VEGF/KDR Signaling in Airway Epithelial Regeneration and Disease

摘要 粘液化生通常与多种肺部疾病的发病率和死亡率有关，包括 纤维化、慢性阻塞性肺病和哮喘。然而，导致粘液化生的细胞和分子机制 这些疾病在很大程度上仍不为人所知。最近的谱系追踪数据表明，俱乐部细胞是起源的细胞 用于化生的粘液上皮。然而，俱乐部细胞是一个异质性的群体，它仍然未知。 哪个俱乐部细胞亚群(S)与粘液化生有关。此外，导致这种现象的分子机制 粘液细胞的分化在很大程度上是不确定的。我们的目标是解决这些悬而未决的问题 求婚。我们的单细胞RNA测序分析确定了三个俱乐部细胞亚群，其中两个是 以表达血管内皮生长因子受体2(也称为Flk1或KDR)为特征。值得注意的是，KDR的删除 导致出生后早期肺内呼吸道上皮粘液化生。此外， 一过性增加的KDR可用于阻断术后呼吸道再生过程中的粘液化生 萘的挑战。上皮性KDR的丢失或配体Vegfa的亚型突变导致丰富的 粘液细胞表达Sox9，已被证明调节肠道粘液细胞的分化。 重要的是，粘液化生也与KDR表达降低伴随着增加有关。 卵白蛋白(OVA)诱导的哮喘肺组织中SOX9蛋白的表达。因此，我们假设血管内皮生长因子/KDR 信号是阻止气道中俱乐部细胞亚群黏液分化的把关人 再生，抑制KDR促进哮喘期间通过Sox9的粘液化生 发病机制。我们制定了三个具体的目标来检验这一假设。目标1：检验假设 上皮性KDR通过ERK信号通路阻断俱乐部细胞向粘液细胞分化。目标2：检验假设 Vegfa/KDR信号通路阻断了俱乐部细胞亚群的粘液化生。目标3：检验假设 Vegfa/KDR信号通路通过抑制Sox9抑制粘液化生。这些研究的结果将 提供对控制正常粘膜细胞分化的细胞和分子机制的重要见解 以及该机制如何出错，导致粘液化生。我们的研究也将提供潜在的 这种常见病理实体的治疗靶点。

ABSTRACT Mucous metaplasia is commonly associated with morbidity and mortality in multiple lung diseases including fibrosis, COPD and asthma. However, the cellular and molecular mechanisms leading to mucous metaplasia in these diseases remain largely unknown. Recent lineage tracing data suggest that club cells are the cell of origin for metaplastic mucous epithelium. However, the club cell is a heterogenous population, and it remains unknown which club cell subpopulation(s) contribute to mucous metaplasia. Moreover, the molecular mechanisms leading to mucous cell differentiation are largely undetermined. We aim to address these outstanding issues in this proposal. Our single-cell RNA sequencing analysis identified three club cell subpopulations, two of which are characterized by the expression of VEGF receptor 2 (also known as Flk1 or Kdr). Significantly, deletion of Kdr leads to mucous metaplasia of the intrapulmonary airway epithelium at the early postnatal stage. Furthermore, transiently increased Kdr is required for blocking mucous metaplasia during airway regeneration following naphthalene challenge. Loss of epithelial Kdr or a hypomorphic mutation for the ligand Vegfa leads to abundant mucous cells expressing Sox9 which has been shown to regulate mucous cell differentiation in the intestine. Importantly, mucous metaplasia is also associated with reduced Kdr expression accompanied by increased SOX9 protein levels in ovalbumin (OVA)-induced asthmatic lungs. We therefore hypothesize that Vegf/Kdr signaling is a gatekeeper blocking mucous differentiation of club cell subpopulations during airway regeneration, and that suppressed Kdr promotes mucous metaplasia via Sox9 during asthma pathogenesis. We formulate three specific aims to test the hypothesis. Aim 1: To test the hypothesis that epithelial Kdr blocks club cell differentiation into mucous cells via Erk signaling. Aim 2: To test the hypothesis that Vegfa/Kdr signaling blocks mucous metaplasia of club cell subpopulations. Aim 3: To test the hypothesis that Vegfa/Kdr signaling blocks mucous metaplasia via inhibition of Sox9. Findings from these studies will provide critical insights into the cellular and molecular mechanisms that govern normal mucous cell differentiation and how the mechanism goes awry, leading to mucous metaplasia. Our study will also provide potential therapeutic targets for this common pathological entity.