Patient-derived iPS cells as model systems for AxD

摘要 亚历山大病（AxD）是由gfap基因突变引起的星形胶质细胞的原发性疾病。 AxD突变如何导致蛋白质聚集和星形胶质细胞功能障碍以及突变如何导致蛋白质聚集和星形胶质细胞功能障碍 表达GFAP的星形胶质细胞导致神经元变性仍然是未知的。从进化的角度来看， 星形胶质细胞在人脑功能中发挥着越来越重要和复杂的作用。所以 直接检查AxD患者星形胶质细胞的能力不仅可以补充现有的模型， 还揭示了人类星形胶质细胞在AxD发病机制中的潜在独特方面。我们已经建立了诱导 来自AxD患者的具有三种不同突变的多能干细胞（iPSC）通过封闭 与梅辛实验室合作（项目3）。我们还制定了一项可复制的战略， 引导hPSC富集星形胶质细胞。我们的初步研究表明，在AxD中存在RF 患者的星形胶质细胞在培养和移植到小鼠脑中后，强调了 在我们的iPSC系统中的关键AxD病理学的重演。然而，来自AxD的星形胶质细胞 患者表现出与非AxD个体相当的GFAP水平，表明GFAP可能不 AxD早期的增加和GFAP蛋白的简单增加可能不是主要的触发因素 用于人类细胞中RF形成和星形胶质细胞功能障碍，正如为模型动物提出的那样。我们的能力 从AxD患者中产生富集的星形胶质细胞也导致发现AxD星形胶质细胞 已经改变了GFAP-d/a亚型的比例，这导致了“重新发现”类似的比例变化， 转基因小鼠我们将使用先进的基因编辑技术来确定GFAP的变化是否 同种型介导GFAP突变对蛋白质聚集和/或星形胶质细胞功能障碍的影响。 通过神经元-星形胶质细胞共培养和神经移植，我们将确定AxD星形胶质细胞是否以及如何 导致神经元退化。作为程序项目的一部分，我们将验证AxD的修改器学习 从AxD患者星形胶质细胞/神经元的动物模型中，为未来的研究奠定了基础。 翻译.

ABSTRACT Alexander disease (AxD) is a primary disease of astrocytes caused by mutations in the gfap gene. How AxD mutations lead to protein aggregation and astrocyte dysfunction as well as how mutant GFAP-expressing astrocytes result in neuronal degeneration remain unknown. Evolutionarily, astrocytes play increasingly more important and complex roles in human brain functions. Hence, the ability to directly examine AxD patients' astrocytes will not only complement the existing models but also reveal potential unique aspects of human astrocytes in AxD pathogenesis. We have built induced pluripotent stem cells (iPSCs) from AxD patients with three different mutations through close collaboration with the Messing lab (project 3). We have also developed a reproducible strategy to guide hPSCs to enriched astrocytes. Our preliminary study revealed the presence of RFs in AxD patients' astrocytes in culture and following transplantation into the mouse brain, highlighting the recapitulation of key AxD pathology in our iPSC system. However, astrocytes derived from AxD patients exhibit comparable GFAP levels as non-AxD individuals, suggesting that GFAP may not increase at early stages of AxD and a simple increase in GFAP protein might not be the major trigger for RF formation and astrocyte dysfunction in human cells as proposed for model animals. Our ability to produce enriched astrocytes from AxD patients has also led to the discovery that AxD astrocytes have altered ratio of GFAP-d/a isoforms, which has led to the “rediscovery” of a similar ratio change in transgenic mice. We will use advanced gene editing technology to determine if change of GFAP isoforms mediates the effects of gfap mutations on protein aggregation and/or astrocyte dysfunction. By neuron-astrocyte co-culture and neural transplantation, we will determine if and how AxD astrocytes cause neuronal degeneration. As part of the program project, we will validate modifiers of AxD learned from animal models in our AxD patients' astrocytes/neurons, setting up the foundation for future translation.