Experimental tools and mathematical models to study electrical-mechanical properties of spatial-temporal patterns in cultured cardiac cells

研究培养心肌细胞时空模式的电机械特性的实验工具和数学模型

• 批准号: RGPIN-2014-04233
• 负责人: Comtois, Philippe
• 金额: $ 1.82万
• 依托单位: Université de Montréal
• 依托单位国家: 加拿大
• 项目类别: Discovery Grants Program - Individual
• 财政年份: 2018
• 资助国家: 加拿大
• 起止时间: 2018-01-01 至 2019-12-31
• 项目状态: 已结题
• 来源: https://www.nserc-crsng.gc.ca/ase-oro/Details-Detailles_eng.asp?id=647974
• 关键词: Experimental; tools; mathematical; models; study

The proposed study is aimed at understanding the link between the biomechanical and electrical properties of cultured cardiac cells and their role on autonomous electrical activity. The high level of complexity of cardiomyocyte (electrically-active cardiac cells) dynamics needs integration of numerous approaches and techniques to uncover the multiscale changes occurring in culture and the functional impact on electrical activity at both the cell and multicellular levels. Here, we proposed a project involving bioinstrumentation development, image acquisition techniques as well as analysis, and modelling works summarized below to better understand the effects of cell deformation on bioelectric characteristics and spatio-temporal electrical self-organization.*1. Develop a combined approach for culture in our bioreactor and data acquisition with electrical and mechanical stimulations for non-terminal experiments and sub-cellular study of electrophysiological properties *We have developed a bioreactor for culture of CMs that provided programmed electrical and mechanical stimuli to cells. In parallel, an acquisition system to record fluorescence changes (for example for intracellular calcium transient with fluo-4) to visualize the effects of electrical stimulation and mechanical deformation of the cells in post-culture has been developed. Three important limitations of the proposed combined systems exist at this stage: displacement of the field of view (FOV) under study when stretched, jitter due to linear stepper motors control, and impossibility to acquire fluorescence data while stretching. We propose to correct these limitations by integrating a feedback control motion to stabilize the FOV and to modify the bioreactor stretching electronic circuit to microstepping.*2. Study of the spatial-temporal autonomous electrical activity of isotropic and patterned cardiomyocyte monolayers following culture on elastic substrates:*Experiments by our group present varying time-dependent behaviors when recording at a single site. The temporal activity can be transiently or permanently affected following acute electrical or mechanical stimulation. However, it is clear that the changes seen locally are limited to explain what could be variation in the spatial-temporal dynamics. We thus propose to study the stability of spatio-temporal activity of topographically patterned cardiomyocytes. More precisely we will study self-organized activity on electrically-coupled CMs and its stability is perturbed by acute stretch to understand the role of cell deformation.*3. Evaluate, with a mathematical model, the role of heterogeneous dispersion of intrinsic frequencies of autonomous electrical activity on global activity of patterned and unpatterned monolayers*Evaluate, with a mathematical model, the role of heterogeneous dispersion of intrinsic frequencies of autonomous electrical activity on electrical self-organization. CMs isolated from neonatal hearts can be either autonomous or non-autonomous cells. In cell culture, initial seeding is random such that how these two populations are distributed within the monolayer is unknown. We propose to look at the effects of having mixture of these two populations on spatio-temporal activity based on a novel approach of mathematical modeling and study how cell topography can influence the autonomous activity. The high level of complexity of CM dynamics needs integration of these innovative approaches and techniques to uncover the multiscale changes occurring in culture and the functional impact on electrical activity at both the cell and multicellular levels.

这项研究旨在了解培养的心脏细胞的生物力学和电学特性及其对自主电活动的作用之间的联系。心肌细胞（电活性心脏细胞）动力学的高度复杂性需要整合多种方法和技术，以揭示培养中发生的多尺度变化以及细胞和多细胞水平上对电活动的功能影响。在这里，我们提出了一个涉及生物仪器开发，图像采集技术以及分析和建模工作的项目，总结如下，以更好地了解细胞变形对生物电特性和时空电自组织的影响。1.开发一种在我们的生物反应器中进行培养的组合方法，并通过电刺激和机械刺激进行数据采集，用于非终末实验和电生理特性的亚细胞研究 * 我们开发了一种用于培养CM的生物反应器，为细胞提供程序化的电刺激和机械刺激。同时，已经开发了一种采集系统，用于记录荧光变化（例如，用fluo-4记录细胞内钙瞬变），以可视化培养后细胞的电刺激和机械变形的影响。在这个阶段，所提出的组合系统存在三个重要的限制：拉伸时研究的视场（FOV）的位移，由于线性步进电机控制的抖动，以及拉伸时不可能获取荧光数据。我们建议通过集成反馈控制运动来稳定FOV并将生物反应器拉伸电子电路修改为微步进来纠正这些限制。2.研究在弹性基质上培养后各向同性和图案化心肌细胞单层的时空自主电活动：* 我们小组的实验在单个部位记录时呈现不同的时间依赖性行为。在急性电刺激或机械刺激后，时间活动可以暂时或永久地受到影响。然而，很明显，局部观察到的变化仅限于解释时空动态的变化。因此，我们建议研究拓扑图案心肌细胞的时空活动的稳定性。更确切地说，我们将研究电耦合CM上的自组织活动，以及它的稳定性受到急性拉伸的干扰，以了解细胞变形的作用。3.用数学模型评估自主电活动的固有频率的非均匀分散对图案化和未图案化单层的全局活动的作用 * 用数学模型评估自主电活动的固有频率的非均匀分散对电自组织的作用。从新生儿心脏分离的CM可以是自主或非自主细胞。在细胞培养中，初始接种是随机的，使得这两个群体如何在单层内分布是未知的。我们建议看看这两个群体的混合物对时空活动的影响，基于一种新的方法的数学建模和研究细胞拓扑结构如何影响自主活动。CM动力学的高度复杂性需要这些创新方法和技术的整合，以揭示培养中发生的多尺度变化以及细胞和多细胞水平上对电活动的功能影响。