Mechanisms controlling epicardial-dependent promotion of heart regeneration in zebrafish.

控制斑马鱼心外膜依赖性促进心脏再生的机制。

• 批准号: 10298630
• 负责人: Jingli Cao
• 金额: $ 42.38万
• 依托单位: WEILL MEDICAL COLL OF CORNELL UNIV
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-07-05 至 2026-06-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10298630
• 关键词: ATAC-seq; Ablation; Address; Adult; Alleles; Bacterial Artificial Chromosomes; Binding Sites; Biological Assay; Biology; Candidate Disease Gene; Cardiac Myocytes; Cell Proliferation; Cell Transplantation; Cells; Chromatin; Cicatrix; Clinical; Collagen; Coronary; Environment; Epicardium; Gene Expression Regulation; Genes; Genetic; Heart; Heart Block; Heart Injuries; Heart failure; Human; Impairment; Incidence; Injury; Knock-in; Knowledge; Label; Lead; Mammals; Mediating; Mesothelial Cell; Mitogens; Modeling; Molecular; Mutate; Myocardial Infarction; Myocardium; Natural regeneration; Nucleic Acid Regulatory Sequences; Outcome; Paracrine Communication; Pathway interactions; Patients; Pericytes; Pharmacology; Proliferating; Regenerative capacity; Regenerative response; Regulatory Element; Reporter; Role; Signal Transduction; Source; Supporting Cell; Surface; Testing; Transcriptional Regulation; Transducers; Transgenic Organisms; Transplantation; Up-Regulation; Work; Zebrafish; angiogenesis; attack victim; cardiac regeneration; cardiac repair; cell type; genetic analysis; genetic manipulation; genetic signature; genome-wide; heart damage; heart function; human stem cells; in vivo; inhibitor/antagonist; insight; multipotent cell; muscle regeneration; mutant; new therapeutic target; novel; novel strategies; paracrine; progenitor; receptor; regenerative; regenerative therapy; response; response to injury; single-cell RNA sequencing; stem cells; therapy development; transcriptome

Abstract The human heart shows little regenerative capacity following an injury such as myocardial infarction (MI). Instead, the heart scars, decreasing cardiac function, and leading to heart failure. There is no clinically meaningful regenerative therapy available for MI patients. By contrast, adult zebrafish regenerate heart muscle after severe cardiac damage without significant scarring. This is achieved through proliferation of existing cardiomyocytes (CMs), aided by the environment provided by non-muscle cells, such as the epicardium, a mesothelial cell sheet covering the surface of the heart. An analogous regenerative machinery of CM proliferation and epicardium contributions also exists in the adult mammalian heart; however, it is not sufficiently activated for significant regeneration. Recent studies demonstrated that restoring epicardial factors through the application of epicardial patches or co-transplantation of human stem cell-derived epicardial cells together with stem cell- derived CMs after an MI benefit heart regeneration. Thus, enhancing the pro-regenerative activation of the epicardium may benefit mammalian heart regeneration after MI. We and others previously found that the zebrafish epicardium is activated by injury and aids muscle regeneration through paracrine effects and as a source of multipotent cells. However, little is known about the cellular and molecular mechanisms controlling epicardial activation that lead to successful heart regeneration. To this end, understanding how regenerative responses of the epicardium are regulated in adult zebrafish will lead to new therapeutic targets that underlie the regenerative deficiencies in mammals. To address this, using single-cell RNA-sequencing, we have identified a transient adult epicardial progenitor cell (aEPC) subpopulation within the epicardium after heart injury. Transplantation assays implicate a capacity of aEPCs to give rise to perivascular cells, which are critical for coronary revascularization. Genetic ablation of these aEPCs blocks heart regeneration, suggesting an indispensable role. Pharmacological manipulations and transcriptome analyses yielded candidate genes that underlie the activation of aEPCs. Further, unbiased genome-wide profiling of chromatin accessibility using ATAC-seq revealed putative regulatory elements that exert transcriptional regulation of these genes. We hypothesize that activation of a progenitor cell state in the epicardium underlies successful heart regeneration. To test this hypothesis, we propose to 1) define the cell fates and functions of the aEPCs in adult zebrafish heart regeneration using genetic fate mapping, genetic ablation, and single-cell transplantation approaches; and 2) define the molecular mechanisms underlying aEPC activation through genetic manipulations and analyzing dozens of transgenic lines and mutants. The outcome of this proposal may ultimately inform approaches for activating the epicardial progenitors to enhance the limited regeneration displayed in humans after MI.

抽象的 损伤（例如心肌梗塞（MI））后，人心脏几乎没有再生能力。 取而代之的是，心脏疤痕，降低心脏功能并导致心力衰竭。没有临床 有意义的再生疗法可用于MI患者。相比之下，成年斑马鱼再生心肌 严重心脏损伤没有明显的疤痕。这是通过现有的扩散来实现的 心肌细胞（CMS），由非肌肉细胞提供的环境，例如心外膜，A 覆盖心脏表面的间皮细胞片。 CM增殖的类似再生机制 成年哺乳动物心脏也存在心外膜贡献。但是，它没有充分激活 为了重大再生。最近的研究表明，通过应用恢复心外膜因子 心外膜斑块或人类干细胞源性心外膜细胞的共移植以及干细胞 MI后，派生的CMS受益于心脏再生。因此，增强了促进性激活 MI后，心外膜可能受益于哺乳动物心脏再生。我们和其他人以前发现 斑马鱼心外膜被损伤激活，并通过旁分泌作用和作为一种 多能细胞的来源。但是，关于控制的细胞和分子机制知之甚少 心外神经激活导致成功的心脏再生。为此，了解再生如何 心外膜的反应在成年斑马鱼中受到调节，将导致新的治疗靶标的 哺乳动物的再生缺陷。为了解决这个问题，使用单细胞RNA-setering，我们已经确定了一个 心脏损伤后心外膜内的瞬态成年心膜祖细胞（AEPC）亚群。 移植测定法暗示了AEPC产生血管周细胞的能力，这对 冠状动脉血运重建。这些AEPC的遗传消融阻碍了心脏再生，这表明 必不可少的角色。药理学操纵和转录组分析产生了候选基因 基于AEPC的激活。此外，使用染色质可及性的无偏基因组分析 ATAC-SEQ揭示了对这些基因进行转录调控的推定调节元件。我们 假设心外膜中祖细胞状态的激活是成功的心脏再生。 为了检验这一假设，我们建议1）定义AEPC在成年斑马鱼心中的细胞命运和功能 使用遗传命运图，遗传消融和单细胞移植方法再生；和2） 通过遗传操作定义AEPC激活的分子机制并分析 数十个转基因线和突变体。该提议的结果最终可能会为 激活心外膜祖细胞，以增强MI后人类显示的有限再生。