Defining the lineage, mechanisms of maintenance, and function of a new injury-resistant airway epithelial structure: the hillock

我们最近在小鼠的气管中发现了一种新的呼吸道上皮结构，丘状结构，我们 目前有证据表明，人类呼吸道上皮中也存在类似的结构。呼吸道上皮细胞 山丘是一种分层的上皮结构，具有多层细胞，与 典型的假复层上皮，排列在大部分气道腔内。他们拥有新奇的 腔上皮细胞为多倍体，但完全缺乏纤毛细胞。我们现在展示的是这些 层状丘状上皮结构可抵抗多种形式的呼吸道损伤。我们假设 丘状细胞的层状结构导致保护性屏障，类似于保护功能 位于表皮基底细胞之上的层状表皮细胞。此外，我们证明了 山丘含有一群独特的基础干细胞，以细胞角蛋白13(KRT13)为标志。 这些基底细胞的特点是非常高的基线周转率，并能够 在裸露的呼吸道上皮上快速迁移。我们假设原子的扩散分布 抗损伤的呼吸道小丘促进已抵抗的干细胞的播散性迁移 伤势严重。我们的初步证据表明，丘状的基础干细胞，这是第一个 小鼠气管中高度定义的呼吸道基底细胞亚群是可塑性的，可以产生 正常假复层上皮损伤后重建为正常上皮。最后，我们展示了 老鼠丘的大小和数量随着年龄的增长而增加。有趣的是，由吸烟引起的 鳞状化生和维生素A缺乏所致的鳞状化生 与复层KRT13上皮相关，提示它们可能起源于丘状细胞。 在这项赠款申请中，我们首先建议定义山丘的谱系和可塑性 动态平衡和损伤后使用两种不同的基因驱动系，这两种驱动系专为 标记山丘，更具体地说，标记山丘基底细胞。我们还将确定功能 丘状细胞在上皮细胞动态平衡和白喉毒素损伤修复后的意义 介入性消融与呼吸道损伤模型的结合，特别关注 评估丘状多倍体细胞的作用。使用小鼠外植体模型的组合 气管再生，双光子实时成像，以及化学和遗传调节 人类丘陵培养系统，我们将剖析丘陵分化的分子机制 和丘介导的损伤修复，重点是维甲酸信号转导和修复的具体作用。 丘状与鳞状化生的关系。

We recently identified a novel airway epithelial structure, the hillock, in the murine trachea and we present evidence that similar structures occur in the human airway epithelium. Airway epithelial hillocks are stratified epithelial structures with multiple layers of cells that are distinct from the classic pseudostratified epithelium that lines most of the airway lumen. They possess novel polyploid luminal epithelial cells, but completely lack ciliated cells. We now show that these stratified hillock epithelial structures resist multiple forms of airway injury. We hypothesize that the stratified structure of hillock cells results in a protective barrier, in analogy to the protective function of the stratified epidermal cells that lie atop epidermal basal cells. Furthermore, we show that hillocks contain a unique population of basal stem cells marked by Cytokeratin 13 (KRT13). These basal cells are characterized by a very high baseline rate of turnover and are capable of rapidly migrating over denuded airway epithelium. We hypothesize that the diffuse distribution of injury-resistant airway hillocks facilitates disseminated migration of stem cells that have resisted severe injury. We show preliminary evidence that hillock basal stem cells, which are the first highly defined subset of airway basal cells in the murine trachea, are plastic and can produce normal pseudostratified epithelium after damage to recreate a normal epithelium. Finally, we show that murine hillocks increase in size and number with age. Interestingly, both smoking-induced squamous metaplasia and vitamin A-deficiency induced squamous metaplasia have both been associated with stratified KRT13+ epithelium suggesting they may originate from hillock-like cells. In this grant application, we first propose to define the lineage and plasticity of hillocks during homeostasis and after injury using 2 distinct genetic driver lines that are designed to specifically label hillocks and even more specifically hillock basal cells. We will also determine the functional significance of hillocks during epithelial homeostasis and after injury repair using diphtheria toxin- mediated ablation in combination with models of airway injury, with special attention focused on assessing the role of hillock polyploid cells. Using a combination of an explant model of murine tracheal regeneration, 2-photon live imaging, and the chemical and genetic modulation of a human hillock culture system, we will dissect the molecular mechanisms of hillock differentiation and hillock-mediated injury repair with a focus on the specific role of retinoic acid signaling and the association of hillocks to squamous metaplasia.