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Cited by 157Hearing, Cochlea, Tinnitus, GeneticsPluripotent Stem Cells ResearchNeurogenesis and neuroplasticity mechanisms

3D mouse embryonic stem cell culture for generating inner ear organoids

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Koehler, Karl R, Hashino, Eri·2014·Nature Protocols
DOI 10.1038/nprot.2014.100PMC PMC6453571
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Abstract

This protocol describes a culture system in which inner-ear sensory tissue is produced from mouse embryonic stem (ES) cells under chemically defined conditions. This model is amenable to basic and translational investigations into inner ear biology and regeneration. In this protocol, mouse ES cells are aggregated in 96-well plates in medium containing extracellular matrix proteins to promote epithelialization. During the first 14 d, a series of precisely timed protein and small-molecule treatments sequentially induce epithelia that represent the mouse embryonic non-neural ectoderm, preplacodal ectoderm and otic vesicle epithelia. Ultimately, these tissues develop into cysts with a pseudostratified epithelium containing inner ear hair cells and supporting cells after 16-20 d. Concurrently, sensory-like neurons generate synapse-like structures with the derived hair cells. We have designated the stem cell-derived epithelia harboring hair cells, supporting cells and sensory-like neurons as inner ear organoids. This method provides a reproducible and scalable means to generate inner ear sensory tissue in vitro.

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  • In the 3D mESC culture, a heterogeneous population of mesenchymal and neural cells is also generated.

    “In the process of culture, a heterogeneous population of mesenchymal and neural cells is also generated (Koehler and Hashino, 2014; Koehler et al., 2013).”

    From Hair Follicle Development in Mouse Pluripotent Stem Cell-Derived Skin Organoids (2018) · cites this paper as Koehler and Hashino, 2014

  • A 3D mouse embryonic stem cell culture can produce cranial surface epithelia (non-neural ectoderm), a precursor tissue of the skin epidermis.

    “Recently, we developed a three-dimensional (3D) mouse embryonic stem cell (mESC) culture capable of producing cranial surface epithelia (also known as non-neural ectoderm), a precursor tissue of the skin epidermis.”

    From Hair Follicle Development in Mouse Pluripotent Stem Cell-Derived Skin Organoids (2018) · cites this paper as Koehler and Hashino, 2014

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Cited by

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Authors
Koehler, K. R., & Hashino, E.
Affiliations
Indiana University Indianapolis (US)
Corresponding author
  • Koehler, Karl R
  • Hashino, Eri
ORCID
Venue
Nature Protocols
Publisher
Springer Science and Business Media LLC
Published
2014-05-01
Volume
9
Issue
6
Pages
1229-1244
ISSN
  • 1750-2799 (ISSN-L)
  • 1754-2189
DOI
10.1038/nprot.2014.100
PMC
PMC6453571
OpenAlex
W2006591300
Cited by
157
References
5
Citations by year
2026: 12 · 2025: 18 · 2024: 12 · 2023: 13 · 2022: 11 · 2021: 15 · 2020: 16 · 2019: 9 · 2018: 10 · 2017: 13 · 2016: 11 · 2015: 14 · 2014: 1 · 2013: 2
FWCI
8.02
Type
journal article
Language
en
Open access
Licence
Locations
  • Nature Protocols (publishedVersion)
  • PubMed (publishedVersion)
  • Europe PMC (PubMed Central) (acceptedVersion)
  • PubMed Central (acceptedVersion)
  • IUScholarWorks (Indiana University) (submittedVersion)
  • PMC
Funders
  • National Institute on Deafness and Other Communication Disorders (R21DC012617, RC1DC010706, R01DC013294)
  • National Center for Research Resources (TL1RR025759)
Topics
  • Life Sciences › Neuroscience › Sensory Systems › Hearing, Cochlea, Tinnitus, Genetics
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Pluripotent Stem Cells Research
  • Life Sciences › Neuroscience › Developmental Neuroscience › Neurogenesis and neuroplasticity mechanisms
Keywords
inner ear organoids, mouse embryonic stem cells, 3D organoid culture, hair cells, supporting cells, otic vesicle, inner ear regeneration, chemically defined culture, extracellular matrix, sensory epithelium
MeSH terms
Animals, Cell Differentiation, Hair Cells, Auditory, Ear, Inner, Organoids, Extracellular Matrix Proteins, Cell Culture Techniques, Mice, Embryonic Stem Cells

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