The role of transcription factor MEOX2 in lipofibroblast function during alveolarization

项目摘要 位于肺远端间充质的肺脂成纤维细胞支持肺泡上皮细胞。 两种成人起病的纤维化肺疾病(如特发性肺纤维化，IPF)的稳态和修复 新生儿肺部疾病(如支气管肺发育不良，BPD)。具体地说，LFS：1)吸收和 在肺泡化过程中将脂肪酸转移到肺泡2型(AT2)上皮细胞以产生表面活性物质，以及 2)提供旁分泌配体，如FGF10，刺激肺泡1型(AT1)和AT2增殖，并 差异化。更好地理解肺脂肪成纤维细胞表型的转录调控 将导致针对BPD和IPF的靶向治疗的开发。转录因子MEOX2是一种签名 由公开可用的RNA测序数据集鉴定的小鼠发育中的脂肪成纤维细胞的基因。在……里面 其他器官系统，MEOX2控制心血管内皮细胞的脂肪酸摄取，是 棕色脂肪的成脂作用，对肢体早期间充质的增殖和分化起关键作用 发展。尽管MEOX2在非小细胞肺癌中表达上调，但MEOX2在肺中的作用 发展是完全未知的。我们预测MEOX2将在脂肪成纤维细胞分化中发挥作用， 因此，我们努力生成MEOX2的S上下游功能的初步数据。PPAR信令是 一种已知的脂肪成纤维细胞分化的激活剂，因此我们处理了人PDGFRA胎肺成纤维细胞(IMR90 用PAN-PPAR激活剂二甲双胍诱导MEOX2的表达。此外，我们还进行了一次 人MEOX2基因座转录因子基序搜索及鉴定可能的过氧化物酶增殖物 响应要素(PPRE)。然后我们在IMR90细胞中过表达了MEOX2，并确定MEOX2是 足以诱导脂成纤维细胞特征基因PLIN2和TCF21的表达。这一点的中心假设是 GRANT是MEOX2由PPARA信号直接调节，是建立的必要条件和充分条件 在肺泡化过程中脂成纤维细胞的谱系和功能。目标1将调查推定的PPRE是否 利用分子遗传学方法诱导MEOX2表达的充要条件 永生化的人肺成纤维细胞中。目标2将调查MEOX2是否必要且足以诱导 脂成纤维细胞基因表达、脂肪酸摄取和转运与肺泡上皮的支持 通过1)结合瞬时MEOX2基因敲除和过表达的体外实验进行分化 构建，以及2)体内条件成纤维细胞特异性的MEOX2在牙槽骨形成早期的敲除。我们的 长期目标是确定MEOX2的S作为转录因子在肺间充质发育中的作用， 希望找到治疗慢性新生儿和成人肺部疾病的靶点。这项培训计划 结合分子生物学、细胞生物学和发育生物学回答有关肺脏的基本问题 成纤维细胞生物学。我的职业目标是成为了解转录因子如何调控的领导者 发育中的细胞分化，以及它们如何成为疾病治疗的靶点。

Project Summary Pulmonary lipofibroblasts (LFs) that reside in the distal lung mesenchyme support alveolar epithelial cell homeostasis and repair in both adult-onset fibrotic lung diseases (such as idiopathic pulmonary fibrosis, IPF) and neonatal lung diseases (such as bronchopulmonary dysplasia, BPD). Specifically, LFs: 1) uptake and transfer fatty acids to alveolar type 2 (AT2) epithelial cells during alveolarization for surfactant production, and 2) provide paracrine ligands such as FGF10 that stimulate alveolar type 1 (AT1) and AT2 proliferation and differentiation. A better understanding of the transcriptional regulation of the pulmonary lipofibroblast phenotype will lead to the development of targeted therapies for BPD and IPF. Transcription factor MEOX2 is a signature gene of the murine developmental lipofibroblast, as identified by publicly available RNA-sequencing datasets. In other organ systems, MEOX2 controls fatty acid uptake in cardiovascular endothelial cells, is necessary for brown fat adipogenesis, and is critical for mesenchymal proliferation and differentiation in early limb development. Although MEOX2 is upregulated in non-small cell lung cancer, the role of MEOX2 in lung development is completely unknown. We predicted that MEOX2 would have a role in lipofibroblast differentiation, so we strove to generate preliminary data on MEOX2’s upstream and downstream function. PPAR signaling is a known activator of lipofibroblast differentiation, so we treated human PDGFRa+ fetal lung fibroblasts (IMR90 cells) with pan-PPAR activator metformin, which induced expression of MEOX2. Additionally, we performed a transcription factor motif search at the human MEOX2 locus and identified a putative peroxisome proliferator response element (PPRE). We then overexpressed MEOX2 in IMR90 cells and determined that MEOX2 is sufficient to induce expression of lipofibroblast signature genes PLIN2 and TCF21. The central hypothesis of this grant is that MEOX2 is directly regulated by PPARA signaling and is necessary and sufficient for establishment of the lipofibroblast lineage and function during alveolarization. Aim 1 will investigate if the putative PPRE is necessary and sufficient for induction of MEOX2 expression through the use of molecular genetic approaches in immortalized human lung fibroblasts. Aim 2 will investigate if MEOX2 is necessary and sufficient to induce lipofibroblast gene expression, fatty acid (FA) uptake and transfer, and support of alveolar epithelial differentiation through 1) in vitro assays combined with transient MEOX2 knockout and overexpression constructs, and 2) an in vivo conditional fibroblast specific MEOX2 knockout during early alveolarization. Our long-term goal is to identify MEOX2’s role as a transcription factor in the development of the lung mesenchyme, with the hope of discovering therapeutic targets for chronic neonatal and adult lung diseases. This training plan combines, molecular, cellular, and developmental biology to answer fundamental questions about pulmonary fibroblast biology. My career goal is to become a leader in understanding how transcription factors regulate cellular differentiation in development, and how they may be targetable for therapies in disease.