iPSC-derived astrocytes to model Vanishing White Matter Disease

消失性白质病(VWM)是最常见的脑白质营养不良之一，由 真核翻译起始因子EIF2B的五个亚基中的任何一个发生突变。即使有很多人 不同种类的EIF2B突变已经被鉴定，目前还不清楚这些突变是如何导致 髓鞘戏剧性丧失。 最大的问题之一是缺乏一个合适的实验系统来测试 EIF2B突变对脑细胞的影响我们对VWM患者来源细胞的初步研究是通过 从VWM患者的大脑中分离活的神经胶质前体细胞的机会非常罕见。本研究 揭示了星形胶质细胞的缺陷。但缺乏合适的患者样本限制了我们研究细胞的能力 导致这种疾病，从而阻止我们找到治疗这种毁灭性疾病的方法。 这项提议的目标是利用可诱导多能干细胞(Ipsc)技术来衍生神经。 来自具有不同EIF2B复合突变的VWM患者的干细胞培养。这些VWM神经干细胞将 用于检测与星形胶质细胞分化和功能有关的基因-表型关系。我们的 初步数据显示，IPSCs可以很容易地从VWM患者的细胞中获得，并且这些患者- 来源的PSCs可以被诱导分化为神经干细胞和星形胶质细胞前体。基因突变 EIF2B似乎导致细胞死亡增加，并削弱一些星形胶质细胞系蛋白的表达， 包括星形胶质细胞谱系决定基因CREB3L1。 由于星形胶质细胞在维持中枢神经系统内稳态中的关键作用，我们的发现提出了两个重要的 问题：(1)具有不同EIF2B突变的VWM患者是否表现出相同的星形胶质细胞缺陷？(2)如何 EIF2B损伤扰乱星形胶质细胞生物学？为了解决这些问题，我们将：(目标1)建立和 鉴定来自不同VWM患者的可诱导多能干细胞(IPS)细胞系，以及(Aim 2)用途 这些细胞用于检测星形胶质细胞分化、反应性胶质增生以及支持生存和 少突胶质细胞的分化。

Vanishing White Matter (VWM) disease is one of the most prevalent leukodystrophies, and is caused by mutations in any of the five subunits of eukaryotic translation initiation factor EIF2B. Even though many different kinds of EIF2B mutations have been identified, it is still unclear how these mutations cause the dramatic loss of myelin. One of the biggest problems has been the lack of a suitable experimental system in which to test the effects of EIF2B mutations on brain cells. Our initial study of VWM patient-derived cells was made possible by the exceedingly rare opportunity to isolate live glial precursor cells from the brain of a VWM patient. This study revealed a defect in astrocytes. But the lack of suitable patient samples limits our ability to study the cellular causes and consequently prevent us from finding a treatment for this devastating disease. It is the goal of this proposal to exploit inducible pluripotent stem cell (iPSC) technology to derive neural stem cell cultures from VWM-patient with distinct EIF2B complex mutations. These VWM-neural stem cells will be used to test the genotype-phenotype relationship with respect to astrocyte differentiation and function. Our preliminary data demonstrate that iPSCs can readily be derived from VWM-patient cells and that these patient- derived PSCs can be induced to differentiate into neural stem cells and astrocyte precursors. Mutations in EIF2B appear to cause increased cell death, and impair expression of some astroglial lineage proteins, including the astroglial lineage determinant CREB3L1. Because of the critical role of astrocytes in maintaining CNS homeostasis, our findings raise two important questions: (1) Do VWM patients with distinct EIF2B mutations exhibit the same astrocyte defect? (2) How does impairment of EIF2B disrupt astrocyte biology? To address these questions, we will: (Aim 1) establish and characterize inducible pluripotent stem (iPS) cell lines from different VWM-disease patients, and (Aim 2) use these cells to examine astroglial differentiation, reactive gliosis and the ability to support survival and differentiation of oligodendrocytes.