Maturation of Normal and Disease-specific Human Stem Cell-derived Cardiomyocytes

正常和疾病特异性人类干细胞来源的心肌细胞的成熟

基本信息

批准号：
8118810
负责人：
HUEI-SHENG Vincent CHEN
金额：
$ 47.75万
依托单位：
SANFORD BURNHAM PREBYS MEDICAL DISCOVERY INSTITUTE
依托单位国家：
美国
项目类别：
财政年份：
2010
资助国家：
美国
起止时间：
2010-07-01 至 2015-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8118810
关键词：
Aborted Fetus Accounting Adult Affect Age Aging Algorithms Animal Model Arrhythmogenic Right Ventricular Dysplasia Back Bioinformatics Biological Models Cardiac Cardiac Myocytes Cardiovascular Diseases Cause of Death Cell Line Cell Proliferation Cell Therapy Cell Transplantation Cells Characteristics Chromosome Mapping Clinical Clinical Pathology Coculture Techniques Cues Data Analyses Data Set Dermal Desmosomes Development Developmental Gene Disease Disease model Elderly Electrodes Embryo Embryonic Heart Endothelial Cells Endothelin A Receptor Endothelin-1 Engineering Environment Fetal Heart Fibroblasts Foundations Future Genes Genetic Genetic Transcription Germ Layers Goals Grouping Heart Heart Diseases Hereditary Disease Human Implant In Vitro Investigation Ion Channel Knowledge Laboratories Lentivirus Vector Maps Measures Medical center Messenger RNA Methods MicroRNAs Microarray Analysis Molecular Mutation Myocardial Nature Pathogenesis Pathway interactions Patients Pattern Phenotype Population Pregnancy Process Puromycin Replacement Therapy Resistance Sampling Signal Pathway Signal Transduction Sodium Somatic Cell Source Staging Stem cells Subfamily lentivirinae System Technology Testing Therapeutic Therapeutic Intervention Tissue Sample Tissues Translating Transplantation Viral Vector Western World Work base cell type clinical application developmental genetics fetal genetic profiling human disease human embryonic stem cell human stem cells improved in vitro Model in vivo induced pluripotent stem cell method development mutant plakophilin 2 protein protein interaction public health relevance repaired stem stem cell technology therapy design transcription factor

项目摘要

DESCRIPTION (provided by applicant): Cardiovascular diseases remain the major cause of death in the US. Stem and progenitor cell- derived cardiomyocytes (SPC-CMs) hold great promise for myocardial repairs. Recent progress in cellular reprogramming of various somatic cell types into induced pluripotent stem cells (iPSCs) opened the door for developing patient-specific, cell-based therapies. However, most SPC-CMs displayed heterogeneous and immature electrophysiological (EP) phenotypes with uncontrollable automaticity. The characteristics and stages of differentiation of cardiomyocytes (CMs) derived from SPCs or iPSCs need to be clearly defined before a safe clinical application could be performed. Furthermore, iPSC technology enables the creation of stem cell lines from patients with known genetic diseases, which has been used to study disease pathogenesis and to design therapy. In this proposal, we plan to create methods of inducing maturation of SPC- or iPSC-CMs by co-culturing endothelial cells (ECs) with these primitive CMs. We have identified that ECs promote Na+ channel expression of primitive CMs via endothelin-1 pathway. Also, we have created 3 human iPSC lines that could differentiate to CMs. We further generated the first cardiac disease-specific iPSC line that produced CMs with pathological signatures of arrhythmogenic right ventricular dysplasia (ARVD). Furthermore, we created human embryonic stem cell (hESC) and iPSC lines with Puromycin resistance by lentiviral vectors to allow rapid isolation of >95% pure iPSC- or hESC-CMs for genetic and EP analysis. Finally, in order to clearly identify the nature and fate of these normal and diseased iPSC-CMs, we have started to generate an extensive genetic map of 6 regions of embryonic and adult human hearts by micro- array technologies. Using bioinformatic analysis of data from genetic arrays, we will create a comprehensive dataset, termed Developmental HeartMatrix, so that we could compare characteristics of iPSC- or SPC-CMs to those of CM subtypes from various stages of human hearts during development, as well as to determine the stages of iPSC-CM differentiation. This project, if completed, will provide the genetic signature maps to develop methods of inducing maturation of primitive iPSC-CMs toward appropriate CM subtypes for a safe cell-based therapy. Most importantly, a human ARVD in vitro disease model could be generated by iPSC technology as the foundation for developing clinical therapy. PUBLIC HEALTH RELEVANCE: Stem and progenitor cell-derived cardiomyocytes (SPC-CMs) hold great promise for myocardial repair. Recent invention of technologies in reprogramming somatic cells to induced pluripotent stem cells (iPSCs) make patient-specific cell-based therapy possible. In addition, iPSC technology enables the creation of cardiac disease-specific lines as human in vitro disease models for developing therapeutic interventions. Also, most SPC- CMs are immature. We will create in vitro method to induce maturation of primitive SPC- CMs. Moreover, we will generate an extensive developmental gene matrix of embryonic and adult human hearts by micro-array technologies to properly classify various iPSC- or SPC-CMs as well as to guide their differentiation and maturation for a safe cell-based therapy in cardiovascular diseases.

