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Engineering Human Heart Tissues with Polyploid Cardiomyocytes

用多倍体心肌细胞改造人类心脏组织

基本信息

批准号：
10467794
负责人：
Nenad Bursac
金额：
$ 54.14万
依托单位：
DUKE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2022
资助国家：
美国
起止时间：
2022-05-01 至 2026-04-30
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10467794
关键词：
3-Dimensional ATAC-seq Adult Affect Biological Assay Biomechanics Biomedical Engineering Bioreactors CRISPR interference CRISPR screen CRISPR-mediated transcriptional activation Cardiac Cardiac Myocytes Cardiac development Cell Cycle Cell Line Cell Size Cells Characteristics Chromatin Clustered Regularly Interspaced Short Palindromic Repeats Cytokinesis DNA Development Diploidy Disease model Dominant-Negative Mutation Dorsal EP300 gene Electrophysiology (science)Engineering Environment Epigenetic Process Exhibits Foundations Future Gene Expression Generations Genes Genetic Genotype Goals Heart Heart Diseases Hormonal Human Human Engineering In Vitro Insulin-Like Growth Factor I Lentivirus Libraries Mechanics Metabolic Metabolism Methodology Microtubules Mitochondria Mitogens Mitosis Molecular Mononuclear Mus Myocardium Names Outcome Study Output Oxidative Stress Pharmacologic Substance Pharmacology Phenotype Physiological Ploidies Polyploidy Property Proteins Regenerative Medicine Reperfusion Injury Reporting Repression Research Rodent Role Signal Transduction Source Stimulus Stress Structure System Testing Thyroid Hormones Tissue Engineering Tissues Workload aurora B kinase base cardiac tissue engineering cardiogenesis catalyst complex IV density drug development drug testing experience genetic inducer genome editing genome-wide human disease improved in vivo indexing induced pluripotent stem cell mechanical load mimetics novel organ on a chip overexpression postnatal postnatal development regenerative therapy response segregation stable cell line three dimensional cell culture tissue culture trait transcriptome sequencing transcriptomics

项目摘要

Human induced pluripotent stem cells (hiPSCs) represent a potentially unlimited source of functional cardiomyocytes (hiPSC-CMs) for use in disease modeling, drug development, and regenerative therapies. In particular, use of hiPSC-CM-derived microtissues in microphysiological (“organ-on-chip”) systems holds promise as the future mainstay of pharmaceutical research and a platform to improve our understanding of genotype-phenotype relationships in mono- and polygenic diseases of the human heart. However, a major obstacle to wide-spread use of hiPSC-CMs in these applications are their immature properties including small cell size, lack of T-tubules, predominantly glycolytic metabolism, and reduced functional output, to name a few. One important aspect of postnatal CM maturation - increased ploidy - has been largely understudied. Namely, in vitro cultured hiPSC-CMs are predominantly mononuclear and diploid, while the adult human myocardium consists of ~90% polyploid CMs. We thus propose to investigate potential roles of polyploidy in hiPSC-CM maturation, and specifically, to explore if engineered polyploidy of hiPSC-CMs can endow human engineered cardiac tissues (hECTs) with increased functionality and maturity compared to control tissues made from primarily diploid CMs. Our preliminary results show that stable hiPSC-CM polyploidy induced genetically or pharmacologically results in increased size and mitochondrial density of hiPSC-CMs, as well as contractile strength and conduction velocity of hECTs. In the proposed studies, we will further examine roles of CM polyploidization in structural, functional, and metabolic maturation of hECTs and investigate polyploidy-induced transcriptomic and epigenetic changes in hiPSC-CMs. Furthermore, using a novel bioreactor with capacity to dynamically control applied mechanical preload and afterload to hECTs, we will investigate the relationships between developmentally-mimetic regimes of mechanical loading and CM polyploidy. We will also determine if polyploidy sensitizes hiPSC-CMs to hypertrophic stimuli in vitro and protects hECTs from oxidative stress in vitro and ischemic damage in vivo. Finally, we will apply CRISPR/Cas9 screening methodologies to identify genetic inducers of terminal maturation in already polyploidy hiPSC-CMs and will perform additional screens in both diploid and polyploid hiPSC-CMs to identify candidate mitogens that can promote CM cell cycle activity. By successfully completing these studies, we expect to improve our understanding of physiological roles of polyploidy in cardiac development and to establish the foundation for the future translational uses of engineered cardiac tissues in disease modeling, drug development, and cardiac therapies.

人诱导多能干细胞（hiPSCs）是一种潜在的无限功能来源