Dissecting epicardial contributions to zebrafish heart regeneration at the single-cell level

摘要 在发生心肌梗死 (MI) 等损伤后，成年人的心脏仅具有极少量的功能 再生能力。与人类和大多数哺乳动物相比，成年斑马鱼具有 具有显着的解决纤维化疤痕和损伤后心肌再生的能力。由于有 目前尚无针对心肌梗死引起的损伤的临床再生疗法，因此仍然非常需要 生物医学研究界了解促再生机制，使强健 斑马鱼心脏再生。对于这个再生过程来说，心外膜是不可或缺的，它是 心脏的最外层组织。通过贡献关键的旁分泌信号和促再生细胞类型， 心外膜促进心肌细胞（CM）增殖和损伤部位的血运重建。那 话虽如此，这些分子和细胞过程的精确机制在很大程度上是未知的，因此 提出了有关斑马鱼心脏再生调节的许多问题。为了解决这些 为了回答这些问题，我们对受伤和受伤的心外膜细胞进行了单细胞 RNA 测序。 未受伤的成年斑马鱼心脏。初步分析鉴定出成人心外膜祖细胞 有潜力分化成血管周围细胞以促进血管生成的亚群。在这里， 通过基因消融和谱系追踪，我们的目标是确定细胞和分子的贡献 这些祖细胞促进心脏再生。进一步的分析表明 Tgfb 通路调节 这些心外膜祖细胞的增殖和分化。为了更好地了解 潜在的调节机制，我们将进一步明确 Tgfb 通路在再生中的作用 使用基因获得/丧失功能测定和改良 RNA 注射来检测心外膜。综上所述，本项目 可能最终为针对心外膜祖细胞的促再生潜力提供策略 改善心肌梗死患者的心脏修复和功能。

Abstract After an injury, such as myocardial infarction (MI), the adult human heart possesses only minimal regenerative capabilities. In contrast to humans and most mammals, the adult zebrafish possesses a remarkable ability to resolve fibrotic scarring and regenerate cardiac muscle after injury. Since there are currently no clinical regenerative therapies for MI-induced injuries, there persists a great need in the biomedical research community to understand the pro-regenerative machinery that enable robust zebrafish heart regeneration. Indispensable to this regenerative process is the epicardium, which is the outermost tissue layer of the heart. By contributing key paracrine signals and pro-regenerative cell types, the epicardium promotes cardiomyocyte (CM) proliferation and revascularization of the injury site. That being said, the precise mechanisms of these molecular and cellular processes are largely unknown, thus raising many questions in regard to the regulation of zebrafish heart regeneration. To address these questions, we performed single-cell RNA sequencing of isolated epicardial cells from injured and uninjured adult zebrafish hearts. Preliminary analysis identified an adult epicardial progenitor cell subpopulation that has the potential to differentiate into perivascular cells to facilitate angiogenesis. Here, through genetic ablation and lineage tracing, we aim to determine the cellular and molecular contributions of these progenitors to heart regeneration. Further analysis suggests that the Tgfb pathway regulates the proliferation and differentiation of these epicardial progenitors. To gain a greater understanding of the underlying regulatory mechanisms, we will further define the regenerative role of the Tgfb pathway in the epicardium using genetic gain/loss-of function assays and modified RNA injections. In sum, this project may ultimately inform strategies to target the pro-regenerative potential of epicardial progenitors to improve heart repair and function in MI patients.