Optimal Derivation of Murine Embryonic Distal Airway Stem Cells

描述(申请人提供)：肺发育不良(PH)的病理包括肺质量减少，由于表面活性物质蛋白生产不足而导致肺保持扩张的能力不足，气囊(肺泡上皮)分化较差，以及气体交换减少。我们的长期目标是使用干细胞治疗来缓解新生儿肺高压，用体外干细胞来源的远端呼吸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来重现关键发育事件将优化远端呼吸道细胞的起源。这包括两个步骤的过程，即获得最终的内胚层，然后分化为远端呼吸道细胞。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呼吸道细胞的产量。在肺中，远端气道的发育和完整性对于适当的气体交换、表面活性物质的产生和生存至关重要。这项拨款提案的重点是探索从胚胎干细胞中提取远端呼吸道细胞的方法。我们的具体目标包括：1：优化从小鼠胚胎干细胞中获得确定的内胚层细胞，特别是通过操纵Wnt和Node途径或提供小分子。2：通过补充生长因子优化远端呼吸道细胞的来源，特别是FGF2，7，10.3：在时间允许的情况下，利用具有体内移植潜力的三维水凝胶支架优化空间线索。

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.