RESUMEN

Introduction: Lipopolysaccharide-responsive and beige-like anchor (LRBA) is a scaffolding protein that interacts with proteins such as CTLA-4 and PKA, the importance of which has been determined in various cell types, including T regulatory cells, B cells, and renal cells. LRBA deficiency is associated with an inborn error in immunity characterized by immunodeficiency and autoimmunity. In addition to defects in T regulatory cells, patients with LRBA deficiency also exhibit B cell defects, such as reduced cell number, low memory B cells, hypogammaglobulinemia, impaired B cell proliferation, and increased autophagy. Although Lrba-/- mice do not exhibit the immunodeficiency observed in humans, responses to B cell receptors (BCR) in B cells have not been explored. Therefore, a murine model is for elucidating the mechanism of Lrba mechanism in B cells. Aim: To compare and evaluate spleen-derived B cell responses to BCR crosslinking in C57BL6 Lrba-/- and Lrba+/+ mice. Materials and methods: Spleen-derived B cells were obtained from 8 to 12-week-old mice. Subpopulations were determined by immunostaining and flow cytometry. BCR crosslinking was assessed by the F(ab')2 anti-µ chain. Activation, proliferation and viability assays were performed using flow cytometry and protein phosphorylation was evaluated by immunoblotting. The nuclear localization of p65 was determined using confocal microscopy. Nur77 expression was evaluated by Western blot. Results: Lrba-/- B cells showed an activated phenotype and a decreased proportion of transitional 1 B cells, and both proliferation and survival were affected after BCR crosslinking in the Lrba-/- mice. The NF-κB pathway exhibited a basal activation status of several components, resulting in increased activation of p50, p65, and IκBα, basal p50 activation was reduced by the Plcγ2 inhibitor U73122. BCR crosslinking in Lrba-/ - B cells resulted in poor p50 phosphorylation and p65 nuclear localization. Increased levels of Nur77 were detected. Discussion: These results indicate the importance of Lrba in controlling NF-κB activation driven by BCR. Basal activation of NF-κB could impact cellular processes, such as, activation, differentiation, proliferation, and maintenance of B cells after antigen encounter.

Asunto(s)

Linfocitos B , FN-kappa B , Animales , Ratones , Proteínas Adaptadoras Transductoras de Señales/genética , Proteínas Adaptadoras Transductoras de Señales/metabolismo , Linfocitos B/inmunología , Linfocitos B/metabolismo , Lipopolisacáridos , Activación de Linfocitos/inmunología , Ratones Endogámicos C57BL , Ratones Noqueados , FN-kappa B/metabolismo , Receptores de Antígenos de Linfocitos B/metabolismo , Transducción de Señal

RESUMEN

The engagement of B cells with surface-tethered antigens triggers the formation of an immune synapse (IS), where the local secretion of lysosomes can facilitate antigen uptake. Lysosomes intersect with other intracellular processes, such as Toll-like Receptor (TLR) signaling and autophagy coordinating immune responses. However, the crosstalk between these processes and antigen presentation remains unclear. Here, we show that TLR stimulation induces autophagy in B cells and decreases their capacity to extract and present immobilized antigens. We reveal that TLR stimulation restricts lysosome repositioning to the IS by triggering autophagy-dependent degradation of GEF-H1, a Rho GTP exchange factor required for stable lysosome recruitment at the synaptic membrane. GEF-H1 degradation is not observed in B cells that lack αV integrins and are deficient in TLR-induced autophagy. Accordingly, these cells show efficient antigen extraction in the presence of TLR stimulation, confirming the role of TLR-induced autophagy in limiting antigen extraction. Overall, our results suggest that resources associated with autophagy regulate TLR and BCR-dependent functions, which can finetune antigen uptake by B cells. This work helps to understand the mechanisms by which B cells are activated by surface-tethered antigens in contexts of subjacent inflammation before antigen recognition, such as sepsis.

