CAREER: Engineering ex vivo Human Cardiogenesis with Optogenetics

人类诱导多能干细胞（human induced pluripotent stem cells，hiPSCs）是一种特殊的细胞类型，在适当的环境下可以转化为任何类型的细胞，这为研究人体组织和器官发育提供了前所未有的机会，并有可能应用于治疗疾病。更狭义地说，hiPSC衍生的体外器官模型（称为类器官）已成为构建精细组织样结构的有力工具，这些结构可以准确模拟许多组织或器官（如肠、肾和脑）的形态。最近，心脏类器官由hiPSC产生，但多个心脏细胞的规格和组织的空间控制仍然具有挑战性。这个CAREER项目旨在利用光遗传学控制hiPSC形成有组织的心脏类器官，光遗传学是一种利用光控制细胞或组织活性的生物技术，然后利用这些技术研究心脏成熟和心脏疾病。教育使命将通过让不同层次的学生参与跨学科干细胞研究和社区外展活动，包括互动课程，研讨会，讲习班，研究经验和普渡美术馆的公共艺术展，融入本研究计划。研究者的长期职业目标是创建一个变革性的跨学科研究和教育计划，将干细胞和基因精准医学的科学和工程方面的卓越成果与未来科学家和工程师的教育相结合。 为了实现这一目标，该CAREER项目将开发新的工具和标准化方法，利用光遗传学精确设计人类类器官形态发生，用于发育研究和疾病建模。以模拟天然心脏发育的方式工程化心脏发生将导致心脏类器官的上级结构和功能特性。本研究的中心假设是，由三维（3D）心脏类器官内的异型细胞-细胞相互作用诱导的旁分泌和旁分泌信号将增强心肌细胞（CM）的成熟和功能。为了验证这一假设，提出了三个任务：（1）建立3D光遗传学平台并验证其在多细胞形态发生的信号传导模式中的功能;（2）使用光遗传学产生心脏类器官以促进和询问3D中hiPSC衍生的CM的成熟;以及（3）研究遗传缺陷对心脏类器官的细胞组成、组织结构和成熟的影响。拟议的研究将揭示实现hiPSC-CM成熟的关键因素，并阐明患病CM中进行性肌无力和死亡的潜在机制。由于细胞成熟是组织工程的一个关键因素，这项研究将在工程其他hiPSC衍生的tissues.This奖项反映了NSF的法定使命，并已被认为是值得通过使用基金会的智力价值和更广泛的影响审查标准进行评估的支持更广泛的分支。

Human induced pluripotent stem cells (hiPSCs) are a special cell type that under the right environment can transform into any type of cell, which provide an unprecedented opportunity to study human tissue and organ development and potentially be applied to treating disease. More narrowly, hiPSC-derived in vitro organ models, termed organoids, have emerged as powerful tools to construct elaborate tissue-like architectures that can accurately mimic the morphology of numerous tissues or organs, such as the intestines, kidney, and brain. More recently, cardiac organoids were created from hiPSCs, but the spatial control of the specification and organization of multiple cardiac cells remain challenging. This CAREER project seeks to control the formation of organized cardiac organoids from hiPSCs using optogenetics, a biological technique to control the activity of cells or tissues with light, and then use these to study cardiac maturation and heart diseases. The educational mission will be integrated into this research program by engaging students at different levels into interdisciplinary stem cell research and community outreach activities, including interactive coursework, seminars, workshops, research experiences, and a public art exhibition at Purdue Galleries. The investigator’s long-term career goal is to create a transformative, interdisciplinary research and education program that integrates excellence in the science and engineering of stem cell- and gene-based precision medicine with education of future scientists and engineers. Towards this goal, this CAREER project will develop novel tools and standardized methodologies to precisely engineer human organoid morphogenesis with optogenetics for developmental studies and disease modeling. Engineering cardiogenesis in a manner mimicking native heart development will result in superior structural and functional properties of the cardiac organoids. The central hypothesis of this research is that the paracrine and juxtacrine signals induced by heterotypic cell-cell interactions within the three-dimensional (3D) cardiac organoids will enhance cardiomyocyte (CM) maturation and function. To test this hypothesis, three tasks are proposed: (1) establish a 3D optogenetic platform and validate its function in signaling patterning for multicellular morphogenesis; (2) generate cardiac organoids using optogenetics to facilitate and interrogate the maturation of hiPSC-derived CMs in 3D; and (3) investigate the influence of genetic defects on cellular composition, tissue structure and maturation of cardiac organoids. The proposed research will reveal critical factors to achieve maturation of the hiPSC-CMs and elucidate underlying mechanisms of progressive muscle weakness and death in diseased CMs. Since cell maturation is a key element of tissue engineering, this research will have broader ramifications in engineering other hiPSC-derived tissues.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.