Interstitial resident fibroblasts direct alveolar epithelial differentiation

项目概述：虽然在理解上皮细胞前体的作用方面取得了巨大进展， 基因在组织修复和纤维化机制中起重要作用，但对基因的起源、相互作用的性质和基因的表达却知之甚少。 成纤维细胞祖细胞的调节程序。肺部研究受到了 成纤维细胞群和间充质干细胞群的复杂性和多样性。我们的长期目标 是了解成纤维细胞群体的复杂性，并确定其对上皮细胞的调节作用， 在肺泡化和上皮修复过程中。我们的主要目标是确定成纤维细胞分化的关键调节因子， 肺里的生殖细胞本申请是基于我们的初步数据，其将GATA 6识别为密钥 基质iReFs中的转录因子，其对于肺泡化是不可缺少的。我们的核心假设是， GATA 6调节iReF中的基因表达，其亚序贯诱导基质iReF特化和帕拉诱导。 在发育和修复过程中与AEC的相互作用。拟议研究的理由是， 了解成纤维细胞亚群的调节，以及它们在上皮-间充质细胞中的独特作用 相互作用，将提供新的答案有关肺内稳态和修复的基本问题。 将用三个特定目的来检验该假设：1）基质iReF指导AEC 1分化。（二） GATA 6转录调节一组基质成纤维细胞标记基因，3）H3 K27 me沉默 标记调节老化iReFs中GATA 6和基质基因的表达。在第一个目标中，肺泡文化将 用于询问特定iRef亚群和野生iRef亚群之间的上皮-间充质串扰。 型肺泡上皮细胞。在第二个目标中，GATA 6的功能获得和丧失研究将用于评估 在成纤维细胞表型和肺泡化之间建立功能性联系。在第三个目标中，H3 K27 me 3组蛋白 将通过ChIP-PCR分析评估分数。使用EZH 2抑制剂的体内和体外研究将确定 EZH 2在抑制基质成纤维细胞标记基因中的作用。这些研究将提供一个更好的下- 成纤维细胞多样性对肺泡化过程中上皮-间充质串扰的影响， 肺泡再生这一贡献将是重大的，因为它将推进有关基因的知识 以及参与肺泡再生的途径，为再生减少的原因提供了新的见解， 并为新疗法提供基础，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- 上皮细胞的作用。关于间质性肺成纤维细胞的新信息对于定义 发育机制、正常肺修复和肺病。

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.