RESUMEN

Alveolar epithelial cell (AEC) injury is central to the pathogenesis of pulmonary fibrosis. Epithelial FGF (fibroblast growth factor) signaling is essential for recovery from hyperoxia- and influenza-induced lung injury, and treatment with FGFs is protective in experimental lung injury. The cell types involved in the protective effect of FGFs are not known. We hypothesized that FGF signaling in type II AECs (AEC2s) is critical in bleomycin-induced lung injury and fibrosis. To test this hypothesis, we generated mice with tamoxifen-inducible deletion of FGFR1-3 (fibroblast growth factor receptors 1, 2, and 3) in surfactant protein C-positive (SPC+) AEC2s (SPC triple conditional knockout [SPC-TCKO]). In the absence of injury, SPC-TCKO mice had fewer AEC2s, decreased Sftpc (surfactant protein C gene) expression, increased alveolar diameter, and increased collagen deposition. After intratracheal bleomycin administration, SPC-TCKO mice had increased mortality, lung edema, and BAL total protein, and flow cytometry and immunofluorescence revealed a loss of AEC2s. To reduce mortality of SPC-TCKO mice to less than 50%, a 25-fold dose reduction of bleomycin was required. Surviving bleomycin-injured SPC-TCKO mice had increased collagen deposition, fibrosis, and ACTA2 expression and decreased epithelial gene expression. Inducible inactivation of individual Fgfr2 or Fgfr3 revealed that Fgfr2, but not Fgfr3, was responsible for the increased mortality and lung injury after bleomycin administration. In conclusion, AEC2-specific FGFR2 is critical for survival in response to bleomycin-induced lung injury. These data also suggest that a population of SPC+ AEC2s require FGFR2 signaling for maintenance in the adult lung. Preventing epithelial FGFR inhibition and/or activating FGFRs in alveolar epithelium may therefore represent a novel approach to treating lung injury and reducing fibrosis.

Asunto(s)

Homeostasis , Lesión Pulmonar/inducido químicamente , Lesión Pulmonar/metabolismo , Receptor Tipo 2 de Factor de Crecimiento de Fibroblastos/metabolismo , Animales , Bleomicina/administración & dosificación , Linaje de la Célula , Colágeno/metabolismo , Eliminación de Gen , Regulación de la Expresión Génica , Lesión Pulmonar/genética , Lesión Pulmonar/patología , Ratones Noqueados , Especificidad de Órganos , Fenotipo , Alveolos Pulmonares/patología , Proteína C Asociada a Surfactante Pulmonar/metabolismo , Receptor Tipo 2 de Factor de Crecimiento de Fibroblastos/deficiencia , Análisis de Supervivencia

RESUMEN

FGFR signaling is critical to development and disease pathogenesis, initiating phosphorylation-driven signaling cascades, notably the RAS-RAF-MEK-ERK and PI3 K-AKT cascades. PTEN antagonizes FGFR signaling by reducing AKT and ERK activation. Mouse lenses lacking FGFR2 exhibit microphakia and reduced ERK and AKT phosphorylation, widespread apoptosis, and defective lens fiber cell differentiation. In contrast, simultaneous deletion of both Fgfr2 and Pten restores ERK and AKT activation levels as well as lens size, cell survival and aspects of fiber cell differentiation; however, the molecular basis of this "rescue" remains undefined. We performed transcriptomic analysis by RNA sequencing of mouse lenses with conditional deletion of Fgfr2, Pten or both Fgfr2 and Pten, which reveal new molecular mechanisms that uncover how FGFR2 and PTEN signaling interact during development. The FGFR2-deficient lens transcriptome demonstrates overall loss of fiber cell identity with deregulated expression of 1448 genes. We find that ~ 60% of deregulated genes return to normal expression levels in lenses lacking both Fgfr2 and Pten. Further, application of customized filtering parameters to these RNA-seq data sets identified 68 high-priority candidate genes. Bioinformatics analyses showed that the cis-binding motif of a high-priority homeodomain transcription factor, NKX6-1, was present in the putative promoters of ~ 78% of these candidates. Finally, biochemical reporter assays demonstrate that NKX6-1 activated the expression of the high-priority candidate Rasgrp1, a RAS-activating protein. Together, these data define a novel regulatory module in which NKX6-1 directly activates Rasgrp1 expression to restore the balance of ERK and AKT activation, thus providing new insights into alternate regulation of FGFR downstream events.

Asunto(s)

Regulación de la Expresión Génica , Factores de Intercambio de Guanina Nucleótido/metabolismo , Proteínas de Homeodominio/metabolismo , Microftalmía/prevención & control , Fosfohidrolasa PTEN/deficiencia , Receptor Tipo 2 de Factor de Crecimiento de Fibroblastos/deficiencia , Transcriptoma , Animales , Diferenciación Celular , Proliferación Celular , Factores de Intercambio de Guanina Nucleótido/genética , Secuenciación de Nucleótidos de Alto Rendimiento , Proteínas de Homeodominio/genética , Ratones , Ratones Noqueados , Microftalmía/etiología , Microftalmía/patología , Fosforilación , Transducción de Señal

RESUMEN

Fibroblast growth factor (FGF) signaling pathways have well-established roles in skeletal development, with essential functions in both chondrogenesis and osteogenesis. In mice, previous conditional knockout studies suggested distinct roles for FGF receptor 1 (FGFR1) signaling at different stages of osteogenesis and a role for FGFR2 in osteoblast maturation. However, the potential for redundancy among FGFRs and the mechanisms and consequences of stage-specific osteoblast lineage regulation were not addressed. Here, we conditionally inactivate Fgfr1 and Fgfr2 in mature osteoblasts with an Osteocalcin (OC)-Cre or Dentin matrix protein 1 (Dmp1)-CreER driver. We find that young mice lacking both receptors or only FGFR1 are phenotypically normal. However, between 6 and 12 weeks of age, OC-Cre Fgfr1/Fgfr2 double- and Fgfr1 single-conditional knockout mice develop a high bone mass phenotype with increased periosteal apposition, increased and disorganized endocortical bone with increased porosity, and biomechanical properties that reflect increased bone mass but impaired material properties. Histopathological and gene expression analyses show that this phenotype is preceded by a striking loss of osteocytes and accompanied by activation of the Wnt/ß-catenin signaling pathway. These data identify a role for FGFR1 signaling in mature osteoblasts/osteocytes that is directly or indirectly required for osteocyte survival and regulation of bone mass during postnatal bone growth. © 2019 American Society for Bone and Mineral Research.

