Differentiating Human Embryonic Stem Cells into Oligodendrocytes

Abstract

Source: Assetta, B., et al. Generation of Human Neurons and Oligodendrocytes from Pluripotent Stem Cells for Modeling Neuron-Oligodendrocyte Interactions. J. Vis. Exp (2020).

This video demonstrates the process of generating oligodendrocyte precursor cells (OPCs) from human embryonic stem cells, followed by the maturation of OPCs to oligodendrocytes. This process of generating oligodendrocytes opens numerous avenues to improve research and treatment of neurodegenerative diseases, including Alzheimer's.

Protocol

1. Human oligodendrocyte precursor cell (OPC) induction from pluripotent stem cells and oligodendrocyte maturation

1. Neural Progenitor Cell (NPC) generation: monolayer protocol (~7 days). See Figure 1A for the flow diagram.
   1. Use commercially available H1 human ES cells at the passage of 52 (see Table of Materials). Culture the cells on extracellular matrix solution-coated 6-well plates (~0.5 mg of matrix solution per 6-well plate; see Table of Materials) using ES cell maintenance medium (see Table of Materials) and incubate the plates at 37 °C with 5% CO₂.
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Results

Figure 1: iOPC (induced oligodendrocyte precursor cell) generation and iOL (induced oligodendrocyte) maturation. (A) Flow diagram of iOPC and iOL generation. (B) Representative bright field and immunofluorescence image.......

Materials

List of materials used in this article

Name	Company	Catalog Number	Comments
Accutase	STEMCELL Technologies	7920
B27 supplement	ThermoFisher	17504044
bFGF	ThermoFisher	PHG 0266
cAMP	MilliporeSigma	A9501
Clemastine	MilliporeSigma	SML0445
DMEM/F12 medium	STEMCELL Technologies	36254
DMSO	ThermoFisher	D12345
Fetal Bovine Serum	ScienCell	10
H1 human ES cells	WiCell	WA01
Matrigel	Corning	354234
mTeSR plus	STEMCELL Technologies	5825
N2 supplement	ThermoFisher	17502001
Neurobasal A medium	ThermoFisher	10888-022
Non Essential Amino Acids	ThermoFisher	11140-050
PDGF-AA	R&D Systems	221-AA-010
pMDLg/pRRE	Addgene	12251
Polybrene	MilliporeSigma	TR-1003-G
pRSV-REV	Addgene	12253
ROCK Inhibitor Y-27632	STEMCELL Technologies	72302
SAG	Tocris	4366
STEMdiff Neural Progenitor Freezing Media	STEMCELL Technologies	5838
STEMdiff SMADi Neural Induction Kit	STEMCELL Technologies	8581
T3 triiodothyronine	MilliporeSigma	T6397
Tempo-iOlogo: Human iPSC-derived OPCs	Tempo BioScience	SKU102
TetO-Ng2-Puro	Addgene	52047
VSV-G	Addgene	12259