RESUMEN
Wound healing in response to acute injury is mediated by the coordinated and transient activation of parenchymal, stromal, and immune cells that resolves to homeostasis. Environmental, genetic, and epigenetic factors associated with inflammation and aging can lead to persistent activation of the microenvironment and fibrosis. Here, we identify opposing roles of interleukin-4 (IL-4) cytokine signaling in interstitial macrophages and type II alveolar epithelial cells (ATIIs). We show that IL4Ra signaling in macrophages promotes regeneration of the alveolar epithelium after bleomycin-induced lung injury. Using organoids and mouse models, we show that IL-4 directly acts on a subset of ATIIs to induce the expression of the transcription factor SOX9 and reprograms them toward a progenitor-like state with both airway and alveolar lineage potential. In the contexts of aging and bleomycin-induced lung injury, this leads to aberrant epithelial cell differentiation and bronchiolization, consistent with cellular and histological changes observed in interstitial lung disease.
RESUMEN
Chronic kidney disease (CKD) is characterized by inflammation and fibrosis in the kidney. Renal biopsies and estimated glomerular filtration rate (eGFR) remain the standard of care, but these endpoints have limitations in detecting the stage, progression, and spatial distribution of fibrotic pathology in the kidney. MRI diffusion tensor imaging (DTI) has emerged as a promising noninvasive technology to evaluate renal fibrosis in vivo both in clinical and preclinical studies. However, these imaging studies have not systematically identified fibrosis particularly deeper in the kidney where biopsy sampling is limited, or completed an extensive analysis of whole organ histology, blood biomarkers, and gene expression to evaluate the relative strengths and weaknesses of MRI for evaluating renal fibrosis. In this study, we performed DTI in the sodium oxalate mouse model of CKD. The DTI parameters fractional anisotropy, apparent diffusion coefficient, and axial diffusivity were compared between the control and oxalate groups with region of interest (ROI) analysis to determine changes in the cortex and medulla. In addition, voxel-based analysis (VBA) was implemented to systematically identify local regions of injury over the whole kidney. DTI parameters were found to be significantly different in the medulla by both ROI analysis and VBA, which also spatially matched with collagen III immunohistochemistry (IHC). The DTI parameters in this medullary region exhibited moderate to strong correlations with histology, blood biomarkers, hydroxyproline, and gene expression. Our results thus highlight the sensitivity of DTI to the heterogeneity of renal fibrosis and importance of whole kidney noninvasive imaging.NEW & NOTEWORTHY Chronic kidney disease (CKD) can be characterized by inflammation and fibrosis of the kidney. Although standard of care methods have been limited in scope, safety, and spatial distribution, MRI diffusion tensor imaging (DTI) has emerged as a promising noninvasive technology to evaluate renal fibrosis in vivo. In this study, we performed DTI in an oxalate mouse model of CKD to systematically identify local kidney injury. DTI parameters strongly correlated with histology, blood biomarkers, hydroxyproline, and gene expression.