Asunto(s)

Desarrollo Óseo , Osteoblastos/metabolismo , Osteocitos/patología , Receptor Tipo 1 de Factor de Crecimiento de Fibroblastos/deficiencia , Receptor Tipo 2 de Factor de Crecimiento de Fibroblastos/deficiencia , Alelos , Animales , Fenómenos Biomecánicos , Remodelación Ósea , Muerte Celular , Supervivencia Celular , Hueso Cortical/patología , Proteínas de la Matriz Extracelular/metabolismo , Femenino , Regulación de la Expresión Génica , Masculino , Ratones Endogámicos C57BL , Ratones Noqueados , Tamaño de los Órganos , Osteoblastos/patología , Osteocitos/metabolismo , Receptor Tipo 1 de Factor de Crecimiento de Fibroblastos/metabolismo , Receptor Tipo 2 de Factor de Crecimiento de Fibroblastos/metabolismo , Vía de Señalización Wnt

RESUMEN

Merkel cells are innervated mechanosensory cells responsible for light-touch sensations. In murine dorsal skin, Merkel cells are located in touch domes and found in the epidermis around primary hairs. While it has been shown that Merkel cells are skin epithelial cells, the progenitor cell population that gives rise to these cells is unknown. Here, we show that during embryogenesis, SOX9-positive (+) cells inside hair follicles, which were previously known to give rise to hair follicle stem cells (HFSCs) and cells of the hair follicle lineage, can also give rise to Merkel Cells. Interestingly, while SOX9 is critical for HFSC specification, it is dispensable for Merkel cell formation. Conversely, FGFR2 is required for Merkel cell formation but is dispensable for HFSCs. Together, our studies uncover SOX9(+) cells as precursors of Merkel cells and show the requirement for FGFR2-mediated epithelial signalling in Merkel cell specification.

Asunto(s)

Células Madre Embrionarias/citología , Células Madre Embrionarias/metabolismo , Folículo Piloso/embriología , Folículo Piloso/metabolismo , Células de Merkel/citología , Células de Merkel/metabolismo , Receptor Tipo 2 de Factor de Crecimiento de Fibroblastos/metabolismo , Factor de Transcripción SOX9/metabolismo , Animales , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/genética , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/metabolismo , Linaje de la Célula , Femenino , Técnicas de Inactivación de Genes , Proteínas Fluorescentes Verdes/genética , Proteínas Fluorescentes Verdes/metabolismo , Folículo Piloso/citología , Proteínas Hedgehog/genética , Proteínas Hedgehog/metabolismo , Proteínas con Homeodominio LIM/metabolismo , Masculino , Ratones , Ratones Noqueados , Ratones Transgénicos , Modelos Biológicos , Factores de Transcripción NFATC/genética , Factores de Transcripción NFATC/metabolismo , Embarazo , Receptor Tipo 2 de Factor de Crecimiento de Fibroblastos/deficiencia , Receptor Tipo 2 de Factor de Crecimiento de Fibroblastos/genética , Factor de Transcripción SOX9/antagonistas & inhibidores , Factor de Transcripción SOX9/genética , Transducción de Señal , Factores de Transcripción/metabolismo

RESUMEN

Hypertrophy, hyperplasia and altered mucus secretion from the respiratory submucosal glands (SMG) are characteristics of airway diseases such as cystic fibrosis, asthma and chronic bronchitis. More commonly, hyper-secretion of the nasal SMGs contributes to allergic rhinitis and upper airway infection. Considering the role of these glands in disease states, there is a significant dearth in understanding the molecular signals that regulate SMG development and patterning. Due to the imperative role of FGF signalling during the development of other branched structures, we investigated the role of Fgf10 during initiation and branching morphogenesis of murine nasal SMGs. Fgf10 is expressed in the mesenchyme around developing SMGs while expression of its receptor Fgfr2 is seen within glandular epithelial cells. In the Fgf10 null embryo, Steno's gland and the maxillary sinus gland were completely absent while other neighbouring nasal glands showed normal duct elongation but defective branching. Interestingly, the medial nasal glands were present in Fgf10 homozygotes but missing in Fgfr2b mutants, with expression of Fgf7 specifically expressed around these developing glands, indicating that Fgf7 might compensate for loss of Fgf10 in this group of glands. Intriguingly the lateral nasal glands were only mildly affected by loss of FGF signalling, while these glands were missing in Eda mutant mice, where the Steno's and maxillary sinus gland developed as normal. This analysis reveals that regulation of nasal gland development is complex with different subsets of glands being regulated by different signalling pathways. This analysis helps shed light on the nasal gland defects observed in patients with hypohidrotic ectodermal dysplasia (HED) (defect EDA pathway) and LADD syndrome (defect FGFR2b pathway).

Asunto(s)

Ectodisplasinas/fisiología , Glándulas Exocrinas/embriología , Factor 10 de Crecimiento de Fibroblastos/fisiología , Receptor Tipo 2 de Factor de Crecimiento de Fibroblastos/fisiología , Transducción de Señal/fisiología , Animales , Ectodisplasinas/deficiencia , Ectodisplasinas/genética , Resección Endoscópica de la Mucosa , Glándulas Exocrinas/metabolismo , Glándulas Exocrinas/ultraestructura , Femenino , Factor 10 de Crecimiento de Fibroblastos/deficiencia , Factor 10 de Crecimiento de Fibroblastos/genética , Factor 7 de Crecimiento de Fibroblastos/fisiología , Masculino , Seno Maxilar/embriología , Seno Maxilar/ultraestructura , Mesodermo/metabolismo , Ratones , Morfogénesis , Mucosa Nasal/embriología , Mucosa Nasal/ultraestructura , Receptor Tipo 2 de Factor de Crecimiento de Fibroblastos/deficiencia , Receptor Tipo 2 de Factor de Crecimiento de Fibroblastos/genética

