Nanowired human cardiac spheroids for heart repair

项目概述：心血管疾病是世界范围内导致死亡和残疾的主要原因。由于 成人心脏再生能力有限，人诱导多能干细胞(HiPSCs)已收到 由于它们被证明有能力获得功能性心肌细胞(HiPSC-CMS)，因此受到了极大的关注。尽管 进展，目前无法有效地将HiPSC-CMS运送和整合到受损心肌中 限制了HiPSC技术在心脏修复中的应用。目前的策略受到1)低单元格的限制 细胞移植后的保留和存活，以及2)当前的非特异性和未成熟的表型 HiPSC-CMS。为了应对这些挑战，我们率先使用了导电硅纳米线 (e-SiNWs)以促进hiPSC-CMS的自组装以形成纳米线的hiPSC心脏球体。加法 研究发现，e-SiNW的存在增强了球体的导电性，改善了其功能。此外, 我们最近发表的文章显示，电刺激与e-SiNW协同作用促进了心室谱系的形成 规范和细胞成熟(即，心室成熟)，并减少自发性心跳 体外培养的HiPSC心脏球体。此外，我们在体内的研究表明，纳米线球体 改善细胞在移植后与宿主心肌的保留和植入，可能是由于其3D 微组织构型和e-SiNW增强了电集成。我们的长期目标是翻译 纳米线HiPSC心脏球体技术应用于心脏修复的临床治疗。这样做的目的是 建议1)研究e-SiNW的内在(电学和表面)性质和外在性质的影响 因素(电刺激)对纳米线球体的心室成熟的影响，以及2)检测其影响 观察植入的纳米线球体的室壁成熟度。这一点的中心假设是 建议是纳米线的hiPSC心脏球体提供了一个强大的平台来加速心室 HIPSC-CMS的体外成熟；2)促进HIPSC-CMS与 活体内宿主心肌。该方案的创新之处在于，我们首次制备了HiPSC心脏 供移植的具有明确的脑室成熟度的显微组织。相应地，我们将追求三个具体的 目的：1)阐明e-SiNW与hiPSC-CMS相互作用的机制；2)进一步研究进展 通过长期电刺激的纳米线HiPSC心脏球体的心室成熟，3) 检测纳米线hiPSC心脏球体在健康(Aim 3a)和损伤(Aim)中的体内效果 3B)大鼠心脏。拟议研究的完成将第一次使我们能够生产 HIPSC-CMS与受控的室性成熟度，2)建立了一套定量标准来评估室性 HIPSC-CMS移植的成熟度；3)确定适合移植的HIPSC-CMS的成熟范围。 用于移植的细胞质雄性不育。也将为建立电刺激的使用奠定坚实的基础， 纳米线hiPSC心脏球体作为提供hiPSC-CMS治疗心力衰竭的创新平台。

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.