CAREER: Engineering Stem Cell-Based Cardiac Organoids

职业：工程基于干细胞的心脏类器官

• 批准号: 1943798
• 负责人: Zhen Ma
• 金额: $ 50万
• 依托单位: Syracuse University
• 依托单位国家: 美国
• 项目类别: Continuing Grant
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-04-01 至 2025-03-31
• 项目状态: 未结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=1943798&HistoricalAwards=false
• 关键词: CAREER; Engineering; Stem; Cell; Based

Human induced pluripotent stem cells (hiPSCs), cells that have the ability to become any cell type, have provided an unprecedented opportunity to study human-specific tissue and organ development. By differentiating hiPSCs into organ-specific cell types, micro/mini tissues, termed organoids, can be developed to have many of the characteristics of the organ of interest. These stem cell-based organoids have been hypothesized to develop in a manner similar to actual organs in an embryo. However, one of the challenges of current 3D organoid technology is lack of spatial physical controls to promote spatial tissue patterning in these self-assembled organoids, which often appear to have a random distribution of cells. This CAREER project seeks to control hiPSC growth and differentiation under geometrical confinement in order to generate beating cardiac organoids with spatially distinct tissue architecture. This cardiac organoid system could provide insights about how physical cues regulate the structural, functional and cellular properties of stem cell organoids. The project aims to synergize the educational mission with the research program to increase the opportunities for students at different levels to participate in interdisciplinary research at the interface of stem cell biology and microsystem engineering. Furthermore, the project is dedicated to community outreach to the area surrounding the Central New York Region, which is demonstrated by plans to develop and donate an exhibit, “The Superpower of Stem Cells,” to the Syracuse Milton J. Rubenstein Museum of Science & Technology (MOST).The investigator’s long-term research goal is to gain in-depth mechanistic understanding of the role of biophysical factors in human heart development and diseases. Towards this goal, the goal of this CAREER project is to engineer a cardiac organoid system with mechano-geometrical inputs to establish a basis for understanding how biophysical cues, particularly physical confinement, affect organoid formation, tissue functionality, and spatial cell differentiation. The Research Plan is organized under three objectives. The FIRST Objective is to investigate the influence of biophysical cues on regulating the structural morphology and contractile functions of cardiac organoids. To create the organoids, hiPSCs will be seeded onto PEG patterned substrata, expanded to near confluence on the individual patterns, and then differentiated into cardiac lineages. To investigate how physical confinement affects cardiac organoid formation and functions, hiPSCs will be seeded into circle-shaped patterns with different diameters to emphasize size differences, and triangle, square, and rectangular shaped patterns with the same geometrical areas as the circles to emphasize shape differences. The organoids will be characterized relative to their contractile function (contractile motion, action potential, calcium transport and heart rate) and structural morphology (wellness, height and width). These characterizations will be used to establish the correlation between organoid structure and cardiac functions, and to study how structure-function relationship will be shifted by the changes on pattern geometry (size and shape). Objective outcomes are expected to enable determination of the optimal physical confinement that might create cardiac organoids with the highest consistency in morphology. THE SECOND Objective is to investigate the influence of biophysical cues on regulating the cellular composition of cardiac organoids, particularly cardiomyocytes (CMs), cardiac fibroblasts (CFs), endocardial cells (EDCs), smooth muscle cells (SMCs), and epicardial cells (EPCs). Cardiac organoids will be differentiated under different pattern geometries with additional differentiation protocols with different cytokines (e.g., VEGF,, retinoic acid (RA) and BMP4)to promote co-differentiation into multiple cardiac-specific lineages. Assessment of the spatial distribution and percentile of the different cell types will enable exploration of how physical confinement interplays with biochemical factors for effective co-differentiation into spatial- organized cardiac organoids with controlled multicellular composition. Expectations are that small pattern geometry will favor the differentiation of stromal cell populations (e.g. CFs, EDs, EPs), while larger pattern geometry will favor the differentiation of muscle cell populations (e.g. CMs, SMCs). The THIRD Objective is to investigate the molecular mechanisms of spatial organization of cardiac organoid associated with the signaling pathways of mechanotransduction and cardiac development. Studies are designed to test the hypothesis that physical confinement will enhance RhoA/ROCK activity, which regulates the nucleocytoplamic shuttling of YAP/TAZ, and nuclear retention of YAP/TAZ in the cells at the pattern perimeter will inhibit endogenous WNT signaling during mesoderm induction, which eventually leads to the spatial differentiation on the micropatterned hiPSC colonies. In summary, research outcomes are expected to: 1) create of an in vitro cardiac organoid model with mechano-geometrical inputs; 2) provide knowledge on the effects of biophysical cues on controlling cardiac cell specification; and 3) provide mechanistic insights into developmental mechanobiology relevant to cardiac tissue morphogenesis.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.