Asunto(s)

Linfocitos B , Receptores de Antígenos de Linfocitos B , Receptores de Antígenos de Linfocitos B/metabolismo , Antígenos/metabolismo , Receptores Toll-Like/metabolismo , Autofagia , Antígenos de Superficie/metabolismo , Factores de Intercambio de Guanina Nucleótido Rho/metabolismo

RESUMEN

Upon interaction with immobilized antigens, B cells form an immune synapse where actin remodeling and re-positioning of the microtubule-organizing center (MTOC) together with lysosomes can facilitate antigen extraction. B cells have restricted cytoplasmic space, mainly occupied by a large nucleus, yet the role of nuclear morphology in the formation of the immune synapse has not been addressed. Here we show that upon activation, B cells re-orientate and adapt the size of their nuclear groove facing the immune synapse, where the MTOC sits, and lysosomes accumulate. Silencing the nuclear envelope proteins Nesprin-1 and Sun-1 impairs nuclear reorientation towards the synapse and leads to defects in actin organization. Consequently, B cells are unable to internalize the BCR after antigen activation. Nesprin-1 and Sun-1-silenced B cells also fail to accumulate the tethering factor Exo70 at the center of the synaptic membrane and display defective lysosome positioning, impairing efficient antigen extraction at the immune synapse. Thus, changes in nuclear morphology and positioning emerge as critical regulatory steps to coordinate B cell activation.

Asunto(s)

Actinas , Receptores de Antígenos de Linfocitos B , Actinas/metabolismo , Antígenos/metabolismo , Linfocitos B , Receptores de Antígenos de Linfocitos B/metabolismo , Sinapsis/metabolismo

RESUMEN

OBJECTIVE: Increasing evidence demonstrates that immune signature plays an important role in the prognosis of gastric cancer (GC). We aimed to develop and validate a robust immune-related gene pair (IRGP) signature for predicting the prognosis of GC patients. METHODS: RNA-Seq data and corresponding clinical information of GC cohort were downloaded from the TCGA (The Cancer Genome Atlas Program) data portal. GSE84437 and GSE15459 microarray datasets were included as independent external cohorts. Least absolute shrinkage and selection operator (LASSO) regression analysis was used to build the best prognostic signature. All patients were classified into the high immune-risk and low immune-risk groups via the optimal cut-off of the signature scores determined by time-dependent receiver-operating characteristic (ROC) curve analysis. The prognostic role of the signature was measured by a log-rank test and a Cox proportional hazard regression model. RESULTS: 14 immune gene pairs consisting of 25 unique genes were identified to construct the immune prognostic signature. High immune-risk groups showed poor prognosis in the TCGA datasets and GSE84437 datasets as well as in the GSE15459 datasets (all P < 0.001). The 14-IRGP signature was an independent prognostic factor of GC after adjusting for other clinical factors (P < 0.05). Functional analysis revealed that DNA integrity checkpoint, DNA replication, T-cell receptor signaling pathway, and B-cell receptor signaling pathway were enriched in the low immune-risk groups. B cells naive and Monocytes were significantly higher in the high-risk group, and B-cell memory and T-cell CD4 memory activated were significantly higher in the low-risk group. The prognostic signature based on IRGP reflected infiltration by several types of immune cells. CONCLUSION: The novel proposed clinical-immune signature is a promising biomarker for prediction overall survival in patients with GC and providing new insights into the treatment strategies.

Asunto(s)

Neoplasias Gástricas/genética , Neoplasias Gástricas/inmunología , Biomarcadores de Tumor/inmunología , Biomarcadores de Tumor/metabolismo , Replicación del ADN , Bases de Datos Genéticas , Bases de Datos de Ácidos Nucleicos , Conjuntos de Datos como Asunto , Expresión Génica , Humanos , Memoria Inmunológica , Linfocitos Infiltrantes de Tumor , Pronóstico , Modelos de Riesgos Proporcionales , Curva ROC , Receptores de Antígenos de Linfocitos B/metabolismo , Receptores de Antígenos de Linfocitos T/metabolismo , Análisis de Regresión , Neoplasias Gástricas/mortalidad

