RESUMEN
B cells are important in mucosal microbial homeostasis through their well-known role in secretory IgA production and their emerging role in mucosal immunoregulation. Several specialized intraintestinal B cell compartments have been characterized, but the nature of conventional B cells in the lamina propria is poorly understood. In this study, we identify a B cell population predominantly composed of surface IgM(+) IgD(+) cells residing in villi of the small intestine and superficial lamina propria of the large intestine, but distinct from the intraepithelial compartment or organized intestinal lymphoid structures. Small intestinal (villous) B cells are diminished in genotypes that alter the strength of BCR signaling (Bruton tyrosine kinase(xid), Galphai2(-/-)), and in mice lacking cognate BCR specificity. They are not dependent on enteric microbial sensing, because they are abundant in mice that are germfree or genetically deficient in TLR signaling. However, villous B cells are reduced in the absence of invariant NK T cells (Jalpha18(-/-) or CD1d(-/-) mice). These findings define a distinct population of conventional B cells in small intestinal villi, and suggest an immunologic link between CD1-restricted invariant NK T cells and this B cell population.
Asunto(s)
Linfocitos B/inmunología , Intestino Delgado/inmunología , Linfocitos T Reguladores/inmunología , Envejecimiento/fisiología , Animales , Presentación de Antígeno/inmunología , Antígenos CD1/genética , Antígenos CD1/inmunología , Antígenos CD1/metabolismo , Linfocitos B/citología , Movimiento Celular/inmunología , Separación Celular , Mucosa Intestinal/citología , Mucosa Intestinal/inmunología , Intestino Delgado/citología , Intestino Delgado/ultraestructura , Ratones , Ratones Noqueados , Microvellosidades/inmunología , Fenotipo , Receptores de Antígenos de Linfocitos B/inmunología , Sensibilidad y Especificidad , Transducción de Señal/inmunologíaRESUMEN
A recent and surprising body of research has linked changes in immune function to biologic and therapeutic targeting of cannabinoid receptors, which prototypically respond to delta-9 tetrahydrocannabinol. The peripheral cannabinoid receptor CB2 is highly expressed in immune cell types (macrophages, dendritic cells, and B cells), and pharmacologically alters their cytokine production and responsiveness. Accordingly, cannabinoid agonists can powerfully alter susceptibility to certain microbial infections, atherosclerosis, and cancer immunotherapy. What is unknown is the physiologic role of natural levels of endocannabinoids and their receptors in normal immune homeostasis. Galphai2-/- mice are deficient in the formation of certain B and T cell subsets and are susceptible to immune dysregulation, notably developing inflammatory bowel disease. A key issue is the identity of the Gi-coupled receptors relevant to this Galphai2-signaling pathway. We find that mice deficient in CB2, the Gi-coupled peripheral endocannabinoid receptor, have profound deficiencies in splenic marginal zone, peritoneal B1a cells, splenic memory CD4+ T cells, and intestinal natural killer cells and natural killer T cells. These findings partially phenocopy and extend the lymphocyte developmental disorder associated with the Galphai2-/- genotype, and suggest that the endocannabinoid system is required for the formation of T and B cell subsets involved in immune homeostasis. This noncompensatable requirement for physiologic function of the endocannabinoid system is novel. Because levels of endocannabinoids are highly restricted microanatomically, local regulation of their production and receptor expression offers a new principle for regional immune homeostasis and disease susceptibility, and extends and refines the rationale for CB2-targeted immunotherapy in immune and inflammatory diseases.
