用于研究肌萎缩侧索硬化症的从人诱导多能干细胞诱导生成人脊髓运动神经元的方案。

Protocol for the induction of human spinal motor neurons from human induced pluripotent stem cells for studying amyotrophic lateral sclerosis.

作者信息

Setsu Selena, Morimoto Satoru, Nakamura Shiho, Ozawa Fumiko, Okano Hideyuki

机构信息

Keio University Regenerative Medicine Research Center, Kanagawa 210-0821, Japan; Laboratory of RNA Function, Institute for Quantitative Biosciences, The University of Tokyo, Tokyo 113-0032, Japan; Department of Computational Biology and Medical Sciences, Graduate School of Frontier Sciences, The University of Tokyo, Tokyo 277-0822, Japan.

Keio University Regenerative Medicine Research Center, Kanagawa 210-0821, Japan; Division of Neurodegenerative Disease Research, Tokyo Metropolitan Institute for Geriatrics and Gerontology, Tokyo 173-0015, Japan.

出版信息

STAR Protoc. 2025 Aug 5;6(3):104016. doi: 10.1016/j.xpro.2025.104016.

DOI:10.1016/j.xpro.2025.104016

PMID:40773352

原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC12348247/

Abstract

Here, we present a protocol for inducing spinal lower motor neurons (LMNs) from human induced pluripotent stem cells (iPSCs). We describe steps for preparation of a chemically induced transitional state (CTraS), transduction with Sendai virus, and LMN differentiation and maintenance. We then detail procedures for live imaging for single-cell-based survival analysis and neurite length of LMNs using BioStation and immunocytochemistry for induction efficiency check. This protocol is optimized for amyotrophic lateral sclerosis (ALS) research and large-scale screening. For complete details on the use and execution of this protocol, please refer to Setsu et al..

摘要

在此，我们展示了一种从人诱导多能干细胞（iPSC）诱导生成脊髓下运动神经元（LMN）的方案。我们描述了化学诱导过渡状态（CTraS）的制备步骤、仙台病毒转导以及LMN的分化和维持。然后，我们详细说明了使用BioStation进行基于单细胞的存活分析和LMN神经突长度的实时成像程序，以及用于诱导效率检查的免疫细胞化学方法。该方案针对肌萎缩侧索硬化症（ALS）研究和大规模筛选进行了优化。有关此方案的使用和执行的完整详细信息，请参考Setsu等人的研究。