RESUMEN

Chronic lymphocytic leukemia (CLL) is a chronic form of leukemia that originates from an abnormal expansion of CD5+ B-1 cells. Deregulation in the BCR signaling is associated with B-cell transformation. Contrariwise to B-2 cells, BCR engagement in B-1 cells results in low proliferation rate and increased apoptosis population, whereas overactivation may be associated with lymphoproliferative disorders. It has been demonstrated that several transcription factors that are involved in the B cell development play a role in the regulation of BCR function. Among them, Ikaros is considered an essential regulator of lymphoid differentiation and activation. Several reports suggest that Ikaros expression is deregulated in different forms of leukemia. Herein, we demonstrated that CLL cells show decreased Ikaros expression and abnormal cytoplasmic cell localization. These alterations were also observed in radioresistant B-1 cells, which present high proliferative activity, suggesting that abnormal localization of Ikaros could determine its loss of function. Furthermore, Ikaros knockdown increased the expression of BCR pathway components in murine B-1 cells, such as Lyn, Blnk, and CD19. Additionally, in the absence of Ikaros, B-1 cells become responsive to BCR stimulus, increasing cell proliferation even in the absence of antigen stimulation. These results suggested that Ikaros is an important controller of B-1 cell proliferation by interfering with the BCR activity. Therefore, altered Ikaros expression in CLL or radioresistant B-1 cells could determine a responsive status of BCR to self-antigens, which would culminate in the clonal expansion of B-1 cells.

Asunto(s)

Transformación Celular Neoplásica/genética , Regulación Leucémica de la Expresión Génica , Factor de Transcripción Ikaros/genética , Leucemia Linfocítica Crónica de Células B/genética , Adulto , Anciano , Anciano de 80 o más Años , Animales , Linfocitos B/inmunología , Transformación Celular Neoplásica/patología , Citoplasma/metabolismo , Femenino , Humanos , Factor de Transcripción Ikaros/metabolismo , Leucemia Linfocítica Crónica de Células B/patología , Masculino , Ratones , Ratones Endogámicos C57BL , Persona de Mediana Edad , Modelos Biológicos , Unión Proteica , Tolerancia a Radiación , Receptores de Antígenos de Linfocitos B/metabolismo , Transducción de Señal

RESUMEN

Autoreactive B cells that bind self-antigen with high avidity in the bone marrow undergo mechanisms of central tolerance that prevent their entry into the peripheral B cell population. These mechanisms are breached in many autoimmune patients, increasing their risk of B cell-mediated autoimmune diseases. Resolving the molecular pathways that can break central B cell tolerance could therefore provide avenues to diminish autoimmunity. Here, we show that B cell-intrinsic expression of a constitutively active form of PI3K-P110α by high-avidity autoreactive B cells of mice completely abrogates central B cell tolerance and further promotes these cells to escape from the bone marrow, differentiate in peripheral tissue, and undergo activation in response to self-antigen. Upon stimulation with T cell help factors, these B cells secrete antibodies in vitro but remain unable to secrete autoantibodies in vivo. Overall, our data demonstrate that activation of the PI3K pathway leads high-avidity autoreactive B cells to breach central, but not late, stages of peripheral tolerance.

Asunto(s)

Autoantígenos/inmunología , Autoinmunidad/inmunología , Linfocitos B/inmunología , Tolerancia Central/inmunología , Fosfatidilinositol 3-Quinasa Clase Ia/metabolismo , Animales , Autoanticuerpos/inmunología , Células de la Médula Ósea/metabolismo , Diferenciación Celular/inmunología , Femenino , Masculino , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Ratones Transgénicos , Modelos Animales , Receptores de Antígenos de Linfocitos B/metabolismo , Receptores de Complemento 3d/metabolismo , Bazo/citología , Linfocitos T/inmunología

RESUMEN

Prolactin has an immunomodulatory effect and has been associated with B-cell-triggered autoimmune diseases, such as systemic lupus erythematosus (SLE). In mice that develop SLE, the PRL receptor is expressed in early bone marrow B-cells, and increased levels of PRL hasten disease manifestations, which are correlated with a reduction in the absolute number of immature B-cells. The aim of this work was to determine the effect of PRL in an in vitro system of B-cell tolerance using WEHI-231 cells and immature B-cells from lupus prone MRL/lpr mice. WEHI-231 cells express the long isoform of the PRL receptor, and PRL rescued the cells from cell death by decreasing the apoptosis induced by the cross-linking of the B-cell antigen receptor (BCR) as measured by Annexin V and active caspase-3. This decrease in apoptosis may have been due to the PRL and receptor interaction, which increased the relative expression of antiapoptotic Bcl-xL and decreased the relative expression of proapoptotic Bad. In immature B-cells from MRL/lpr mice, PRL increased the viability and decreased the apoptosis induced by the cross-linking of BCR, which may favor the maturation of self-reactive B-cells and contribute to the onset of disease.

