Optically Promoting Cardiac Maturation Using Engineered Peptides

项目总结 人类干细胞来源的心肌细胞(HSC-CMS)的前景为可行性打开了大门 用于治疗心脏病的个性化药物和进行更准确的药物发现研究。 此外，HSC-CMS克服了使用动物模型进行高通量处理时物种差异的问题 筛查研究。然而，扩大使用HSC-CMS的瓶颈之一是它们能够准确地 反映成人心肌细胞的天然结构和功能。目前为解决这一关键问题所做的努力 挑战涉及使用生物物理线索的成熟方案，如电刺激或底物- 诱导组织排列。电刺激的方法通常利用电极触点进行场刺激， 用于将电刺激传递与机械或持续化学刺激相结合的笨重仪器， 或者对细胞进行基因改造，使其对光有反应。尽管我们通过这些归纳已经看到了成功 和刺激方法，该领域将受益于一种文化接触最少的刺激方法 在长期培养过程中降低感染风险，以及基于光线的方法具有更高的时空 分辨率优于基于电极的刺激。在这里，我们提出了一种新的范式来刺激HSC-CMS 通过将这些细胞与基于多肽的底物相结合来成熟，这种底物可以诱导组织各向异性和 被设计成将光转化为刺激信号。我们的团队将开发具有功能化的多肽 生色团和细胞结合表位作为可用于光刺激HSC的材料 CMS，结合组织排列的诱导，走向成熟。这个项目的长期目标是 通过工程多肽建立光刺激作为一种可行的方法来刺激心肌细胞和 以无电极、非遗传方式促进HSC-CM成熟。我们假设那是短暂的 心肌细胞-生物材料界面的充电和其他相关的光诱导过程可以影响 胞外电位，导致HSC-CMS走向成熟的光刺激。我们的理由是 提出了一种基于材料的方法来刺激HSC-CMS，这源于先前关于共轭的 聚合物用作触发其他可兴奋细胞的动作电位的光活性底物。为了测试 根据我们的假设，我们提出了以下具体目标：(1)建立工程设计参数 具有最佳光刺激效率的多肽基质；(2)测试细胞和组织水平的影响 与各向异性线索相结合的多肽介导的光刺激；以及(3)阐明 提出的光刺激方法，以及各向异性线索，对HSC-CM的成熟。通过建立 用于诱导组合线索刺激HSC-CMS的光可激肽的设计规则 确保它们有能力兴奋心肌细胞，这一创新的方法提供了一种新的战略，为 刺激心脏组织走向成熟，因此可显著有助于解决 干细胞来源的心肌细胞未成熟的巨大挑战。

PROJECT SUMMARY The promise of human stem cell-derived cardiomyocytes (hSC-CMs) opens doors towards the feasibility of personalized medicine against cardiac diseases and for performing more accurate drug discovery studies. Moreover, hSC-CMs overcome the issue of species differences when using animal models for high throughput screening studies. However, one of the bottlenecks for scaling up the use of hSC-CMs is their ability to accurately reflect the native structure and function of adult human cardiomyocytes. Current efforts to address this critical challenge involve maturation protocols that use biophysical cues such as electrical stimulation or substrate- induced tissue alignment. Methods for electrical stimulation often utilize electrode contacts for field stimulation, bulky instrumentation for combining electrical stimuli delivery with mechanical or sustained chemical stimulation, or genetically modifying cells to be light-responsive. Although we have seen successes through these induction and stimulation approaches, the field would benefit from a stimulation approach with minimal culture contact to reduce risk of infection during long-term cultures, as well as a light-based approach with higher spatiotemporal resolution than electrode-based stimulation. Here, we propose a new paradigm for stimulating hSC-CMs towards maturation by interfacing these cells with peptide-based substrates that can induce tissue anisotropy and are engineered to convert light to stimulatory cues. Our team will develop peptides functionalized with chromophore units and cell-binding epitopes as materials that can be used for light-based stimulation of hSC- CMs, in combination with induction of tissue alignment, towards maturation. The long-term goal of this project is to establish light stimulation via engineered peptides as a viable method to stimulate cardiomyocytes and promote hSC-CM maturation in an electrodeless and non-genetic manner. We hypothesize that transient charging and other associated light-induced processes at the cardiomyocyte-biomaterial interface can influence extracellular potential, resulting in the photostimulation of hSC-CMs towards maturation. Our rationale for proposing a materials-based approach for stimulating hSC-CMs stems from previous reports of conjugated polymers being used as a photoactive substrate for triggering action potentials of other excitable cells. To test our hypothesis, we propose the following specific aims: (1) establishing design parameters for engineered peptide substrates with optimal photostimulation efficiency; (2) test the cellular- and tissue-level impact of peptide-mediated photostimulation in combination with anisotropic cues; and (3) elucidate the effect of the proposed photostimulation method, along with anisotropy cues, on hSC-CM maturation. By establishing the design rules for the proposed photoexcitable peptides for eliciting combinatorial cues to stimulate hSC-CMs and ensure their capability to excite cardiac cells, this innovative approach offers a new strategy for a “wireless” stimulation of cardiac tissues towards maturation, and can therefore significantly contribute towards addressing the grand challenge of immaturity of stem cell-derived cardiomyocytes.