Defining Characteristics of Cortical Progenitor Cells over Time in Mouse and Human

定义小鼠和人类皮质祖细胞随时间变化的特征

• 批准号: 10532479
• 负责人: SALLY TEMPLE
• 金额: $ 8.7万
• 依托单位: REGENERATIVE RESEARCH FOUNDATION
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-12-15 至 2024-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10532479
• 关键词: Address; Adult; Architecture; Candidate Disease Gene; Cell Cycle; Cell division; Cells; Cerebral cortex; Cerebrum; Characteristics; Code; Complex; Data; Development; Disease; Embryo; Environmental Risk Factor; Generations; Genes; Heterogeneity; Human; Image Analysis; In Vitro; Knowledge; Life; Methods; Mus; Neurodegenerative Disorders; Neuroglia; Neurons; Pregnancy; Process; Production; RNA-Binding Proteins; Radial; Role; Specific qualifier value; Structure; Structure of choroid plexus; Testing; Time; Translations; Untranslated RNA; Vascular Endothelial Cell; Work; blastomere structure; daughter cell; developmental disease; gene network; in vivo; nerve stem cell; novel; progenitor; receptor; regenerative therapy; release factor; screening; stem cell technology; stem cells; stem-like cell; therapy development; time use; transcriptome sequencing

Cerebral cortical development is a highly orchestrated process, with production of neurons destined for the cortical layers produced in order, deep to superficial, followed by glial generation. The timing of this process is very different between species. For example, mouse corticogenesis occurs over approximately a week of gestation, while in humans the process takes several months, resulting in a much larger and more complex cortex. Lineage studies have functionally defined the major types of neural progenitor cells (NPCs) contributing to corticogenesis, including stem cell-like radial glial cells (RGCs) and intermediate progenitor cells (IPCs). However, much remains to be discovered regarding how RGCs and IPCs are specified over time. We have discovered that during asymmetric RGC-IPC cell divisions, the RNA binding protein Stau2 segregates a complex cargo of coding and non-coding RNA specifically into the IPC daughter. Analysis of this cargo at different embryonic stages by RNA-sequencing has revealed networks of genes that are candidates for controlling proliferation and temporal specification of the IPC fate. Here we propose to test these candidates in functional studies, using high-throughput automated time-lapse image analysis for in vitro studies, as well as a novel lentiviral in vivo screening method, to define their roles in specifying IPCs and timing corticogenesis. In contrast to the progress made in understanding the characteristics of mouse cortical progenitor cells, less is understood regarding human cortical progenitors. Fundamental knowledge about how human RGCs and IPCs produce diverse progeny over time, their division mode, cell cycle times and lineages, remains unknown. Here we will address these gaps in knowledge using long-term time-lapse lineage analysis in vitro. In addition, by identifying genes expressed in human cortical progenitor cells, including at the single cell level and via analysis of the Stau2 cargo, we will reveal human cortical progenitor subtypes and heterogeneity. Further, a comparison of human and mouse cortical progenitor cell data will help illuminate key differences to address a major mystery: the difference in timing of mouse and human cortical development. Our lab continues to explore the interaction of environmental factors on cortical progenitor cells, aided by the ability to rapidly quantify changes in proliferation, division mode and differentiation using time-lapse analysis. Soluble factors released by structures in the germinal niche such as vascular endothelial cells and the choroid plexus, act on cortical progenitors to regulate the numbers and types of progeny they produce. Our recent work has identified a panel of candidate niche molecules secreted by the choroid plexus that could interact with receptors expressed on neural progenitors, which we propose to examine in vitro and in vivo, in mouse and human. Defining niche factors and their specific actions paves the way to address diseases that involve degeneration of stem cell zones which are normally active throughout life. Furthermore, defining environmental factors that act on human NPCs is important for translation towards regenerative therapy development.

大脑皮层发育是一个高度精心安排的过程，神经元的产生按顺序从深层到浅层产生，然后是神经胶质细胞的产生。这个过程的时间在不同物种之间有很大不同。例如，小鼠的皮质生成发生在妊娠期大约一周的时间，而人类的这一过程需要几个月的时间，从而产生更大、更复杂的皮质。谱系研究在功能上定义了有助于皮质生成的神经祖细胞（NPC）的主要类型，包括干细胞样放射状胶质细胞（RGC）和中间祖细胞（IPC）。然而，关于如何随着时间的推移指定 RGC 和 IPC，还有很多事情有待发现。我们发现，在不对称 RGC-IPC 细胞分裂过程中，RNA 结合蛋白 Stau2 将编码和非编码 RNA 的复杂货物特异性地分离到 IPC 子细胞中。通过 RNA 测序对不同胚胎阶段的这种货物进行分析，揭示了控制增殖和 IPC 命运时间规范的候选基因网络。在这里，我们建议在功能研究中测试这些候选药物，使用高通量自动延时图像分析进行体外研究，以及一种新型慢病毒体内筛选方法，以确定它们在指定 IPC 和计时皮质生成中的作用。与了解小鼠皮质祖细胞特征方面取得的进展相比，对人类皮质祖细胞的了解较少。关于人类 RGC 和 IPC 如何随着时间的推移产生不同的后代、它们的分裂模式、细胞周期时间和谱系的基本知识仍然未知。在这里，我们将使用体外长期延时谱系分析来解决这些知识空白。此外，通过鉴定人类皮质祖细胞中表达的基因，包括在单细胞水平上以及通过对Stau2货物的分析，我们将揭示人类皮质祖细胞亚型和异质性。此外，人类和小鼠皮质祖细胞数据的比较将有助于阐明关键差异，从而解开一个重大谜团：小鼠和人类皮质发育时间的差异。我们的实验室借助延时分析快速量化增殖、分裂模式和分化变化的能力，继续探索环境因素对皮质祖细胞的相互作用。生发生态位中的结构（例如血管内皮细胞和脉络丛）释放的可溶性因子作用于皮质祖细胞，调节它们产生的后代的数量和类型。我们最近的工作确定了一组由脉络丛分泌的候选利基分子，它们可以与神经祖细胞上表达的受体相互作用，我们建议在小鼠和人类体内进行体外和体内检查。定义利基因素及其具体作用，为解决涉及干细胞区域退化的疾病铺平了道路，而干细胞区域通常在一生中都很活跃。此外，定义作用于人类 NPC 的环境因素对于再生疗法的发展非常重要。