Abstract
BackgroundLungs 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.MethodsWe 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.ResultsWe 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.DiscussionIn 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.
Highlights
Lung development and critical aspects of pulmonary epithelial differentiation is mostly studied through the use of animal models [1]
We describe a three dimensional (3D) epithelial culture model using a recently described human basal-like airway epithelial cell line (VA10) cultured in reconstituted basement membrane [17]
Endothelial cells stimulate branching morphogenesis of airway epithelial cells In the light of recent data from various organs demonstrating the importance of vascular endothelium in stem cell niche and organogenesis [21,22,23], and due to the fact that bronchioles and alveoli are adjacent to the vascular endothelium in vivo, we hypothesized that vascular endothelium might induce a distal airway phenotype in bronchial epithelial cells
Summary
Lung development and critical aspects of pulmonary epithelial differentiation is mostly studied through the use of animal models [1]. Due to a lack of good experimental in vitro models, much less is known about development and stem cell biology in human lungs. Many human airway epithelial cell lines have been established, most of them have not been defined with respect to their cellular origin and lack critical characterization in terms of expression of differentiation markers [2]. The human bronchial cell lines 16HBE14o-, Calu-3, and BEAS-2B have been successfully applied to study drug transport, metabolism, and drug delivery due to their ability to form tight junctions (TJ) [2]. 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
Sign-in/Register to view highlights & summary 
Published Version (
Free)
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call