期刊
NATURE PROTOCOLS
卷 15, 期 11, 页码 -出版社
NATURE RESEARCH
DOI: 10.1038/s41596-020-0395-4
关键词
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资金
- Fellowship from Alfonso Martin Escudero Foundation (Madrid, Spain)
- I Plan Propio of the University of Malaga (Spain)
- CIBERNED [CB06/05/1116]
- IWT-iPSCAF grant [150031]
- KUL-PF Stem Cells
- Instituto de Salud Carlos III (ISCiii) of Spain
- FEDER funds from the European Union [PI18/01557]
- Junta de Andalucia
- Programa Operativo FEDER
- Consejeria de Salud of Junta de Andalucia [PI-0276-2018]
- Progressive MS Alliance (PMSA) [PA-1604-08492]
- INSERM
- ICM grants
- European Commission under the Seventh Framework Program of the European Union [291730, TAHUB-II-107]
- ICM animal, genomic
Oligodendrocytes (OLs) are responsible for myelin production and metabolic support of neurons. Defects in OLs are crucial in several neurodegenerative diseases including multiple sclerosis (MS) and amyotrophic lateral sclerosis (ALS). This protocol describes a method to generate oligodendrocyte precursor cells (OPCs) from human pluripotent stem cells (hPSCs) in only similar to 20 d, which can subsequently myelinate neurons, both in vitro and in vivo. To date, OPCs have been derived from eight different hPSC lines including those derived from patients with spontaneous and familial forms of MS and ALS, respectively. hPSCs, fated for 8 d toward neural progenitors, are transduced with an inducible lentiviral vector encoding for SOX10. The addition of doxycycline for 10 d results in >60% of cells being O4-expressing OPCs, of which 20% co-express the mature OL marker myelin basic protein (MBP). The protocol also describes an alternative for viral transduction, by incorporating an inducible SOX10 in the safe harbor locus AAVS1, yielding similar to 100% pure OPCs. O4(+) OPCs can be purified and either cryopreserved or used for functional studies. As an example of the type of functional study for which the derived cells could be used, O4(+) cells can be co-cultured with maturing hPSC-derived neurons in 96/384-well-format plates, allowing the screening of pro-myelinating compounds.
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