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Electrical integration, maturation and proarrhythmic risk of induced pluripotent stem cell-derived cardiomyocytes after transplantation in infarcted mouse hearts

梗塞小鼠心脏移植后诱导多能干细胞来源的心肌细胞的电整合、成熟和致心律失常风险

基本信息

批准号：
325112617
负责人：
Professor Dr. Marcel Halbach
金额：
--
依托单位：
Klinik III für Innere Medizin: Kardiologie, Angiologie, Pneumologie und internistische Intensivmedizin
依托单位国家：
德国
项目类别：
Research Grants
财政年份：
2017
资助国家：
德国
起止时间：
2016-12-31 至 2021-12-31
项目状态：
已结题

来源：
https://gepris.dfg.de/gepris/projekt/325112617?language=en
关键词：
Electrical integration maturation proarrhythmic risk

项目摘要

State-of-the-art therapy of myocardial infarction does not result in regeneration of lost myocardium. Cell therapy using pluripotent stem cell-derived cardiomyocytes is regarded as promising strategy to overcome this hurdle. A potential arrhythmic risk is an important safety concern against cardiac cell therapy. Previous studies provided conflicting results on pro- or antiarrhythmic effects of transplantated cells of different origin, but did not include induced pluripotent stem cell-derived cardiomyocytes (iPSCM), which are regarded as one of the most promising cell types. The aim of this project is to investigate cell persistence, functional integration, maturation and arrhythmic potential of iPSCM after transplantation in infarcted mouse hearts. Murine iPSCM expressing enhanced green fluorescent protein and a puromycin resistance under control of the alpha-myosin heavy chain promoter will be injected into adult mouse hearts after induction of myocardial infarction by coronary artery ligation. iPSCM at different developmental stages will be compared regarding efficacy and safety. Another experimental group will be clusters of iPSCM and mesenchymal stem cells, since there is evidence of a higher persistence of the graft as compared with pure iPSCM. Saline will be injected in control animals. 6 days, 6 weeks and 3 months after transplantation, programmed stimulation will be applied by a right ventricular electrophysiological catheter. External ECG recordings will reveal number and duration of induced ventricular fibrillation episodes. Simultaneously, in vivo multi electrode array recordings of the left ventricle will be performed to display excitation spread. Afterwards, hearts will be resected and viable ventricular tissue slices (150 µm thick) will be prepared to enable in vitro multi electrode array measurements of field potentials as well as action potential recordings with glass microelectrodes in transplanted iPSCM and surrounding host cardiomyocytes. These experiments will provide information on electrical integration and maturation of transplanted cells. Preliminary experiments have already demonstrated the feasibility of the planned project.The expected results will reveal, if iPSCM are suitable for application in cardiac cell therapy based on their quality of electrical integration and arrhythmic potential. The four electrophysiological methods implemented in the project allow a direct correlation of single cell integration and maturation with the arrhythmic risk on tissue and organ levels, since all measurements are performed with the same preparations. This will enable a thorough evaluation of mechanisms of arrhythmogenesis, which will be an important step towards a higher efficacy and safety of cardiac cell therapy.

最先进的心肌梗死治疗方法不会导致丢失的心肌再生。使用多能干细胞来源的心肌细胞进行细胞治疗被认为是克服这一障碍的有希望的策略。一个潜在的心律失常风险是心脏细胞治疗的一个重要的安全问题。以前的研究对不同来源的移植细胞的促或抗心律失常作用提供了相互矛盾的结果，但没有包括被认为是最有前途的细胞类型之一的诱导多能干细胞来源的心肌细胞(IPSCM)。本项目的目的是研究IPSCM在小鼠心肌梗死后移植后的细胞持久性、功能整合、成熟度和心律失常潜能。在冠状动脉结扎诱导心肌梗死后，在α-肌球蛋白重链启动子的控制下，表达增强型绿色荧光蛋白和扑霉素抗性的小鼠ipscm将被注射到成年小鼠的心脏中。不同发育阶段的IPSCM将在有效性和安全性方面进行比较。另一组实验将是IPSCM和间充质干细胞集群，因为有证据表明，与纯粹的IPSCM相比，移植的持久性更高。对照组动物将注射生理盐水。移植后6天、6周和3个月，用右室电生理导管进行程序性刺激。体外心电记录将显示诱导性室颤发作的次数和持续时间。同时，进行在体左心室多电极阵列记录，以显示兴奋扩散。然后，将切除心脏，并制备可存活的心室组织切片(150微米厚)，以便能够在体外用玻璃微电极阵列测量移植的IPSCM和周围宿主心肌细胞的场电位和动作电位。这些实验将提供有关移植细胞的电学整合和成熟的信息。初步实验已经证明了计划项目的可行性。预期的结果将揭示，基于IPSCM的电整合和心律失常电位的质量，IPSCM是否适合应用于心肌细胞治疗。该项目中实施的四种电生理方法允许单细胞整合和成熟与组织和器官水平上的心律失常风险直接相关，因为所有测量都是用相同的准备进行的。这将使对心律失常发生机制的彻底评估成为迈向更高效率和更安全的心肌细胞治疗的重要一步。