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

描述(申请人提供)：尽管肺气肿的对症治疗取得了进展，但这种毁灭性的肺部疾病在全球范围内的患病率正在增加，除了肺移植之外，仍然没有治愈的方法。我们提出了利用胚胎干细胞(ESCs)、成人间充质干细胞(MSCs)或诱导多能干细胞(IPS)在三维培养基质中培养的新的组织工程方法，可以再生功能性肺组织来替代肺气肿组织。我们发现，骨髓间充质干细胞植入去细胞的小鼠肺中，使呼吸道和实质肺区重新细胞化，并在实质肺区获得2型肺泡上皮细胞的表型标记。这些发现增加了在脱细胞肺中培养的胚胎或成体干细胞可用于体外工程新肺组织的可能性，这可能是一种在肺气肿肺中再生新肺泡的治疗策略。此外，在正常呼吸过程中，肺细胞会受到周期性拉伸和其他力的作用。我们发现，循环拉伸培养的MSCs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的表达，如成纤维细胞或肌肉细胞。这些初步研究提出了一种可能性，即通过施加与正常肺相似的机械力来特定地操纵培养环境，可以进一步促进从胚胎或成年干细胞发育成功能肺组织。三维培养和循环拉伸，结合特定的呼吸道生长介质，对MSCs、ESCs和iPS的肺细胞发育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人干细胞进行肺组织工程提供坚实的科学基础，作为治疗肺气肿和其他肺部疾病的可能方法。 公共卫生相关性：慢性阻塞性肺疾病(COPD)是全球第四大最常见的死亡原因，它仍然是一种毁灭性的致命肺部疾病，是发病率上升的仅有的几种重大疾病之一，除了肺移植之外，没有治愈的方法。我们最近的工作提出了一种新的基于细胞的治疗方法，表明在三维支架中培养的成人间充质干细胞可以形成有活力的肺组织，用于肺气肿新肺的再生。目前建议的重点是进一步探索三维支架和机械拉伸是否可以用于从间充质干细胞以及胚胎和诱导的多能干细胞直接生成具有功能的三维肺组织。这些研究将为未来干细胞治疗肺气肿的翻译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.