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.

拟议的研究旨在了解培养的心肌细胞的生物力学和电特性之间的联系及其对自主电活动的作用。心肌细胞（电活性心肌细胞）动力学的高度复杂性需要整合多种方法和技术，以揭示培养物中发生的多尺度变化以及对细胞和多细胞水平上电活动的功能影响。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，概述如下，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。开发一种组合方法，用于在我们的生物反应器中进行培养以及通过电和机械刺激进行数据采集，用于非终端实验和电生理特性的亚细胞研究*我们开发了一种用于 CM 培养的生物反应器，可为细胞提供程序化的电和机械刺激。与此同时，还开发了一种记录荧光变化（例如，fluo-4 的细胞内钙瞬变）的采集系统，以可视化培养后细胞的电刺激和机械变形的影响。现阶段所提出的组合系统存在三个重要的局限性：拉伸时所研究的视场（FOV）的位移、线性步进电机控制引起的抖动以及拉伸时无法获取荧光数据。我们建议通过集成反馈控制运动来稳定视场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位点记录时呈现出不同的时间依赖性行为。急性电或机械刺激后，时间活动可能会受到短暂或永久的影响。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时空活动的稳定性。更准确地说，我们将研究电耦合 CM 上的自组织活动及其稳定性受到急性拉伸的干扰，以了解细胞变形的作用。*3。使用数学模型评估自主电活动固有频率的异质色散对图案化和无图案单层的全局活动的作用*使用数学模型评估自主电活动固有频率的异质色散对电自体的作用-组织。从新生儿心脏分离的 CM 可以是自主细胞或非自主细胞。在细胞培养中，初始接种是随机的，因此这两个群体在单层内的分布方式未知。我们建议基于一种新颖的数学建模方法来研究这两个群体的混合对时空活动的影响，并研究细胞拓扑如何影响自主活动。 CM 动力学的高度复杂性需要整合这些创新方法和技术，以揭示培养物中发生的多尺度变化以及对细胞和多细胞水平电活动的功能影响。