Asunto(s)
Imagen de Difusión Tensora , Modelos Animales de Enfermedad , Fibrosis , Ratones Endogámicos C57BL , Insuficiencia Renal Crónica , Animales , Insuficiencia Renal Crónica/patología , Insuficiencia Renal Crónica/diagnóstico por imagen , Masculino , Oxalatos/metabolismo , Riñón/patología , Riñón/diagnóstico por imagen , Riñón/metabolismo , RatonesRESUMEN
BACKGROUND: Transforming growth factor ß (TGF-ß) is implicated as a key mediator of pathological fibrosis, but its pleiotropic activity in a range of homeostatic functions presents challenges to its safe and effective therapeutic targeting. There are three isoforms of TGF-ß, TGF-ß1, TGF-ß2, and TGF-ß3, which bind to a common receptor complex composed of TGF-ßR1 and TGF-ßR2 to induce similar intracellular signals in vitro. We have recently shown that the cellular expression patterns and activation thresholds of TGF-ß2 and TGF-ß3 are distinct from those of TGF-ß1 and that selective short-term TGF-ß2 and TGF-ß3 inhibition can attenuate fibrosis in vivo without promoting excessive inflammation. Isoform-selective inhibition of TGF-ß may therefore provide a therapeutic opportunity for patients with chronic fibrotic disorders. METHODS: Transcriptomic profiling of skin biopsies from patients with systemic sclerosis (SSc) from multiple clinical trials was performed to evaluate the role of TGF-ß3 in this disease. Antibody humanization, biochemical characterization, crystallization, and pre-clinical experiments were performed to further characterize an anti-TGF-ß3 antibody. FINDINGS: In the skin of patients with SSc, TGF-ß3 expression is uniquely correlated with biomarkers of TGF-ß signaling and disease severity. Crystallographic studies establish a structural basis for selective TGF-ß3 inhibition with a potent and selective monoclonal antibody that attenuates fibrosis effectively in vivo at clinically translatable exposures. Toxicology studies suggest that, as opposed to pan-TGF-ß inhibitors, this anti-TGF-ß3 antibody has a favorable safety profile for chronic administration. CONCLUSION: We establish a rationale for targeting TGF-ß3 in SSc with a favorable therapeutic index. FUNDING: This study was funded by Genentech, Inc.
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Esclerodermia Sistémica , Factor de Crecimiento Transformador beta3 , Humanos , Factor de Crecimiento Transformador beta3/metabolismo , Factor de Crecimiento Transformador beta1/metabolismo , Factor de Crecimiento Transformador beta2/metabolismo , Fibrosis , Esclerodermia Sistémica/tratamiento farmacológico , Isoformas de Proteínas/metabolismoRESUMEN
Wnt ligands oligomerize Frizzled (Fzd) and Lrp5/6 receptors to control the specification and activity of stem cells in many species. How Wnt signaling is selectively activated in different stem cell populations, often within the same organ, is not understood. In lung alveoli, we show that distinct Wnt receptors are expressed by epithelial (Fzd5/6), endothelial (Fzd4), and stromal (Fzd1) cells. Fzd5 is uniquely required for alveolar epithelial stem cell activity, whereas fibroblasts utilize distinct Fzd receptors. Using an expanded repertoire of Fzd-Lrp agonists, we could activate canonical Wnt signaling in alveolar epithelial stem cells via either Fzd5 or, unexpectedly, non-canonical Fzd6. A Fzd5 agonist (Fzd5ag) or Fzd6ag stimulated alveolar epithelial stem cell activity and promoted survival in mice after lung injury, but only Fzd6ag promoted an alveolar fate in airway-derived progenitors. Therefore, we identify a potential strategy for promoting regeneration without exacerbating fibrosis during lung injury.
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Lesión Pulmonar , Ratones , Animales , Proteínas Wnt , Receptores Frizzled , Vía de Señalización Wnt , Células Epiteliales Alveolares , Células MadreRESUMEN
Inhibition of NF-κB inducing kinase (NIK) has been pursued as a promising therapeutic target for autoimmune disorders due to its highly regulated role in key steps of the NF-κB signaling pathway. Previously reported NIK inhibitors from our group were shown to be potent, selective, and efficacious, but had higher human dose projections than desirable for immunology indications. Herein we report the clearance-driven optimization of a NIK inhibitor guided by metabolite identification studies and structure-based drug design. This led to the identification of an azabicyclo[3.1.0]hexanone motif that attenuated in vitro and in vivo clearance while maintaining NIK potency and increasing selectivity over other kinases, resulting in a greater than ten-fold reduction in predicted human dose.
