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Transcription Factor Programming of Human Pluripotent Stem Cells to Functionally Mature Astrocytes for Monocultures and Cocultures with Neurons

Quist, Ella LU orcid ; Ahlenius, Henrik LU and Canals, Isaac LU (2021) In Methods in Molecular Biology 2352. p.133-148
Abstract

Astrocytes are essential cells for normal brain functionality and have recently emerged as key players in many neurological diseases. However, the limited availability of human primary astrocytes for cell culture studies hinders our understanding of their physiology and precise role in disease development and progression. Here, we describe a detailed step-by-step protocol to rapidly and efficiently generate functionally mature induced astrocytes (iAs) from human embryonic and induced pluripotent stem cells (hES/iPSCs). Astrocyte induction is accomplished by ectopic lentiviral expression of two gliogenic transcription factors, Sox9 and Nfib. iAs exhibit morphology features as well as gene and protein expression similar to human mature... (More)

Astrocytes are essential cells for normal brain functionality and have recently emerged as key players in many neurological diseases. However, the limited availability of human primary astrocytes for cell culture studies hinders our understanding of their physiology and precise role in disease development and progression. Here, we describe a detailed step-by-step protocol to rapidly and efficiently generate functionally mature induced astrocytes (iAs) from human embryonic and induced pluripotent stem cells (hES/iPSCs). Astrocyte induction is accomplished by ectopic lentiviral expression of two gliogenic transcription factors, Sox9 and Nfib. iAs exhibit morphology features as well as gene and protein expression similar to human mature astrocytes and display important astrocytic functions, such as glutamate uptake, propagation of calcium waves, expression of various cytokines after stimulation, and support of synapse formation and function, making them suitable models for studying the role of astrocytes in health and disease. Moreover, we describe a procedure for cryopreservation of iAs for long-term storage or shipping. Finally, we provide the required information needed to set up cocultures with human induced neurons (iNs, also described in this book), generated from hES/iPSCs, to generate cocultures, allowing studies on astrocyte-neuron interactions and providing new insights in astrocyte-associated disease mechanisms.

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Please use this url to cite or link to this publication:
author
; and
organization
publishing date
type
Chapter in Book/Report/Conference proceeding
publication status
published
subject
keywords
Astrocyte-neuron cocultures, Astrocytes, Direct conversion, Reprogramming, Stem cells
host publication
Neural Reprogramming: Methods and Protocols
series title
Methods in Molecular Biology
volume
2352
pages
133 - 148
publisher
Humana Press
external identifiers
  • scopus:85111817182
  • pmid:34324185
ISSN
1064-3745
1940-6029
ISBN
978-1-0716-1600-0
978-1-0716-1601-7
DOI
10.1007/978-1-0716-1601-7_10
language
English
LU publication?
yes
additional info
Publisher Copyright: © 2021, Springer Science+Business Media, LLC, part of Springer Nature.
id
cdeae008-8a10-47a7-9ec8-e63cc162f75f
date added to LUP
2021-12-10 09:26:11
date last changed
2024-02-20 18:11:59
@inbook{cdeae008-8a10-47a7-9ec8-e63cc162f75f,
  abstract     = {{<p>Astrocytes are essential cells for normal brain functionality and have recently emerged as key players in many neurological diseases. However, the limited availability of human primary astrocytes for cell culture studies hinders our understanding of their physiology and precise role in disease development and progression. Here, we describe a detailed step-by-step protocol to rapidly and efficiently generate functionally mature induced astrocytes (iAs) from human embryonic and induced pluripotent stem cells (hES/iPSCs). Astrocyte induction is accomplished by ectopic lentiviral expression of two gliogenic transcription factors, Sox9 and Nfib. iAs exhibit morphology features as well as gene and protein expression similar to human mature astrocytes and display important astrocytic functions, such as glutamate uptake, propagation of calcium waves, expression of various cytokines after stimulation, and support of synapse formation and function, making them suitable models for studying the role of astrocytes in health and disease. Moreover, we describe a procedure for cryopreservation of iAs for long-term storage or shipping. Finally, we provide the required information needed to set up cocultures with human induced neurons (iNs, also described in this book), generated from hES/iPSCs, to generate cocultures, allowing studies on astrocyte-neuron interactions and providing new insights in astrocyte-associated disease mechanisms.</p>}},
  author       = {{Quist, Ella and Ahlenius, Henrik and Canals, Isaac}},
  booktitle    = {{Neural Reprogramming: Methods and Protocols}},
  isbn         = {{978-1-0716-1600-0}},
  issn         = {{1064-3745}},
  keywords     = {{Astrocyte-neuron cocultures; Astrocytes; Direct conversion; Reprogramming; Stem cells}},
  language     = {{eng}},
  pages        = {{133--148}},
  publisher    = {{Humana Press}},
  series       = {{Methods in Molecular Biology}},
  title        = {{Transcription Factor Programming of Human Pluripotent Stem Cells to Functionally Mature Astrocytes for Monocultures and Cocultures with Neurons}},
  url          = {{http://dx.doi.org/10.1007/978-1-0716-1601-7_10}},
  doi          = {{10.1007/978-1-0716-1601-7_10}},
  volume       = {{2352}},
  year         = {{2021}},
}