Use of 3-D Culture and Stretch to Develop Lung from MSCs, ESCs, and iPS

描述（由申请人提供）：尽管肺气肿的对症治疗取得了进展，但这种毁灭性肺部疾病的患病率在全球范围内不断增加，除了肺移植外，仍然没有治愈方法。我们提出，新的组织工程方法，利用胚胎干细胞（ESC），成人间充质干细胞（MSC），或诱导多能干细胞（iPS）培养在三维培养基质可以再生功能肺组织的肺气肿肺组织的替代。我们已经发现，MSC浸渍到去细胞化的小鼠肺再细胞化气道和实质肺区域，并获得实质肺区域中的2型肺泡上皮细胞的表型标志物。这些发现提高了在去细胞化的肺中培养的胚胎或成体干细胞可用于体外工程化新的肺组织的可能性，并且这可能是在肺气肿肺中再生新肺泡的治疗策略。此外，肺细胞在正常呼吸期间经受周期性拉伸和其他力。我们已经发现，培养的MSC的周期性拉伸特异性地诱导肺上皮蛋白的表达，同时降低其他细胞类型的标志物（例如成纤维细胞或肌肉细胞）的表达。这些初步研究提出了这样一种可能性，即通过施加机械力对培养环境进行特定操作，使人联想到在正常肺中发现的那些机械力，可以进一步促进从胚胎或成体干细胞发育功能性肺组织。三维培养和循环拉伸，结合特定的气道生长培养基，对肺细胞从MSC，ESC和iPS的发展的影响将在本提案中进行探索和定义。这些研究将为利用胚胎和成体干细胞作为肺气肿和其他肺部疾病的可能治疗方法的肺组织工程提供坚实的科学基础。 公共卫生相关性：慢性阻塞性肺疾病（COPD）是全球第四大常见死因，仍然是一种毁灭性的致命肺病，是患病率增加的唯一主要疾病之一，除了肺移植外，没有治愈方法。我们最近的工作表明，在三维支架中培养的成体间充质干细胞可能形成有活力的肺组织，用于肺气肿的新肺再生，这提出了一种新的基于细胞的治疗方法。目前建议的重点是进一步探索三维支架以及机械拉伸是否可以用于从间充质干细胞以及从胚胎和诱导多能干细胞直接生成功能性三维肺组织。这些研究将为将来干细胞治疗肺气肿的转化和临床试验提供坚实的科学基础。

DESCRIPTION (provided by applicant): Despite advances in the symptomatic treatment of emphysema, the prevalence of this devastating lung disease is increasing worldwide and there remains no cure save for lung transplantation. We propose that novel tissue engineering approaches utilizing embryonic stem cells (ESCs), adult mesenchymal stem cells (MSCs), or induced pluripotent stem cells (iPS) cultured in three-dimensional culture matrices can regenerate functional lung tissue for replacement of emphysematous lung tissue. We have found that MSCs impregnated into de-cellularized mouse lungs re-cellularize both airways and parenchymal lungs regions and acquire phenotypic markers of type 2 alveolar epithelial cells in the parenchymal lung regions. These findings raise the possibility that embryonic or adult stem cells cultured in de-cellularized lungs can be utilized to engineer new lung tissue ex vivo and that this might be a therapeutic strategy for regenerating new alveoli in emphysematous lungs. Further, lung cells are subjected to cyclic stretch and other forces during normal breathing. We have found that cyclic stretch of cultured MSCs specifically induces expression of lung epithelial proteins while decreasing expression of markers of other cell types such as fibroblasts or muscle cells. These preliminary studies raise the possibility that specific manipulation of the culture environment by applying mechanical forces reminiscent of those found in normal lung can further promote development of functional lung tissue from embryonic or adult stem cells. The influences of 3-dimensional culture and of cyclic stretch, in combination with a specific airways growth medium, on development of lung cells from MSCs, ESCs, and iPS will be explored and defined in this proposal. These studies will provide a firm scientific basis for lung tissue engineering utilizing embryonic and adult stem cells as a possible therapeutic approach for emphysema and other lung diseases. PUBLIC HEALTH RELEVANCE: Chronic obstructive pulmonary disease (COPD), the fourth most common cause of death worldwide, remains a devastating fatal lung disease that is one of the only major diseases increasing in prevalence and for which there is no cure save lung transplantation. A new cell-based therapy approach is suggested by our recent work demonstrating that adult mesenchymal stem cells cultured in three-dimensional scaffolds may form viable lung tissue for potential use in regenerating new lung in emphysema. The focus of the current proposal is to further explore whether 3 dimensional scaffolds as well as mechanical stretch may be used to direct generation of functional 3- dimensional lung tissue from mesenchymal stem cells as well as from embryonic and induced pluripotent stem cells. These studies will provide a firm scientific basis for future translational and clinical trials of stem cells for treatment of emphysema.