RESUMEN
Mast cell activation involves the rapid release of inflammatory mediators, including histamine, from intracellular granules. The cells are capable of regranulation and multiple rounds of activation. The goal of this study was to determine if there are changes in the content of pre-formed mast cell mediators after a round of activation. After 24 h, the histamine content of bone marrow-derived mast cells (BMMC), but not that of peritoneal mast cells, exceeded the amount in resting cells. Accumulation of histamine in BMMC peaked at 72 h of activation, and returned toward preactivation levels by 96 h. The increase in histamine content was accompanied by an increase in the gene expression of histidine decarboxylase. No increases in beta hexosaminidase or murine mast cell protease-6 were observed. These findings indicate that BMMC respond to activation by increasing total cell-associated histamine content. This increase may be important to the response of these cells upon subsequent exposure to antigens.
Asunto(s)
Diferenciación Celular , Histamina/metabolismo , Mastocitos/metabolismo , Mastocitos/fisiología , Animales , Células de la Médula Ósea/citología , Células de la Médula Ósea/fisiología , Recuento de Células , Diferenciación Celular/inmunología , Proliferación Celular , Gránulos Citoplasmáticos/metabolismo , Inducción Enzimática , Femenino , Histidina Descarboxilasa/biosíntesis , Histidina Descarboxilasa/genética , Mastocitos/citología , Mastocitos/inmunología , Ratones , Ratones Endogámicos C57BL , Cavidad Peritoneal/citologíaRESUMEN
PD-1 is an immunoinhibitory receptor expressed by activated T cells, B cells, and myeloid cells. Mice deficient in PD-1 exhibit a breakdown of peripheral tolerance and demonstrate multiple autoimmune features. We report here that the ligand of PD-1 (PD-L1) is a member of the B7 gene family. Engagement of PD-1 by PD-L1 leads to the inhibition of T cell receptor-mediated lymphocyte proliferation and cytokine secretion. In addition, PD-1 signaling can inhibit at least suboptimal levels of CD28-mediated costimulation. PD-L1 is expressed by antigen-presenting cells, including human peripheral blood monocytes stimulated with interferon gamma, and activated human and murine dendritic cells. In addition, PD-L1 is expressed in nonlymphoid tissues such as heart and lung. The relative levels of inhibitory PD-L1 and costimulatory B7-1/B7-2 signals on antigen-presenting cells may determine the extent of T cell activation and consequently the threshold between tolerance and autoimmunity. PD-L1 expression on nonlymphoid tissues and its potential interaction with PD-1 may subsequently determine the extent of immune responses at sites of inflammation.
Asunto(s)
Antígenos CD/inmunología , Antígenos de Superficie/inmunología , Antígeno B7-1/inmunología , Glicoproteínas de Membrana/inmunología , Secuencia de Aminoácidos , Animales , Células Presentadoras de Antígenos/inmunología , Antígenos CD/clasificación , Antígenos CD/genética , Antígenos de Superficie/genética , Antígenos de Superficie/metabolismo , Proteínas Reguladoras de la Apoptosis , Antígeno B7-1/clasificación , Antígeno B7-1/genética , Antígeno B7-2 , Secuencia de Bases , Antígenos CD28/inmunología , Complejo CD3/inmunología , División Celular , ADN Complementario , Expresión Génica , Humanos , Ligandos , Glicoproteínas de Membrana/clasificación , Glicoproteínas de Membrana/genética , Ratones , Datos de Secuencia Molecular , Receptor de Muerte Celular Programada 1 , Transducción de Señal/inmunología , Linfocitos T/citologíaRESUMEN
The TLS-CHOP oncoprotein, found in the majority of human myxoid liposarcomas, consists of a fusion between the transcription factor CHOP/GADD153 and the N terminus of an RNA-binding protein TLS/FUS. Clinical correlation and in vitro transformation assays indicate that the N terminus of TLS plays an important role in oncogenesis by TLS-CHOP. Until now, however, the only activity attributed to the oncoprotein is that of inhibiting the binding of transcription factors of the C/EBP class to certain adipogenic target genes, a function that TLS-CHOP shares with the nononcogenic CHOP protein. Here we report the isolation of a gene, DOL54, that is activated in primary fibroblasts by the expression of TLS-CHOP. DOL54 is expressed in the neoplastic component of human myxoid liposarcomas and increases the tumorigenicity of cells injected in nude mice. Activation of DOL54 requires an intact DNA-binding and dimerization domain in TLS-CHOP, a suitable cellular dimerization partner, and depends on the TLS N terminus. Normal adipocytic differentiation is associated with an early and transient expression of DOL54, and the gene encodes a secreted protein that is tightly associated with the cell surface or extracellular matrix. TLS-CHOP thus leads to the unscheduled expression of a gene that is normally associated with adipocytic differentiation.
