为肌萎缩侧索硬化症和其他神经退行性疾病研究生成人类诱导多能干细胞(hiPSC)衍生星形胶质细胞

Katarina Stoklund Dittlau, Abinaya Chandrasekaran, Kristine Freude, L. Van Den Bosch
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引用次数: 0

摘要

星形胶质细胞在肌萎缩性脊髓侧索硬化症(ALS)等神经退行性疾病中的重要作用日益得到认可。在 ALS 中,星形胶质细胞从提供神经元平衡支持的主要功能转变为反应性和毒性作用,这在总体上导致了神经元毒性和细胞死亡。目前,我们对这些过程的了解还不全面,因此需要在人类背景下建立高效、可重复的模型系统,以了解和治疗毒性星形胶质细胞反应,为未来的治疗提供选择。在这里,我们提出了一种高效、直接的方案,利用基于小分子的分化方案生成人类诱导多能干细胞(hiPSC)衍生的星形胶质细胞。经过最初的 25 天,hiPSC 分化为星形胶质细胞,并成熟 4 周以上。在分化和成熟过程中,hiPSC 衍生的星形胶质细胞可在每个阶段进行低温保存。这就为该方案提供了方便的暂停和细胞库机会,从而限制了从 hiPSCs 开始的连续性。该方案已被证明在 ALS 研究中很有价值,但也可适用于任何对星形胶质细胞感兴趣的研究领域。主要特点 - 本方案需要预先具备培养 hiPSC 的经验,才能取得成功。- 该方案依赖于小分子分化方案和简单易学的方法,可在多个时间点暂停。- 该方案每次分化可产生 >50 × 106 个星形胶质细胞,每次分化均可低温保存,确保大规模实验产出。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
Generation of Human Induced Pluripotent Stem Cell (hiPSC)-Derived Astrocytes for Amyotrophic Lateral Sclerosis and Other Neurodegenerative Disease Studies
Astrocytes are increasingly recognized for their important role in neurodegenerative diseases like amyotrophic lateral sclerosis (ALS). In ALS, astrocytes shift from their primary function of providing neuronal homeostatic support towards a reactive and toxic role, which overall contributes to neuronal toxicity and cell death. Currently, our knowledge on these processes is incomplete, and time-efficient and reproducible model systems in a human context are therefore required to understand and therapeutically modulate the toxic astrocytic response for future treatment options. Here, we present an efficient and straightforward protocol to generate human induced pluripotent stem cell (hiPSC)-derived astrocytes implementing a differentiation scheme based on small molecules. Through an initial 25 days, hiPSCs are differentiated into astrocytes, which are matured for 4+ weeks. The hiPSC-derived astrocytes can be cryopreserved at every passage during differentiation and maturation. This provides convenient pauses in the protocol as well as cell banking opportunities, thereby limiting the need to continuously start from hiPSCs. The protocol has already proven valuable in ALS research but can be adapted to any desired research field where astrocytes are of interest. Key features • This protocol requires preexisting experience in hiPSC culturing for a successful outcome. • The protocol relies on a small molecule differentiation scheme and an easy-to-follow methodology, which can be paused at several time points. • The protocol generates >50 × 106 astrocytes per differentiation, which can be cryopreserved at every passage, ensuring a large-scale experimental output.
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