Micropattern differentiation and morphogenesis of the human ectoderm

项目摘要/摘要 在动态组织规模的背景下如何准确地指定细胞命运 重排是发育生物学中最令人兴奋的问题之一。在……里面 我们的这项提议旨在破译命运获取之间的相互作用， 外胚层胚层的图案化和形态发生 人类的神经形成。我们最近开发了一种强大的协议，它允许 用于产生极具再生性的人类神经元：自组织 人类胚胎干细胞(HESC)重塑组织的组件 在神经形成阶段通过组织神经、神经 冠、胎盘和表皮种群在同一菌落内粘着 微图案。这种自组织是非常可重现的，并且可以 以亚细胞分辨率进行量化，并在数百个菌落上实时进行。 在这项新技术的武装下，我们提出了三个具体目标。第一个是 解开细胞-细胞信号驱动自组织的机制。第二 是将信号与命运的获得和形态发生结合在一起 报告成像和时间依赖的单细胞RNAseq。最后，第三个目标 重点研究外胚层的起源和亚群的完整特征 衍生物及其体内验证通过与 雏鸡阶段配型绒猴胎儿标本及移植试验 胚胎。大量同质人类神经细胞的产生， 在那里，外胚层命运的自组织可以动态地跟踪一段时间 一周，具有亚细胞分辨率，不仅解决了固有的 在脑器官中观察到的异质性，但为我们提供了一个独特的 在人类胚胎模型中研究这些事件的机会。这将会有一个 在基础研究和临床应用方面都有很高的影响力，前景广阔 已经在地平线上了。 好了！

Project Summary/Abstract How accurate cell fate specification occurs in the context of dynamic tissue-scale rearrangements is one of the most exciting questions in developmental biology. In this proposal we aim at deciphering the interplay between fate acquisition, patterning and morphogenesis of the ectodermal germ layer in the context of human neurulation. We have recently developed a robust protocol which allows for the generation of extremely reproducible human neuruloids: self-organized human Embryonic Stem Cell (hESC) assemblies that recapitulate the organization of the ectodermal compartment at neurulation stages by organizing neural, neural crest, placodes and epidermis populations within the same colony on adhesive micropatterns. This self-organization is extremely reproducible and can be quantified with sub-cellular resolution and in real time over hundreds of colonies. Armed with this novel technology, we propose three specific aims. The first is to unravel the mechanism of cell-cell signaling driving self-organization. The second is to integrate signaling with fate acquisition and morphogenesis through live reporter imaging and time dependent single cell RNAseq. Finally, the third aim focuses on the full characterization of the origin and sub-populations of ectodermal derivatives and their in vivo validation by performing side by side comparisons with stage-matched marmoset fetal samples and grafting experiments in chick embryos. The generation of large numbers of homogenous human neuruloids, where self-organization of ectodermal fate can be followed dynamically for a period of one week, with sub-cellular resolution, not only solves the inherent heterogeneity observed in cerebral organoids, but provides us a unique opportunity to study these events in models of human embryos. This will have a high impact in both basic research as well as clinical application, a prospect already on the horizon. !