Cellular Cardiomyoplasty in Vitro: Host-Donor Cell Interactions

描述(由申请人提供)：将外源性供体细胞植入心脏(细胞心肌成形术)是治疗缺血后心室重塑的一种新兴方法。虽然自体骨骼肌成肌细胞和骨髓细胞的初步临床移植前景看好，但这些细胞影响周围心脏组织机电功能的内在潜力仍未阐明。因此，这项建议的主要目的是系统地研究不同供体细胞(即骨骼肌成肌细胞和间充质干细胞，与对照心肌成纤维细胞相比)在体外通过宿主心脏网络繁殖机电活动的能力。我们假设：1)通过供体细胞的电活动的传播取决于它们的分化类型和阶段，以及供体-供体和宿主-供体细胞对之间不仅存在电连接，而且还有机械连接的存在；2)通过特定的生长因子上调细胞间的通讯，可以改善供体细胞植入体内的机电连接。为了验证这些假设，我们建议使用几何简化的、可重复的一维设置来研究供体细胞的电导，即带有供体细胞插入件的微图案化心肌细胞链。在各种分化因子和生长因子存在的情况下，供体细胞的细胞膜电位和细胞内钙瞬变的传播将被光学映射。所获得的结果将与免疫组织化学和分子分析的结果相关联。这项提议的发现有望阐明不同供体细胞在心脏中功能整合的潜力。最终，所提出的实验框架将允许我们进行高通量的体外分析，这些因素可以改善不同供体细胞的机电繁殖。最终目的是帮助开发有效和安全的基于细胞的方法来治疗因缺血、梗死或先天性缺陷而造成的区域性心脏损伤。

DESCRIPTION (provided by applicant): Implantation of exogenous donor cells into the heart (cellular cardiomyoplasty) is an emerging methodology for the treatment of post-ischemic ventricular remodeling. While initial clinical implantations of autologous skeletal myoblasts and bone marrow cells have been promising, the intrinsic potential of these cells to affect electro-mechanical function of surrounding heart tissue has still not been elucidated. Therefore, the main goal of this proposal is to systematically study the ability of different donor cells (i.e. skeletal myoblasts and mesenchymal stem cells, as compared to control cardiac myofibroblasts) to propagate electro-mechanical activity through a host cardiac network in vitro. We hypothesize that: 1) propagation of electrical activity through donor cells depends on their type and stage of differentiation, as well as the presence of not only electrical but also mechanical junctions between the donor-donor and host-donor cell pairs, and 2) electro- mechanical propagation in the donor cell implant can be improved through upregulation of the intercellular communication by specific growth factors. To test these hypotheses we propose to study electrical conduction through donor cells using a geometrically simplified, reproducible one-dimensional setting, i.e. the micropatterned cardiomyocyte strands with inserts made of donor cells. The propagation of cell membrane potentials and intracellular calcium transients in donor cells will be optically mapped in the presence of various differentiation agents and growth factors. Obtained results will be correlated with those from immunohistochemical and molecular analyses. The findings from this proposal are expected to elucidate potential of different donor cells to functionally integrate in the heart. Eventually, the proposed experimental framework will allow us to perform high throughput in vitro analysis of the factors that can improve electromechanical propagation of different donor cells. The final aim is to aid in the development of efficient and safe cell-based approaches for the treatment of regional heart injury due to ischemia, infarction, or congenital defects.