Engineered Cardiac Niche Arrays for Exploring and Optimizing Stem Cell Therapies

描述(申请人提供)：由于成人心肌细胞不能增殖，通常情况下不可能自发修复或再生心肌。因此，心肌梗死等病理事件会导致永久性损害，往往会导致心力衰竭和死亡。作为一种潜在的治疗策略，干细胞提供了巨大的潜力，因为它们能够在局部利基微环境中根据提示分化为组织特异性细胞类型，可能为心脏修复提供细胞来源。然而，包括低细胞存留率和存活率在内的并发症，再加上心肌梗死后严苛和不断变化的环境，阻碍了对基于干细胞的心肌梗死治疗机制的确定，导致临床试验失败。工程化的心脏组织提供了替代的实验模型系统，结合了比标准培养皿更具生理性的3D环境，具有长期生存能力和改进的实验控制，这是天然心肌制剂所不可能的。然而，工程化心脏组织主要用于外科修复应用，而不是用于研究心肌损伤、修复和再生的体外活系统。这项提议旨在开发结合软光刻和组织工程的创新工具和方法，总体目标是提高基于干细胞的心脏修复方法的理解和有效性。一个指导性的假设是，三维微环境的力学是控制人骨髓间充质干细胞(MSC)和心肌细胞(CM)之间相互作用的关键因素。我们设计了两个具体的目标来建立这个新的模型体系，检验这个假说，并为这一领域的未来研究提供一个跳板。目的1：建立高通量工程心脏组织(ECT)阵列系统，以评估和优化干细胞共培养策略，以在受控的三维微环境中增强心肌细胞的收缩功能。本研究的目的将集中于研究被动拉伸、机械硬度和驻留心肌细胞类型对MSC使用基于独特的模块化软光刻的力传感器阵列来改善工程心脏组织的收缩功能的能力的影响。目的：建立组织工程化三维冷冻心肌梗死模型，研究局灶性细胞损伤对干细胞迁移、分化和心脏修复的影响。这一目标将把上述工程心脏组织阵列系统与一种新的冷冻梗死方法结合起来，在受控的3D模型损伤环境中检测MSC的功能，帮助将Aim 1的发现转化为心肌梗死或ECT植入的实验动物模型。这一高风险、高收益的建议的成功应该会导致在理解干细胞机械生物学方面取得新的进展，促进向更复杂的实验动物和临床环境的转换。 公共卫生相关性：由于心肌的自发修复或再生通常是不可能的，心肌梗死等病理事件会导致永久性损害，通常会导致心力衰竭和死亡。尽管干细胞为心脏修复提供了巨大的潜力，但由于缺乏能够长期研究损伤和修复过程的良好控制的心肌梗死实验模型，我们对干细胞引导分化机制的理解受到阻碍。这项建议旨在开发结合软光刻和组织工程的创新工具和方法，总体目标是了解和指导干细胞分化用于心脏修复应用，这将有望在理解干细胞机械生物学方面取得新的进展，并促进翻译到更复杂的实验动物和人类患者。

DESCRIPTION (provided by applicant): Due to the inability of adult cardiac myocytes to proliferate, spontaneous repair or regeneration of heart muscle is not normally possible. Consequently, pathological events such as myocardial infarction result in permanent damage that often leads to heart failure and death. As a potential therapeutic strategy, stem cells offer great potential due to their ability to differentiate into tissue-specific cell types guided by cues within the local niche microenvironment, possibly providing a cell source for cardiac repair. However, complications including low cell retention and viability, combined with a demanding and evolving post-infarction environment, have impeded the identification of mechanisms governing stem cell based treatments for myocardial infarction, resulting in failed clinical trials. Engineered cardiac tissues offer alternative experimental model systems combining a more physiologic 3D environment than the standard Petri dish, with long-term viability and improved experimental control not possible with natural heart muscle preparations. However, engineered cardiac tissues have been developed primarily for surgical repair applications, rather than as living in vitro systems designed for investigating myocardial injury, repair, and regeneration. This proposal aims to develop innovative new tools and approaches combining soft lithography and tissue engineering, with the overall objective of improving the understanding and efficacy of stem cell based approaches for cardiac repair. A guiding hypothesis is that mechanics of the 3D microenvironment is a key factor governing the interaction between human mesenchymal stem cells (MSC) and cardiac myocytes (CM). Two specific aims are designed to establish this new model system, test this hypothesis, and provide a springboard for future studies in this area. Aim 1: To develop a high-throughput engineered cardiac tissue (ECT) array system to evaluate and optimize stem cell co-culture strategies for enhancing cardiomyocyte contractile function in a controlled 3D microenvironment. This aim will focus on studying the effects of passive stretch, mechanical stiffness, and resident cardiac cell types on the ability of MSC to improve the contractile function of engineered cardiac tissues using a unique modular soft lithography based force sensor array. Aim 2: To establish a tissue engineered 3D cryo-infarct model to examine the effects of focal cell injury on stem cell migration, differentiation, and cardiac repair. This aim will combine the above engineered cardiac tissue array system with a novel cryo-infarct approach for examining MSC function in a controlled 3D model injury environment, helping to translate the findings of Aim 1 to experimental animal models of myocardial infarction or ECT implantation. Success of this high- risk, high-yield proposal should lead to new advances in understanding stem cell mechanobiology, facilitating translation to more complex experimental animal and clinical settings. PUBLIC HEALTH RELEVANCE: Because spontaneous repair or regeneration of heart muscle is not normally possible, pathological events such as myocardial infarction result in permanent damage that often leads to heart failure and death. Although stem cells offer great potential for cardiac repair, our understanding of the mechanisms guiding differentiation of stem cells is hampered by a lack of well-controlled experimental models of myocardial infarction that allow long term study of injury and repair processes. This proposal aims to develop innovative new tools and approaches combining soft lithography and tissue engineering, with the overall objective of understanding and directing stem cell differentiation for cardiac repair applications, which will hopefully lead to new advances in understanding stem cell mechanobiology and facilitate translation to more complex experimental animals and human patients.