The Fetal Lung Mesothelial Differentiation Program

描述(由申请人提供)：间皮细胞在多大程度上促进肺发育和出生后修复是该领域的一个开放的和基本的问题。这笔赠款的目标是解决这一根本问题，并评估Wilm的肿瘤1转录因子(WT1)在这些事件中的关键作用。使用携带带有敲入Cre重组酶和GFP基因的WT1等位基因的小鼠，我们产生了初步数据，从而得出了3个假设，这些假设将被检验：1)胎儿间皮含有分化的间充质肺细胞的前体细胞；2)WT1控制关键基因的表达，如控制间皮细胞迁移到胎肺的Hedgehog(HH)途径成分，以及3)间皮衍生细胞促进出生后肺修复和再生长。综上所述，我们发现WT1在E11.5-E16的肺间皮细胞中选择性表达，在成人中检测不到。我们发现在灵长类动物肺中WT1的表达具有相似的时间模式，这表明在哺乳动物中存在保守的间皮WT1程序。谱系追踪表明，间皮源性细胞产生大量的支气管平滑肌细胞(BSM)，以及其他将被确定其身份的实质肺细胞(目标1)。我们观察到WT1的表达与间皮细胞进入下层肺和活跃的HH信号相一致。在机制上，我们发现WT1与间皮细胞中多个HH途径基因的启动子结合，并且选择性地丢失间皮细胞HH信号显著减弱进入下肺的能力，这与EMT基因的表达减少有关。这些数据指出WT1在胎儿间皮细胞中的关键作用，控制着参与迁移和EMT的HH信号通路，这一点将进一步探索(目标2)。有趣的是，初步数据表明，WT1在全肺切除后肺重新生长过程中被重新激活，而WT1在炎症性肺损伤中不被重新激活，例如哮喘和纤维化。这些发现为间皮细胞如何在出生后生活中促进肺重塑提供了两个模型。在模型1中，胎儿WT1调节的间皮程序被重新激活。在模型2中，来自发育中的胎儿间皮的实质细胞有助于修复。这些模型在多大程度上参与了出生后生活中肺的再生长和重塑，将进一步研究(目标3)。 我们期望这些研究的完成将为今后这方面的工作奠定坚实的基础。 肺部生物学的新领域。

DESCRIPTION (provided by applicant): To what extent mesothelial-derived cells contribute to lung development and post-natal repair is an open and basic question for the field. The objectives of this grant are to address this fundamental issue and to assess the key role of the Wilm's tumor 1 transcription factor (WT1) in these events. Using mouse lines that carry WT1 alleles with a knock-in Cre recombinase and GFP genes, we generated preliminary data leading to 3 hypotheses that will be examined: 1) the fetal mesothelium contains progenitors for differentiated mesenchymal lung cells 2) WT1 controls the expression of key genes, such as hedgehog (Hh) pathway constituents that control mesothelial migration into the fetal lung, and 3) mesothelium-derived cells contribute to post-natal lung repair and re-growth. To summarize, we found that WT1 is selectively expressed in the lung mesothelium from E11.5 to E16 and is undetectable in the adult. We identified a similar temporal pattern of WT1 expression in the primate lung, suggesting a conserved mesothelial WT1 program across mammalian species. Lineage tracing showed that mesothelium-derived cells give rise to a substantial number of bronchial smooth muscle cells (BSM), along with other parenchymal lung cells whose identities will be established (Aim 1). We observed that WT1 expression coincides with mesothelial cell entry into the underlying lung and active Hh signaling. Mechanistically, we found that WT1 binds to the promoters of multiple Hh pathway genes in mesothelial cells, and that selective loss of mesothelial Hh signaling markedly attenuates entry into the underlying lung in association with diminished expression of EMT genes. These data point to a key role for WT1 in the fetal mesothelium, controlling pathways such as Hh signaling that are involved in migration and EMT, which will be further explored (Aim 2). Interestingly, preliminary data indicate that WT1 is reactivated during lung re-growth post-pneumonectomy whereas WT1 is not re-activated in inflammatory lung injuries, such as asthma and fibrosis. These findings suggest 2 models for how the mesothelium may contribute to lung remodeling in post-natal life. In model 1, the fetal WT1-regulated mesothelial program is re-activated. In model 2, parenchymal cells that arise from the fetal mesothelium in development contribute to repair. To what degree these models are involved in lung re- growth and remodeling in post-natal life will be further examined (Aim 3). We expect that completion of these studies will establish a firm foundation for future work in this new area of lung biology.