RESUMEN

Fibroblast growth factor receptor 2 (FGFR2) signaling is critical for proper craniofacial development. A gain-of-function mutation in the 2c splice variant of the receptor's gene is associated with Crouzon syndrome, which is characterized by craniosynostosis, the premature fusion of one or more of the cranial vault sutures, leading to craniofacial maldevelopment. Insight into the molecular mechanism of craniosynostosis has identified the ERK-MAPK signaling cascade as a critical regulator of suture patency. The aim of this study is to investigate the role of FGFR2c-induced ERK-MAPK activation in the regulation of coronal suture development. Loss-of-function and gain-of-function Fgfr2c mutant mice have overlapping phenotypes, including coronal synostosis and craniofacial dysmorphia. In vivo analysis of coronal sutures in loss-of-function and gain-of-function models demonstrated fundamentally different pathogenesis underlying coronal suture synostosis. Calvarial osteoblasts from gain-of-function mice demonstrated enhanced osteoblastic function and maturation with concomitant increase in ERK-MAPK activation. In vitro inhibition with the ERK protein inhibitor U0126 mitigated ERK protein activation levels with a concomitant reduction in alkaline phosphatase activity. This study identifies FGFR2c-mediated ERK-MAPK signaling as a key mediator of craniofacial growth and coronal suture development. Furthermore, our results solve the apparent paradox between loss-of-function and gain-of-function FGFR2c mutants with respect to coronal suture synostosis.

Asunto(s)

Suturas Craneales/embriología , Disostosis Craneofacial/embriología , Sistema de Señalización de MAP Quinasas/fisiología , Receptor Tipo 2 de Factor de Crecimiento de Fibroblastos/fisiología , Fosfatasa Alcalina/biosíntesis , Fosfatasa Alcalina/genética , Animales , Butadienos/farmacología , Células Cultivadas , Suturas Craneales/anomalías , Activación Enzimática/efectos de los fármacos , Ratones , Ratones Noqueados , Mutación , Nitrilos/farmacología , Osteoblastos/metabolismo , Osteoblastos/patología , Osteoclastos/metabolismo , Osteogénesis/fisiología , Fenotipo , Isoformas de Proteínas/antagonistas & inhibidores , Isoformas de Proteínas/fisiología , Receptor Tipo 2 de Factor de Crecimiento de Fibroblastos/antagonistas & inhibidores , Receptor Tipo 2 de Factor de Crecimiento de Fibroblastos/deficiencia , Receptor Tipo 2 de Factor de Crecimiento de Fibroblastos/genética , Sinostosis/genética , Sinostosis/patología

RESUMEN

Fibroblast growth factor (FGF) signaling is cardioprotective in various models of myocardial infarction. FGF receptors (FGFRs) are expressed in multiple cell types in the adult heart, but the cell type-specific FGFR signaling that mediates different cardioprotective endpoints is not known. To determine the requirement for FGFR signaling in endothelium in cardiac ischemia-reperfusion injury, we conditionally inactivated the Fgfr1 and Fgfr2 genes in endothelial cells with Tie2-Cre (Tie2-Cre, Fgfr1(f/f), Fgfr2(f/f) DCKO mice). Tie2-Cre, Fgfr1(f/f), Fgfr2(f/f) DCKO mice had normal baseline cardiac morphometry, function, and vessel density. When subjected to closed-chest, regional cardiac ischemia-reperfusion injury, Tie2-Cre, Fgfr1(f/f), Fgfr2(f/f) DCKO mice showed a significantly increased hypokinetic area at 7 days, but not 1 day, after reperfusion. Tie2-Cre, Fgfr1(f/f), Fgfr2(f/f) DCKO mice also showed significantly worsened cardiac function compared with controls at 7 days but not 1 day after reperfusion. Pathophysiological analysis showed significantly decreased vessel density, increased endothelial cell apoptosis, and worsened tissue hypoxia in the peri-infarct area at 7 days following reperfusion. Notably, Tie2-Cre, Fgfr1(f/f), Fgfr2(f/f) DCKO mice showed no impairment in the cardiac hypertrophic response. These data demonstrate an essential role for FGFR1 and FGFR2 in endothelial cells for cardiac functional recovery and vascular remodeling following in vivo cardiac ischemia-reperfusion injury, without affecting the cardiac hypertrophic response. This study suggests the potential for therapeutic benefit from activation of endothelial FGFR pathways following ischemic injury to the heart.

Asunto(s)

Células Endoteliales/metabolismo , Infarto del Miocardio/metabolismo , Daño por Reperfusión Miocárdica/metabolismo , Receptor Tipo 1 de Factor de Crecimiento de Fibroblastos/metabolismo , Receptor Tipo 2 de Factor de Crecimiento de Fibroblastos/metabolismo , Remodelación Vascular , Animales , Apoptosis , Capilares/metabolismo , Capilares/patología , Capilares/fisiopatología , Cardiomegalia/genética , Cardiomegalia/metabolismo , Cardiomegalia/patología , Cardiomegalia/fisiopatología , Modelos Animales de Enfermedad , Células Endoteliales/patología , Femenino , Masculino , Ratones de la Cepa 129 , Ratones Endogámicos C57BL , Ratones Noqueados , Infarto del Miocardio/genética , Infarto del Miocardio/patología , Infarto del Miocardio/fisiopatología , Daño por Reperfusión Miocárdica/genética , Daño por Reperfusión Miocárdica/patología , Daño por Reperfusión Miocárdica/fisiopatología , Miocardio/metabolismo , Miocardio/patología , Neovascularización Fisiológica , Receptor Tipo 1 de Factor de Crecimiento de Fibroblastos/deficiencia , Receptor Tipo 1 de Factor de Crecimiento de Fibroblastos/genética , Receptor Tipo 2 de Factor de Crecimiento de Fibroblastos/deficiencia , Receptor Tipo 2 de Factor de Crecimiento de Fibroblastos/genética , Recuperación de la Función , Transducción de Señal , Volumen Sistólico , Factores de Tiempo , Función Ventricular Izquierda

RESUMEN

Chronic skin inflammation resulting from a defective epidermal barrier is a hallmark of atopic dermatitis (AD). We previously demonstrated that mice lacking FGF receptors 1 and 2 in keratinocytes (K5-R1/R2 mice) develop an AD-like chronic dermatitis as a result of an impaired epidermal barrier. Here, we show that γδ T cells, which rapidly respond to various insults, accumulate in the epidermis of K5-R1/R2 mice before the development of histological abnormalities. Their number and activation further increase as the phenotype progresses, most likely as a consequence of increased expression of Il-2 and Il-7 and the stress-induced proteins Rae-1, H60c, Mult1, PlexinB2, and Skint1. To determine the role of γδ T cells in the skin phenotype, we generated quadruple mutant K5-R1/-R2 mice lacking γδ T cells. Surprisingly, loss of γδ T cells did not or only marginally affect keratinocyte proliferation, epidermal thickness, epidermal barrier function, and accumulation and activation of different immune cells in the skin of K5-R1/R2 mice, possibly due to partial compensation by αß T cells. These results demonstrate that γδ T cells do not contribute to the development or maintenance of chronic inflammation in response to a defect in the epidermal barrier.

