Optimal Derivation of Murine Embryonic Distal Airway Stem Cells

描述（由申请方提供）：肺发育不全（PH）的病理包括肺质量减少、由于表面活性蛋白产生不足导致肺保持扩张的能力不足、气囊（肺泡上皮）衬里分化不良和气体交换减少。我们的长期目标是使用干细胞疗法来减轻新生儿PH，以体外干细胞衍生的远端气道细胞来增加早产儿肺组织。具体的假设是，通过提供转录和生长因子重演关键发育事件将优化远端气道细胞的衍生。这包括两步过程，即定形内胚层的衍生，然后分化成远端气道细胞。此外，提供三维支架将通过提供重要的空间线索来提高远端气道细胞的产量。在肺中，远端气道的发育和完整性对于适当的气体交换、表面活性剂的产生和存活至关重要。这项拨款计划的重点是探索从胚胎干细胞中获得远端气道细胞的方法。我们的具体目标包括：一曰：优化小鼠胚胎干细胞的定形内胚层细胞的衍生，特别是通过操纵Wnt和nodal途径或通过提供小分子。2：通过补充生长因子（特别是FGF 2）优化远端气道细胞的衍生，7，10。3：利用具有潜在体内移植的三维水凝胶支架优化空间线索，时间允许。 公共卫生相关性：早产导致肺发育不全是产科的主要问题，占围产期死亡率的70%以上。2004年，美国有12.5%的新生儿是早产儿。这些未成熟的肺具有减少的成熟上皮细胞数量，这导致表面活性剂产生不足（用于保持远端气道开放的蛋白质）以及减少的脉管系统。我们建议利用小鼠胚胎干细胞来获得远端气道细胞，以用于肺发育不良。这项探索性研究可能有助于确定获得远端气道细胞的最佳方法，这些细胞可能用于治疗肺部疾病，如肺发育不全。

DESCRIPTION (provided by applicant): The pathology of Pulmonary Hypoplasia (PH) includes reduced lung mass, insufficient ability of the lungs to stay expanded due to a deficiency in surfactant protein production, a poorly differentiated lining of the air sac (alveolar epithelium), and a reduction of gas exchange. Our long term goal is to alleviate neonatal PH using stem cell therapy to augment premature lung tissue with in vitro stem cell derived distal airway cells. The specific hypothesis is that recapitulation of key developmental events by providing transcription and growth factors will optimize derivation of distal airway cells. This includes a 2 step process with derivation of definitive endoderm followed by differentiation into distal airway cells. Furthermore, providing a 3-dimensional scaffold will enhance our yield of distal airway cells by providing important spatial cues. In the lung, distal airway development and integrity is essential for proper gas exchange, surfactant production and survival. The focus of this grant proposal is to explore methods of deriving distal airway cells from embryonic stem cells. Our specific aims include: 1: Optimize derivation of definitive endoderm cells from murine embryonic stem cells, specifically by manipulating the Wnt and nodal pathway or by providing small molecules. 2: Optimize derivation of distal airway cells through growth factor supplementation, specifically FGF2, 7,10. 3: Optimize spatial cues utilizing 3-dimensional hydrogel scaffolds with potential transplantation in vivo, time permitting. PUBLIC HEALTH RELEVANCE: Preterm delivery with resultant immature lungs (pulmonary hypoplasia) is a major problem in obstetrics and accounts for more than 70% of perinatal mortality. In 2004, 12.5% of births in the United States were preterm. These immature lungs have reduced numbers of mature epithelial cells which results in insufficient surfactant production (a protein used to keep the distal airways open) as well as decreased vasculature. We propose to utilize murine embryonic stem cells to derive distal airway cells for possible utilization in pulmonary hypoplasia. This exploratory study may help identify optimal methods of deriving distal airway cells that can potentially be utilized for treatment of pulmonary diseases such as pulmonary hypoplasia.