Patient-Specific Induced Pluripotent Stem Cells for Modeling Single Ventricle Congenital Heart Disease

用于模拟单心室先天性心脏病的患者特异性诱导多能干细胞

• 批准号: 9980713
• 负责人: Sharon Lynn Paige
• 金额: $ 8.04万
• 依托单位: STANFORD UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-07-19 至 2021-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9980713
• 关键词: Advisory Committees; Affect; Age; Apoptosis; Area; Bioinformatics; Blood flow; CRISPR/Cas technology; Cardiac; Cardiac Myocytes; Cardiac development; Cardiomyopathies; Cell Death; Cell Proliferation; Common Ventricle; Congenital Abnormality; Data; Development; Development Plans; Disease; Disease model; Embryonic Development; Evaluation; Exposure to; Foundations; Functional disorder; Funding; Gel; Gender; Gene Abnormality; Gene Expression; Gene Expression Profile; Generations; Glucose; Goals; Grant; Growth; Heart; Heart Abnormalities; Heart Valves; Human; Impairment; In Vitro; Individual; Investigation; K-Series Research Career Programs; Knowledge; Left; Left ventricular structure; Live Birth; Lung; Mediating; Mentorship; Modeling; Morbidity - disease rate; Muscle Cells; Myocardial dysfunction; Nutrient; Oxygen; Pathogenesis; Pathway interactions; Patients; Pediatric Cardiac Genomics Consortium; Pediatric cardiology; Phenotype; Physicians; Positioning Attribute; Predisposition; Prevention strategy; Reporter; Research; Research Personnel; Resources; Right ventricular structure; Scientist; Secure; Single ventricle congenital heart disease; Somatic Cell; Stress; Syndrome; System; Testing; Therapeutic; Tissues; Training; Training Programs; Translating; Universities; Ventricular; Writing; base; cardiogenesis; career; career development; congenital heart disorder; deprivation; disorder prevention; effective therapy; experimental study; genome editing; improved; induced pluripotent stem cell; infant death; malformation; mortality; novel; novel therapeutics; prevent; response; single-cell RNA sequencing; skills; stem cell model; theories; therapeutic target; therapy development; transcriptome; transcriptome sequencing; treatment strategy

PROJECT SUMMARY The purpose of this five-year proposal is to provide an integrative and personalized training program for the applicant to transition into an independent academic position as a basic scientist focused on understanding the underlying causes of congenital heart disease. The career development plan will provide additional training in induced pluripotent stem cell-derived cardiomyocyte (iPSC-CM) phenotyping, CRISPR/Cas9 genome editing, and bioinformatics. The applicant will also receive a wealth of informal and didactic training at Stanford University in specialized areas such as professional development and grant writing skills, which will be critical for the applicant to gain autonomy and launch a productive career as an independent investigator. Under the expert mentorship of Dr. Sean Wu and Dr. Euan Ashley, along with the assembled advisory committee (Dr. Daniel Bernstein, Dr. Marlene Rabinovitch, Dr. James Priest, and Dr. Matthew Porteus), the applicant will receive the necessary guidance and resources to accomplish these goals and efficiently transition to independence following the K08 training period. The research topic of this proposal fulfills a significant knowledge gap in the field by identifying myocyte- intrinsic contributions to single ventricle congenital heart disease. By using patient-specific iPSCs, intrinsic abnormalities of early cardiac development that contribute to human congenital heart disease can be studied. Among the most severe and lethal forms of congenital heart disease are single ventricle diseases, which involve profound underdevelopment of either the left or right ventricle, with resulting insufficient systemic or pulmonary blood flow. While the prevailing theory of disease pathogenesis involves impaired ventricular growth due to reduced blood flow in the developing ventricle, myocyte-intrinsic abnormalities are also suspected but largely unexplored. In Aim 1, intrinsic deficiencies in ventricular cardiomyocyte generation and function will be assessed. By using an elegant reporter system, specific abnormalities will be characterized in a chamber- specific manner and attributed to left or right ventricular iPSC-CMs. In Aim 2, the mechanisms behind myocyte- intrinsic perturbations will be investigated using single cell RNA sequencing analysis to identify abnormal gene expression patterns. Additionally, this data will be compared to ventricular tissue RNA sequencing data from the Pediatric Cardiac Genomics Consortium (PCGC) to validate the ability of iPSC-CMs to reflect diseased myocardium. Finally, in Aim 3 iPSC-CMs will be exposed to glucose and oxygen deprivation to mimic the effects of impaired blood flow during development, followed by an assessment of cell death and proliferation.

项目摘要 这一五年计划的目的是提供一个综合和个性化的培训计划， 申请人过渡到一个独立的学术立场，作为一个基础科学家，专注于了解 先天性心脏病的病因职业发展计划将提供以下方面的额外培训： 诱导多能干细胞衍生的心肌细胞（iPSC-CM）表型分析，CRISPR/Cas9基因组编辑， 和生物信息学。申请人还将在斯坦福大学接受丰富的非正式和教学培训 在大学的专业领域，如专业发展和赠款写作能力，这将是至关重要的 申请人获得自主权，并作为独立调查员开展富有成效的职业生涯。下 Sean Wu博士和Euan阿什利博士的专家指导，沿着聚集的咨询委员会（Sean Wu博士和Euan博士）。 丹尼尔伯恩斯坦，博士玛琳Rabinovitch，博士詹姆斯牧师，博士马修Porteus），申请人将 获得必要的指导和资源，以实现这些目标，并有效地过渡到 在K 08训练期之后的独立性。 该提案的研究主题通过识别肌细胞来填补该领域的重大知识空白， 单心室先天性心脏病的内在原因通过使用患者特异性iPSC， 可以研究导致人类先天性心脏病的早期心脏发育异常。 其中最严重和致命的形式的先天性心脏病是单心室疾病， 涉及左心室或右心室严重发育不全，导致全身或 肺血流虽然流行的疾病发病机制理论涉及心室发育受损 由于发育中的心室血流减少，也怀疑肌细胞内在异常， 大部分未开发。在目标1中，心室心肌细胞生成和功能的内在缺陷将被发现。 评估。通过使用优雅的报告系统，将在室内描述特定异常情况- 特定的方式和归因于左或右心室iPSC-CM。在目标2中，肌细胞- 将使用单细胞RNA测序分析研究内在扰动，以鉴定异常基因 表达模式此外，该数据将与来自以下的心室组织RNA测序数据进行比较： 儿科心脏基因组学联盟（PCGC）验证iPSC-CM反映疾病的能力， 心肌最后，在目标3中，iPSC-CM将暴露于葡萄糖和氧剥夺以模拟iPSC-CM的生长。 发育过程中血流受损的影响，然后评估细胞死亡和增殖。