Asunto(s)

Dermatitis/inmunología , Queratinocitos/inmunología , Receptor Tipo 1 de Factor de Crecimiento de Fibroblastos/deficiencia , Receptor Tipo 2 de Factor de Crecimiento de Fibroblastos/deficiencia , Receptores de Antígenos de Linfocitos T gamma-delta/deficiencia , Subgrupos de Linfocitos T/inmunología , Animales , Proteínas Portadoras/genética , Proteínas Portadoras/inmunología , Proliferación Celular , Enfermedad Crónica , Dermatitis/genética , Dermatitis/patología , Modelos Animales de Enfermedad , Epidermis/inmunología , Epidermis/patología , Regulación de la Expresión Génica , Antígenos de Histocompatibilidad Clase I/genética , Antígenos de Histocompatibilidad Clase I/inmunología , Inmunoglobulinas/genética , Inmunoglobulinas/inmunología , Interleucina-2/genética , Interleucina-2/inmunología , Interleucina-7/genética , Interleucina-7/inmunología , Queratinocitos/patología , Activación de Linfocitos , Depleción Linfocítica , Proteínas de la Membrana , Ratones , Ratones Transgénicos , Antígenos de Histocompatibilidad Menor/genética , Antígenos de Histocompatibilidad Menor/inmunología , Proteínas del Tejido Nervioso/genética , Proteínas del Tejido Nervioso/inmunología , Proteínas Asociadas a Matriz Nuclear/genética , Proteínas Asociadas a Matriz Nuclear/inmunología , Proteínas de Transporte Nucleocitoplasmático/genética , Proteínas de Transporte Nucleocitoplasmático/inmunología , Receptor Tipo 1 de Factor de Crecimiento de Fibroblastos/genética , Receptor Tipo 1 de Factor de Crecimiento de Fibroblastos/inmunología , Receptor Tipo 2 de Factor de Crecimiento de Fibroblastos/genética , Receptor Tipo 2 de Factor de Crecimiento de Fibroblastos/inmunología , Receptores de Antígenos de Linfocitos T gamma-delta/genética , Receptores de Antígenos de Linfocitos T gamma-delta/inmunología , Transducción de Señal , Subgrupos de Linfocitos T/patología

RESUMEN

BACKGROUND: The developmental processes of the genital tubercle (GT), the anlage of the external genitalia, possess several developmental aspects, including GT outgrowth, urethral tube formation, and epithelial differentiation of the urethra. The GT comprises the mesenchyme derived from the lateral mesoderm, ectodermal epithelium, and endodermal epithelium (embryonic urethral epithelium). The three tissue layers develop the GT coordinately. RESULTS: Around the initial stage of GT outgrowth (E11.5), FGF signaling was detected in the mesenchyme of the GT. FGF signaling was detected in the three tissue layers of the GT around the early stage of urethral formation (E13.5). Subsequently, FGF signaling was predominantly detected in the urethral epithelium (E14.5). Tissue-specific roles of FGF signaling in GT development were revealed by conditional Fgfr gene knockout approaches. Mesenchymal FGF signaling in the early-stage GT is required for its outgrowth. Ectodermal FGF signaling in the GT is required for the differentiation of the ectoderm and urethral epithelium at their junction to form the proper urethral tube. Endodermal FGF signaling in the GT is required for the stratification and cell adhesive characteristics of the urethral epithelium. CONCLUSIONS: The current study suggests that spatiotemporally regulated FGF signaling plays tissue-specific roles in multiple processes of external genitalia development.

Asunto(s)

Factores de Crecimiento de Fibroblastos/fisiología , Genitales Femeninos/embriología , Genitales Masculinos/embriología , Receptor Tipo 1 de Factor de Crecimiento de Fibroblastos/fisiología , Receptor Tipo 2 de Factor de Crecimiento de Fibroblastos/fisiología , Animales , Proteínas Morfogenéticas Óseas/fisiología , Adhesión Celular , Supervivencia Celular , Femenino , Regulación del Desarrollo de la Expresión Génica , Genes Reporteros , Genitales Femeninos/fisiología , Genitales Masculinos/fisiología , Estratos Germinativos/citología , Estratos Germinativos/metabolismo , Proteínas Hedgehog/fisiología , Masculino , Mesodermo/fisiología , Ratones , Ratones Noqueados , Especificidad de Órganos , Receptor Tipo 1 de Factor de Crecimiento de Fibroblastos/deficiencia , Receptor Tipo 1 de Factor de Crecimiento de Fibroblastos/genética , Receptor Tipo 2 de Factor de Crecimiento de Fibroblastos/deficiencia , Receptor Tipo 2 de Factor de Crecimiento de Fibroblastos/genética , Proteínas Recombinantes de Fusión/metabolismo , Uretra/embriología , Uretra/fisiología , Proteínas Wnt/fisiología , Vía de Señalización Wnt

