Derivation of functional neurons from induced pluripotent stem cells using a simple neuromesodermal progenitor generation and rapid spinal cord neuron differentiation process.

IF 3.4 3区 生物学 Q3 CELL BIOLOGY
Selinay Şenkal-Turhan, Ezgi Bulut-Okumuş, Fikrettin Şahin, Yavuz Yavuz, Bayram Yılmaz, Hatice Burcu Şişli, Sadık Kalaycı, Hüseyin Buğra Özgün, Zehra Ömeroğlu Ulu, Pınar Akkuş Süt, Ayşegül Doğan
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引用次数: 0

Abstract

To generate spinal cord neurons from pluripotent stem cells via neuromesodermal progenitors (NMPs) is not only an important step for regenerative purposes but also required for human developmental research. This study describes a protocol to obtain spinal cord neurons in culture using induced pluripotent stem-cell-derived NMPs. The protocol starts with a 3D culture of NMPs and continues with the transfer of 3D NMPs to monolayer culture in which retinoic acid and sonic hedgehog pathways were triggered sequentially. The established protocol enabled generation of spinal cord neurons with active calcium signaling, electrophysiological activity, axon elongation capacity, and synaptic vesicle trafficking. The expression profile of marker proteins, including β-Tubulin, NeuroD1, Pax6, NeuN, Mnx-1, Isl1, Isl2, Map2, NF, Sox2 was detected to explore the production of developmental regulatory transcription factors and terminal differentiation markers in a time-dependent manner. Cells during differentiation process acquired a fully neural phenotype, which was confirmed by RNA sequencing at the molecular level. The protein expression profile showed neural differentiation induction pathways based on LS-MS/MS analysis. Since NMPs differentiate into spinal cord neuron cells at the developmental stage, the results of this study highlight the further potential of NMP-derived spinal cord neurons in disease modeling and treatment in the clinics.

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来源期刊
Human Cell
Human Cell CELL BIOLOGY-
CiteScore
5.90
自引率
2.30%
发文量
176
审稿时长
4.5 months
期刊介绍: Human Cell is the official English-language journal of the Japan Human Cell Society. The journal serves as a forum for international research on all aspects of the human cell, encompassing not only cell biology but also pathology, cytology, and oncology, including clinical oncology. Embryonic stem cells derived from animals, regenerative medicine using animal cells, and experimental animal models with implications for human diseases are covered as well. Submissions in any of the following categories will be considered: Research Articles, Cell Lines, Rapid Communications, Reviews, and Letters to the Editor. A brief clinical case report focusing on cellular responses to pathological insults in human studies may also be submitted as a Letter to the Editor in a concise and short format. Not only basic scientists but also gynecologists, oncologists, and other clinical scientists are welcome to submit work expressing new ideas or research using human cells.
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