The ECM as a coordinator of cardiac function, growth and morphogenesis during heart development

心脏是一个复杂的器官，其形状与功能紧密相连。在胚胎发育过程中，心脏最初是一根线形的管子，经过重新排列形成最终的形状。这一过程被称为心脏形态发生，如果发生不当，可能会导致先天性心脏缺陷。当心脏正在经历形态发生时，另外两个事件也会发生，这两个事件对心脏发育至关重要。首先，心脏的大小是通过向心脏两极添加细胞来实现的，这些细胞来自周围中胚层的祖细胞池，称为第二心区(SHF)。尽管SHF细胞加入心脏是心脏形态发生的重要组成部分，但我们对这些细胞如何融入心脏知之甚少。其次，心脏在形态发生过程中已经在跳动。我们以前的工作表明，心脏收缩控制着SHF细胞如何添加到心脏，并将心脏生长与形态发生结合起来。细胞和组织并不是孤立存在的-细胞周围的环境是称为细胞外基质(ECM)的各种蛋白质和糖分子，其组成因不同组织而异。细胞外基质调节周围细胞的生化信号，提供机械支持和生物力学信号，是细胞迁移、组织形态发生和器官功能的关键调节因子。因此，ECM是心管和周围SHF的重要组成部分。我们已经证明，ECM成分层粘连蛋白调节SHF的增加，可能是通过调节SHF细胞的收缩能力的影响，在心脏功能和生长之间提供了新的联系。然而，心脏功能、ECM和SHF添加在生长和形态发生耦合过程中的关系尚不清楚。在这个建议中，我们使用斑马鱼胚胎心脏发育的实时成像来了解心脏收缩能力和ECM在调节SHF添加方面的关系。斑马鱼代表了一种独特的模式生物，使我们能够定义这种关系。胚胎是透明的，并在外部发育，所以我们可以在活胚胎中可视化心脏的发育。此外，斑马鱼可以在没有心脏功能的情况下在发育的最初几天存活下来，这意味着我们可以研究心脏功能如何影响心脏发育，这在其他生物中是具有挑战性的。我们将通过三个目标来定义这种关系，所有这些目标都将利用这种能力来对发育中的斑马鱼心脏和SHF进行活体成像。首先，我们将停止或加快斑马鱼胚胎心脏的速度，并评估收缩能力如何影响细胞外基质本身。我们将可视化在心脏形态发生过程中ECM的数量和分布，并将标记特定的ECM成分，以可视化收缩能力改变时ECM的组织和组成。其次，我们将通过对跳动的心管成像和随着时间的推移迁移SHF细胞来确定ECM成分层粘连蛋白的丢失如何影响心管收缩和SHF细胞的运动。第三，我们将结合前两个目标中的实验方法来确定收缩性和/或ECM的变化如何共同影响心脏生长的速度。我们将在目标1和目标2的实验模型中生成最先进的心脏发育的3D重建，使我们能够首次定义收缩能力和细胞外基质如何通过调节生长来共同塑造心脏。定义ECM如何将心脏功能、生长和形态联系起来有很多应用。它将有助于了解推动人类心脏发育的过程，以及细胞外基质在发育过程中如何塑造体内其他组织。细胞外基质最近也被确定为心脏再生的关键驱动因素，因此这项工作对提高我们改善损伤后组织再生的能力具有重要意义。

The heart is a complex organ whose shape is tightly linked to function. During embryonic development the heart is initially a linear tube which undergoes rearrangement to generate its final form. This process is called heart morphogenesis, and if it does not occur properly can lead to birth abnormalities known as congenital heart defects. While the heart is undergoing morphogenesis, two additional events also occur which are crucial for heart development. First, the heart grows in size through the addition of cells to the poles of the heart from a progenitor pool of cells in the surrounding mesoderm called the Second Heart Field (SHF). Although the addition of SHF cells to the heart is an essential part of heart morphogenesis, we understand very little about how these cells become incorporated into the heart. Second, the heart is already beating during morphogenesis. Our previous work suggests that heart contraction controls how SHF cells are added to the heart, and couples heart growth with morphogenesis.Cells and tissues do not exist in isolation - the environment around cells is a diverse assortment of protein and sugar molecules called the extracellular matrix (ECM), the composition of which is specific to different tissues. The ECM regulates biochemical signalling to surrounding cells, provides mechanical support and biomechanical cues, and is a key regulator of cell migration, tissue morphogenesis, and organ function. The ECM is therefore a crucial component of the heart tube and surrounding SHF. We have shown that the ECM component laminin regulates SHF addition, likely by regulating the impact of contractility on SHF cells, providing a new link between heart function and growth. However, the relationship between cardiac function, the ECM, and SHF addition in coupling growth and morphogenesis is unknown.In this proposal we use live imaging of heart development in zebrafish embryos where heart function is lost or increased to understand the relationship between contractility and the ECM in regulating SHF addition. Zebrafish represent a unique model organism allowing us to define this relationship. The embryos are transparent and develop externally, so we can visualise heart development in a live embryo. Furthermore, zebrafish can survive the first few days of development without a functioning heart, meaning we can investigate how heart function impacts heart development in a way that is challenging in other organisms.We will define this relationship via three objectives, all of which will exploit this ability to perform live in vivo imaging of the developing zebrafish heart and SHF. First, we will stop or speed up the embryonic zebrafish heart and assess how contractility affects the ECM itself. We will visualise the amount and distribution of ECM in the heart during morphogenesis, and will label specific ECM components to visualise ECM organisation and composition upon altered contractility. Second, we will define how loss of the ECM component laminin affects heart tube contractility and SHF cell movement by imaging the beating heart tube and migrating SHF cells over time. Third we will combine the experimental approaches from the first two objectives to define how changes in contractility and/or the ECM together affect the rate of heart growth. We will generate state-of-the-art 3D reconstructions of the developing heart in the experimental models from objectives 1 and 2, enabling us to define for the first time how contractility and the ECM together shape the heart by mediating growth. Defining how the ECM links heart function, growth, and morphology has many applications. It will help understand the processes driving heart development in humans, as well as how the ECM shapes other tissues in the body during development. The ECM has also been recently identified as a key driver of heart regeneration, so this work has important implications in advancing our ability to improve tissue regeneration after injury.