RESUMEN

While urothelial signals, including sonic hedgehog (Shh), drive bladder mesenchyme differentiation, it is unclear which pathways within the mesenchyme are critical for its development. Studies have shown that fibroblast growth factor receptor (Fgfr)2 is necessary for kidney and ureter mesenchymal development. The objective of the present study was to determine the role of Fgfr2 in the bladder mesenchyme. We used Tbx18cre mice to delete Fgfr2 in the bladder mesenchyme (Fgfr2(BM-/-)). We performed three-dimensional reconstructions, quantitative real-time PCR, in situ hybridization, immunolabeling, ELISAs, immunoblot analysis, void stain on paper, ex vivo bladder sheet assays, and in vivo decerebrated cystometry. Compared with control bladders, embryonic day 16.5 (E16.5) Fgfr2(BM-/-) bladders had thin muscle layers with less α-smooth muscle actin and thickened lamina propria with increased collagen type Ia and IIIa that intruded into the muscle. The reciprocal changes in mutant layer thicknesses appeared partly due to a cell fate switch. From postnatal days 1 to 30, Fgfr2(BM-/-) bladders demonstrated progressive muscle loss and increased collagen expression. Postnatal Fgfr2(BM-/-) bladder sheets exhibited decreased agonist-mediated contractility and increased passive stretch tension versus control bladder sheets. Cystometry revealed high baseline and threshold pressures and shortened intercontractile intervals in Fgfr2(BM-/-) versus control bladders. Mechanistically, whereas Shh expression appeared normal, mRNA and protein readouts of hedgehog activity were increased in E16.5 Fgfr2(BM-/-) versus control bladders. Moreover, E16.5 Fgfr2(BM-/-) bladders exhibited higher levels of Cdo and Boc, hedgehog coreceptors that enhance sensitivity to Shh, compared with control bladders. In conclusion, loss of Fgfr2 in the bladder mesenchyme leads to abnormal bladder morphology and decreased compliance and contractility.

Asunto(s)

Tipificación del Cuerpo , Mesodermo/metabolismo , Músculo Liso/metabolismo , Receptor Tipo 2 de Factor de Crecimiento de Fibroblastos/metabolismo , Vejiga Urinaria/metabolismo , Animales , Apoptosis , Moléculas de Adhesión Celular/metabolismo , Diferenciación Celular , Linaje de la Célula , Proliferación Celular , Adaptabilidad , Fibrosis , Regulación del Desarrollo de la Expresión Génica , Genotipo , Edad Gestacional , Proteínas Hedgehog/metabolismo , Inmunoglobulina G/metabolismo , Masculino , Mesodermo/anomalías , Ratones Noqueados , Contracción Muscular , Músculo Liso/anomalías , Músculo Liso/fisiopatología , Tamaño de los Órganos , Fenotipo , Receptor Tipo 2 de Factor de Crecimiento de Fibroblastos/deficiencia , Receptor Tipo 2 de Factor de Crecimiento de Fibroblastos/genética , Receptores de Superficie Celular/metabolismo , Transducción de Señal , Vejiga Urinaria/anomalías , Vejiga Urinaria/fisiopatología , Urodinámica

RESUMEN

FGF signaling is essential for mammary gland development, yet the mechanisms by which different members of the FGF family control stem cell function and epithelial morphogenesis in this tissue are not well understood. Here, we have examined the requirement of Fgfr2 in mouse mammary gland morphogenesis using a postnatal organ regeneration model. We found that tissue regeneration from basal stem cells is a multistep event, including luminal differentiation and subsequent epithelial branching morphogenesis. Basal cells lacking Fgfr2 did not generate an epithelial network owing to a failure in luminal differentiation. Moreover, Fgfr2 null epithelium was unable to undergo ductal branch initiation and elongation due to a deficiency in directional migration. We identified FGF10 and FGF2 as stromal ligands that control distinct aspects of mammary ductal branching. FGF10 regulates branch initiation, which depends on directional epithelial migration. By contrast, FGF2 controls ductal elongation, requiring cell proliferation and epithelial expansion. Together, our data highlight a pleiotropic role of Fgfr2 in stem cell differentiation and branch initiation, and reveal that different FGF ligands regulate distinct aspects of epithelial behavior.

Asunto(s)

Epitelio/crecimiento & desarrollo , Factor 10 de Crecimiento de Fibroblastos/metabolismo , Factor 2 de Crecimiento de Fibroblastos/metabolismo , Glándulas Mamarias Animales/citología , Glándulas Mamarias Animales/metabolismo , Morfogénesis , Animales , Animales Recién Nacidos , Muerte Celular/efectos de los fármacos , Diferenciación Celular/efectos de los fármacos , Diferenciación Celular/genética , Movimiento Celular/efectos de los fármacos , Movimiento Celular/genética , Polaridad Celular/efectos de los fármacos , Polaridad Celular/genética , Proliferación Celular/efectos de los fármacos , Células Epiteliales/citología , Células Epiteliales/efectos de los fármacos , Células Epiteliales/enzimología , Epitelio/efectos de los fármacos , Epitelio/metabolismo , Femenino , Factor 10 de Crecimiento de Fibroblastos/genética , Factor 10 de Crecimiento de Fibroblastos/farmacología , Regulación del Desarrollo de la Expresión Génica/efectos de los fármacos , Ligandos , Glándulas Mamarias Animales/crecimiento & desarrollo , Metaloproteinasas de la Matriz/metabolismo , Ratones , Ratones Desnudos , Quinasas de Proteína Quinasa Activadas por Mitógenos/metabolismo , Morfogénesis/efectos de los fármacos , Morfogénesis/genética , Fosfatidilinositol 3-Quinasas/metabolismo , Receptor Tipo 2 de Factor de Crecimiento de Fibroblastos/deficiencia , Receptor Tipo 2 de Factor de Crecimiento de Fibroblastos/metabolismo , Regeneración/efectos de los fármacos , Regeneración/genética , Transducción de Señal/efectos de los fármacos , Transducción de Señal/genética , Células Madre/citología , Células Madre/efectos de los fármacos , Células Madre/metabolismo , Células del Estroma/citología , Células del Estroma/efectos de los fármacos , Células del Estroma/metabolismo

RESUMEN

The optic fissure (OF) is a transient opening on the ventral side of the developing vertebrate eye that closes before nearly all retinal progenitor cell differentiation has occurred. Failure to close the OF results in coloboma, a congenital disease that is a major cause of childhood blindness. Although human genetic studies and animal models have linked a number of genes to coloboma, the cellular and molecular mechanisms driving the closure of the OF are still largely unclear. In this study, we used Cre-LoxP-mediated conditional removal of fibroblast growth factor (FGF) receptors, Fgfr1 and Fgfr2, from the developing optic cup (OC) to show that FGF signaling regulates the closing of the OF. Our molecular, cellular and transcriptome analyses of Fgfr1 and Fgfr2 double conditional knockout OCs suggest that FGF signaling controls the OF closure through modulation of retinal progenitor cell proliferation, fate specification and morphological changes. Furthermore, Fgfr1 and Fgfr2 double conditional mutant retinal progenitor cells fail to initiate retinal ganglion cell (RGC) genesis. Taken together, our mouse genetic studies reveal that FGF signaling is essential for OF morphogenesis and RGC development.

