Integrated System for Mechanoelectrical Studies of Cardiac Myofibroblasts

描述（由申请人提供）：在不同的细胞类型中，如肌肉、成纤维细胞和骨骼，细胞可以产生收缩或牵引力，同时对电生理和机械信号做出反应。虽然这些相互作用的子集已经在机械转导和兴奋-收缩耦合的主题下进行了研究，但要充分理解细胞力学和细胞电生理之间的相互关系，需要在施加外力期间同时测量细胞力、细胞电压和细胞电流。该项目的目标是首次组装一个集成的测量和力应用系统，该系统将能够在具有不同细胞骨架结构的细胞中检查机械敏感通道，跨膜电压和特定膜电流的功能作用。即将研究的模型系统是心肌成纤维细胞，这种细胞在心肌梗死和疤痕形成中起着重要作用，能够感知力、产生力和电生理活动。近年来，微图案柔性衬底，如弹性微米级柱阵列，在细胞牵引力下偏转，已成为测量细胞收缩性的新系统，具有亚细胞分辨率。此外，通过在一些柱子中嵌入磁性纳米颗粒，现在可以对附着的细胞施加力，同时通过周围的非磁性柱子测量它们的机械反应。在这个项目中，基于磁微柱的力产生和测量系统将与电生理技术相结合，以实现对单个细胞的同时电和机械刺激和读出。该项目将由两位研究者的实验室共同努力，他们在心脏电生理学、实验物理学、细胞生长模式、磁学、微加工和细胞力学方面具有互补的专业知识。Specific Aim 1将通过微柱阵列整合单细胞牵引力测量与跨膜电位和膜离子电流的电生理测量。微接触打印将用于改变肌成纤维细胞的形状，以操纵肌动蛋白应力纤维的内部分布，并探索亚细胞力分布对跨膜电位和电流的影响。具体目标2将把机械力和应变的磁驱动集成到目标1中开发的机电读出系统中，并将研究细胞对外力和应变的机械和电生理反应的相互作用。通过创建细胞机电功能研究的统一技术，该项目将为阐明生物学和临床重要系统开辟新的方向。

DESCRIPTION (provided by applicant): In diverse cell types such as muscle, fibroblasts and bone, the cells can generate contractile or traction forces while being responsive to electrophysiological and mechanical signals. Although subsets of these interactions have been studied under the themes of mechanotransduction and excitation-contraction coupling, a full understanding of the interrelationship between cell mechanics and cell electrophysiology requires the simultaneous measurement of cell force, cell voltage and cell current during the application of external forces. The goal of this project is to assemble, for the first time, an integrated measurement and force application system that will enable the functional role of mechanosensitive channels, transmembrane voltage, and specific membrane currents to be examined in cells having different cytoskeletal architectures. The model system that will be studied is the cardiac myofibroblast, a cell that plays a prominent role in myocardial infarcts and scar formation and is capable of force sensing, force production, and electrophysiological activity. In recent years, micropatterned flexible substrates, such as arrays of elastomeric micrometer-scale posts that deflect under cellular traction forces have emerged as novel systems for measuring cellular contractility with sub-cellular resolution. Further, by embedding magnetic nanoparticles in some of the posts, it is now possible to apply forces to adherent cells while measuring their mechanical responses via the surrounding non-magnetic posts. In this project, magnetic micropost-based force generation and measurement systems will be combined with electrophysiological techniques to enable simultaneous electrical and mechanical stimulation and readout of single cells. This project will be a joint effort between the laboratories of two Investigators with complementary expertise in cardiac electrophysiology, experimental physics, patterned cell growth, magnetism, microfabrication, and cell mechanics. Specific Aim 1 will integrate single cell traction force measurements via the micropost arrays with electrophysiological measurements of transmembrane potential and membrane ionic currents. Micro contact printing will be used to vary the shape of the myofibroblasts to manipulate their internal distribution of actin stress fibers, and to explore the effect of sub-cellular force distributions on transmembrane potential and currents. Specific Aim 2 will integrate magnetic actuation of mechanical forces and strains into the electromechanical readout system developed in Aim 1, and will study the mutual interactions of cellular mechanical and electrophysiological responses to external force and strain. By creating a unified technology for the study of cellular mechanoelectrical function, this project will open new directions for the elucidation of biologically and clinically important systems. ) PUBLIC HEALTH RELEVANCE: This research will combine electrophysiology techniques with recent, advanced developments in magnetic nanotechnology to develop a new platform to study the interplay between mechanical and electrical processes in heart, muscle, and other tissues. A number of diseases have been attributed to abnormal interactions between mechanical and electrical functiontransduction of mechanical cues at the cellular level, including cardiac arrhythmias, polycystic kidney disease, glioma, glaucoma, Duchenne muscular dystrophy and tumorigenesis. This research will enable new insights into cellular mechano-electrical processes at the cellular level, and will have the potential to contribute significantly to the understanding of the progression of these diseases.