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FN-kappa B , Transducción de Señal , Humanos , FN-kappa B/metabolismo , Semivida , Diseño de FármacosRESUMEN
Non-alcoholic fatty liver disease (NAFLD), represents an unmet medical need that can progress to non-alcoholic steatohepatitis (NASH), which, without intervention, can result in the development of cirrhosis and hepatocellular carcinoma (HCC). Inflammation is a pathological hallmark of NASH, and targeting key inflammatory mediators of NASH may lead to potential therapeutics for the disease. Herein, we aimed to investigate the role of IL-23 signaling in NASH progression in murine models. We showed that recombinant IL-23 can promote IL-17 producing cell expansion in the liver and that these cells are predominately γδ T cells and Mucosal Associated Invariant T cells (MAITs). Reciprocally, we found that IL-23 signaling is necessary for the expansion of γδ T cells and MAIT cells in the western diet (WD) diet induced NASH model. However, we did not observe any significant differences in liver inflammation and fibrosis between wild type and Il23r-/- mice in the same NASH model. Furthermore, we found that Il23r deletion does not impact liver inflammation and fibrosis in the choline-deficient, L-amino acid-defined and high-fat diet (CDA-HFD) induced NASH model. Based on these findings, we therefore propose that IL-23 signaling is not necessary for NASH pathogenesis in preclinical models and targeting this pathway alone may not be an effective therapeutic approach to ameliorate the disease progression in NASH patients.
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Carcinoma Hepatocelular , Hepatitis , Neoplasias Hepáticas , Enfermedad del Hígado Graso no Alcohólico , Aminoácidos/uso terapéutico , Animales , Carcinoma Hepatocelular/patología , Colina , Modelos Animales de Enfermedad , Hepatitis/complicaciones , Mediadores de Inflamación , Interleucina-17/genética , Interleucina-23 , Cirrosis Hepática/patología , Neoplasias Hepáticas/patología , Ratones , Ratones Endogámicos C57BL , Enfermedad del Hígado Graso no Alcohólico/patologíaRESUMEN
Bone morphogenetic protein 1 (BMP1) belongs to the astacin/BMP1/tolloid-like family of zinc metalloproteinases, which play a fundamental role in the development and formation of extracellular matrix (ECM). BMP1 mediates the cleavage of carboxyl terminal (C-term) propeptides from procollagens, a crucial step in fibrillar collagen fiber formation. Blocking BMP1 by small molecule or antibody inhibitors has been linked to anti-fibrotic activity in the preclinical models of skin, kidney and liver fibrosis. Therefore, we reason that BMP1 may be important for the pathogenesis of lung fibrosis and BMP1 could be a potential therapeutic target for progressive fibrotic disease such as idiopathic pulmonary fibrosis (IPF). Here, we observed the increased expression of BMP1 in both human IPF lungs and mouse fibrotic lungs induced by bleomycin. Furthermore, we developed an inducible Bmp1 conditional knockout (cKO) mouse strain. We found that Bmp1 deletion does not protect mice from lung fibrosis triggered by bleomycin. Moreover, we found no significant impact of BMP1 deficiency upon C-term propeptide of type I procollagen (CICP) production in the fibrotic mouse lungs. Based on these results, we propose that BMP1 is not required for lung fibrosis in mice and BMP1 may not be considered a candidate therapeutic target for IPF.
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Proteína Morfogenética Ósea 1 , Fibrosis Pulmonar Idiopática , Animales , Bleomicina/metabolismo , Proteína Morfogenética Ósea 1/genética , Proteína Morfogenética Ósea 1/metabolismo , Matriz Extracelular/metabolismo , Fibrosis Pulmonar Idiopática/genética , Fibrosis Pulmonar Idiopática/metabolismo , Ratones , Procolágeno/genéticaRESUMEN
Transforming growth factor-ß (TGFß) is a key driver of fibrogenesis. Three TGFß isoforms (TGFß1, TGFß2, and TGFß3) in mammals have distinct functions in embryonic development; however, the postnatal pathological roles and activation mechanisms of TGFß2 and TGFß3 have not been well characterized. Here, we show that the latent forms of TGFß2 and TGFß3 can be activated by integrin-independent mechanisms and have lower activation thresholds compared to TGFß1. Unlike TGFB1, TGFB2 and TGFB3 expression is increased in human lung and liver fibrotic tissues compared to healthy control tissues. Thus, TGFß2 and TGFß3 may play a pathological role in fibrosis. Inducible conditional knockout mice and anti-TGFß isoform-selective antibodies demonstrated that TGFß2 and TGFß3 are independently involved in mouse fibrosis models in vivo, and selective TGFß2 and TGFß3 inhibition does not lead to the increased inflammation observed with pan-TGFß isoform inhibition. A cocrystal structure of a TGFß2-anti-TGFß2/3 antibody complex reveals an allosteric isoform-selective inhibitory mechanism. Therefore, inhibiting TGFß2 and/or TGFß3 while sparing TGFß1 may alleviate fibrosis without toxicity concerns associated with pan-TGFß blockade.
