Morphogenetic Self-Assembly of Human Heart Organoids

项目摘要 出于实验目的而无法获得人类胚胎是一个长期存在的障碍 人类器官形成的体外研究为了打破这一僵局，R21的总体目标是 建议是建立一个强大的新的三维实验模型系统，概括关键结构和 胚胎生态位环境的生物物理要素，使人类心脏的研究 形态发生和发育我们提出了一种协同组合的创新生物反应器设计， 最先进的人类多能干细胞生物学和尖端的人类心脏组织工程 技术，并根据健全的生物工程原则的定量方法。战略是发展 一种基于微流体的生物反应器系统，用于使用人类细胞生长内皮管、心肌和心脏胶质， 多能干细胞（hPSC）衍生的心肌细胞、成纤维细胞和内皮细胞，以研究其在心肌细胞中的作用。 体外心管形成的早期发育阶段。通过提供流体剪切力，机械 限制，和形态梯度类似的发展中的心管所经历的， 在模拟原始细胞的起始位置用空间分隔的细胞类型接种生物反应器 解剖学，我们的目标是提供一个利基环境，他们可以开始建立一个心脏，因为他们将在 自然本提案的两个具体目标与R21探索性/开发性 生物工程研究资助机制（PA-16-040）：目标1是开发微流体生物反应器， 工程形态发生人的创造和评价的边界约束和流程控制 心脏类器官目的2建立人先天性心管畸形的体外模型， 探索已知会导致体内异位的化学和遗传干预。这种探索性和 发展提案提供了一个迫切需要且可能具有破坏性的飞跃， 现有体外人体模型系统的复杂性。研究团队的多学科专业知识， 结合现有的最先进的资源和设施，提供了一个独特的机会，以克服 预期的技术挑战，并成功地实现了拟议的目标。因此，这被认为 一个高风险，高影响的建议，可能会产生新的工具和见解，了解心脏的过程， 管形成，用于创建更逼真的多组织心脏类器官，并最终研究疾病 使用独特的解剖学体外3-D人类心脏模型系统来研究心脏结构缺陷。

PROJECT SUMMARY A justifiable lack of access to human embryos for experimental purposes has presented a longstanding barrier to the in vitro investigation of human organogenesis. To circumvent this impasse, the overall goal of this R21 proposal is to establish a powerful new 3-D experimental model system that recapitulates key structural and biophysical elements of the embryonic niche environment to enable investigation of human cardiac morphogenesis and development. We propose a synergistic combination of innovative bioreactor design, state-of-the-art human pluripotent stem cell biology, and cutting-edge human cardiac tissue engineering technology, with a quantitative approach based on sound bioengineering principles. The strategy is to develop a microfluidics based bioreactor system to grow endocardial tubes, myocardium, and cardiac jelly using human pluripotent stem cell (hPSC) derived cardiomyocytes, fibroblasts and endocardial cells in order to study the early developmental stages of heart tube formation in vitro. By providing fluid shear forces, mechanical constraints, and morphogen gradients similar to those experienced by the developing heart tube, and by seeding the bioreactor with spatially compartmentalized cell types in starting positions that mimic the primitive anatomy, we aim to provide a niche environment in which they can begin building a heart as they would in nature. The two specific aims of this proposal are consistent with the R21 Exploratory/Developmental Bioengineering Research Grant mechanism (PA-16-040): Aim 1 is to develop a microfluidic bioreactor with boundary constraints and flow control for the creation and evaluation of engineered morphogenetic human heart organoids. Aim 2 is to establish an in vitro model of human congenital heart tube malformation, exploring chemical and genetic interventions known to cause heterotaxy in vivo. This exploratory and developmental proposal offers a desperately needed and potentially disruptive leap forward in the sophistication of available in vitro human model systems. The multidisciplinary expertise of the research team, combined with the available state-of-the-art resources and facilities, provides a unique opportunity to overcome the anticipated technical challenges and successfully achieve the proposed aims. This is therefore considered a high-risk, high-impact proposal, likely to yield new tools and insights for understanding the process of cardiac tube formation, for creating more realistic multi-tissue heart organoids, and for eventually studying diseases related to cardiac structural defects using a unique anatomical in vitro 3-D human heart model system.