Dou Wenkun, Wang Li, Malhi Manpreet, Liu Haijiao, Zhao Qili, Plakhotnik Julia, Xu Zhensong, Huang Zongjie, Simmons Craig A, Maynes Jason T, Sun Yu
Department of Mechanical and Industrial Engineering, University of Toronto, Toronto, M5S 3G8, Canada.
Department of Mechanical and Industrial Engineering, University of Toronto, Toronto, M5S 3G8, Canada; School of Mechanical & Automotive Engineering, Qilu University of Technology (Shandong Academy of Sciences), Jinan, 250353, China.
Biosens Bioelectron. 2021 Mar 1;175:112875. doi: 10.1016/j.bios.2020.112875. Epub 2020 Dec 3.
The use of human induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs) as an in vitro model of the heart is limited by their structurally and functionally immature phenotypes. During heart development, mechanical stimuli from in vivo microenvironments are known to regulate cardiomyocyte gene expression and maturation. Accordingly, protocols for culturing iPSC-CMs have recently incorporated mechanical or electromechanical stimulation to induce cellular maturation in vitro; however, the response of iPSC-CMs to different mechanical strain magnitudes is unknown, and existing techniques lack the capability to dynamically measure changes to iPSC-CM contractility in situ as maturation progresses. We developed a microdevice platform which applies cyclical strains of varying magnitudes (5%, 10%, 15% and 20%) to a monolayer of iPSC-CMs, coincidentally measuring contractile stress during mechanical stimulation using fluorescent nanobeads embedded in the microdevice's suspended membrane. Cyclic strain was found to induce circumferential cell alignment on the actuated membranes. In situ contractility measurements revealed that cyclic stimulation gradually increased cardiomyocyte contractility during a 10-day culture period. The contractile stress of iPSC-CM monolayers was found to increase with a higher strain magnitude and plateaued at 15% strain. Cardiomyocyte contractility positively correlated with the elongation of sarcomeres and an increased expression of β-myosin heavy chain (MYH7) in a strain magnitude-dependent manner, illustrating how mechanical stress can be optimized for the phenotypic and proteomic maturation of the cells. iPSC-CMs with improved maturity have the potential to create a more accurate heart model in vitro for applications in disease modeling and therapeutic discovery.
将人诱导多能干细胞衍生的心肌细胞（iPSC-CMs）用作心脏的体外模型受到其结构和功能不成熟表型的限制。在心脏发育过程中，已知来自体内微环境的机械刺激会调节心肌细胞基因表达和成熟。因此，培养iPSC-CMs的方案最近已纳入机械或机电刺激，以在体外诱导细胞成熟；然而，iPSC-CMs对不同机械应变幅度的反应尚不清楚，并且现有技术缺乏在成熟过程中原位动态测量iPSC-CM收缩性变化的能力。我们开发了一种微器件平台，该平台将不同幅度（5%、10%、15%和20%）的周期性应变施加到单层iPSC-CMs上，同时使用嵌入微器件悬浮膜中的荧光纳米珠在机械刺激期间测量收缩应力。发现周期性应变会在驱动膜上诱导细胞周向排列。原位收缩性测量表明，在10天的培养期内，周期性刺激逐渐增加心肌细胞的收缩性。发现iPSC-CM单层的收缩应力随着应变幅度的增加而增加，并在15%应变时达到稳定状态。心肌细胞收缩性与肌节伸长以及β-肌球蛋白重链（MYH7）表达的增加呈正相关，且呈应变幅度依赖性，这说明了如何针对细胞的表型和蛋白质组成熟优化机械应力。成熟度提高了的iPSC-CMs有潜力在体外创建一个更准确的心脏模型，用于疾病建模和治疗发现等应用。
