The zebrafish tectal stem cell niche - a new model for in vivo analysis of neural stem cell biology

胚胎后生物体中的干细胞调节生长、再生和修复。哺乳动物中最典型的两个例子是肠道干细胞和造血干细胞，前者每隔几天更新一次肠道上皮，后者确保新血细胞的稳定供应。神经干细胞因其带来的治疗前景而获得了突出地位。从理论上讲，这种细胞提供了再生受损神经组织的可能性，从而为治疗神经退行性疾病(如阿尔茨海默氏症和帕金森氏病)和神经系统损伤提供了治疗。部分出于这个原因，人们正在做出重大努力来识别和描述调节神经干细胞维持和分化的因素。哺乳动物的神经系统只包含有限的神经干细胞群体，这些干细胞居住在不易到达的利基环境中。因此，在活体动物的自然环境中研究这种细胞几乎是不可能的。与哺乳动物相比，包括鱼类在内的其他脊椎动物拥有许多神经干细胞群体，这使它们成为补充哺乳动物和组织培养实验的良好模型。在本项目中，我们建议建立斑马鱼视顶盖作为研究神经干细胞生物学的新的模型系统。视顶盖是鱼类一生中生长和增加神经元的重要视觉中心。干细胞使这种生长成为可能，并且有能力产生十几种类型的顶盖神经元，它们位于大脑表面附近。再加上斑马鱼胚胎的透明性，这些细胞可以在体内被跟踪。对于完好动物中神经干细胞和祖细胞的行为，人们知之甚少，我们的主要目标之一将是观察这些细胞在分裂、移动和分化时的行为。为了做到这一点，我们将使用先进的转基因工具，使我们能够可视化细胞形状，跟踪细胞分裂，并在祖细胞生成神经元时跟踪细胞运动。我们的第二个目标是描述影响神经干细胞生物学的信号。关于控制造血干细胞的信号已经了解了很多，我们发现顶盖和造血祖细胞之间有一些令人惊讶的相似之处。Wnt途径在这两个位置都被激活，我们将确定这一途径如何影响顶盖干细胞的维持、祖细胞的增殖和神经元的产生。其次，最近的数据表明前列腺素是造血干细胞生态位的新的、关键的调节剂。我们有初步数据表明，前列腺素可能在顶盖干龛中具有保守功能，并将探索这一可能性。这些研究将有助于确定调节干细胞的机制的保守程度，并将阐明神经干细胞生物学基础的信号机制。另一个目标是解决眼睛和视盖之间的生长是如何协调的，我们将通过评估眼睛的顶盖神经如何调节干细胞和祖细胞的行为来解决这个问题。我们的工作的好处包括更好地理解神经干细胞生物学，这是从对完整动物的研究中得出的。斑马鱼的透明度和顶盖干细胞的易获性为在这一背景下研究神经干细胞提供了明显的优势。我们的研究将使其他致力于不同干细胞模型的人在帮助设计实验和操纵干细胞用于治疗目的方面受益。调控干细胞的信号通路可以通过药物和/或基因操作来调节，这将提供操纵干细胞的途径。这项拟议的研究还将建立一个新的干细胞研究模型系统，我们希望这一系统将被其他研究人员利用。

Stem cells in post-embryonic organisms regulate growth, regeneration and repair. The two best-characterized examples in mammals are intestinal stem cells, which renew the gut epithelium every few days, and haematopoietic stem cells that ensure the steady supply of new blood cells. Neural stem cells have acquired prominence due to the therapeutic promise they carry. Theoretically, such cells provide the possibility to regenerate damaged neural tissue thereby providing treatment for neurodegenerative disorders (such as Alzheimer's and Parkinson's disease) and nervous system injuries. In part for this reason, significant efforts are being made to identify and characterise the factors that regulate the maintenance and differentiation of neural stem cells. The mammalian nervous system contains only limited populations of neural stem cells, and these reside in niches that are not easily accessible. It is therefore almost impossible to study such cells in their natural context of the living animal. In contrast to mammals, other vertebrates, including fish, have many populations of neural stem cells making them good models that can complement mammalian and tissue culture experiments. In this project, we propose to establish the zebrafish optic tectum as a new model system to study the biology of neural stem cells. The optic tectum is an important visual centre that grows and adds neurons throughout the life of the fish. The stem cells that make this growth possible, and that have the capacity to generate more than a dozen tectal neuron types, reside close to the superficial surface of the brain. Coupled with the transparency of the zebrafish embryo, these cells can be followed in vivo. Relatively little is known about the behaviour of neural stem cells and progenitors in the intact animal and one of our key goals will be to observe these cells as they divide, move and differentiate. To do this, we will use sophisticated transgenic tools that allow us to visualise cell shape, to follow cell divisions and to track cell movements as progenitor cells generate neurons. Our second goal is to characterise the signals that influence the biology of the neural stem cells. Much has been learned about the signals that control haematopoietic stem cells, and we have found some surprising similarities between tectal and haematopoietic progenitor cells. The Wnt pathway is activated in both locations and we will determine how this pathway influences the maintenance of tectal stem cells, the proliferation of progenitors and the production of neurons. Secondly, recent data has implicated prostaglandins as being novel, critical regulators of the haematopoietic stem niche. We have preliminary data that prostaglandins may have conserved function in the tectal stem niche and will explore this possibility. These studies will help to determine the extent of conservation of the mechanisms that regulate stem cells and will elucidate the signalling mechanisms that underlie neural stem cell biology.A further goal is to resolve how growth is coordinated between the eyes and the optic tectum and we will address this issue by assessing how tectal innervation by the eyes regulates stem cell and progenitor behaviour. The benefits from our work include a better understanding of neural stem cell biology drawn from studies in the intact animal. The transparency of the zebrafish coupled with the ease of accessibility of the tectal stem cells provides a clear advantage for studying neural stem cells in this context. Our research will benefit others working on different stem cell models in helping to design experiments and manipulate stem cells for therapeutic purposes. The signalling pathways that regulate stem cells can be modulated by drugs and/or genetic manipulation and this will provide routes to manipulate stem cells. The proposed research will also establish a new model system to study stem cells that we hope will be exploited by other researchers.