas de Unión al ADN/metabolismo , Activación Enzimática/inmunología , Feto , Janus Quinasa 1 , Janus Quinasa 2 , Janus Quinasa 3 , Hígado/citología , Ratones , Fosforilación , Proteínas Tirosina Quinasas/metabolismo , Receptores de Interleucina-7/química , Receptores de Interleucina-7/fisiología , Factor de Transcripción STAT5 , Células del Estroma/metabolismo , Timo/citología , Factores de Tiempo , Transactivadores/metabolismo , Tirosina/metabolismo , Linfopoyetina del Estroma TímicoRESUMEN
Severe combined immunodeficiency (SCID) is caused by multiple genetic defects. The most common form of SCID, X-linked SCID (XSCID), results from mutations in IL2RG (ref. 4), which encodes the common cytokine receptor gamma chain (gamma(c)) that is shared by the IL-2, IL-4, IL-7, IL-9 and IL-15 receptors. In XSCID and SCID resulting from mutations in JAK3, which encodes a Janus family tyrosine kinase that couples to gamma(c) and is required for gamma(c)-dependent signalling, T- and natural killer (NK)-cells are decreased but B-cell numbers are normal (T(-)B(+)NK(-)SCID). Some SCID patients lack T cells but retain NK cells. Given diminished T-cell development in Il7- or Il7r-deficient mice and that Il/7r-deficient mice have NK cells, we hypothesized that T(-)B(+)NK(+) SCID might result from defective IL-7 signalling, although apparent differences in the role of the IL-7/IL-7R pathway in humans and mice in T-cell and B-cell development have been suggested. We now demonstrate that defective IL7R expression causes T(-)B(+)NK(+) SCID, indicating that the T-cell, but not the NK-cell, defect in XSCID results from inactivation of IL-7Ralpha signalling.
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Receptores de Interleucina-7/genética , Inmunodeficiencia Combinada Grave/genética , Animales , Linfocitos B/inmunología , Secuencia de Bases , Cartilla de ADN , Humanos , Lactante , Recién Nacido , Células Asesinas Naturales/inmunología , Ratones , Datos de Secuencia Molecular , Mutagénesis Sitio-Dirigida , Reacción en Cadena de la Polimerasa , Receptores de Interleucina-7/metabolismo , Transducción de Señal/genética , Linfocitos T/inmunologíaRESUMEN
A cDNA clone encoding a human homolog of the Caenorhabditis elegans unc-18 gene was identified following random sequencing of clones from IL-2-activated human NK cells. This cDNA clone is related to the nonneuronal Munc-18b and so has been called Hunc-18b. The Hunc-18b transcripts were found in most human tissues, with the exception of brain and skeletal muscle, and in cells from all lymphoid lineages. The Hunc-18b gene was localized to human chromosome 19p13.2-p13.3, and the mouse homolog, Munc-18b, was mapped to the proximal region of mouse Chromosome 8.
Asunto(s)
Mapeo Cromosómico , Cromosomas Humanos Par 19 , Proteínas del Tejido Nervioso/genética , Proteínas de Transporte Vesicular , Secuencia de Aminoácidos , Animales , Clonación Molecular , ADN Complementario , Humanos , Hibridación Fluorescente in Situ , Ratones , Datos de Secuencia Molecular , Proteínas Munc18 , Homología de Secuencia de AminoácidoRESUMEN
Binding of granulocyte colony-stimulating factor (G-CSF) to normal myeloid cells activates the protein tyrosine kinases Lyn and Syk and results in the immediate early upregulation of G-CSF receptor (R) mRNA. In our studies of the signaling pathways activated by G-CSF that are coupled to proliferation and differentiation of myeloid cells, we examined whether G-CSF activated a latent transcription factor belonging to the STAT protein family. Electrophoretic mobility shift assays (EMSAs) of nuclear extracts from G-CSF-stimulated human myeloid cells showed the rapid activation of a DNA-binding protein that bound to the high-affinity serum-inducible element (hSIE) and migrated with mobility similar to serum inducible factor (SIF)-A (Stat3 homodimer). The G-CSF-stimulated SIF-A complex (G-SIF-A) did not bind to duplex oligonucleotides used to purify and characterize other Stat proteins (Stat1-6). In addition, antibodies raised against Stat1-6 failed to supershift the G-SIF-A complex or interfere with its formation. Based on its binding to the hSIE and lack of antigenic cross-reactivity with other known STAT proteins that bind to this element, it is likely that G-SIF-A is composed of a distinct member of the STAT protein family. EMSAs of whole-cell extracts prepared from cell lines containing full-length and truncated mutants of the G-CSFR showed that activation of G-SIF-A did not correlate with proliferation; rather, optimal activation requires the distal half of the cytosolic domain of the G-CSFR that is essential for differentiation. Activation of G-SIF-A, therefore, may be an early G-CSFR-coupled event that is critical for myeloid maturation.
Asunto(s)
Linfocitos B/efectos de los fármacos , Proteínas de Unión al ADN/metabolismo , Regulación de la Expresión Génica/efectos de los fármacos , Factor Estimulante de Colonias de Granulocitos/farmacología , Células Madre Hematopoyéticas/efectos de los fármacos , Neutrófilos/efectos de los fármacos , Transducción de Señal/fisiología , Transactivadores/metabolismo , Secuencia de Bases , Diferenciación Celular/efectos de los fármacos , Línea Celular , Células Cultivadas , Proteínas de Unión al ADN/aislamiento & purificación , Precursores Enzimáticos/metabolismo , Humanos , Interferón gamma/farmacología , Péptidos y Proteínas de Señalización Intracelular , Sustancias Macromoleculares , Datos de Secuencia Molecular , Unión Proteica , Proteínas Tirosina Quinasas/metabolismo , Proteínas Recombinantes/farmacología , Quinasa Syk , Transactivadores/aislamiento & purificación , Transcripción Genética/efectos de los fármacos , Familia-src Quinasas/metabolismoRESUMEN
The protein tyrosine kinases JAK1 and JAK2 are phosphorylated tyrosine after the interaction of granulocyte colony-stimulating factor (G-CSF) with its transmembrane receptor. So too is Stat3, a member of the STAT family of transcriptional activators thought to be activated by the JAK kinases. Truncated G-CSF receptor (G-CSF-R) mutants were used to determine the different regions of the cytoplasmic domain necessary for tyrosine phosphorylation of the signaling molecules JAK2, Stat3, and p42, p44MAPK. We have shown that G-CSF-induced tyrosine phosphorylation and kinase activation of JAK2 requires the membrane proximal 57 amino acids of the cytoplasmic domain. In contrast, maximal Stat3 tyrosine phosphorylation required amino acids 96 to 183 of the G-CSF-R cytoplasmic domain, Stat3 DNA binding could occur with a receptor truncated 96 amino acids from the transmembrane domain and containing a single tyrosine residue, but was reduced in comparison with the full-length receptor. Together with the tyrosine phosphorylation of Stat3, this finding suggests that additional Stat3 does not appear to be required for proliferation. MAP kinase tyrosine phosphorylation correlated with both the proliferative response and JAK2 activation.