Developing a patient-specific organoid model of pulmonary fibrosis using iPSCs

项目摘要/摘要 特发性肺纤维化(IPF)仍然是一种致命的间质性肺疾病(ILD)，正在接受治疗 由于对启动和延续机制的不完全了解，各种选择受到限制 疾病。现在越来越多的文献表明肺上皮功能障碍在这些事件中发挥了作用。 这会导致下游成纤维细胞的激活，最终导致无情的纤维化。这些研究， 再加上观察到所有形式的IPF的肺上皮细胞都显示端粒缩短， 提示肺上皮功能障碍可能引发IPF，并加速衰老表型或 端粒酶途径异常可能在此发病机制中起重要作用。但是，如果不能访问 特定于患者的人类上皮-间充质模型系统，用于测试的选项有限 基因多态或端粒酶紊乱引起上皮细胞改变的假说 可能对IPF有机械上的贡献。在这里，我们建议开发一种基于人类有机化合物的体外 IPF研究的模型系统。我们已经建立了诱导多能干细胞的生物库 来自散发性或家族性肺纤维化患者的细胞(IPSCs)。在目标1中，我们 应用该文库指导携带端粒酶的IPF IPSCs的体外分化 突变(与正常IPSCs相比)进入不同的呼吸道和肺泡上皮细胞，目的是 生成一种简化的、仅限于上皮的固有上皮功能障碍的3D培养模型 我们推测可能会引发肺纤维化。在目标2中，我们通过以下方式增加该模型的复杂性 介绍由IPSC来源的肺上皮细胞与人并列组成的人器官类化合物 间充质谱系，以模拟假设的上皮-间充质相互作用 使IPF永久化。最后，在目标3中，我们测试了不同端粒酶途径突变的假设 导致共有的肺上皮细胞紊乱，包括短端粒和P53激活，然后 导致下游间充质细胞激活。

Project Summary/Abstract Idiopathic pulmonary fibrosis (IPF) remains a deadly interstitial lung disease (ILD) with treatment options limited by an incomplete understanding of the mechanisms that initiate and perpetuate disease. A growing literature now implicates lung epithelial dysfunction as playing a role in the events that lead to downstream fibroblast activation, culminating in relentless fibrosis. These studies, together with the observation that lung epithelial cells in all forms of IPF display shortened telomeres, suggests that lung epithelial dysfunction may initiate IPF, and accelerated aging phenotypes or telomerase pathway abnormalities likely contribute to this pathogenesis. However, without access to patient-specific human epithelial-mesenchymal model systems, there are limited options for testing hypotheses of how epithelial changes induced by gene polymorphisms or telomerase perturbations might mechanistically contribute to IPF. Here we propose to develop a human organoid-based in vitro model system for the study of IPF. We have established a biorepository of induced pluripotent stem cells (iPSCs) generated from individuals with sporadic or familial pulmonary fibrosis. In aim 1 we apply this repository by directing the in vitro differentiation of banked IPF iPSCs carrying telomerase mutations (vs normal iPSCs) into various airway and alveolar lung epithelial cells for the purpose of generating a reductionist, epithelial-only 3D culture model of the intrinsic epithelial dysfunction that we posit may initiate pulmonary fibrosis. In aim 2 we augment the complexity of this model by introducing human organoids composed of iPSC-derived lung epithelia juxtaposed with human mesenchymal lineages in order to model the epithelial-mesenchymal interactions hypothesized to perpetuate IPF. Finally, in aim 3 we test the hypothesis that different telomerase pathway mutations result in shared lung epithelial perturbations, including short telomeres and p53 activation, that then leads to downstream mesenchymal activation.