Tracheal occlusion increases the rate of epithelial branching of embryonic mouse lung via the FGF10-FGFR2b-Sprouty2 pathway (original) (raw)
Related papers
Evidence that SPROUTY2 functions as an inhibitor of mouse embryonic lung growth and morphogenesis
Mechanisms of Development, 2001
Experimental evidence is rapidly emerging that the coupling of positive regulatory signals with the induction of negative feedback modulators is a mechanism of ®ne regulation in development. Studies in Drosophila and chick have shown that members of the SPROUTY family are inducible negative regulators of growth factors that act through tyrosine kinase receptors. We and others have shown that Fibroblast Growth Factor 10 (FGF10) is a key positive regulator of lung branching morphogenesis. Herein, we provide direct evidence that mSprouty2 is dynamically expressed in the peripheral endoderm in embryonic lung and is downregulated in the clefts between new branches at E12.5. We found that mSprouty2 was expressed in a domain restricted in time and space, adjacent to that of Fgf10 in the peripheral mesenchyme. By E14.5, Fgf10 expression was restricted to a narrow domain of mesenchyme along the extreme edges of the individual lung lobes, whereas mSprouty2 was most highly expressed in the subjacent epithelial terminal buds. FGF10 beads upregulated the expression of mSprouty2 in adjacent epithelium in embryonic lung explant culture. Lung cultures treated with exogenous FGF10 showed greater branching and higher levels of mSpry2 mRNA. Conversely, Fgf10 antisense oligonucleotides reduced branching and decreased mSpry2 mRNA levels. However, treatment with exogenous FGF10 or antisense Fgf10 did not change Shh and FgfR2 mRNA levels in the lungs. We investigated Sprouty2 function during lung development by two different but complementary approaches. The targeted overexpression of mSprouty2 in the peripheral lung epithelium in vivo, using the Surfactant Protein C promoter, resulted in a low level of branching, lung lobe edges abnormal in appearance and the inhibition of epithelial proliferation. Transient high-level overexpression of mSpry2 throughout the pulmonary epithelium by intra-tracheal adenovirus microinjection also resulted in a low level of branching. These results indicate for the ®rst time that mSPROUTY2 functions as a negative regulator of embryonic lung morphogenesis and growth. q
Molecular Mechanisms of Early Lung Specification and Branching Morphogenesis
Pediatric Research, 2005
The "hard wiring" encoded within the genome that determines the emergence of the laryngotracheal groove and subsequently early lung branching morphogenesis is mediated by finely regulated, interactive growth factor signaling mechanisms that determine the automaticity of branching, interbranch length, stereotypy of branching, left-right asymmetry, and finally gas diffusion surface area. The extracellular matrix is an important regulator as well as a target for growth factor signaling in lung branching morphogenesis and alveolarization. Coordination not only of epithelial but also endothelial branching morphogenesis determines bronchial branching and the eventual alveolar-capillary interface. Improved prospects for lung protection, repair, regeneration, and engineering will depend on more detailed understanding of these processes. Herein, we concisely review the functionally integrated morphogenetic signaling network comprising the critical bone morphogenetic protein, fibroblast growth factor, Sonic hedgehog, transforming growth factor-, vascular endothelial growth factor, and Wnt signaling pathways that specify and drive early embryonic lung morphogenesis. (Pediatr Res 57: 1-12, 2005) Abbreviations BMP, bone morphogenetic protein DKK, Dickkopf EGF (R), epidermal growth factor (receptor) ERK, extracellular regulated kinase FGF (R), fibroblast growth factor (receptor) FN, fibronectin LRP, lipoprotein receptor-related proteins MAP, membrane-associated protein PDGF, platelet-derived growth factor RAR, retinoic acid receptor sFRP, secreted Frizzled-related protein SHH, Sonic hedgehog Sp-C, surfactant protein C TGF-␣ (), transforming growth factor alpha (beta) VEGF (R), vascular endothelial growth factor (receptor)
The molecular basis of lung morphogenesis
Mechanisms of Development, 2000
To form a diffusible interface large enough to conduct respiratory gas exchange with the circulation, the lung endoderm undergoes extensive branching morphogenesis and alveolization, coupled with angiogenesis and vasculogenesis. It is becoming clear that many of the key factors determining the process of branching morphogenesis, particularly of the respiratory organs, are highly conserved through evolution. Synthesis of information from null mutations in Drosophila and mouse indicates that members of the sonic hedgehog/patched/ smoothened/Gli/FGF/FGFR/sprouty pathway are functionally conserved and extremely important in determining respiratory organogenesis through mesenchymal±epithelial inductive signaling, which induces epithelial proliferation, chemotaxis and organ-speci®c gene expression. Transcriptional factors including Nkx2.1, HNF family forkhead homologues, GATA family zinc ®nger factors, pou and hox, helix-loop-helix (HLH) factors, Id