Asunto(s)

Coloboma/metabolismo , Factores de Crecimiento de Fibroblastos/metabolismo , Retina/citología , Transducción de Señal , Animales , Coloboma/patología , Perfilación de la Expresión Génica , Ratones , Ratones Noqueados , Neurogénesis , Receptor Tipo 1 de Factor de Crecimiento de Fibroblastos/deficiencia , Receptor Tipo 1 de Factor de Crecimiento de Fibroblastos/genética , Receptor Tipo 1 de Factor de Crecimiento de Fibroblastos/metabolismo , Receptor Tipo 2 de Factor de Crecimiento de Fibroblastos/deficiencia , Receptor Tipo 2 de Factor de Crecimiento de Fibroblastos/genética , Receptor Tipo 2 de Factor de Crecimiento de Fibroblastos/metabolismo , Células Ganglionares de la Retina/citología , Células Ganglionares de la Retina/metabolismo , Células Madre/citología , Células Madre/metabolismo

RESUMEN

BACKGROUND: Homozygous null mutation of the fibroblast growth factor receptor 2IIIb (Fgfr2IIIb) gene in mice results in 42% of embryos developing duodenal atresias. Retinaldehyde dehydrogenase 2 (Raldh2, a gene critical for the generation of retinoic acid) is expressed in the mouse duodenum during the temporal window when duodenal atresias form. Raldh2 is critical for the normal development of the pancreatoduodenal region; therefore, we were interested in the effect of a Raldh2 mutation on duodenal atresia formation. To test this, we rendered Fgfr2IIIb(-/-) embryos haploinsufficient for the Raldh2 and examined these embryos for the incidence and severity of duodenal atresia. METHODS: Control embryos, Fgfr2IIIb(-/-) mutants, and Fgfr2IIIb(-/-); Raldh2(+/-) mutants were harvested at embryonic day 18.5, genotyped, and fixed overnight. Intestinal tracts were isolated. The type and severity of duodenal atresia was documented. RESULTS: A total of 97 Fgfr2IIIb(-/-) embryos were studied; 44 had duodenal atresias, and 41 of these presented as type III. In the 70 Fgfr2IIIb(-/-); Raldh2(+/-) embryos studied, a lesser incidence of duodenal atresia was seen (15 of 70; P = .0017; Fisher exact test). Atresia severity was also decreased; there were 12 embryos with type I atresias, 3 with type II atresias, and 0 with type III atresias (P < 2.81E-013; Fisher exact test). CONCLUSION: Haploinsufficiency of Raldh2 decreases the incidence and severity of duodenal atresia in the Fgfr2IIIb(-/-) model. The ability to alter defect severity through manipulation of a single gene in a specific genetic background has potentially important implications for understanding the mechanisms by which intestinal atresias arise.

Asunto(s)

Aldehído Oxidorreductasas/deficiencia , Aldehído Oxidorreductasas/genética , Obstrucción Duodenal/congénito , Obstrucción Duodenal/genética , Atresia Intestinal/genética , Receptor Tipo 2 de Factor de Crecimiento de Fibroblastos/deficiencia , Receptor Tipo 2 de Factor de Crecimiento de Fibroblastos/genética , Animales , Obstrucción Duodenal/embriología , Obstrucción Duodenal/metabolismo , Femenino , Haploinsuficiencia , Imagenología Tridimensional , Hibridación in Situ , Atresia Intestinal/embriología , Atresia Intestinal/metabolismo , Masculino , Ratones , Ratones Noqueados , Penetrancia , Embarazo

RESUMEN

Fibroblast growth factor receptor (FGFR) signaling determines the size of the cerebral cortex by regulating the amplification of radial glial stem cells, and participates in the formation of midline glial structures. We show that Fgfr1 and Fgfr2 double knockouts (FGFR DKO) generated by Cre-mediated recombination driven by the human GFAP promoter (hGFAP) have reduced cerebellar size due to reduced proliferation of radial glia and other glial precursors in late embryonic and neonatal FGFR DKO mice. The proliferation of granule cell progenitors (GCPs) in the EGL was also reduced, leading to reduced granule cell numbers. Furthermore, both inward migration of granule cells into the inner granule cell layer (IGL) and outward migration of GABA interneurons into the molecular layer (ML) were arrested, disrupting layer and lobular morphology. Purkinje neurons and their dendrites, which were not targeted by Cre-mediated recombination of Fgf receptors, were also misplaced in FGFR DKO mice, possibly as a consequence of altered Bergmann glia orientation or reduced granule cell number. Our findings indicate a dual role for FGFR signaling in cerebellar morphogenesis. The first role is to amplify the number of granule neuron precursors in the external granular layer and glial precursor cells throughout the cerebellum. The second is to establish the correct Bergmann glia morphology, which is crucial for granule cell migration. The disrupted cerebellar size and laminar architecture resulting from loss of FGFR signaling impair motor learning and coordination in FGFR DKO mice.

Asunto(s)

Enfermedades Cerebelosas/genética , Enfermedades Cerebelosas/patología , Trastornos de la Destreza Motora/patología , Trastornos de la Destreza Motora/fisiopatología , Receptor Tipo 1 de Factor de Crecimiento de Fibroblastos/deficiencia , Receptor Tipo 2 de Factor de Crecimiento de Fibroblastos/deficiencia , Animales , Enfermedades Cerebelosas/fisiopatología , Femenino , Humanos , Masculino , Ratones , Ratones Noqueados , Ratones Mutantes Neurológicos , Trastornos de la Destreza Motora/genética , Receptor Tipo 1 de Factor de Crecimiento de Fibroblastos/metabolismo , Receptor Tipo 2 de Factor de Crecimiento de Fibroblastos/metabolismo

RESUMEN

RATIONALE: Although the fibroblast growth factor (FGF) signaling axis plays important roles in heart development, the molecular mechanism by which the FGF regulates cardiogenesis is not fully understood. OBJECTIVE: To investigate the mechanism by which FGF signaling regulates cardiac progenitor cell differentiation. METHODS AND RESULTS: Using mice with tissue-specific ablation of FGF receptors and FGF receptor substrate 2α (Frs2α) in heart progenitor cells, we demonstrate that disruption of FGF signaling leads to premature differentiation of cardiac progenitor cells in mice. Using embryoid body cultures of mouse embryonic stem cells, we reveal that FGF signaling promotes mesoderm differentiation in embryonic stem cells but inhibits cardiomyocyte differentiation of the mesoderm cells at later stages. Furthermore, we also report that inhibiting FRS2α-mediated signals increases autophagy and that activating autophagy promotes myocardial differentiation and vice versa. CONCLUSIONS: The results indicate that the FGF/FRS2α-mediated signals prevent premature differentiation of heart progenitor cells through suppressing autophagy. The findings provide the first evidence that autophagy plays a role in heart progenitor differentiation.

