Characterization of a New Epithelial Cell in the Developing Human Lung

项目摘要 肺芽经历一个称为分支形态发生的过程， 肺的网络1 -7.在分支过程中，在分支的尖端有一群特化的肺祖细胞， 芽尖祖细胞（BTP）产生肺的所有上皮细胞类型，包括近端气道的细胞 （气管、支气管）和远端肺细胞（肺泡）1，8 -16。 我们最近对10 - 21周的人胎肺进行了单细胞RNA测序（scRNAseq）， 妊娠16，并鉴定了一种新的上皮细胞群，其特征在于独特的基因表达谱： SCGB 3A 2 HI/SFTPB HI/CFTRHI，我们将其称为胎儿气道分泌（FAS）细胞。我们已经证明 在培养物中生长的人BTP可以产生FAS细胞，并且它们已经在小气道中原位验证。 我的初步数据表明，FAS细胞是短暂的，在整个发育过程中数量减少。此外，我们的 数据表明FAS细胞是人类独有的，并且不存在于小鼠中。由于对此一无所知 新确定的人口，这项建议的目标是询问功能，分化 潜力和FAS细胞的调节。 我的初步数据表明：（a）FAS细胞可能是一种发育前体状态， 在成为近端气道的途中，和（B）FAS细胞发育受NOTCH信号传导调节，a 关键调节细胞命运的决策在发展中的肺17，18.根据初步数据，我 假设FAS细胞是产生所有气道细胞类型过渡祖细胞群， NOTCH信号调节从BTP到FAS到定型气道细胞类型的转变。使用一种新的细胞 在人肺类器官模型中的条形码谱系追踪方法中，我的目的是1）测试FAS细胞的功能， 特别是如果它们分化成近端气道细胞。我还打算2）研究NOTCH信号转导在 在FAS细胞中使用功能获得和功能丧失慢病毒构建体、NOTCH阻断抗体和 在类器官模型中激活或抑制NOTCH信号传导的药理学抑制剂。

PROJECT ABSTRACT The lung buds undergo a process known as branching morphogenesis, which establishes the complex, arborized network of the lung1–7. During branching, a specialized population of lung progenitors at the tips of the branches, bud tip progenitors (BTPs), give rise to all epithelial cell types of the lung including cells of the proximal airway (trachea, bronchi) and cells of the distal lung (alveoli)1,8–16. We recently performed single-cell RNA sequencing (scRNAseq) on the human fetal lung from 10 to 21 weeks gestation16, and identified a novel epithelial cell population characterized by a unique gene expression profile: SCGB3A2HI/SFTPB HI/CFTRHI which we refer to as Fetal Airway Secretory (FAS) cells. We have shown that human BTPs grown in culture can give rise to FAS cells, and they have been verified in the small airways in situ. My preliminary data suggest that FAS cells are transient, dwindling in number across development. Further, our data suggests that FAS cells are uniquely human, and do not exist in mice. Since nothing is known about this newly identified population, the goal of this proposal is to interrogate the function, differentiation potential, and regulation of FAS cells. My preliminary data suggests that: (a) FAS cells may be a developmental precursor state that exists while BTPs are en route to becoming proximal airway and (b) FAS cell development is regulated by NOTCH signaling, a critical regulator of cell fate decision making in the developing lung17,18. Based on my preliminary data, I hypothesize that FAS cells are a transitional progenitor population that give rise to all airway cell types and NOTCH signaling regulates the transition from BTP-to-FAS to committed airway cell types. Using a novel cell barcoding lineage tracing approach in a human lung organoid model, I aim to 1) test the function of FAS cells, specifically if they differentiate into proximal airway cells. I also aim to 2) investigate the role of NOTCH signaling in FAS cells using gain-of-function and loss-of-function lentiviral constructs, NOTCH blocking antibodies, and pharmacologic inhibitors that activate or inhibit NOTCH signaling in an organoid model.