Asunto(s)
Proteínas Potenciadoras de Unión a CCAAT , Regulación Neoplásica de la Expresión Génica , Liposarcoma Mixoide/genética , Proteínas de Neoplasias/genética , Proteínas Nucleares/genética , Proteínas de Fusión Oncogénica/genética , Proteína FUS de Unión a ARN , Animales , Células Cultivadas , Clonación Molecular , Proteínas de Unión al ADN/genética , Proteínas de Unión al ADN/metabolismo , Fibroblastos/metabolismo , Humanos , Liposarcoma Mixoide/metabolismo , Ratones , Datos de Secuencia Molecular , Proteínas de Neoplasias/metabolismo , Factor de Transcripción CHOPRESUMEN
Two components of a receptor complex for IL-13, the IL-4R and a low affinity IL-13-binding chain, IL-13R alpha 1, have been cloned in mice and humans. An additional high affinity binding chain for IL-13, IL-13R alpha 2, has been described in humans. We isolated a cDNA from the thymus that encodes the murine orthologue of the human IL-13R alpha 2. The predicted protein sequence of murine IL-13R alpha 2 (mIL-13R alpha 2) has 59% overall identity to human IL-13R alpha 2 and is closely related to the murine low affinity IL-13-binding subunit, IL-13R alpha 1. The genes for both mIL-13-binding chains map to the X chromosome. A specific interaction between mIL-13R alpha 2.Fc protein and IL-13 was demonstrated by surface plasmon resonance using a BIACORE instrument. Ba/F3 cells that were transfected with mIL-13R alpha 2 expressed 5000 molecules per cell and bound IL-13 with a single Kd of 0.5 to 1.2 nM. However, these cells did not proliferate in response to IL-13, and the IL-4 dose response was unaffected by high concentrations of IL-13. In contrast, the expression of mIL-13R alpha 1 by Ba/F3 cells resulted in a sensitive proliferative response to IL-13. Consistent with its lower affinity for IL-13, IL-13R alpha 1.Fc was 100-fold less effective than IL-13R alpha 2.Fc in neutralizing IL-13 in vitro. These results show that mIL-13R alpha 2 and mIL-13R alpha 1 are not functionally equivalent and predict distinct roles for each polypeptide in IL-13R complex formation and in the modulation of IL-13 signal transduction.
Asunto(s)
Interleucina-13/metabolismo , Receptores de Interleucina/química , Receptores de Interleucina/genética , Homología de Secuencia de Aminoácido , Secuencia de Aminoácidos , Animales , Linfocitos B/inmunología , Línea Celular , Mapeo Cromosómico , ADN Complementario/aislamiento & purificación , Electroforesis en Gel de Poliacrilamida , Humanos , Interleucina-13/antagonistas & inhibidores , Interleucina-13/farmacología , Subunidad alfa1 del Receptor de Interleucina-13 , Activación de Linfocitos/efectos de los fármacos , Ratones , Datos de Secuencia Molecular , Especificidad de Órganos/genética , Especificidad de Órganos/inmunología , Polimorfismo Conformacional Retorcido-Simple , Unión Proteica/inmunología , ARN Mensajero/biosíntesis , Receptores de Interleucina/aislamiento & purificación , Receptores de Interleucina/fisiología , Receptores de Interleucina-13 , Transfección/inmunologíaRESUMEN
Flt4 is a receptor protein tyrosine kinase that is expressed in the adult lymphatic endothelium and high endothelial venules. We have used a BIAcore assay to identify rodent and human cell conditioned media containing the ligand of Flt4 (Flt4-L). Receptor-based affinity chromatography was used to purify this growth factor, followed by amino acid sequencing and molecular cloning of the murine cDNA, the orthologue of human vascular endothelial growth factor-C and vascular endothelial growth factor related protein. The murine flt4-L gene was localized to chromosome 8 and demonstrated to be widely expressed. Flt4-L was found to have a hydrophobic signal sequence and a pro-peptide-like sequence that is removed to generate the mature N-terminus. In addition, the C-terminal region of Flt4-L has four repeats of a cysteine-rich motif that is presumably also proteolytically processed to generate the 21000 Mr polypeptide subunit of the Flt4-L homodimer. Recombinant Flt4-L activated Flt4 as judged by induction of tyrosyl phosphorylation, and induced mitogenesis in vitro of lymphatic endothelial cells.