Cellular Cardiomyoplasty in Vitro: Host-Donor Cell Interactions

描述（由申请人提供）：外源性供体细胞植入心脏（细胞心肌成形术）是一种新兴的治疗缺血性心室重构的方法。虽然自体骨骼肌母细胞和骨髓细胞的初步临床植入很有希望，但这些细胞影响周围心脏组织机电功能的内在潜力仍未得到阐明。因此，本提案的主要目标是系统地研究不同供体细胞（即骨骼肌母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传播机电活动的能力。我们假设：1)电活动在供体细胞中的传播取决于它们的类型和分化阶段，以及供体-供体和宿主-供体细胞对之间不仅存在电连接，还存在机械连接；2)特定生长因子可以通过上调细胞间通讯来改善供体细胞植入物中的机电传播。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传导，即微图案心肌细胞链与供体细胞制成的插入物。在各种分化剂和生长因子的存在下，供细胞的细胞膜电位和细胞内钙瞬态的繁殖将被光学绘制。所得结果将与免疫组织化学和分子分析结果相关联。这一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局部心脏损伤，由于缺血，梗死，或先天性缺陷。

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.