Molecular Regulation of Human Glial Progenitor Cell-Based Remyelination

描述（由申请人提供）：髓鞘疾病包括遗传性脑白质营养不良和脑瘫，以及成人血管性、创伤性和炎性脱髓鞘综合征。为了解决这一庞大而多样的疾病组，我们建立了一种中枢髓鞘再生的细胞治疗方法，通过该方法将分离的人胶质祖细胞（GPC）移植物脑内递送给新生儿受体，然后使其成熟至成年。当受体是低髓鞘化突变体时，例如shiverer小鼠，移植的细胞主要成熟为髓鞘少突胶质细胞，并且可以挽救治疗动物的神经表型和寿命。然而，值得注意的是，大量的人类祖细胞整合到受体大脑中，在那里它们有效地胜过小鼠祖细胞，产生了具有基本上人源化的白色物质的小鼠，以及灰质中的主要的人类神经胶质祖细胞-以及最终的星形胶质细胞。由此产生的人类神经胶质嵌合小鼠脑为我们提供了各种迄今为止无法获得的机会，在体内研究人类神经胶质细胞及其祖细胞，包括他们的反应，不能充分模拟在体外损伤和疾病的过程。在拟议的实验中，我们将使用这些小鼠来评估体内毒性脱髓鞘对人类GPCs的影响，以确定它们对损伤诱导的动员和补偿性髓鞘再生过程中少突胶质细胞分化的分子反应。特别是，我们将使用表型特异性细胞分选和基因表达分析，以确定这些异种移植的人GPCs在体内脱髓鞘的反应。这些数据，有史以来第一次获得专门从人类GPCs在脱髓鞘和髓鞘再生在体内，应该给我们提供基本的新的见解与髓鞘再生相关的信号事件，其潜在的目标点的监管控制。为了实现这一目标，我们提出了以下目标：在目标1中，我们将用cuprizone处理神经胶质嵌合小鼠，以便更好地理解人GPCs对脱髓鞘的反应，评估它们的动员、分化、对重复诱导的反应，以及它们的有丝分裂衰老阈值及其调控。在目标2中，我们将研究人类GPC在体内的表达模式，在基线和脱髓鞘反应中将它们从嵌合颤抖小鼠中分选出来，以确定GPC动员和髓鞘再生过程中差异调节的基因和途径。在目标3中，我们将比较共同居住的人类和小鼠GPCs对cuprizone脱髓鞘的反应，以确定那些可能成为药物开发中高价值靶点的共享途径，以及那些在小鼠中研究可能无法预测人类治疗结果的物种特异性途径。总之，这些实验有望为我们确定治疗脱髓鞘性脑损伤或脊髓损伤的新策略提供信息。此外，在这项工作的过程中产生的数据库，作为免费提供的资源，该领域应该证明催化推进我们的理解髓鞘再生在体内。

DESCRIPTION (provided by applicant): Disorders of myelin include the hereditary leukodystrophies and cerebral palsies, as well as adult vascular, traumatic and inflammatory demyelination syndromes. To address this large and diverse group of disease, we established a cell-therapeutic approach to central remyelination, by which transplants of isolated human glial progenitor cells (GPCs) are delivered intracerebrally to neonatal recipients, which are then allowed to mature to adulthood. When the recipients are hypomyelinated mutants, such as the shiverer mouse, the transplanted cells mature largely as myelinating oligodendrocytes, and can rescue both the neurological phenotype and lifespan of the treated animals. Remarkably though, large numbers of human progenitors integrate into the recipient brains, wherein they effectively out-compete mouse progenitors, yielding mice with a substantially humanized white matter, and a major contingent of human glial progenitors - and ultimately astrocytes - in the gray matter as well. The resultant human glial-chimeric mouse brains provide us a variety of hitherto unavailable opportunities for studying human glial cells and their progenitors in vivo, including their responses to injury and disease processes that cannot be adequately modeled in vitro. In the proposed experiments, we will use these mice to assess the effects of toxic demyelination on human GPCs in vivo, so as to identify their molecular responses to injury-induced mobilization and oligodendrocytic differentiation during compensatory remyelination. In particular, we will use phenotype-specific cell sorting and gene expression analysis, to define the responses of these xenografted human GPCs to demyelination in vivo. These data, the first ever obtained specifically from human GPCs during demyelination and remyelination in vivo, should afford us fundamental new insights into the signaling events associated with remyelination, and their potentially targetable points of regulatory control. To achieve that end, we propose the following Aims: In Aim 1, we will treat glial-chimeric mice with cuprizone so as to better understand the responses of human GPCs to demyelination, assessing their mobilization, differentiation, responses to repetitive induction, as well as their thresholds for mitotic senescence and the regulatory control thereof. In Aim 2, we will examine the expression patterns of human GPCs in vivo, sorting them from chimeric shiverer mice both at baseline and in response to demyelination, so as to define those genes and pathways differentially regulated during GPC mobilization and remyelination. In Aim 3 we will compare the responses of co-resident human and mouse GPCs to cuprizone demyelination, so as to identify those shared pathways likely to be high-value targets in drug development, as well as those species-specific pathways whose investigation in mice might not predict human therapeutic outcome. Together, these experiments promise to inform our efforts to define new strategies for treating demyelinating brain or spinal cord injury. In addition, the databases to be generated in the course of this work, as freely available resources to the field should prove catalytic in advancing our understanding of remyelination in vivo.