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Factor de Crecimiento Transformador beta2 , Factor de Crecimiento Transformador beta3 , Animales , Modelos Animales de Enfermedad , Femenino , Fibrosis , Humanos , Ratones , Isoformas de Proteínas/metabolismo , Factor de Crecimiento Transformador beta2/metabolismo , Factor de Crecimiento Transformador beta3/metabolismoRESUMEN
Metabolic reprograming toward aerobic glycolysis is a pivotal mechanism shaping immune responses. Here we show that deficiency in NF-κB-inducing kinase (NIK) impairs glycolysis induction, rendering CD8+ effector T cells hypofunctional in the tumor microenvironment. Conversely, ectopic expression of NIK promotes CD8+ T cell metabolism and effector function, thereby profoundly enhancing antitumor immunity and improving the efficacy of T cell adoptive therapy. NIK regulates T cell metabolism via a NF-κB-independent mechanism that involves stabilization of hexokinase 2 (HK2), a rate-limiting enzyme of the glycolytic pathway. NIK prevents autophagic degradation of HK2 through controlling cellular reactive oxygen species levels, which in turn involves modulation of glucose-6-phosphate dehydrogenase (G6PD), an enzyme that mediates production of the antioxidant NADPH. We show that the G6PD-NADPH redox system is important for HK2 stability and metabolism in activated T cells. These findings establish NIK as a pivotal regulator of T cell metabolism and highlight a post-translational mechanism of metabolic regulation.
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Linfocitos T CD8-positivos/enzimología , Neoplasias del Colon/enzimología , Metabolismo Energético , Activación de Linfocitos , Linfocitos Infiltrantes de Tumor/enzimología , Melanoma Experimental/enzimología , Proteínas Serina-Treonina Quinasas/metabolismo , Animales , Linfocitos T CD8-positivos/inmunología , Linfocitos T CD8-positivos/trasplante , Línea Celular Tumoral , Neoplasias del Colon/inmunología , Neoplasias del Colon/patología , Neoplasias del Colon/terapia , Citotoxicidad Inmunológica , Estabilidad de Enzimas , Femenino , Glucosafosfato Deshidrogenasa/metabolismo , Glucólisis , Hexoquinasa/genética , Hexoquinasa/metabolismo , Inmunoterapia Adoptiva , Linfocitos Infiltrantes de Tumor/inmunología , Linfocitos Infiltrantes de Tumor/trasplante , Masculino , Melanoma Experimental/inmunología , Melanoma Experimental/patología , Melanoma Experimental/terapia , Ratones Endogámicos C57BL , Ratones Noqueados , NADP/metabolismo , Fenotipo , Proteínas Serina-Treonina Quinasas/deficiencia , Proteínas Serina-Treonina Quinasas/genética , Especies Reactivas de Oxígeno/metabolismo , Transducción de Señal , Microambiente Tumoral , Quinasa de Factor Nuclear kappa BRESUMEN
The dysregulation of multiple signaling pathways, including those through endosomal Toll-like receptors (TLRs), Fc gamma receptors (FcγR), and antigen receptors in B cells (BCR), promote an autoinflammatory loop in systemic lupus erythematosus (SLE). Here, we used selective small-molecule inhibitors to assess the regulatory roles of interleukin-1 receptor (IL-1R)-associated kinase 4 (IRAK4) and Bruton's tyrosine kinase (BTK) in these pathways. The inhibition of IRAK4 repressed SLE immune complex- and TLR7-mediated activation of human plasmacytoid dendritic cells (pDCs). Correspondingly, the expression of interferon (IFN)-responsive genes (IRGs) in cells and in mice was positively regulated by the kinase activity of IRAK4. Both IRAK4 and BTK inhibition reduced the TLR7-mediated differentiation of human memory B cells into plasmablasts. TLR7-dependent inflammatory responses were differentially regulated by IRAK4 and BTK by cell type: In pDCs, IRAK4 positively regulated NF-κB and MAPK signaling, whereas in B cells, NF-κB and MAPK pathways were regulated by both BTK and IRAK4. In the pristane-induced lupus mouse model, inhibition of IRAK4 reduced the expression of IRGs during disease onset. Mice engineered to express kinase-deficient IRAK4 were protected from both chemical (pristane-induced) and genetic (NZB/W_F1 hybrid) models of lupus development. Our findings suggest that kinase inhibitors of IRAK4 might be a therapeutic in patients with SLE.
