Sources and Regulation of Epithelial Stem/Progenitor Cells in Alveolar Regeneration

项目总结/摘要 严重急性损伤后正常肺功能的重现意味着肺组织固有的再生能力， 肺然而，肺上皮干/祖细胞的来源和相对再生能力仍然存在 不清楚，尤其是在人类的肺部。根据损伤类型和严重程度， 被激活并通过增殖和分化来响应，以帮助接近完全恢复。气道和 肺泡干/祖细胞被激活并有助于严重损伤后的肺泡修复， 流感或博来霉素。为此，我们实验室最近的研究发现了一种气道上皮祖细胞， 以主要组织相容性复合物（MHC）I类蛋白H2-K1水平升高为标志的细胞。尽管 H2-K1 high祖细胞具有与成熟俱乐部细胞高度相似的转录组，与成熟俱乐部细胞不同， 细胞，选择性增殖损伤后，并帮助改善氧合损伤小鼠后原位 移植然而，这些祖细胞的早期和选择性激活的机制仍然存在， 未知此外，远端肺再生还有几个方面尚不清楚。首席 其中之一是我们在小鼠肺中观察到的远端上皮祖细胞等级是否得以维持， 在人类肺部的远端远端人肺气道具有更高比例的基底细胞，并且具有更多的 与小鼠气道上皮细胞相比，小鼠气道上皮细胞具有异质性分泌细胞群。此外，人类2型 肺泡上皮细胞（AEC 2）具有显著的体外增殖和再生能力。因此，我们认为， 存在理解远端人体气道和肺泡的识别和调节的未满足的需要 祖先为此，我们的初步数据表明，一个远端的人类分泌亚群， 类似于小鼠H2-K1 high的棒状祖细胞可以在体外产生AEC 2s。相反，我们有 揭示了成熟人AEC 2向气道谱系分化的新的和意想不到的能力， 体外和体内，表明人体远端上皮再生至少有两种来源。因此有 关键是要澄清远侧小鼠和人上皮祖细胞的身份和特征， 进行牙槽修复本提案旨在通过三个目标来回答这些问题：1）确定 肺泡修复中H2-K1 high祖细胞活化的潜在机制2)为了识别远端分泌细胞 作为人肺远端损伤后肺泡细胞的来源。3)确定是否有一个亚群 成熟的人AEC 2具有可逆的双向分化为肺泡基底细胞的潜能。这些 目的是利用单细胞mRNA和开放染色质测序，在体外和体内操作的建议， 信号通路和祖细胞的原位移植，以澄清上皮干/祖细胞 小鼠和人肺远端的细胞层次结构。这些研究将为审讯几个 新的信号通路，最终帮助我们努力操纵再生机制， 更好的疾病结果。

PROJECT SUMMARY/ABSTRACT Recapitulation of normal lung function following a severe acute injury implies an inherent regenerative ability of the lung. However, sources and relative regenerative capacities of lung epithelial stem/progenitor cells remain unclear, especially in the human lung. Depending on the injury type and severity, several distinct progenitors are activated and respond by proliferating and differentiating to aid in near complete recovery. Both airway and alveolar stem/progenitor cells are activated and contribute to alveolar repair following severe injuries such as influenza or bleomycin. To this end, recent studies from our lab have uncovered an airway epithelial progenitor cell marked by elevated levels of Major Histocompatibility Complex (MHC) Class I protein, H2-K1. Despite having a transcriptome highly similar to the mature club cells, the H2-K1high progenitors, unlike mature club cells, selectively proliferate post injury and aid in improved oxygenation in injured mice after orthotopic transplantation. However, mechanisms underlying early and selective activation of these progenitors remain unknown. In addition, there are several more aspects of distal lung regeneration that are yet unclear. The chief among them is whether the distal epithelial progenitor hierarchy that we observe in mouse lungs is maintained in distal human lungs. The distal human lung airways have a higher proportion of basal cells and have more heterogeneous secretory cell populations than the mouse airway epithelium. Furthermore, the human type 2 alveolar epithelial cells (AEC2s) have remarkable in vitro proliferative and regenerative capacity. Therefore, there is an unmet need to understand the identity and regulation of distal human airway and alveolar progenitors. To this end, our preliminary data show that a distal human secretory subpopulation that is analogous to the mouse H2-K1high club-like progenitors can give rise to AEC2s in vitro. Conversely, we have uncovered a novel and unexpected ability of mature human AEC2s to differentiate towards airway lineages in vitro and in vivo, suggesting at least two sources of distal epithelial regeneration in the human. Therefore, it is critical to clarify the identity and characteristics of both distal mouse and human epithelial progenitors primed for alveolar repair. This proposal seeks to answer these questions through three aims: 1) Determine the mechanisms underlying activation of H2-K1high progenitors in alveolar repair. 2) To identify distal secretory cells as a source of alveolar cells post injury in the distal human lung. 3) Determine whether a subpopulation of mature human AEC2s has reversible bi-directional potential to differentiate into alveolar basal cells. These aims will utilize single cell mRNA and open chromatin sequencing, in vitro and in vivo manipulation of proposed signaling pathways, and orthotopic transplantation of progenitor cells to clarify the epithelial stem/progenitor cell hierarchy in distal mouse and human lungs. These studies will lay the foundation for interrogating several novel signaling pathways to ultimately aid in our efforts to manipulate regenerative mechanisms to achieve better disease outcomes.