Asunto(s)

Apoptosis , Células Precursoras de Linfocitos B/citología , Células Precursoras de Linfocitos B/metabolismo , Prolactina/metabolismo , Receptores de Antígenos de Linfocitos B/metabolismo , Animales , Apoptosis/efectos de los fármacos , Apoptosis/genética , Apoptosis/inmunología , Caspasa 3/metabolismo , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , Supervivencia Celular/inmunología , Expresión Génica , Ratones , Ratones Endogámicos MRL lpr , Células Precursoras de Linfocitos B/efectos de los fármacos , Prolactina/farmacología , Unión Proteica , Receptores de Prolactina/genética , Receptores de Prolactina/metabolismo

RESUMEN

The aim of this study is to analyze gene expression data to identify key genes and pathways associated with resistance to platinum-based chemotherapy in epithelial ovarian cancer (EOC) and to improve clinical treatment strategies. The gene expression data set was downloaded from Gene Expression Omnibus and included 12 chemotherapy-resistant EOC samples and 16 chemotherapy-sensitive EOC samples. A differential analysis was performed to screen out differentially expressed genes (DEGs). A functional enrichment analysis was conducted for the DEGs using the database for annotation, visualization, and integration discovery. A protein-protein interaction (PPI) network was constructed with information from the human protein reference database. Pathway-pathway interactions were determined with a test based on the hypergeometric distribution. A total of 1564 DEGs were identified in chemotherapy-sensitive EOC, including 654 upregulated genes and 910 downregulated genes. The top three upregulated genes were HIST1H3G, AKT3, and RTN3, while the top three downregulated genes were NBLA00301, TRIM62, and EPHA5. A Gene Ontology enrichment analysis showed that cell adhesion, biological adhesion, and intracellular signaling cascades were significantly enriched in the DEGs. A KEGG pathway enrichment analysis revealed that the calcium, mitogen-activated protein kinase, and B cell receptor signaling pathways were significantly over-represented in the DEGs. A PPI network containing 101 interactions was acquired. The top three hub genes were RAC1, CAV1, and BCL2. Five modules were identified from the PPI network. Taken together, these findings could advance the understanding of the molecular mechanisms underlying intrinsic chemotherapy resistance in EOC.

Asunto(s)

Resistencia a Antineoplásicos/genética , Regulación Neoplásica de la Expresión Génica , Redes Reguladoras de Genes , Proteínas de Neoplasias/genética , Neoplasias Glandulares y Epiteliales/genética , Neoplasias Ováricas/genética , Antineoplásicos/uso terapéutico , Señalización del Calcio , Carcinoma Epitelial de Ovario , Adhesión Celular , Bases de Datos de Proteínas , Femenino , Perfilación de la Expresión Génica , Ontología de Genes , Humanos , Análisis por Micromatrices , Proteínas Quinasas Activadas por Mitógenos/genética , Proteínas Quinasas Activadas por Mitógenos/metabolismo , Anotación de Secuencia Molecular , Proteínas de Neoplasias/metabolismo , Neoplasias Glandulares y Epiteliales/tratamiento farmacológico , Neoplasias Glandulares y Epiteliales/metabolismo , Neoplasias Glandulares y Epiteliales/patología , Compuestos Organoplatinos/uso terapéutico , Neoplasias Ováricas/tratamiento farmacológico , Neoplasias Ováricas/metabolismo , Neoplasias Ováricas/patología , Mapeo de Interacción de Proteínas , Receptores de Antígenos de Linfocitos B/genética , Receptores de Antígenos de Linfocitos B/metabolismo

RESUMEN

The treatment of patients with chronic lymphocytic leukemia (CLL), an indolent B-cell lymphoma is in the midst of a transformation. There are a large number of promising new therapeutic agents and cellular therapies being studied which exhibit remarkable activity, favorable toxicity profiles, convenient administration schedules, and treatment options are rapidly expanding. The recent advances in the management of CLL exemplify the value of translational medicine. This review highlights key aspects of B-cell receptor (BCR) signaling in relation to novel inhibitors of the BCR signaling pathway, currently at various stages of preclinical and clinical development.

