De-Cellularized Human Lungs for Ex Vivo Lung Regeneration

描述（由申请人提供）： 永远不会有足够的供体肺来满足当前和未来的移植需求。相反，整个尸体肺的去细胞化将产生完整的支架，其可以用源自个体患者的成体干细胞（包括诱导多能干细胞（iPS））再细胞化，并随后用于自体移植。值得注意的是，去细胞化过程去除了导致免疫排斥的细胞抗原，并且去细胞化的肺保持了天然气道和肺泡结构、细胞外基质蛋白组成和肺血管网络。这将为肺气肿和其他目前无法有效治愈的疾病提供潜在的无限量供应无关供体尸体肺。然而，使用人肺作为支架有几个基本问题需要解决。其中的关键是去细胞化和再细胞化过程的可靠性和可重复性。与同质实验室小鼠不同，供体人肺将来自不同年龄和各种临床和疾病背景的患者，包括吸烟史，这可能会影响脱细胞化后支架的性质或再细胞化过程本身。因此，该提案将集中于确定来自不同供体的去细胞化肺支架之间的关键差异，并发现决定用人类间充质基质细胞（hMSC）和诱导多能干细胞（iPS）成功再细胞化肺支架的关键环境条件。我们的初步数据表明，人肺可以成功地去细胞化，并通过各种组织学和功能评估，包括肺力学，表面活性剂的生产，血管灌注和上皮屏障功能，作为肺再细胞化的研究。此外，初步数据表明，人间充质基质细胞（hMSC）和人肺泡上皮细胞（A549细胞）可以成功地接种并生长在去细胞化的人肺中。这些数据为拟议的两个具体目标提供了坚实的基础。1)确定从不同供体获得的去细胞化肺支架之间的关键差异。2)优化hMSC和人iPS细胞在接种到去细胞化的人肺切片中时生长和分化为功能性三维肺组织的条件，并将该技术应用于整个去细胞化的人肺。(End摘要）

DESCRIPTION (provided by applicant): There will never be enough donor lungs available to meet current and future transplantation needs. In contrast, de-cellularization of whole cadaveric lungs will result in an intact scaffold that can be re- cellularized with adult stem cells, including induced pluripotent stem cells (iPS), derived from individual patients and subsequently utilized for autologous transplantation. Notably, the de-cellularization process removes cellular antigens responsible for immune rejection and the de-cellularized lungs maintain native airway and alveolar architecture, extracellular matrix protein composition, and pulmonary vascular network. This will provide a potentially limitless supply of unrelated donor cadaveric lungs for use in emphysema and other diseases for which there is currently no effective cure. However, there are several fundamental questions to be addressed for use of human lungs as scaffolds. Key among these is the reliability and reproducibility of the de-cellularization and re-cellularization processes. Unlike homogenous laboratory mice, donor human lungs will come from patients of different ages and from a variety of clinical and disease backgrounds, including a history of smoking, which might influence the nature of the scaffold following de-cellularization or the re-cellularization process itself. Therefore this proposal will focus on determining key differences between de-cellularized lung scaffolds originating from different donors and on discovering key environmental conditions that determine successful re-cellularization of lung scaffolds with human mesenchymal stromal cells (hMSCs) and induced pluripotent stem cells (iPS). Our preliminary data demonstrates that human lung can be successfully de-cellularized and studied by a variety of histological and functional assessments including lung mechanics, surfactant production, vascular perfusion, and epithelial barrier function, as the lungs re-cellularize. Further, preliminary data demonstrates that human mesenchymal stromal cells (hMSCs) and human alveolar epithelial cells (A549 cells) can be successfully inoculated and grown in de-cellularized human lung. These data provide a firm platform for the proposed two Specific Aims. 1) To determine key differences between de-cellularized lung scaffolds obtained from different donors. 2) To optimize conditions for growth and differentiation of hMSCs, and human iPS cells into functional three dimensional lung tissue when inoculated into de-cellularized human lung slices and apply this technology to whole de-cellularized human lungs. (End of Abstract)