Asunto(s)

Autofagia , Diferenciación Celular , Factores de Crecimiento de Fibroblastos/metabolismo , Corazón/embriología , Proteínas de la Membrana/metabolismo , Miocitos Cardíacos/metabolismo , Transducción de Señal , Células Madre/metabolismo , Animales , Proliferación Celular , Células Cultivadas , Técnicas de Cultivo de Embriones , Regulación del Desarrollo de la Expresión Génica , Genotipo , Proteínas de la Membrana/deficiencia , Proteínas de la Membrana/genética , Mesodermo/metabolismo , Mesodermo/patología , Ratones , Ratones Noqueados , Miocitos Cardíacos/patología , Fenotipo , Receptor Tipo 1 de Factor de Crecimiento de Fibroblastos/deficiencia , Receptor Tipo 1 de Factor de Crecimiento de Fibroblastos/genética , Receptor Tipo 2 de Factor de Crecimiento de Fibroblastos/deficiencia , Receptor Tipo 2 de Factor de Crecimiento de Fibroblastos/genética , Células Madre/patología , Factores de Tiempo , Técnicas de Cultivo de Tejidos

RESUMEN

Fibroblast growth factors (FGFs) comprise a family of developmental regulators implicated in a wide variety of neurological functions. FGF receptors 1, 2, and 3 (Fgfrs) are expressed in the embryonic forebrain, including regions overlapping with ventral sites of oligodendrocyte progenitor (OLP) generation. Although FGF signaling is known to influence the proliferation of OLPs in vitro, functions of different Fgfrs in vivo are lacking. Here, we examined single and double mutants with conditional disruption of Fgfrs, specifically in the embryonic forebrain, to investigate the effect of FGFs on the generation and proliferation of OLPs in vivo. FGF signaling, through cooperation between Fgfr1 and Fgfr2 but not Fgfr3, is required for the initial generation of OLPs in the mouse ventral forebrain, with Fgfr1 being a stronger inducer than Fgfr2. In cultures derived from embryonic mutant forebrains or from normal forebrains grown in the presence of Fgfr inhibitor, a strong attenuation of OLP generation was observed, supporting the role of FGF signaling in vivo. Contrary to in vitro findings, Fgfr1 and Fgfr2 signaling is not required for the proliferation of OLPs in vivo. Finally, failure of OLP generation in the Fgfr mutants occurred without loss of sonic hedgehog (Shh) signaling; and pharmacological inhibition of either Fgfr or hedgehog signaling in parallel cultures strongly inhibited OLP generation, suggesting that Fgfrs cooperate with Shh to generate OLPs. Overall, our results reveal for the first time an essential role of FGF signaling in vivo, where the three Fgfrs differentially control the normal generation of OLPs from the embryonic ventral forebrain.

Asunto(s)

Células Madre Embrionarias/fisiología , Neurogénesis/genética , Oligodendroglía/fisiología , Prosencéfalo/embriología , Receptor Tipo 1 de Factor de Crecimiento de Fibroblastos/fisiología , Receptor Tipo 2 de Factor de Crecimiento de Fibroblastos/fisiología , Transducción de Señal/genética , Animales , Células Cultivadas , Células Madre Embrionarias/citología , Células Madre Embrionarias/metabolismo , Factores de Crecimiento de Fibroblastos/fisiología , Ratones , Ratones Noqueados , Ratones Transgénicos , Oligodendroglía/citología , Prosencéfalo/citología , Prosencéfalo/fisiología , Receptor Tipo 1 de Factor de Crecimiento de Fibroblastos/deficiencia , Receptor Tipo 1 de Factor de Crecimiento de Fibroblastos/genética , Receptor Tipo 2 de Factor de Crecimiento de Fibroblastos/deficiencia , Receptor Tipo 2 de Factor de Crecimiento de Fibroblastos/genética , Células Madre/citología , Células Madre/metabolismo , Células Madre/fisiología

RESUMEN

To understand the role of specific fibroblast growth factor receptors (FGFRs) in cortical development, we conditionally inactivated Fgfr2 or both Fgfr1 and Fgfr2 [Fgfr2 conditional knock-out (cKO) or double knock-out mice, respectively] in radial glial cells of the dorsal telencephalon. Fgfr1 and Fgfr2 are necessary for the attainment of a normal number of excitatory neurons in the cerebral cortex. The action of FGF receptors appears to be through increasing self-renewal of neuronal precursors within the ventricular zone. Volume measurements, assessments of excitatory neuron number, and areal marker expression suggested that the proper formation of the medial prefrontal cortex (mPFC) depends on the function of Fgfr2, whereas Fgfr1 together with Fgfr2 control excitatory cortical neuron development within the entire cerebral cortex. Fgfr2 cKO mice had fewer and smaller glutamate synaptic terminals in the bed nuclei of the stria terminalis (BST), a projection area for mPFC cortical neurons. Furthermore, Fgfr2 cKO mice showed secondary decreases in GABAergic neurons in the BST and septum. These data demonstrate that FGFR2 signaling expands the number of excitatory neurons in the mPFC and secondarily influences target neurons in subcortical stations of the limbic system.

