Endothelial hiPSC-Cardiomyocyte Interactions Relevant to Model Disease and Aging

描述（由申请人提供）：原则上，来源于hESC和hiPSC的分化心肌细胞是研究疾病和衰老的生理效应的理想模型系统。然而，hESC/hiPSC衍生的心肌细胞特征性地表现出早期胎儿细胞而不是成体细胞的电和机械特性。尽管有报道称一些生长因子样分子具有促成熟作用，但关于其确切的作用机制却知之甚少。 生理作用或分子如何可用于改善hiPSC/hESC衍生的心肌细胞对衰老和疾病建模的效用。这项R21提案的目标是根据内皮细胞通常产生成熟诱导因子的初步数据来鉴定和表征这些分子。数据显示的高通量评估电生理成熟的仪器和软件的发展。使用这种技术，我们提供了初步的证据表明，内皮细胞（EC）通过产生在条件培养基中积累的因子，促进hiPSC/ESC衍生的心肌细胞的电和离子通道特征成熟。这项建议的具体目标是：1）通过离子通道和泵的功能表达、药物敏感性分析、动作电位和钙处理度量来定义EC诱导的成熟，2）确定EC是否也促进收缩器官结构和功能的成熟，和3）使用来自目的1和2的电和机械度量作为定量度量来鉴定和表征EC衍生的成熟因子的作用。促进心肌细胞成熟的信号的识别应该导致用于获得功能性心肌细胞的改进方法，这将使衰老和疾病的研究成为可能。除了R21项目的高风险/高产量范围之外，我们还建立了合作，以评估成熟因子是否确实增强了hiPSC-心肌细胞作为药物风险评估和疾病模型的效用。

DESCRIPTION (provided by applicant): In principle, differentiated cardiomyocytes derived from hESCs and hiPSCs are ideal model systems in which to study the physiological effects of disease and aging. However, hESC/hiPSC-derived cardiomyocytes characteristically exhibit electrical and mechanical properties of early fetal, rather than adult, cells. Although a few growt factor-like molecules are reported to have a maturational effect, little is known about the precise physiological effects or how the molecules can be used to improve the utility of hiPSC/hESC-derived cardiomyocytes to model aging and disease. The goal of this R21 proposal is to identify and characterize such molecules, based on preliminary data that endothelial cells normally produce maturation-inducing factors. Data are presented showing the development of instrumentation and software for high-throughput assessment of electrophysiological maturation. Using this technology, we provide preliminary evidence that endothelial cells (ECs) promote electrical and ion channel profile maturation of hiPSC/ESC-derived cardiomyocytes through the production of factors that accumulate in conditioned media. The specific aims of this proposal are to: 1) define EC-induced maturation by functional expression of ion channels and pumps, drug sensitivity profiling, action potential and calcium handling metrics, 2) determine if ECs also promote maturation of contractile apparatus structure and function, and 3) use the electrical and mechanical metrics from Aims 1 and 2 as quantitative metrics to identify and characterize the effects of EC-derived maturation factor(s). The identification of signals that promote cardiomyocyte maturation should lead to improved methods for deriving functional cardiomyocytes that will be enabling for studies of aging and disease. Beyond the high- risk/high-yield scope of the R21 project, we have established collaborations to evaluate whether the maturational factors indeed enhance the utility of hiPSC-cardiomyocytes as models for drug risk assessment and disease.