Asunto(s)

Antineoplásicos/uso terapéutico , Leucemia Linfocítica Crónica de Células B/tratamiento farmacológico , Inhibidores de las Quinasa Fosfoinosítidos-3 , Proteínas Tirosina Quinasas/antagonistas & inhibidores , Receptores de Antígenos de Linfocitos B/antagonistas & inhibidores , Agammaglobulinemia Tirosina Quinasa , Anticuerpos Monoclonales/uso terapéutico , Antimetabolitos Antineoplásicos/uso terapéutico , Linfocitos B/efectos de los fármacos , Linfocitos B/metabolismo , Linfocitos B/patología , Regulación de la Expresión Génica , Humanos , Leucemia Linfocítica Crónica de Células B/genética , Leucemia Linfocítica Crónica de Células B/metabolismo , Leucemia Linfocítica Crónica de Células B/patología , Fosfatidilinositol 3-Quinasas/genética , Fosfatidilinositol 3-Quinasas/metabolismo , Inhibidores de Proteínas Quinasas/uso terapéutico , Proteínas Tirosina Quinasas/genética , Proteínas Tirosina Quinasas/metabolismo , Receptores de Antígenos de Linfocitos B/genética , Receptores de Antígenos de Linfocitos B/metabolismo , Transducción de Señal/efectos de los fármacos

RESUMEN

Colostrum plays an important role in protecting newborn infants against acute gastrointestinal and respiratory infections. IgA antibodies have been considered the major effector component; however, the role of their receptors on colostral phagocytes, especially neutrophils, has not been studied. Here, we demonstrate that CD15+ colostrum neutrophils express IgA Fc receptors (Fc alphaR, CD89) at levels similar to those of blood neutrophils. Most colostral cells (70%) bear secretory IgA (SIgA) on their surface (and intracellularly), whereas blood cells do not. The Fc alphaR on colostral neutrophils was identified as the a.1 isoform with a similar molecular mass (55-75 kDa) as that identified for blood neutrophils. Removal of N-linked carbohydrates revealed a major protein core of 32 kDa for both cell types. In contrast, co-immunoprecipitation and immunoblot experiments using a mild detergent, digitonin, revealed a lack of gamma chain association with Fc alphaR (gamma-less) exclusively on colostral neutrophils. The functional role of these gamma-less Fc alphaR cells was evaluated by measuring superoxide release and killing of SIgA-coated enteropathogenic E. coli. No increase in superoxide release was observed in colostral cells compared with blood neutrophils, whereas optimal release was obtained with PMA stimulation. Furthermore, despite similar bacterial phagocytosis index between both cell types, IgA-mediated bacterial-killing was not detectable with colostral neutrophils, whereas killing was detectable on blood cells. These results reveal exclusive expression of gamma-less Fc alphaR on colostral neutrophils associated with receptor hyperoccupation by IgA and with low, bacterial-killing activity, which suggest that this receptor may mediate noninflammatory effects of SIgA.

Asunto(s)

Antígenos CD/biosíntesis , Calostro/inmunología , Calostro/metabolismo , Inmunoglobulina A Secretora/metabolismo , Inmunoglobulina A/metabolismo , Neutrófilos/inmunología , Neutrófilos/metabolismo , Receptores Fc/biosíntesis , Adolescente , Adulto , Antígenos CD/sangre , Actividad Bactericida de la Sangre/inmunología , Preescolar , Calostro/citología , Calostro/microbiología , Endocitosis/inmunología , Escherichia coli/inmunología , Escherichia coli/patogenicidad , Femenino , Humanos , Inmunoglobulina A/sangre , Lactante , Inflamación/inmunología , Inflamación/metabolismo , Neutrófilos/microbiología , Proteínas Opsoninas/inmunología , Fagocitosis/inmunología , Isoformas de Proteínas/biosíntesis , Receptores de Antígenos de Linfocitos B/metabolismo , Receptores Fc/sangre , Superóxidos/metabolismo