factors, glucocorticoid and retinoic acid receptors mediate and integrate the developmental genetic instruction of lung morphogenesis and cell lineage determination. Signaling by the IGF, EGF and TGF-b/BMP pathways, extracellular matrix components and integrin signaling pathways also directs lung morphogenesis as well as proximo-distal lung epithelial cell lineage differentiation. Soluble factors secreted by lung mesenchyme comprise a`compleat' inducer of lung morphogenesis. In general, peptide growth factors signaling through cognate receptors with tyrosine kinase intracellular signaling domains such as FGFR, EGFR, IGFR, PDGFR and c-met stimulate lung morphogenesis. On the other hand, cognate receptors with serine/threonine kinase intracellular signaling domains, such as the TGF-b receptor family are inhibitory, although BMP4 and BMPR also play key inductive roles. Pulmonary neuroendocrine cells differentiate earliest in gestation from among multipotential lung epithelial cells. MASH1 null mutant mice do not develop PNE cells. Proximal and distal airway epithelial phenotypes differentiate under distinct transcriptional control mechanisms. It is becoming clear that angiogenesis and vasculogenesis of the pulmonary circulation and capillary network are closely linked with and may be necessary for lung epithelial morphogenesis. Like epithelial morphogenesis, pulmonary vascularization is subject to a ®ne balance between positive and negative factors. Angiogenic and vasculogenic factors include VEGF, which signals through cognate receptors¯k and¯t, while novel anti-angiogenic factors include EMAP II. q
Temporal effects of Sprouty on lung morphogenesis
Developmental Biology, 2003
Paracrine signaling mediated by FGF-10 and the FGF-R2IIIb receptor is required for formation of the lung. To determine the temporal requirements for FGF signaling during pulmonary morphogenesis, Sprouty-4 (Spry-4), an intracellular FGF receptor antagonist, was expressed in epithelial cells of the fetal lung under control of a doxycycline-inducible system. Severe defects in lobulation and severe lung hypoplasia were observed when Spry-4 was expressed throughout fetal lung development (E6.5-E18.5) or from E6.5 until E13.5. Effects of Spry-4 on branching were substantially reversed by removal of doxycycline from the dam at E12.5, but not at E13.5. In contrast, when initiated late in development (E12.5 to birth), Spry-4 caused less severe pulmonary hypoplasia. Expression of Spry-4 from E16.5 to E18.5 reduced lung growth and resulted in perinatal death due to respiratory failure. Expression of Spry-4 during the saccular and alveolar stages, from E18.5 to postnatal day 21, caused mild emphysema. These findings demonstrate that the embryonic-pseudoglandular stage is a critical time period during which Spry-sensitive pathways are required for branching morphogenesis, lobulation, and formation of the peripheral lung parenchyma.
Airway branching morphogenesis in three dimensional culture
Respiratory Research, 2010
Background: Lungs develop from the fetal digestive tract where epithelium invades the vascular rich stroma in a process called branching morphogenesis. In organogenesis, endothelial cells have been shown to be important for morphogenesis and the maintenance of organ structure. The aim of this study was to recapitulate human lung morphogenesis in vitro by establishing a three dimensional (3D) co-culture model where lung epithelial cells were cultured in endothelial-rich stroma. Methods: We used a human bronchial epithelial cell line (VA10) recently developed in our laboratory. This cell line cell line maintains a predominant basal cell phenotype, expressing p63 and other basal markers such as cytokeratin-5 and-14. Here, we cultured VA10 with human umbilical vein endothelial cells (HUVECs), to mimic the close interaction between these cell types during lung development. Morphogenesis and differentiation was monitored by phase contrast microscopy, immunostainings and confocal imaging. Results: We found that in co-culture with endothelial cells, the VA10 cells generated bronchioalveolar like structures, suggesting that lung epithelial branching is facilitated by the presence of endothelial cells. The VA10 derived epithelial structures display various complex patterns of branching and show partial alveolar type-II differentiation with pro-Surfactant-C expression. The epithelial origin of the branching VA10 colonies was confirmed by immunostaining. These bronchioalveolar-like structures were polarized with respect to integrin expression at the cell-matrix interface. The endothelial-induced branching was mediated by soluble factors. Furthermore, fibroblast growth factor receptor-2 (FGFR-2) and sprouty-2 were expressed at the growing tips of the branching structures and the branching was inhibited by the FGFR-small molecule inhibitor SU5402. Discussion: In this study we show that a human lung epithelial cell line can be induced by endothelial cells to form branching bronchioalveolar-like structures in 3-D culture. This novel model of human airway morphogenesis can be used to study critical events in human lung development and suggests a supportive role for the endothelium in promoting branching of airway epithelium.