Asunto(s)

Sistema Límbico/metabolismo , Red Nerviosa/metabolismo , Corteza Prefrontal/metabolismo , Receptor Tipo 2 de Factor de Crecimiento de Fibroblastos/fisiología , Animales , Proliferación Celular , Femenino , Humanos , Sistema Límbico/embriología , Sistema Límbico/crecimiento & desarrollo , Ratones , Ratones Noqueados , Ratones Transgénicos , Red Nerviosa/embriología , Red Nerviosa/crecimiento & desarrollo , Vías Nerviosas/embriología , Vías Nerviosas/crecimiento & desarrollo , Vías Nerviosas/metabolismo , Corteza Prefrontal/embriología , Corteza Prefrontal/crecimiento & desarrollo , Embarazo , Receptor Tipo 2 de Factor de Crecimiento de Fibroblastos/deficiencia , Receptor Tipo 2 de Factor de Crecimiento de Fibroblastos/genética

RESUMEN

During corticogenesis, the balance between the self-renewal of radial glial stem cells and the production of their descendent progenitor cells is essential in generating the correct size and cell composition of the neocortex. How the stem-to-progenitor cell transition is regulated is poorly understood. FGFs are commonly implicated in promoting proliferation of neural precursor cells, but it is unclear how they exert their effects on stem cells, progenitor cells, or both in vivo. Here, three FGF receptor genes are simultaneously deleted during cortical neurogenesis. In these mutants, radial glia are depleted due to an increased transition from an uncommitted state to a more differentiated one, initially causing an increase in progenitors, but ultimately resulting in a smaller cortex. The proliferation rate of progenitors themselves, however, is unchanged. These results indicate that FGFs normally repress the radial glia to progenitor cell transition during corticogenesis.

Asunto(s)

Corteza Cerebral/fisiología , Neurogénesis , Neuroglía/fisiología , Receptor Tipo 1 de Factor de Crecimiento de Fibroblastos/fisiología , Receptor Tipo 2 de Factor de Crecimiento de Fibroblastos/fisiología , Receptor Tipo 3 de Factor de Crecimiento de Fibroblastos/fisiología , Transducción de Señal , Células Madre/fisiología , Animales , Diferenciación Celular/genética , Proliferación Celular , Supervivencia Celular/genética , Corteza Cerebral/citología , Corteza Cerebral/embriología , Femenino , Ratones , Ratones Noqueados , Ratones Transgénicos , Neurogénesis/genética , Neuroglía/citología , Embarazo , Receptor Tipo 1 de Factor de Crecimiento de Fibroblastos/deficiencia , Receptor Tipo 1 de Factor de Crecimiento de Fibroblastos/genética , Receptor Tipo 2 de Factor de Crecimiento de Fibroblastos/deficiencia , Receptor Tipo 2 de Factor de Crecimiento de Fibroblastos/genética , Receptor Tipo 3 de Factor de Crecimiento de Fibroblastos/deficiencia , Receptor Tipo 3 de Factor de Crecimiento de Fibroblastos/genética , Transducción de Señal/genética , Células Madre/citología

RESUMEN

Current techniques to morphologically characterize the processes of nephrogenesis and ureteric branching during kidney development have many limitations. Here, we used in vivo three-dimensional analysis to study renal development in mice lacking fibroblast growth factor receptor 2 in the ureteric bud (Fgfr2(UB-/-)) and in littermate controls. We found that Fgfr2(UB-/-) mice have more severe defects in ureteric branching morphogenesis than previously reported, including significantly fewer branches and tips than control mice. Furthermore, these mice had decreased ureteric volume and surface area and longer ureteric segments than control mice. We also observed previously unrecognized abnormalities in nephrogenesis, including a gradual increase in volume and surface area during maturation from renal vesicles to mature nephrons, in the mutant mice. Finally, we quantified many events of normal renal development that are either difficult or impossible to measure without this three-dimensional technique. In summary, the three-dimensional approach is a powerful and quantitative means to characterize branching morphogenesis and nephrogenesis.

Asunto(s)

Imagenología Tridimensional/métodos , Riñón/anomalías , Mesodermo/anomalías , Receptor Tipo 2 de Factor de Crecimiento de Fibroblastos/deficiencia , Receptor Tipo 2 de Factor de Crecimiento de Fibroblastos/genética , Uréter/anomalías , Animales , Femenino , Edad Gestacional , Procesamiento de Imagen Asistido por Computador/métodos , Riñón/embriología , Mesodermo/embriología , Ratones , Ratones Noqueados , Ratones Transgénicos , Nefronas/anomalías , Nefronas/embriología , Embarazo , Uréter/embriología

RESUMEN

Fibroblast growth factor 10 (FGF10) signaling through FGF receptor 2 (FGFR2) is required for lung initiation. While studies indicate that Fgf10 and Fgfr2 are also important at later stages of lung development, their roles in early branching events remain unclear. We addressed this question through conditional inactivation of both genes in mouse subsequent to lung initiation. Inactivation of Fgf10 in lung mesenchyme resulted in smaller lobes with a reduced number of branches. Inactivation of Fgfr2 in lung epithelium resulted in disruption of lobes and small epithelial outgrowths that arose arbitrarily along the main bronchi. In both mutants, there was an increase in cell death. Also, the expression patterns of key signaling molecules implicated in branching morphogenesis were altered and a proximal lung marker was expanded distally. Our results indicate that both Fgf10 and Fgfr2 are required for a normal branching program and for proper proximal-distal patterning of the lung.

Asunto(s)

Factor 10 de Crecimiento de Fibroblastos/fisiología , Pulmón/embriología , Receptor Tipo 2 de Factor de Crecimiento de Fibroblastos/fisiología , Animales , Secuencia de Bases , Tipificación del Cuerpo/genética , Tipificación del Cuerpo/fisiología , Muerte Celular/genética , Muerte Celular/fisiología , Supervivencia Celular/genética , Supervivencia Celular/fisiología , Cartilla de ADN/genética , Epitelio/embriología , Epitelio/fisiología , Femenino , Factor 10 de Crecimiento de Fibroblastos/deficiencia , Factor 10 de Crecimiento de Fibroblastos/genética , Regulación del Desarrollo de la Expresión Génica , Marcación de Gen , Pulmón/anomalías , Pulmón/fisiología , Mesodermo/embriología , Mesodermo/fisiología , Ratones , Ratones Noqueados , Ratones Mutantes , Ratones Transgénicos , Embarazo , Receptor Tipo 2 de Factor de Crecimiento de Fibroblastos/deficiencia , Receptor Tipo 2 de Factor de Crecimiento de Fibroblastos/genética , Factor de Transcripción SOX9/genética , Factor de Transcripción SOX9/fisiología , Factores de Transcripción SOXB1/genética , Factores de Transcripción SOXB1/fisiología , Transducción de Señal