Probing the efficacy and potency of cardiomyocyte regeneration in the zebrafish heart

与哺乳动物相比，斑马鱼的再生能力更强。在对损伤的反应中，斑马鱼的心脏不会形成永久性的疤痕，心肌细胞会重新进入细胞周期。然而，尽管斑马鱼心脏再生经常被描述为“完全”，但实际上并不清楚该器官是否恢复到损伤前的状态。特别是，它是未知的，在何种程度上损伤心肌细胞失去再生。我们发现，心肌细胞的细胞周期活动恢复到基线水平之前，伤口组织已完全解决，这表明没有永久性疤痕不能作为证据，充分心肌再生。考虑到斑马鱼心脏再生模型的普及，我们认为最终澄清心肌细胞实际再生的程度是很重要的。我们已经建立了可靠的方法来评估心肌细胞的数量，使用体视学原理，从组织切片的数量。我们将在心脏冷冻损伤后的几个时间点确定心肌细胞的数量，因此我们不仅能够回答心肌细胞是否完全再生，而且还能够回答心肌细胞完全再生的时间范围以及这与伤口组织消退的关系。此外，我们计划解决斑马鱼心脏再生研究中另一个有趣但尚未回答的基本问题。在许多再生系统中，细胞显示出有趣的细胞可塑性，包括成熟细胞去分化为多能状态的例子。虽然分化的心肌细胞已通过遗传谱系追踪显示为心脏再生期间新形成的心肌细胞的前体，但尚不清楚它们是否也可以在再生期间产生其他细胞类型。这是因为迄今为止用于基于Cre-lox的心肌细胞谱系追踪的所有转基因系仅在心肌细胞中表达。因此，可能会错过向其他细胞类型的转分化。我们已经创建了一个新的转基因Cre reponder线，在心脏的主要细胞类型中表达。使用这个工具，我们将问是否心肌细胞可以有助于内皮细胞，心外膜或成纤维细胞谱系在regeneration.Together的工作提出这里将解决两个基本的开放问题有关的斑马鱼心脏再生的细胞机制，从而将允许一个大大改善的评估这个模型的有用性。

Compared to mammals, zebrafish display much elevated regenerative capacities. In response to injury, zebrafish hearts do not form permanent scars, and cardiomyocytes re-enter the cell cycle. Yet, although zebrafish heart regeneration is frequently described as "complete", it is actually unclear whether the organ recovers to a pre-injury status. In particular, it is unknown to which extent cardiomyocytes lost to injury are regenerated. We found that cardiomyocyte cell cycle activity returns to baseline levels much before wound tissue has been fully resolved, suggesting that absence of a permanent scar cannot be used as evidence for full myocardial regeneration. Considering the popularity of the zebrafish heart regeneration model, we think it is important to finally clarify to which extent cardiomyocytes actually regenerate. We have established reliable methods for assessment of cardiomyocyte numbers, using stereological principles to derive the number from tissue sections. We will determine cardiomyocyte numbers at several time points after heart cryoinjury, and thus we will not only be able to answer whether cardiomyocytes fully regenerate but also the time frame in which this happens and how this relates to wound tissue resolution.In addition, we plan to tackle another intriguing yet unanswered fundamental question in zebrafish heart regeneration research. In many regenerating systems, cells display intriguing cellular plasticity, including examples of dedifferentiation of mature cells to a multipotent state. While differentiated cardiomyocytes have been shown by genetic lineage tracing to be the precursors of newly forming cardiomyocytes during heart regeneration, it is unknown whether they can also give rise to other cells types during regeneration. This is because all transgenic lines that have so far been used for Cre-lox based cardiomyocyte lineage tracing were only expressed in cardiomyocytes. Thus, potential transdifferentiation to other cell types would have been missed. We have created a new transgenic Cre reponder line that is expressed in the major cell types of the heart. Using this tool we will ask whether cardiomyocytes can contribute to the endothelial, epicardial or fibroblast lineage during regeneration.Together the work proposed here will settle two fundamental open questions related to the cellular mechanisms of zebrafish heart regeneration, and thus will allow for a much-improved evaluation of the usefulness of this model.