描述（由申请人提供）：心血管疾病仍然是美国的主要死亡原因。茎和祖细胞衍生的心肌细胞（SPC-CMS）对心肌维修具有很大的希望。各种体细胞类型的细胞重编程中的最新进展为诱导的多能干细胞（IPSC）打开了开发患者特异性，基于细胞的疗法的大门。但是，大多数SPC-CM显示出具有不可控制的自动化的异质和未成熟电生理（EP）表型。在进行安全临床应用之前，需要明确定义源自SPCS或IPSC的心肌细胞（CMS）的特征和阶段。此外，IPSC技术可以从已知遗传疾病的患者中创建干细胞系，该患者已用于研究疾病发病机理和设计治疗。在此提案中，我们计划通过与这些原始CMS共同培养的内皮细胞（EC）来创建诱导SPC或IPSC-CM成熟的方法。我们已经确定EC通过内皮素-1途径促进原始CM的Na+通道表达。另外，我们创建了3种可以与CMS区分开的人IPSC线路。我们进一步产生了第一条心脏病特异性IPSC系，该系列产生了CMS具有心律失常右心室发育不良（ARVD）的病理特征。此外，我们创建了具有慢病毒载体抗毒素抗性的人类胚胎干细胞（HESC）和IPSC线，以允许快速分离> 95％的纯IPSC或HESC-CMS进行遗传和EP分析。最后，为了清楚地识别这些正常和患病的IPSC-CM的性质和命运，我们已经通过微阵列技术开始生成6个胚胎和成人人类心脏区域的广泛遗传图。使用来自遗传阵列的数据的生物信息学分析，我们将创建一个称为发育性HeartMatrix的综合数据集，以便我们可以将IPSC-或SPC-CM的特征与开发过程中人类各个阶段的CM子类型的特征进行比较，并确定IPSC-CM差异化的阶段。该项目（如果完成）将提供遗传标志图，以开发诱导原始IPSC-CM成熟的方法，以适当的CM亚型用于基于安全的细胞治疗。最重要的是，IPSC技术可以为开发临床治疗的基础而产生人类ARVD体外疾病模型。公共卫生相关性：STEM和祖细胞衍生的心肌细胞（SPC-CMS）对心肌修复具有很大的希望。在重编程体细胞中对诱导多能干细胞（IPSC）进行重编程的技术的最新发明使基于患者特异性细胞的治疗成为可能。此外，IPSC技术可以创建心脏病特异性线，作为人类体外疾病模型，用于开发治疗性干预措施。同样，大多数SPC-CM不成熟。我们将创建体外方法来诱导原始SPC-CM的成熟。此外，我们将通过微阵列技术生成广泛的胚胎和成人人心脏的发育基因基质，以适当地对各种IPSC或SPC-CMS进行分类，并指导其在心血管疾病中基于安全细胞的疗法的分化和成熟。