A three-dimensional study of alveologenesis in mouse lung.
A three-dimensional study of alveologenesis in mouse lung.
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DOI：
10.1016/j.ydbio.2015.11.017
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发表时间：
2016-01-15
影响因子：
2.7
通讯作者：
Sun X
中科院分区：
文献类型：
--
作者：
Branchfield K;Li R;Lungova V;Verheyden JM;McCulley D;Sun X
肺泡发生是肺成熟的最后一步，它将肺的肺泡区域细分为称为肺泡的更小单元。每一个新生的分隔物都是一个新的气体交换表面，它们共同大大增加了呼吸的表面积。肺泡发生中断导致肺泡简化，如在诊断为支气管肺发育不良（BPD）的早产儿中所见，BPD是一种常见的肺部疾病，通常与终身呼吸缺陷相关。迄今为止，大多数肺泡发生的研究依赖于组织切片的二维（2D）分析。考虑到肺泡发生的首要主题是上皮和间充质的变薄和延伸以促进气体交换，通常在单个2D平面中仅表示细胞或细胞结构的一小部分。在这里，我们使用一个三维（3D）的方法来检查正常以及BPD样小鼠肺的肌成纤维细胞，肺泡2型细胞，弹性蛋白和脂滴的结构架构和细胞组成。我们发现，通常用于描绘生长的肺泡隔的2D手指状间隔峰通常是通过完全建立的肺泡壁进行切片的伪影。相反，生长中的隔膜的更准确表示是由血小板衍生生长因子受体α（PDGFRA）和α平滑肌肌动蛋白（α-SMA）表达的肌成纤维细胞以及它们产生的弹性蛋白纤维排列的3D脊。因此，在3D中，α-SMA和弹性蛋白均存在于3D间隔嵴下的连接网络中，而不是在2D间隔嵴尖端的孤立点中。通过肺泡发生的代表性阶段的分析揭示了这些模式的不受重视的动态变化。表达PDGFRA的细胞仅在肺泡发生的第一阶段呈α-SMA阳性，而在第二阶段则不呈阳性，这表明隔膜形成的两个阶段可能由不同的机制驱动。细的弹性蛋白纤维在肺泡形成之前已经存在于肺泡区域中，这表明在肺泡形成期间，不仅存在新的弹性蛋白沉积，而且还存在广泛的重塑，以将细的且均匀分布的纤维转变成环绕新生隔膜的粗的缆线。对几种遗传和高氧诱导的BPD模型的分析表明，无论缺陷的起源是上皮、间充质、内皮还是环境，肌成纤维细胞组织都受到干扰。最后，PDGFRA阳性细胞和肺泡2型细胞的相对位置的分析表明，在肺泡发生，这两种细胞类型并不总是彼此相邻。这一结果表明，在成人肺中，由它们的紧密并列所提供的生态位和祖细胞关系可能是后来获得的特性。隔膜3D重建揭示的这些见解为未来研究驱动正常肺泡发生的机制以及BPD肺泡简化的原因奠定了基础。
Alveologenesis is the final step of lung maturation, which subdivides the alveolar region of the lung into smaller units called alveoli. Each of the nascent dividers serves as a new gas-exchange surface, and collectively they drastically increase the surface area for breathing. Disruption of alveologenesis results in simplification of alveoli, as is seen in premature infants diagnosed with bronchopulmonary dysplasia (BPD), a prevalent lung disease that is often associated with lifelong breathing deficiencies. To date, a majority of studies of alveologenesis rely on two-dimensional (2D) analysis of tissue sections. Given that an overarching theme of alveologenesis is thinning and extension of the epithelium and mesenchyme to facilitate gas exchange, often only a small portion of a cell or a cellular structure is represented in a single 2D plane. Here, we use a three-dimensional (3D) approach to examine the structural architecture and cellular composition of myofibroblasts, alveolar type 2 cells, elastin and lipid droplets in normal as well as BPD-like mouse lung. We found that 2D finger-like septal crests, commonly used to depict growing alveolar septae, are often artifacts of sectioning through fully established alveolar walls. Instead, a more accurate representation of growing septae are 3D ridges that are lined by platelet-derived growth factor receptor alpha (PDGFRA) and alpha smooth muscle actin (α-SMA)-expressing myofibroblasts, as well as the elastin fibers that they produce. Accordingly in 3D, both α-SMA and elastin were each found in connected networks underlying the 3D septal ridges rather than as isolated dots at the tip of 2D septal crests. Analysis through representative stages of alveologenesis revealed unappreciated dynamic changes in these patterns. PDGFRA-expressing cells are only α-SMA-positive during the first phase of alveologenesis, but not in the second phase, suggesting that the two phases of septae formation may be driven by distinct mechanisms. Thin elastin fibers are already present in the alveolar region prior to alveologenesis, suggesting that during alveologenesis, there is not only new elastin deposition, but also extensive remodeling to transform thin and uniformly distributed fibers into thick cables that rim the nascent septae. Analysis of several genetic as well as hyperoxia-induced models of BPD revealed that the myofibroblast organization is perturbed in all, regardless of whether the origin of defect is epithelial, mesenchymal, endothelial or environmental. Finally, analysis of relative position of PDGFRA-positive cells and alveolar type 2 cells reveal that during alveologenesis, these two cell types are not always adjacent to one another. This result suggests that the niche and progenitor relationship afforded by their close juxtaposition in the adult lung may be a later acquired property. These insights revealed by 3D reconstruction of the septae set the foundation for future investigations of the mechanisms driving normal alveologenesis, as well as causes of alveolar simplification in BPD.