RESUMEN

Las inmunoglobulinas o anticuerpos son glicoproteínas que se encuentran en la superficie de los linfocitos B y son secretadas por las células plasmáticas, linfocitos B terminalmente diferenciados, en respuesta a un antígeno y, como tal, representan la inmunidad humoral de los vertebrados. Existen 5 formas o isotipos principales: IgG, IgM, IgA, IgD e IgE. Las Igs presentan una estructura básica que posee dos cadenas pesadas (H) idénticas y dos cadenas livianas (L) idénticas, unidas entre sí por puentes disulfuro e interacciones no covalentes. Ambos tipos de cadenas presentan un patrón estructural que consiste de segmentos o dominios de 110 aminoácidos. El análisis de su secuencia de aminoácidos revela la existencia de un dominio variable (V) hacia el extremo aminoterminal y varios dominios constantes (C) hacia el extremo carboxilo terminal. Las cadenas pesadas también poseen un dominio variable (VL) y uno constante (CL). Las cadenas pesadas también poseen un dominio variable (VH) pero tienen 3 ó 4 dominios constantes (CH). Los dominios variables de mabas cadenas contienen zonas de alta variabilidad no contiguas en la secuencia de aminoácidos, son las denominadas regiones hipervariables o CRD (determinantes de complementariedad) y son las principales responsables de la diversidad de los anticuerpos...

Asunto(s)

Humanos , Linfocitos B , Formación de Anticuerpos/inmunología , Inmunoglobulinas/inmunología , Diversidad de Anticuerpos/genética , Formación de Anticuerpos/inmunología , Genes de Inmunoglobulinas , Inmunoglobulinas/biosíntesis , Inmunoglobulinas/clasificación , Receptores de Antígenos de Linfocitos B/metabolismo , Transcripción Genética

RESUMEN

The B cell antigen receptor complex (BCR) is composed of membrane Ig and heterodimers of Ig-alpha and Ig-beta/gamma. Recent findings indicate that Ig-alpha associates with Src-family kinases, including Fyn and Lyn, via an approximately 26 amino acid motif termed ARH1. Studies reported here (i) define two mechanisms whereby this motif binds Fyn and (ii) reveal an important functional consequence of binding, i.e. kinase activation. Mutational analysis indicates that specific low-affinity binding is determined by a short sequence, -DCSM-, in the motif and is not dependent on motif tyrosine residues. In contrast, the doubly tyrosine phosphorylated motif binds independently of DCSM and with high affinity. Importantly, this binding leads to Fyn activation. Taken together with studies which map low-affinity binding of Fyn or Lyn to the kinase's N-terminal unique region and high-affinity binding to the kinase's SH2 domain, these results suggest a mechanism of BCR activation in which the non-phosphorylated resting receptor is associated with Src-family kinases and, upon stimulation, tyrosine phosphorylation of Ig-alpha leads to reorientation and activation of receptor-associated kinases.

Asunto(s)

Proteínas Tirosina Quinasas/metabolismo , Proteínas Proto-Oncogénicas/metabolismo , Receptores de Antígenos de Linfocitos B/metabolismo , Células 3T3 , Secuencia de Aminoácidos , Animales , Secuencia de Bases , Línea Celular , Análisis Mutacional de ADN , Activación Enzimática , Ratones , Datos de Secuencia Molecular , Fosforilación , Unión Proteica , Proteínas Proto-Oncogénicas c-fyn , Receptores de Antígenos de Linfocitos B/genética , Relación Estructura-Actividad , Tirosina/metabolismo

Asunto(s)

Linfocitos/inmunología , Infecciones por Nematodos/inmunología , Receptores de Antígenos de Linfocitos B/metabolismo , Animales , Separación Celular , Femenino , Inmunidad Celular , Inmunización Pasiva , Infecciones por Nematodos/prevención & control , Nippostrongylus/inmunología , Ratas , Conducto Torácico , Factores de Tiempo