Abstract
The human stomach contains two primary domains: the corpus, which contains the fundic epithelium, and the antrum. Each of these domains has distinct cell types and functions, and therefore each presents with unique disease pathologies. Here, we detail two protocols to differentiate human pluripotent stem cells (hPSCs) into human gastric organoids (hGOs) that recapitulate both domains. Both protocols begin with the differentiation of hPSCs into definitive endoderm (DE) using activin A, followed by the generation of free-floating 3D posterior foregut spheroids using FGF4, Wnt pathway agonist CHIR99021 (CHIR), BMP pathway antagonist Noggin, and retinoic acid. Embedding spheroids in Matrigel and continuing 3D growth in epidermal growth factor (EGF)-containing medium for 4 weeks results in antral hGOs (hAGOs). To obtain fundic hGOs (hFGOs), spheroids are additionally treated with CHIR and FGF10. Induced differentiation of acid-secreting parietal cells in hFGOs requires temporal treatment of BMP4 and the MEK inhibitor PD0325901 for 48 h on protocol day 30. In total, it takes ~34 d to generate hGOs from hPSCs. To date, this is the only approach that generates functional human differentiated gastric cells de novo from hPSCs.
Publication types
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Research Support, N.I.H., Extramural
MeSH terms
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Activins / pharmacology
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Benzamides / pharmacology
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Carrier Proteins / pharmacology
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Cell Culture Techniques*
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Cell Differentiation
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Collagen / chemistry
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Culture Media / chemistry
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Culture Media / pharmacology
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Diphenylamine / analogs & derivatives
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Diphenylamine / pharmacology
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Drug Combinations
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Endoderm / cytology*
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Endoderm / drug effects
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Endoderm / metabolism
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Epidermal Growth Factor / pharmacology
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Epithelial Cells / cytology*
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Epithelial Cells / drug effects
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Epithelial Cells / metabolism
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Fibroblast Growth Factor 10 / pharmacology
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Fibroblast Growth Factor 4 / pharmacology
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Gastric Fundus / cytology*
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Gastric Fundus / metabolism
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Humans
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Laminin / chemistry
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Organ Specificity
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Organoids / cytology*
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Organoids / drug effects
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Organoids / metabolism
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Pluripotent Stem Cells / cytology*
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Pluripotent Stem Cells / drug effects
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Pluripotent Stem Cells / metabolism
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Proteoglycans / chemistry
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Pyloric Antrum / cytology*
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Pyloric Antrum / metabolism
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Pyridines / pharmacology
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Pyrimidines / pharmacology
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Tretinoin / pharmacology
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Wnt Signaling Pathway / drug effects
Substances
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Benzamides
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Carrier Proteins
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Chir 99021
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Culture Media
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Drug Combinations
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FGF10 protein, human
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FGF4 protein, human
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Fibroblast Growth Factor 10
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Fibroblast Growth Factor 4
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Laminin
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Proteoglycans
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Pyridines
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Pyrimidines
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activin A
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Activins
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matrigel
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noggin protein
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Tretinoin
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Epidermal Growth Factor
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mirdametinib
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Collagen
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Diphenylamine