Stem Cell Generation of Transport Epithelia for the Brain's Ventricular System

本研究的目的是研究胎儿神经上皮干细胞是如何支持神经发生的， 和室管膜发生，并确定脑室扩大或脑积水如何影响 室管膜发生和相关的干细胞功能。室管膜细胞沿着 脑室壁，并作为脑脊髓液（CSF）和间质之间的运输功能单位 流体（ISF）。一个健康的室管膜循环脑脊液和促进清除毒素从ISF的大脑 薄壁组织在妊娠中期，足够数量的室管膜细胞由神经上皮细胞产生， 干细胞覆盖整个脑室表面，但在脑积水中，脑室周围干细胞龛是 产生更多的室管膜细胞来覆盖扩张的心室。我们将测试 假设心室扩张耗尽了干细胞库并导致心室的覆盖不足， 心室表面、室周神经胶质增生和关键的经室管膜转运/清除功能丧失。 实验旨在记录干细胞和干细胞之间的动态供需关系， 其后代室管膜细胞，在人类组织和小鼠脑积水模型中。在目标1中，我们 创建室管膜细胞生成的时空图，并确定 正常胎儿及出生后人室管膜细胞分布及干细胞数量/组织学研究 脑室周围脑组织将在小鼠中进行类似的研究，补充延时干细胞 使用piggyBac子宫内电穿孔系统进行细胞谱系追踪，以追踪干细胞命运并测试 室管膜发生是否导致干细胞退出细胞周期。由此产生的3D时空地图 室管膜发生将作为评价异常室管膜发育和干细胞生态位的工具 脑积水的变化。在目标2中，我们将检查几种脑积水模型，以确定 心室表面的干细胞龛和室管膜细胞覆盖受到影响。细胞组织，细胞-细胞 连接复合体、室管膜血流改变和心室表面清除机制， 将阐明干细胞动力学。所产生的数据将提供对再生潜力的深入了解， 神经上皮干细胞修复心室表面，以及干细胞功能缺陷如何 导致与脑积水相关的多种神经病理。

The goals of this proposal are to examine how the fetal neuroepithelial stem cells support both neurogenesis and ependymogenesis, and to determine how ventricular enlargement, or hydrocephalus, affects ependymogenesis and associated stem cell functions. Ependymal cells are arranged as a monolayer along the ventricle walls and act as the functional units of transport between the cerebrospinal fluid (CSF) and interstitial fluid (ISF). A healthy ependyma circulates CSF and facilitates the clearance of toxins from the ISF of the brain parenchyma. During mid-gestation, sufficient numbers of ependymal cells are generated from neuroepithelial stem cells to cover the entire ventricle surface, but in hydrocephalus, the periventricular stem cell niche is called upon to generate many more ependymal cells to cover the expanding ventricles. We will test the hypothesis that ventricle expansion exhausts the stem cell pool and results in insufficient coverage of the ventricle surface, periventricular gliosis, and loss of critical transependymal transport/clearance functions. Experiments are designed to document the dynamic supply and demand relationship between stem cells and their progeny ependymal cells, in both human tissue and mouse models of hydrocephalus. In Aim 1, we will create spatiotemporal maps of ependymal cell generation and determine the relationship between the distribution of ependymal cells and stem cell numbers/organization in normal fetal and postnatal human periventricular brain tissue. Similar studies will be performed in mouse, supplemented with time-lapse stem cell lineage tracing using the piggyBac in utero electroporation system, to track stem cell fates and test whether ependymogenesis results in stem cell exit from the cell cycle. The resulting 3D spatiotemporal maps of ependymogenesis will serve as tools for evaluating abnormal ependyma development and stem cell niche changes in hydrocephalus. In Aim 2, we will examine several models of hydrocephalus to determine how the stem cell niche and ependymal cell coverage of the ventricle surface are affected. Cell organization, cell-cell junction complexes, alterations in transependymal flow and clearance mechanisms at the ventricle surface and stem cell dynamics will be elucidated. Data generated will provide insight into the regenerative potential of neuroepithelial stem cells for repair of the ventricle surface and how deficits in stem cell functions may contribute to the multiple neurologic pathologies associated with hydrocephalus.