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Células Dendríticas/metabolismo , Endosomas/metabolismo , Quinasas Asociadas a Receptores de Interleucina-1/metabolismo , Glicoproteínas de Membrana/metabolismo , Células Plasmáticas/metabolismo , Transducción de Señal , Receptor Toll-Like 7/metabolismo , Agammaglobulinemia Tirosina Quinasa , Animales , Endosomas/genética , Humanos , Quinasas Asociadas a Receptores de Interleucina-1/genética , Glicoproteínas de Membrana/genética , Ratones , Receptor Toll-Like 7/genéticaRESUMEN
Dendritic cells (DCs) play an integral role in regulating mucosal immunity and homeostasis, but the signaling network mediating this function of DCs is poorly defined. We identified the noncanonical NF-κB-inducing kinase (NIK) as a crucial mediator of mucosal DC function. DC-specific NIK deletion impaired intestinal immunoglobulin A (IgA) secretion and microbiota homeostasis, rendering mice sensitive to an intestinal pathogen, Citrobacter rodentium. DC-specific NIK was required for expression of the IgA transporter polymeric immunoglobulin receptor (pIgR) in intestinal epithelial cells, which in turn relied on the cytokine IL-17 produced by TH17 cells and innate lymphoid cells (ILCs). NIK-activated noncanonical NF-κB induced expression of IL-23 in DCs, contributing to the maintenance of TH17 cells and type 3 ILCs. Consistent with the dual functions of IL-23 and IL-17 in mucosal immunity and inflammation, NIK deficiency also ameliorated colitis induction. Thus, our data suggest a pivotal role for the NIK signaling axis in regulating DC functions in intestinal immunity and homeostasis.
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Células Dendríticas/inmunología , Homeostasis/inmunología , Inmunidad Mucosa/inmunología , Mucosa Intestinal/inmunología , Proteínas Serina-Treonina Quinasas/inmunología , Animales , Colitis/inmunología , Inmunidad Innata , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Transducción de Señal/inmunología , Quinasa de Factor Nuclear kappa BRESUMEN
NF-κB-inducing kinase (NIK) mediates non-canonical NF-κB signaling downstream of multiple TNF family members, including BAFF, TWEAK, CD40, and OX40, which are implicated in the pathogenesis of systemic lupus erythematosus (SLE). Here, we show that experimental lupus in NZB/W F1 mice can be treated with a highly selective and potent NIK small molecule inhibitor. Both in vitro as well as in vivo, NIK inhibition recapitulates the pharmacological effects of BAFF blockade, which is clinically efficacious in SLE. Furthermore, NIK inhibition also affects T cell parameters in the spleen and proinflammatory gene expression in the kidney, which may be attributable to inhibition of OX40 and TWEAK signaling, respectively. As a consequence, NIK inhibition results in improved survival, reduced renal pathology, and lower proteinuria scores. Collectively, our data suggest that NIK inhibition is a potential therapeutic approach for SLE.