Growth Factor Control of Growth and Epithelial Differentiation in Embryonic Lungs
Biochemical and Molecular Medicine, 1997
Beginning on Day 10 of mouse embryonic develop-Branching morphogenesis and epithelial differen-We conclude that both EGF and TGFb1 affect the mones such as glucocorticoids, thyroid hormone, esdevelopment of branching morphogenesis and of trogen, and androgen (9-12), as well as by growth epithelial differentiation in the embryonic lung. factors which also in early lung development influ-᭧ 1997 Academic Press ence branching morphogenesis (13-17). Specifically, 38
Novel Role for Netrins in Regulating Epithelial Behavior during Lung Branching Morphogenesis
Current Biology, 2004
The development of many organs, including the lung, depends upon a process known as branching morphogenesis, in which a simple epithelial bud gives rise to a complex tree-like system of tubes specialized for the transport of gas or fluids. Previous studies on lung development have highlighted a role for fibroblast growth factors (FGFs), made by the mesodermal cells, in promoting the proliferation, budding, and chemotaxis of the epithelial endoderm [1-3]. Here, by using a three-dimensional culture system, we provide evidence for a novel role for Netrins, best known as axonal guidance molecules [4,5], in modulating the morphogenetic response of lung endoderm to exogenous FGFs. This effect involves inhibition of localized changes in cell shape and phosphorylation of the intracellular mitogen-activated protein kinase(s) (ERK1/2, for extracellular signal-regulated kinase-1 and-2), elicited by exogenous FGFs. The temporal and spatial expression of netrin 1, netrin 4, and Unc5b genes and the localization of Netrin-4 protein in vivo suggest a model in which Netrins in the basal lamina locally modulate and fine-tune the outgrowth and shape of emergent epithelial buds. Results and Discussion The branching morphogenesis of the mouse lung starts at embryonic day (E)9.5 (E9.5), when two primordial buds composed of an inner endodermal epithelium and an outer mesenchymal jacket appear in the ventral-lateral wall of the foregut. Between E9.5 and E16.5 (the pseudo-glandular stage), the primordial buds give rise to the respiratory tree [1
Mouse Embryonic Lung Culture, A System to Evaluate the Molecular Mechanisms of Branching
Journal of Visualized Experiments, 2010
Lung primordial specification as well as branching morphogenesis, and the formation of various pulmonary cell lineages requires a specific interaction of the lung endoderm with its surrounding mesenchyme and mesothelium. Lung mesenchyme has been shown to be the source of inductive signals for lung branching morphogenesis. Epithelial-mesenchymal-mesothelial interactions are also critical to embryonic lung morphogenesis. Early embryonic lung organ culture is a very useful system to study epithelial-mesenchymal interactions. Both epithelial and mesenchymal morphogenesis proceeds under specific conditions that can be readily manipulated in this system (in the absence of maternal influence and blood flow). More importantly this technique can be readily done in a serumless, chemically defined culture media. Gain and loss of function can be achieved using expressed proteins, recombinant viral vectors and/or analysis of transgenic mouse strains, antisense RNA, as well as RNA interference gene knockdown.
Frontiers in Zoology, 2012
Gas exchangers fundamentally form by branching morphogenesis (BM), a mechanistically profoundly complex process which derives from coherent expression and regulation of multiple genes that direct cell-to-cell interactions, differentiation, and movements by signaling of various molecular morphogenetic cues at specific times and particular places in the developing organ. Coordinated expression of growth-instructing factors determines sizes and sites where bifurcation occurs, by how much a part elongates before it divides, and the angle at which branching occurs. BM is essentially induced by dualities of factors where through feedback-or feed forward loops agonists/antagonists are activated or repressed. The intricate transactions between the development orchestrating molecular factors determine the ultimate phenotype. From the primeval time when the transformation of unicellular organisms to multicellular ones occurred by systematic accretion of cells, BM has been perpetually conserved. Canonical signalling, transcriptional pathways, and other instructive molecular factors are commonly employed within and across species, tissues, and stages of development. While much still remain to be elucidated and some of what has been reported corroborated and reconciled with rest of existing data, notable progress has in recent times been made in understanding the mechanism of BM. By identifying and characterizing the morphogenetic drivers, and markers and their regulatory dynamics, the elemental underpinnings of BM have been more precisely explained. Broadening these insights will allow more effective diagnostic and therapeutic interventions of developmental abnormalities and pathologies in pre-and postnatal lungs. Conservation of the molecular factors which are involved in the development of the lung (and other branched organs) is a classic example of nature's astuteness in economically utilizing finite resources. Once purposefully formed, well-tested and tried ways and means are adopted, preserved, and widely used to engineer the most optimal phenotypes. The material and time costs of developing utterly new instruments and routines with every drastic biological change (e.g. adaptation and speciation) are circumvented. This should assure the best possible structures and therefore functions, ensuring survival and evolutionary success.