Asunto(s)
Subgrupos de Linfocitos B/metabolismo , Receptor Cannabinoide CB2/metabolismo , Subgrupos de Linfocitos T/metabolismo , Animales , Subgrupos de Linfocitos B/inmunología , Secuencia de Bases , ADN/genética , Subunidad alfa de la Proteína de Unión al GTP Gi2/deficiencia , Subunidad alfa de la Proteína de Unión al GTP Gi2/genética , Subunidad alfa de la Proteína de Unión al GTP Gi2/metabolismo , Expresión Génica , Memoria Inmunológica , Mucosa Intestinal/citología , Mucosa Intestinal/inmunología , Mucosa Intestinal/metabolismo , Células Asesinas Naturales/clasificación , Células Asesinas Naturales/inmunología , Células Asesinas Naturales/metabolismo , Ratones , Ratones Endogámicos C3H , Ratones Endogámicos C57BL , Ratones Noqueados , Especificidad de Órganos , Fenotipo , Receptor Cannabinoide CB2/deficiencia , Receptor Cannabinoide CB2/genética , Transducción de Señal , Bazo/citología , Bazo/inmunología , Bazo/metabolismo , Subgrupos de Linfocitos T/inmunologíaRESUMEN
PURPOSE: As kinase inhibitors transition from the laboratory to patients, it is imperative to develop biomarkers that can be used in the clinic. The primary objectives are to identify patients most likely to benefit from molecularly targeted therapies and to document modulation of the drug target. Constitutive activation of the phosphoinositide 3-kinase (PI3K) pathway and its downstream effectors, as a result of PTEN loss or by other mechanisms, occurs in a high proportion of prostate cancers, making it an ideal template for the design of clinical trials involving PI3K pathway inhibitors. Prostate cancers also present unique organ-specific challenges, in that tumors are heterogeneous and diagnostic tissue is extremely limited. EXPERIMENTAL DESIGN: Working within these limitations, we have developed a set of immunohistochemical assays that define activation of the PI3K pathway in clinical samples. RESULTS AND CONCLUSIONS: Using both univariate and multivariate analyses, we show that loss of PTEN is highly correlated with the activation of AKT, and this, in turn, is associated with the phosphorylation of S6, one of its main effectors. These three antibodies are potentially able to define a molecular signature of PTEN loss and/or AKT pathway activation in prostate cancer.
Asunto(s)
Anticuerpos Monoclonales , Regulación Neoplásica de la Expresión Génica , Fosfatidilinositol 3-Quinasas/metabolismo , Neoplasias de la Próstata/enzimología , Transducción de Señal , Regulación hacia Abajo , Activación Enzimática , Regulación Enzimológica de la Expresión Génica , Humanos , Masculino , Fosfohidrolasa PTEN , Monoéster Fosfórico Hidrolasas/antagonistas & inhibidores , Monoéster Fosfórico Hidrolasas/metabolismo , Fosforilación , Próstata/enzimología , Próstata/inmunología , Próstata/patología , Neoplasias de la Próstata/inmunología , Neoplasias de la Próstata/patología , Proteínas Serina-Treonina Quinasas/metabolismo , Proteínas Proto-Oncogénicas/metabolismo , Proteínas Proto-Oncogénicas c-akt , Proteínas Quinasas S6 Ribosómicas/metabolismo , Proteínas Supresoras de Tumor/antagonistas & inhibidores , Proteínas Supresoras de Tumor/metabolismoRESUMEN
Null mutation of the Galphai2 trimeric G protein results in a discrete and profound mucosal disorder, including inflammatory bowel disease (IBD), attenuation of IL-10 expression, and immune function polarized to Th1 activity. Genetic and adoptive transfer experiments have established a role for B cells and IL-10 in mucosal immunologic homeostasis and IBD resistance. In this study, we addressed the hypothesis that Galphai2 is required for the development of IL-10-producing B cells. Galphai2(-/-) mice were reduced in the relative abundance of marginal zone (MZ), transitional type 2 (T2), and B-1a B cells and significantly increased in follicular mature and B-1b B cells. Reconstitution of RAG2(-/-) mice with Galphai2(-/-) bone marrow induced an IBD-like colitis and a deficiency in absolute numbers of MZ, T2, and B-1 B cells. Thus, the Galphai2(-/-) genotype in colitis susceptibility and B cell development involved a cis effect within the hemopoietic compartment. In vitro, the B cell population of Galphai2(-/-) mice was functionally deficient in LPS-induced proliferation and IL-10 production, consistent with the exclusive capacity of T2 and MZ cell subpopulations for LPS responsiveness. In vivo, Galphai2(-/-) mice were selectively impaired for the IgM response to T-independent type II, consistent with the relative depletion of MZ and peritoneal B-1 subpopulations. Collectively, these results reveal a selective role for Galphai2 in MZ and B-1 B cell development. Disorders of this Galphai2-dependent process in B cell development may represent a mechanism for IBD susceptibility.