Culture medium for epithelial stem cells and organoids comprising the stem cells
AssigneeKoninklijke Nederlandse Akademie van Wetenschappen
InventorsToshiro Sato, Johannes Carolus Clevers
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Abstract
The invention relates to a method for culturing epithelial stem cells, isolated tissue fragments comprising the epithelial stem cells, or adenoma cells, and culturing the cells or fragments in the presence of a Bone Morphogenetic Protein (BMP) inhibitor, a mitogenic growth factor, and a Wnt agonist when culturing epithelial stem cells and isolated tissue fragments. The invention further relates to a cell culture medium comprising a BMP inhibitor, a mitogenic growth factor, and a Wnt agonist, to the use of the culture medium, and to crypt-villus organoids, gastric organoids and pancreatic organoids that are formed in the culture medium.
Claims35
An in vitro method for obtaining a crypt villus organoid or a colon crypt organoid, the method comprising:
- incubating an epithelial stem cell or an isolated tissue fragment comprising an epithelial stem cell with an extracellular matrix, and
- culturing the stem cell or isolated tissue fragment for a length of time sufficient to be able to observe formation of a crypt villus organoid or a colon crypt organoid, in an animal or human cell culture medium comprising:
- a Bone Morphogenetic Protein (BMP) inhibitor,
- between 5 and 500 ngram/ml of a mitogenic growth factor, and
- a Wnt agonist, and
- obtaining from the culture medium a crypt villus organoid or a colon crypt organoid.
An in vitro method for obtaining and/or culturing a colon crypt, the method comprising:
- culturing epithelial stem cells or isolated tissue fragments comprising epithelial stem cells in contact with an extracellular matrix for a length of time sufficient to be able to observe formation of a colon crypt, in a medium comprising Noggin, EGF, any one of R-spondin 1 through R-spondin 4 and/or Wnt3, B27, N2, and N-Acetylcysteine;
- and
- obtaining from the culture medium a colon crypt.
An in vitro method for obtaining a crypt villus organoid or colon crypt organoid, the method comprising:
- culturing epithelial stem cells or isolated tissue fragments comprising epithelial stem cells on an extracellular matrix for a length of time sufficient to be able to observe formation of a crypt villus organoid or a colon crypt organoid, in an animal or human cell culture medium comprising:
- a Bone Morphogenetic Protein (BMP) inhibitor;
- a Wnt agonist;
- and
- between 5 and 500 nanogram/ml of Epidermal Growth Factor (EGF);
- and
- obtaining from the culture medium a crypt villus organoid or a colon crypt organoid.
An in vitro method for obtaining a crypt villus organoid or a colon crypt organoid the method comprising:
- incubating an epithelial stem cell or an isolated tissue fragment comprising an epithelial stem cell with an extracellular matrix, wherein the epithelial stem cell is an adenoma cell, and
- culturing the stem cell or isolated tissue fragment for a length of time sufficient to be able to observe formation of a crypt villus organoid or a colon crypt organoid, in an animal or human cell culture medium comprising:
- a Bone Morphogenetic Protein (BMP) inhibitor, and
- between 5 and 500 ngram/ml of a mitogenic growth factor, and
- obtaining from the culture medium a crypt villus organoid or a colon crypt organoid.
Dependencies are inferred from the claim text.