In Vitro Production of Foregut Stem Cells
AssigneeCambridge Enterprise Limited
InventorsLudovic Vallier, Nicholas Hannan
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Abstract
This invention relates to the differentiation of pluripotent cells (PSCs) into foregut stem cells (FSCs) using a definitive endoderm induction medium comprising a TGFfi ligand, fibroblast growth factor (FGF), bone morphogenetic protein (BMP) and a PI3K inhibitor to differentiate the pluripotent cells into definitive endoderm cells and a foregut induction medium comprising a TGFβ ligand to differentiate the definitive endoderm cells into foregut stem cells (FSCs). Methods of differentiation, populations of foregut stem cells, culture media and kits are provided.
Claims39
A method for producing a population of foregut stem cells (FSCs) comprising:
- i) providing a population of definitive endoderm cells (DECs),
- ii) culturing the DECs in a foregut induction medium comprising a TGFβ ligand to produce a population of foregut stem cells (FSCs).
A method according to any one of the preceding claims wherein the population of DECs is a homogeneous population or a heterogeneous population.
A method according to any one of the preceding claims wherein the DECs express one or more of the following endoderm associated markers:
- Sox17, foxA2, GSC, Mixl1, Lhx1, CXCR4, GATA4, eomesodermin (EOMES), Mixl1, HNF-3 beta, Cerberus, OTX4, goosecoid, C-kit, CD99, and Hex.
A method according to any one of the preceding claims wherein the foregut induction medium consists of a chemically defined nutrient medium supplemented with a TGFβ ligand.
A method according to any one of the preceding claims wherein the TGFβ ligand is activin.
A method according to any one of the preceding claims wherein the FSCs express one or more of the following markers:
- SOX2, HHEX, HOXA3, HNF4α, SOX17, CXCR4, EpCAM, HNF1β, GATA4, Cer, HNF6, and HNF1bet.
A method according to any one of the preceding claims comprising culturing the population of FSCs in an FSC maintenance medium comprising an effective amount of TGFβ ligand, fibroblast growth factor (FGF), bone morphogenetic protein (BMP), HGF, EGF, and heparin.
A method according to any one of the preceding claims wherein the FSCs are multipotent and self-renewing in vitro.
A method according to any one of the preceding claims wherein the FSCs are differentiated into endoderm cells.
A method of producing pulmonary progenitor cells comprising;
- i) providing a population of FSCs
- ii) culturing the population of FSCs in a first pulmonary induction medium comprising RA and FGF;
- iii) culturing the cells from step i) in a second pulmonary induction medium comprising FGF and HGF;
- thereby producing a population of pulmonary progenitor cells.
A method for maintaining a population of foregut stem cells (FSCs), the method comprising:
- i) providing a population of foregut stem cells (FSCs);
- ii) culturing the population of FSCs in an FSC maintenance medium comprising a TGFβ ligand, fibroblast growth factor (FGF), bone morphogenetic protein (BMP), HGF, EGF, and Heparin.
A kit for production of FSCs comprising;
- a foregut induction medium consisting of a chemically defined nutrient medium supplemented with a TGFβ ligand.
Use of a foregut induction medium for the production of a population of FSCs,
- said foregut induction medium consisting of a chemically defined nutrient medium supplemented with a TGFβ ligand.
Use of first and a second pulmonary induction media for the production of a population of pulmonary progenitor cells, the first pulmonary induction medium consisting of a chemically defined nutrient medium supplemented with retinoic acid and fibroblast growth factor (FGF) and the second pulmonary induction medium consists of a chemically defined nutrient medium supplemented with hepatocyte growth factor (HGF) and fibroblast growth factor (FGF).
Dependencies are inferred from the claim text.