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Spinal dI4 Interneuron Differentiation From Human Pluripotent Stem Cells
Spinal interneurons (INs) form intricate local networks in the spinal cord and regulate not only the ascending and descending nerve transduction but also the central pattern generator function. They are therefore potential therapeutic targets in spinal cord injury and diseases. In this study, we dev...
Autores principales: | , , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
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Frontiers Media S.A.
2022
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Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9026311/ https://www.ncbi.nlm.nih.gov/pubmed/35465095 http://dx.doi.org/10.3389/fnmol.2022.845875 |
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author | Xu, Jia Huang, Liang-Jiang Fang, Zhengyu Luo, Hong-Mei Chen, Yun-Qiang Li, Ya-Jie Gong, Chen-Zi Chen, Hong |
author_facet | Xu, Jia Huang, Liang-Jiang Fang, Zhengyu Luo, Hong-Mei Chen, Yun-Qiang Li, Ya-Jie Gong, Chen-Zi Chen, Hong |
author_sort | Xu, Jia |
collection | PubMed |
description | Spinal interneurons (INs) form intricate local networks in the spinal cord and regulate not only the ascending and descending nerve transduction but also the central pattern generator function. They are therefore potential therapeutic targets in spinal cord injury and diseases. In this study, we devised a reproducible protocol to differentiate human pluripotent stem cells (hPSCs) from enriched spinal dI4 inhibitory GABAergic INs. The protocol is designed based on developmental principles and optimized by using small molecules to maximize its reproducibility. The protocol comprises induction of neuroepithelia, patterning of neuroepithelia to dorsal spinal progenitors, expansion of the progenitors in suspension, and finally differentiation into mature neurons. In particular, we employed both morphogen activators and inhibitors to restrict or “squeeze” the progenitor fate during the stage of neural patterning. We use retinoic acid (RA) which ventralizes cells up to the mid-dorsal region, with cyclopamine (CYC), an SHH inhibitor, to antagonize the ventralization effect of RA, yielding highly enriched dI4 progenitors (90% Ptf1a(+), 90.7% Ascl1(+)). The ability to generate enriched spinal dI4 GABAergicINs will likely facilitate the study of human spinal IN development and regenerative therapies for traumatic injuries and diseases of the spinal cord. |
format | Online Article Text |
id | pubmed-9026311 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2022 |
publisher | Frontiers Media S.A. |
record_format | MEDLINE/PubMed |
spelling | pubmed-90263112022-04-23 Spinal dI4 Interneuron Differentiation From Human Pluripotent Stem Cells Xu, Jia Huang, Liang-Jiang Fang, Zhengyu Luo, Hong-Mei Chen, Yun-Qiang Li, Ya-Jie Gong, Chen-Zi Chen, Hong Front Mol Neurosci Molecular Neuroscience Spinal interneurons (INs) form intricate local networks in the spinal cord and regulate not only the ascending and descending nerve transduction but also the central pattern generator function. They are therefore potential therapeutic targets in spinal cord injury and diseases. In this study, we devised a reproducible protocol to differentiate human pluripotent stem cells (hPSCs) from enriched spinal dI4 inhibitory GABAergic INs. The protocol is designed based on developmental principles and optimized by using small molecules to maximize its reproducibility. The protocol comprises induction of neuroepithelia, patterning of neuroepithelia to dorsal spinal progenitors, expansion of the progenitors in suspension, and finally differentiation into mature neurons. In particular, we employed both morphogen activators and inhibitors to restrict or “squeeze” the progenitor fate during the stage of neural patterning. We use retinoic acid (RA) which ventralizes cells up to the mid-dorsal region, with cyclopamine (CYC), an SHH inhibitor, to antagonize the ventralization effect of RA, yielding highly enriched dI4 progenitors (90% Ptf1a(+), 90.7% Ascl1(+)). The ability to generate enriched spinal dI4 GABAergicINs will likely facilitate the study of human spinal IN development and regenerative therapies for traumatic injuries and diseases of the spinal cord. Frontiers Media S.A. 2022-04-08 /pmc/articles/PMC9026311/ /pubmed/35465095 http://dx.doi.org/10.3389/fnmol.2022.845875 Text en Copyright © 2022 Xu, Huang, Fang, Luo, Chen, Li, Gong and Chen. https://creativecommons.org/licenses/by/4.0/This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms. |
spellingShingle | Molecular Neuroscience Xu, Jia Huang, Liang-Jiang Fang, Zhengyu Luo, Hong-Mei Chen, Yun-Qiang Li, Ya-Jie Gong, Chen-Zi Chen, Hong Spinal dI4 Interneuron Differentiation From Human Pluripotent Stem Cells |
title | Spinal dI4 Interneuron Differentiation From Human Pluripotent Stem Cells |
title_full | Spinal dI4 Interneuron Differentiation From Human Pluripotent Stem Cells |
title_fullStr | Spinal dI4 Interneuron Differentiation From Human Pluripotent Stem Cells |
title_full_unstemmed | Spinal dI4 Interneuron Differentiation From Human Pluripotent Stem Cells |
title_short | Spinal dI4 Interneuron Differentiation From Human Pluripotent Stem Cells |
title_sort | spinal di4 interneuron differentiation from human pluripotent stem cells |
topic | Molecular Neuroscience |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9026311/ https://www.ncbi.nlm.nih.gov/pubmed/35465095 http://dx.doi.org/10.3389/fnmol.2022.845875 |
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