人类诱导多能干细胞(HiPSCs)是一种能够成为任何细胞类型的细胞，它为研究人类特有的组织和器官发育提供了前所未有的机会。通过将HiPSCs分化为器官特有的细胞类型，被称为类器官的微型/微型组织可以被培养成具有感兴趣器官的许多特征。这些以干细胞为基础的有机化合物被假设为以类似于胚胎中实际器官的方式发育。然而，当前3D有机体技术的挑战之一是缺乏空间物理控制来促进这些自组装有机体中的空间组织图案，这些自组装有机体通常看起来具有随机的细胞分布。这一职业项目旨在控制HiPSC在几何限制下的生长和分化，以产生具有空间不同组织结构的跳动心脏器官。这个心脏类器官系统可以提供关于物理线索如何调节干细胞类器官的结构、功能和细胞属性的见解。该项目旨在将教育使命与研究计划相结合，以增加不同水平的学生参与干细胞生物学和微系统工程交界处的跨学科研究的机会。此外，该项目致力于将社区推广到纽约中心地区周围地区，并计划开发并向锡拉丘兹米尔顿·J·鲁宾斯坦科学与技术博物馆(MOST)捐赠一个名为“干细胞的超能力”的展览。研究人员的长期研究目标是从机制上深入了解生物物理因素在人类心脏发育和疾病中的作用。为了实现这一目标，这个职业项目的目标是设计一个具有机械几何输入的心脏器官系统，为理解生物物理线索，特别是物理限制，如何影响器官形成、组织功能和空间细胞分化奠定基础。研究计划是按照三个目标组织的。第一个目标是研究生物物理线索对调节心脏器官的结构形态和收缩功能的影响。为了创造有机类化合物，HiPSCs将被种植到聚乙二醇图案的底物上，在单个图案上扩展到接近融合，然后分化为心脏谱系。为了研究物理限制如何影响心脏类器官的形成和功能，将hPSCs种植成不同直径的圆形图案以强调大小差异，并将三角形、正方形和矩形图案与圆具有相同的几何面积以强调形状差异。这些有机化合物将根据其收缩功能(收缩运动、动作电位、钙转运和心率)和结构形态(健全性、高度和宽度)进行表征。这些特征将被用来建立器官结构和心脏功能之间的关系，并研究结构-功能关系将如何随着图案几何形状(大小和形状)的变化而改变。客观的结果有望确定最佳的物理限制条件，以产生形态上具有最高一致性的心脏器官。第二个目标是研究生物物理线索对心脏类器官细胞组成的影响，特别是心肌细胞(CMS)、心脏成纤维细胞(CFs)、心内膜细胞(EDCs)、平滑肌细胞(SMC)和心外膜细胞(EPC)。心脏类器官将在不同的模式几何形状下通过附加的分化方案与不同的细胞因子(如血管内皮生长因子1、维甲酸(RA)和骨形态发生蛋白4)进行分化，以促进共分化为多个心脏特异的谱系。对不同细胞类型的空间分布和百分位数的评估将有助于探索物理限制如何与生化因素相互作用，以有效地共分化为具有受控多细胞组成的空间组织的心脏器官。预期较小的图案几何形状将有利于基质细胞群(例如CFS、EDs、EPs)的分化，而较大的图案几何形状将有利于肌肉细胞群(例如CMS、SMC)的分化。第三个目标是研究与机械转导和心脏发育的信号通路相关的心脏器官空间组织的分子机制。研究旨在验证这一假说，即物理限制将增强RhoA/ROCK的活性，从而调节YAP/TAZ的核质粒穿梭，而YAP/TAZ在图案周长的细胞内的核保留将抑制中胚层诱导过程中的内源性WNT信号，最终导致微图案化的HiPSC克隆的空间分化。总而言之，研究成果有望：1)创建具有机械几何输入的体外心脏器官模型；2)提供关于生物物理线索对控制心脏细胞规格的影响的知识；以及3)提供与心脏组织形态发生相关的发育机械生物学的机械见解。该奖项反映了NSF的法定使命，并通过使用基金会的智力优势和更广泛的影响审查标准进行评估，被认为值得支持。

项目成果

Progressive Myofibril Reorganization of Human Cardiomyocytes on a Dynamic Nanotopographic Substrate

动态纳米拓扑基质上人心肌细胞的渐进性肌原纤维重组

• DOI: 10.1021/acsami.0c03464
• 发表时间: 2020
• 期刊: ACS Applied Materials & Interfaces
• 影响因子: 9.5
• 作者: Sun, Shiyang;Shi, Huaiyu;Moore, Sarah;Wang, Chenyan;Ash-Shakoor, Ariel;Mather, Patrick T.;Henderson, James H.;Ma, Zhen
• 通讯作者: Ma, Zhen

Organoid intelligence: Integration of organoid technology and artificial intelligence in the new era of in vitro models

类器官智能：类器官技术与人工智能在体外模型新时代的融合

• DOI: 10.1016/j.medntd.2023.100276
• 发表时间: 2024
• 期刊: Medicine in Novel Technology and Devices
• 作者: Shi, Huaiyu;Kowalczewski, Andrew;Vu, Danny;Liu, Xiyuan;Salekin, Asif;Yang, Huaxiao;Ma, Zhen
• 通讯作者: Ma, Zhen

Stimuli-responsive biomaterials for cardiac tissue engineering and dynamic mechanobiology.

• DOI: 10.1063/5.0025378
• 发表时间: 2021-03
• 期刊: APL bioengineering
• 影响因子: 6
• 作者: Shi H;Wang C;Ma Z
• 通讯作者: Ma Z

Micro-engineered architected metamaterials for cell and tissue engineering

用于细胞和组织工程的微工程超材料

• DOI: 10.1016/j.mtadv.2022.100206
• 发表时间: 2022
• 期刊: Materials Today Advances
• 影响因子: 10
• 作者: Wang, Chenyan;Vangelatos, Zacharias;Grigoropoulos, Costas P.;Ma, Zhen
• 通讯作者: Ma, Zhen