Myocardial Repair with a Novel Engineered Cardiac Muscle Patch

项目摘要 细胞移植是一种很有前途的心肌修复治疗方法。然而， 仍有几个关键问题需要解决，包括但不限于：1)供体细胞植入率低 税率(在以前的出版物中从0.1%到10%不等)；2)缺乏对基础机制的了解 移植细胞的活体受益效应。了解移植细胞的体内效应可能会促进 制定更有效的心脏保护策略。 几个小组报告说，应用预制的心脏组织，一种由 HiPSC来源的心肌细胞，有效地提高了植入率。我们最近制定了一项新的战略 这已被证明显著提高了植入率。具体来说，我们建立了一个人类 携带编码人CCND2(Cyclin D2)转基因的诱导多能干细胞(HiPSC)系 由心肌细胞特异性α-肌球蛋白重链(α-MHC)启动子驱动。CCND2-高表达HiPSC- 来源的心肌细胞(HiPSC-CCND2OECMs)表现出细胞周期活性和细胞增殖的增强 与基因单纯表达野生型CCND2的hiPSC-CMS(hiPSC-CCND2WTCms)相比。在……里面 在小鼠心肌梗死(MI)模型上，移植细胞的数量增加了两倍 HIPSC-CCND2OECMS与移植后4周接受HIPSC-CCND2WTCMS的患者相比， 从而显著缩小梗塞面积并改善心功能。这些数据表明，转基因 CCND2在HiPSC-CM移植物中的过表达是增强植入和修复的可行途径 在缺血性心脏病中的作用。 该计划将开发一种新型的含有hPSC-CCND2OECMs的人类心肌贴片(命名为hCMP- CCND2OECMs)，评估其长期持续使受伤心脏肌肉化的能力 最终替换疤痕组织，并在大型动物模型中测试其安全性和翻译潜力( 猪MI模型)。在移植了hCMPS的情况下，改善了慢性脑梗塞猪心脏的心功能， 该提案还确定这种功能改善是否归因于肌肉化或其他 机械装置。具体地说，我们的目标是解决尚未得到回答，但具有深远意义的问题，即hCMPS 通过供体-宿主直接偶联改善猪MI模型的心功能。我们的长期目标是培育一颗心脏 可以转化为人类的再生策略。提出了两个具体目标。 特定目的1将检验这种新的hCMP持续使受损心肌重新肌化的假设， 替代跨壁疤痕组织，改善梗死猪心脏的心功能。我们将决定(I) 补片移植对心脏结构和功能的影响以及(Ii)补片移植的安全性 (易诱发心律失常和肿瘤形成的风险)。 特定目标2检验功能改变的大小与数量正相关的假设 供体细胞来源的CMS。我们将确定(I)是否需要供体细胞存活才能持续改善 以及(Ii)心功能的改善是否至少部分地通过 供体心肌细胞移植后的机电耦合。

Project Summary Cellular transplantation has emerged as a promising therapeutic approach for myocardial repair. However, several critical issues remain to be addressed which include, but are not limited to: 1) low donor cell engraftment rate (ranging from 0.1-10% in previous publications); 2) lack of knowledge on the mechanisms underlying the in vivo beneficiary effects of grafted cells. Understanding the in vivo effects of grafted cells may promote the development of more effective cardioprotective strategies. Several groups reported that applying prefabricated cardiac tissue, a "cardiac muscle patch" (CMP) made of hiPSC-derived cardiac cells, effectively increased engraftment rate. We recently established a novel strategy which has been demonstrated to significantly enhance engraftment rate. Specifically, we established a human induced pluripotent stem cell (hiPSC) line which carries a transgene encoding for the human CCND2 (Cyclin D2) driven by the cardiomyocyte specific α-myosin heavy chain (α-MHC) promoter. CCND2-overexpressing hiPSC- derived cardiomyocytes (hiPSC-CCND2OECMs) exhibits increased cell cycle activity and cell proliferation compared with genetically naïve hiPSC-CMs expressing wild-type levels of CCND2 (hiPSC-CCND2WTCMs). In a mouse model of myocardial infarction (MI), the number of engrafted cells was tripled in hearts injected with hiPSC-CCND2OECMs compared to those receiving hiPSC-CCND2WTCMs 4 weeks post MI and transplantation, resulting in significantly smaller infarct size and improved cardiac function. These data suggests that transgenic CCND2 overexpression in hiPSC-CM grafts constitutes a viable approach to enhance engraftment and restore function in ischemic heart disease. The proposal will develop a novel human cardiac muscle patch with hiPSC-CCND2OECMs (designated as hCMP- CCND2OECMs), assess their capability to continuously remuscularize the injured heart in the long term and ultimately replace the scar tissue, and test their safety and translational potential in a large animal model (the pig MI model). In the case transplanted hCMPs improve cardiac function in the chronically infarcted pig hearts, the proposal also determine if this functional improvement is attributable to remuscularization or other mechanisms. Specifically, we aimed to address the yet unanswered, but profoundly important, question whether hCMPs improve cardiac function via direct donor-host coupling in pig MI model. Our long term goal is to develop a heart regeneration strategy that can be translated to humans. Two Specific Aims are proposed. Specific Aim 1 will test the hypothesis that this novel hCMP continuously remuscularize injured myocardium, replacing transmural scar tissue, and improve cardiac function in infarcted pig hearts. We will determine (i) the impact of patch transplantation on cardiac structure and function, and (ii) safety of patch transplantation (susceptibility to inducible arrhythmias, and risk of tumor formation). Specific Aim 2 test the hypothesis that the magnitude of functional changes positively correlates with the number of donor cell-derived CMs. We will determine whether (i) donor cell survival is required for sustained improvement of cardiac function, and (ii) whether improvement in cardiac function is mediated, at least in part, by electromechanical coupling of transplanted donor myocytes.