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Linfocitos B/efectos de los fármacos , Riñón/efectos de los fármacos , Lupus Eritematoso Sistémico/inmunología , Inhibidores de Proteínas Quinasas/farmacología , Proteínas Serina-Treonina Quinasas/antagonistas & inhibidores , Linfocitos T/efectos de los fármacos , Animales , Linfocitos B/inmunología , Proliferación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Citocina TWEAK/metabolismo , Células Dendríticas/efectos de los fármacos , Células Dendríticas/inmunología , Modelos Animales de Enfermedad , Expresión Génica/efectos de los fármacos , Humanos , Técnicas In Vitro , Inflamación/genética , Subunidad p40 de la Interleucina-12/efectos de los fármacos , Subunidad p40 de la Interleucina-12/inmunología , Riñón/inmunología , Riñón/patología , Lupus Eritematoso Sistémico/tratamiento farmacológico , Nefritis Lúpica/inmunología , Nefritis Lúpica/patología , Ratones , Ratones Endogámicos NZB , Terapia Molecular Dirigida , Proteinuria/inmunología , Receptores OX40/metabolismo , Transducción de Señal , Bazo/efectos de los fármacos , Bazo/inmunología , Linfocitos T/inmunología , Quinasa de Factor Nuclear kappa BRESUMEN
NF-κB inducing kinase (NIK) is a central component of the noncanonical NF-κB signaling pathway. Although NIK has been extensively studied for its function in the regulation of lymphoid organ development and B-cell maturation, the role of NIK in regulating T cell functions remains unclear and controversial. Using T cell-conditional NIK knockout mice, we here demonstrate that although NIK is dispensable for thymocyte development, it has a cell-intrinsic role in regulating the homeostasis and function of peripheral T cells. T cell-specific NIK ablation reduced the frequency of effector/memory-like T cells and impaired T cell responses to bacterial infection. The T cell-conditional NIK knockout mice were also defective in generation of inflammatory T cells and refractory to the induction of a T cell-dependent autoimmune disease, experimental autoimmune encephalomyelitis. Our data suggest a crucial role for NIK in mediating the generation of effector T cells and their recall responses to antigens. Together, these findings establish NIK as a cell-intrinsic mediator of T cell functions in both immune and autoimmune responses.
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Infecciones Bacterianas/inmunología , Encefalomielitis Autoinmune Experimental/inmunología , Proteínas Serina-Treonina Quinasas/metabolismo , Linfocitos T/fisiología , Timocitos/fisiología , Animales , Autoinmunidad/genética , Diferenciación Celular , Células Cultivadas , Homeostasis , Inmunidad Celular/genética , Memoria Inmunológica , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Ratones Transgénicos , FN-kappa B/metabolismo , Proteínas Serina-Treonina Quinasas/genética , Transducción de Señal/genética , Quinasa de Factor Nuclear kappa BRESUMEN
NF-κB-inducing kinase (NIK) is a primary regulator of the noncanonical NF-κB signaling pathway, which plays a vital role downstream of BAFF, CD40L, lymphotoxin, and other inflammatory mediators. Germline deletion or inactivation of NIK in mice results in the defective development of B cells and secondary lymphoid organs, but the role of NIK in adult animals has not been studied. To address this, we generated mice containing a conditional allele of NIK. Deletion of NIK in adult mice results in decreases in B cell populations in lymph nodes and spleen, similar to what is observed upon blockade of BAFF. Consistent with this, B cells from mice in which NIK is acutely deleted fail to respond to BAFF stimulation in vitro and in vivo. In addition, mice with induced NIK deletion exhibit a significant decrease in germinal center B cells and serum IgA, which is indicative of roles for NIK in additional pathways beyond BAFF signaling. Our conditional NIK-knockout mice may be broadly useful for assessing the postdevelopmental and cell-specific roles of NIK and the noncanonical NF-κB pathway in mice.
