Nanowired human cardiac spheroids for heart repair

项目概述：心血管疾病是全球死亡和残疾的主要原因。由于 成人心脏再生能力有限，人类诱导多能干细胞（hiPSC）已经接受了 由于其已被证明具有衍生功能性心肌细胞（hiPSC-CM）的能力而受到显著关注。尽管 尽管有进展，但目前无法有效地将hiPSC-CM递送并整合到受损心肌中， 限制了hiPSC技术在心脏修复中的应用。目前的策略受到以下限制：1）低细胞 细胞移植后的保留和存活，以及2）电流的非特异性和不成熟表型 hiPSC-CM。为了应对这些挑战，我们率先使用导电硅纳米线 在一些实施方案中，将hiPSC-CM与e-SiNW结合以促进hiPSC-CM的自组装以形成交联的hiPSC心脏球状体。添加 发现e-SiNWs增强了球状体中的导电性并改善了它们的功能。此外，本发明还提供了一种方法， 我们最近的出版物显示电刺激与e-SiNW协同作用以促进心室谱系 特化和细胞成熟（即，心室成熟），并减少 hiPSC心脏球状体体外培养。此外，我们的体内研究表明， 移植后改善细胞保留和与宿主心肌的植入，可能是由于它们的3D 微组织配置和e-SiNW增强的电集成。我们的长期目标是翻译 将hiPSC心脏球体技术应用于心脏修复的临床治疗。这个目标 建议是1）研究e-SiNW的本征（电学和表面）性质和非本征性质的影响。 因素（电刺激）对心室成熟的毛球体，和2）检查的影响 心室成熟度的测定。这个问题的核心假设是 建议的是，被染色的hiPSC心脏球体提供了一个强大的平台，以1）加速心室 2）改善hiPSC-CM的保留、植入和整合， 体内宿主心肌。该提案具有创新性，因为我们首次制备了hiPSC心脏 用于移植的具有确定心室成熟度的微组织。因此，我们将采取三项具体措施， 目的：1）阐明e-SiNW与hiPSC-CM的相互作用机制; 2）进一步研究e-SiNW与hiPSC-CM的相互作用机制 通过长期电刺激使成熟的hiPSC心脏球体心室成熟，以及3） 检查健康（Aim 3a）和损伤（Aim 3b）两者中的去毛的hiPSC心脏球状体的体内功效。 3b）大鼠心脏。这项研究的完成将使我们第一次能够生产出 具有受控心室成熟度的hiPSC-CM，2）开发一组定量标准来评估心室成熟度， 用于移植的hiPSC-CM的心室成熟度的合适范围，以及3）鉴定hiPSC-CM的心室成熟度的合适范围。 CMs用于移植。此外，它将奠定坚实的基础，建立使用电刺激， 将hiPSC心脏球体作为一种创新平台，用于提供hiPSC-CM治疗心力衰竭。

Project Summary: Cardiovascular disease is the leading cause of death and disability worldwide. Due to the limited regenerative capacity of adult hearts, human induced pluripotent stem cells (hiPSCs) have received significant attention due to their proven ability to derive functional cardiomyocytes (hiPSC-CMs). Despite the progress, the current inability to effectively deliver and integrate hiPSC-CMs into damaged myocardium has limited the applications of hiPSC technology in cardiac repair. The current strategies are limited by 1) low cell retention and survival after cell transplantation, and 2) unspecific and immature phenotype of the current hiPSC-CMs. To address these challenges, we pioneered the use of electrically conductive silicon nanowires (e-SiNWs) to facilitate self-assembly of hiPSC-CMs to form nanowired hiPSC cardiac spheroids. The addition of e-SiNWs was found to enhance electrical conduction in the spheroids and improve their function. In addition, our recent publication showed electrical stimulation synergizes with e-SiNWs to promote ventricular lineage specification and cellular maturation (i.e., ventricular maturation) and reduce the spontaneous beating of the hiPSC cardiac spheroids in in vitro culture. Further, our in vivo studies showed the nanowired spheroids improve cell retention and engraftment with host myocardium after transplantation, presumably due to their 3D microtissue configuration and the e-SiNW enhanced electrical integration. Our long-term goal is to translate nanowired hiPSC cardiac spheroid technology into a clinical therapy for heart repair. The goal of this proposal is to 1) study the effects of the intrinsic (electrical and surface) properties of e-SiNWs and extrinsic factor (electrical stimulation) on ventricular maturation of the nanowired spheroids, and 2) examine the effects of ventricular maturity of the nanowired spheroids on their engraftment. The central hypothesis of this proposal is the nanowired hiPSC cardiac spheroids provide a powerful platform to 1) accelerate ventricular maturation of hiPSC-CMs in vitro and 2) improve the retention, engraftment and integration of hiPSC-CMs with host myocardium in vivo. The proposal is innovative in that, for the first time, we prepare hiPSC cardiac microtissues with defined ventricular maturity for transplantation. Accordingly, we will pursue three specific aims: 1) Elucidate the mechanisms of the interactions between e-SiNWs and hiPSC-CMs, 2) Further advance ventricular maturation of nanowired hiPSC cardiac spheroids through long-term electrical stimulations, and 3) Examine in vivo efficacy of the nanowired hiPSC cardiac spheroids in both healthy (Aim 3a) and injured (Aim 3b) rat hearts. The completion of the proposed research would, for the first time, allow us to 1) produce hiPSC-CMs with controlled ventricular maturity, 2) develop a set of quantitative criteria to assess ventricular maturity of hiPSC-CMs for transplantation, and 3) identify a suitable range of ventricular maturity of hiPSC- CMs for transplantation. Also, it will lay down a solid foundation to establish the use of electrically stimulated, nanowired hiPSC cardiac spheroids as an innovative platform to deliver hiPSC-CMs to treat heart failure.