Engineered Cardiac Niche Arrays for Exploring and Optimizing Stem Cell Therapies

描述（由申请人提供）：由于成人心肌细胞不能增殖，心肌的自发修复或再生通常是不可能的。因此，病理事件，如心肌梗死，导致永久性损伤，往往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生态位微环境的提示下分化成组织特异性细胞类型，可能为心脏修复提供细胞来源。然而，包括低细胞保留和活力在内的并发症，加上梗死后环境的苛刻和不断变化，阻碍了对心肌梗死干细胞治疗机制的识别，导致临床试验失败。工程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生理的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、修复和再生的活体体外系统。本提案旨在结合软光刻和组织工程开发创新的新工具和方法，总体目标是提高对基于干细胞的心脏修复方法的理解和疗效。一个指导性假设是，三维微环境的机制是控制人间充质干细胞（MSC）和心肌细胞（CM）之间相互作用的关键因素。为了建立这一新的模型体系，验证这一假设，并为该领域未来的研究提供一个跳板，本文设计了两个具体目标。目的1：开发一种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，在受控的3D微环境中增强心肌细胞收缩功能。该目标将集中研究被动拉伸、机械刚度和驻留心脏细胞类型对MSC改善工程心脏组织收缩功能的能力的影响，使用独特的模块化软光刻力传感器阵列。目的2：建立组织工程三维冷冻梗死模型，研究局灶细胞损伤对干细胞迁移、分化和心脏修复的影响。该目标将结合上述工程心脏组织阵列系统和一种新的冷冻梗死方法，在受控的3D模型损伤环境中检测MSC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、高产的建议的成功将导致干细胞力学生物学理解的新进展，促进更复杂的实验动物和临床环境的转化。

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.