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Factor Activador de Células B/genética , Linfocitos B/inmunología , Activación de Linfocitos/genética , Subunidad p52 de NF-kappa B/biosíntesis , Proteínas Serina-Treonina Quinasas/genética , Animales , Linfocitos B/citología , Diferenciación Celular/inmunología , Supervivencia Celular/genética , Supervivencia Celular/inmunología , Mutación de Línea Germinal , Quinasa I-kappa B/metabolismo , Inmunoglobulina A/sangre , Ganglios Linfáticos/citología , Activación de Linfocitos/inmunología , Ratones , Ratones Noqueados , Subunidad p52 de NF-kappa B/genética , Eliminación de Secuencia , Transducción de Señal/genética , Transducción de Señal/inmunología , Bazo/citología , Tamoxifeno/farmacología , Quinasa de Factor Nuclear kappa BRESUMEN
MicroRNAs (miRNAs) exert powerful effects on immunological function by tuning networks of target genes that orchestrate cell activity. We sought to identify miRNAs and miRNA-regulated pathways that control the type 2 helper T cell (TH2 cell) responses that drive pathogenic inflammation in asthma. Profiling miRNA expression in human airway-infiltrating T cells revealed elevated expression of the miRNA miR-19a in asthma. Modulating miR-19 activity altered TH2 cytokine production in both human and mouse T cells, and TH2 cell responses were markedly impaired in cells lacking the entire miR-17â¼92 cluster. miR-19 promoted TH2 cytokine production and amplified inflammatory signaling by direct targeting of the inositol phosphatase PTEN, the signaling inhibitor SOCS1 and the deubiquitinase A20. Thus, upregulation of miR-19a in asthma may be an indicator and a cause of increased TH2 cytokine production in the airways.
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Asma/inmunología , Citocinas/biosíntesis , MicroARNs/inmunología , Células Th2/inmunología , Animales , Asma/genética , Asma/metabolismo , Líquido del Lavado Bronquioalveolar/citología , Ensayos Clínicos como Asunto , Citometría de Flujo , Ensayos Analíticos de Alto Rendimiento , Humanos , Inflamación/genética , Inflamación/inmunología , Inflamación/metabolismo , Ratones , Ratones Transgénicos , Reacción en Cadena de la Polimerasa Multiplex , Células Th2/metabolismo , Regulación hacia ArribaAsunto(s)
Linfocitos B/inmunología , Centro Germinal/inmunología , Inmunoglobulina E/inmunología , Memoria Inmunológica/inmunología , Células Plasmáticas/inmunología , Traslado Adoptivo , Animales , Linfocitos B/citología , Linfocitos B/metabolismo , Diferenciación Celular/genética , Diferenciación Celular/inmunología , Citometría de Flujo , Centro Germinal/citología , Centro Germinal/metabolismo , Proteínas Fluorescentes Verdes/genética , Proteínas Fluorescentes Verdes/metabolismo , Interacciones Huésped-Parásitos/inmunología , Humanos , Cambio de Clase de Inmunoglobulina/genética , Cambio de Clase de Inmunoglobulina/inmunología , Inmunoglobulina E/sangre , Inmunoglobulina G/sangre , Inmunoglobulina G/inmunología , Ratones , Ratones de la Cepa 129 , Ratones Endogámicos C57BL , Ratones Noqueados , Ratones Transgénicos , Microscopía Confocal , Nippostrongylus/inmunología , Nippostrongylus/fisiología , Células Plasmáticas/citología , Células Plasmáticas/metabolismo , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Infecciones por Strongylida/inmunología , Infecciones por Strongylida/parasitologíaRESUMEN
Immunoglobulin E (IgE) antibodies are pathogenic in asthma and allergic diseases, but the in vivo biology of IgE-producing (IgE(+)) cells is poorly understood. A model of the differentiation of IgE(+) B cells proposes that IgE(+) cells develop through a germinal-center IgG1(+) intermediate and that IgE memory resides in the compartment of IgG1(+) memory B cells. Here we have used a reporter mouse expressing green fluorescent protein associated with membrane IgE transcripts (IgE-GFP) to assess in vivo IgE responses. In contrast to the IgG1-centered model of IgE switching and memory, we found that IgE(+) cells developed through a germinal-center IgE(+) intermediate to form IgE(+) memory B cells and plasma cells. Our studies delineate a new model for the in vivo biology of IgE switching and memory.
