Interstitial resident fibroblasts direct alveolar epithelial differentiation

项目摘要：虽然在理解上皮亲组织的作用方面已经取得了巨大的进展。 基因在组织修复和纤维化机制中的作用，对其起源、性质和基因的相互作用知之甚少 成纤维细胞前体细胞的调控程序。肺部研究受到了引人注目的 成纤维细胞群体和间充质干细胞群体的复杂性和多样性。我们的长期目标 是了解成纤维细胞群体的复杂性，并确定它们对上皮细胞的调节作用 在牙槽化和上皮修复期间。我们的主要目标是确定成纤维细胞差异表达的关键调节因子。 在肺里发酵。本申请基于我们将GATA6标识为关键字的初步数据 基质iRef中的转录因子，它们是牙槽骨形成所必需的。我们的中心假设是 GATA6调节iRef中的基因表达，从而亚序列诱导基质IREF规范和类... 在发育和修复过程中与血管内皮细胞的相互作用。建议进行这项研究的理由是 了解成纤维细胞亚群的调节及其在上皮-间充质细胞中的不同作用 相互作用，将为有关肺内稳态和修复的基本问题提供新的答案。 这一假说将通过三个具体的目标进行检验：1)基质iRef引导AEC1分化。2) GATA6转录调控一组基质成纤维细胞特征基因和3)H3K27me沉默 标记调节老年iREF中GATA6和基质基因的表达。在第一个目标中，肺泡球培养将 用于检测特定的IREF亚群与野生型之间的上皮-间充质串扰 型肺泡上皮细胞。在第二个目标中，GATA6的得失功能研究将被用于ES。 建立成纤维细胞表型和牙槽化之间的功能联系。第三个目标是H3K27me3组蛋白 分数将通过芯片-聚合酶链式反应分析进行评估。使用EZH2抑制剂的体内和体外研究将确定 EZH2在抑制基质成纤维细胞特征基因中的作用。这些研究将提供一个更好的- 牙槽骨成纤维细胞多样性对上皮-间充质串扰的影响 肺泡再生。这一贡献将是重大的，因为它将促进对基因的了解 和参与牙槽骨再生的途径，使人们对再生减少的原因有了新的见解 并为新的治疗方法提供基础，以克服目前人类肺部再生的局限性。 我们对肺成纤维细胞在肺损伤和再生中的作用的理解远远落后于我们的理解。 上皮性细胞的作用。有关间质肺成纤维细胞的新信息将对确定 发育机制、正常肺修复和肺部疾病。

PROJECT SUMMARY: While tremendous progress has been made to understand the roles of epithelial pro- genitors in tissue repair and fibrotic mechanisms, little is known about the origin, nature of interaction and gene regulatory programs of fibroblast progenitors. Pulmonary research has been challenged by the remarkable complexity and diversity of fibroblast populations and mesenchymal stem cell populations. Our long-term goal is to understand the complexity of fibroblast populations and to identify their regulatory role on the epithelium during alveolarization and epithelial repair. Our primary objective is to identify key regulators of fibroblast dif- ferentiation in the lung. The present application is based on our preliminary data that identifies GATA6 as a key transcription factor in matrix iReFs, which are indispensable for alveolarization. Our central hypothesis is that GATA6 regulates gene expression in iReFs that sub sequentially induce matrix iReF specification and para- crine interactions with AECs during development and repair. The rationale for the proposed research is that understanding the regulation of fibroblast subpopulations, and their distinct roles in epithelial-mesenchymal cell interactions, will provide new answers for fundamental questions regarding lung homeostasis and repair. This hypothesis will be tested with three specific aims: 1) that matrix iReFs instruct AEC1 differentiation. 2) that GATA6 transcriptionally regulates a set of matrix fibroblast signature genes and 3) that H3K27me silencing marks regulate GATA6 and matrix gene expression in aged iReFs. In the first aim, alveolosphere cultures will be used to interrogate the epithelial-mesenchymal crosstalk between specific iRef subpopulations and wild type alveolar epithelial cells. In the second aim, gain and loss of function studies of GATA6 will be used to es- tablish a functional link between fibroblast phenotype and alveolarization. In the third aim, H3K27me3 histone marks will be assessed by ChIP-PCR analysis. In vivo and in vitro studies using EZH2 inhibitors will determine a role of EZH2 in suppressing matrix fibroblast signature genes. These studies will provide a better under- standing of the impact of fibroblast diversity on epithelial-mesenchymal crosstalk during alveolarization and alveolar regeneration. This contribution will be significant because it will advance the knowledge about genes and pathways involved in alveolar regeneration, give new insights into the cause of decreased regrowth with age and provide the basis for new therapies to overcome current limitations of regeneration in the human lung. Our understanding of the role of lung fibroblasts in lung injury and regeneration lags far behind our understand- ing of the role of epithelial cells. New information about interstitial lung fibroblasts will be invaluable for defining mechanisms of development, normal lung repair, and lung disease.