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Linfocitos B/citología , Diferenciación Celular/inmunología , Centro Germinal/citología , Inmunoglobulina E/inmunología , Memoria Inmunológica/inmunología , Células Plasmáticas/inmunología , Traslado Adoptivo , Animales , Linfocitos B/inmunología , Separación Celular , Ensayo de Inmunoadsorción Enzimática , Citometría de Flujo , Técnica del Anticuerpo Fluorescente , Técnicas de Sustitución del Gen , Centro Germinal/inmunología , Humanos , Cambio de Clase de Inmunoglobulina/inmunología , Ratones , Ratones Endogámicos C57BL , Microscopía Confocal , Células Plasmáticas/citología , Reacción en Cadena de la Polimerasa de Transcriptasa InversaRESUMEN
IgE-mediated hypersensitivity is central to the pathogenesis of asthma and other allergic diseases. Although neutralization of serum IgE with IgE-specific antibodies is in general an efficacious treatment for allergic asthma, one limitation of this approach is its lack of effect on IgE production. Here, we have developed a strategy to disrupt IgE production by generating monoclonal antibodies that target a segment of membrane IgE on human IgE-switched B cells that is not present in serum IgE. This segment is known as the M1' domain, and using genetically modified mice that contain the human M1' domain inserted into the mouse IgE locus, we demonstrated that M1'-specific antibodies reduced serum IgE and IgE-producing plasma cells in vivo, without affecting other immunoglobulin isotypes. M1'-specific antibodies were effective when delivered prophylactically and therapeutically in mouse models of immunization, allergic asthma, and Nippostrongylus brasiliensis infection, likely by inducing apoptosis of IgE-producing B cells. In addition, we generated a humanized M1'-specific antibody that was active on primary human cells in vivo, as determined by its reduction of serum IgE levels and IgE plasma cell numbers in a human PBMC-SCID mouse model. Thus, targeting of human IgE-producing B cells with apoptosis-inducing M1'-specific antibodies may be a novel treatment for asthma and allergy.
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Anticuerpos Monoclonales/farmacología , Anticuerpos/inmunología , Linfocitos B/efectos de los fármacos , Hipersensibilidad Inmediata/inmunología , Ratones SCID/inmunología , Animales , Anticuerpos Monoclonales/inmunología , Asma/inmunología , Linfocitos B/inmunología , Humanos , Hipersensibilidad/inmunología , Inmunización , Ratones , Ratones Transgénicos , Nippostrongylus/efectos de los fármacos , Nippostrongylus/inmunologíaRESUMEN
One of the factors that contributes to the pathogenesis of acne is Propionibacterium acnes; yet, the molecular mechanism by which P. acnes induces inflammation is not known. Recent studies have demonstrated that microbial agents trigger cytokine responses via Toll-like receptors (TLRs). We investigated whether TLR2 mediates P. acnes-induced cytokine production in acne. Transfection of TLR2 into a nonresponsive cell line was sufficient for NF-kappa B activation in response to P. acnes. In addition, peritoneal macrophages from wild-type, TLR6 knockout, and TLR1 knockout mice, but not TLR2 knockout mice, produced IL-6 in response to P. acnes. P. acnes also induced activation of IL-12 p40 promoter activity via TLR2. Furthermore, P. acnes induced IL-12 and IL-8 protein production by primary human monocytes and this cytokine production was inhibited by anti-TLR2 blocking Ab. Finally, in acne lesions, TLR2 was expressed on the cell surface of macrophages surrounding pilosebaceous follicles. These data suggest that P. acnes triggers inflammatory cytokine responses in acne by activation of TLR2. As such, TLR2 may provide a novel target for treatment of this common skin disease.