Method of Differentiating Stem Cells
First Claim
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1. A method for differentiating stem cells into cells expressing C-peptide comprising the steps of:
- (a) culturing stem cells in the presence of at least one extracellular matrix;
(b) culturing the stem cells obtained from step (a) in at least one first medium comprising at least BMP4 and Activin A to obtain definitive endoderm cells;
(c) culturing the cells obtained from step (b) in at least one second medium comprising at least Activin A;
(d) culturing the cells obtained from step (c) in at least one third medium comprising at least one fibroblast growth factor (FGF), retinoic acid (RA) and at least one inhibitor of hedgehog signalling to obtain cells expressing Pdx-1; and
(e) culturing the cells obtained from step (d) in at least one fourth medium comprising at least one inhibitor of notch signalling to differentiate cells expressing Pdx-1 to C-peptide producing cells.
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Abstract
There is provided an improved efficient method for differentiating stem cells into pancreatic endoderm cells and pancreatic hormone expressing and secreting cells which express Pdx-1 and C-peptide. The invention further provides screening methods for detecting factors of interest that will affect the differentiation of the stem cells into pancreatic endoderm cells.
11 Citations
18 Claims
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1. A method for differentiating stem cells into cells expressing C-peptide comprising the steps of:
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(a) culturing stem cells in the presence of at least one extracellular matrix; (b) culturing the stem cells obtained from step (a) in at least one first medium comprising at least BMP4 and Activin A to obtain definitive endoderm cells; (c) culturing the cells obtained from step (b) in at least one second medium comprising at least Activin A; (d) culturing the cells obtained from step (c) in at least one third medium comprising at least one fibroblast growth factor (FGF), retinoic acid (RA) and at least one inhibitor of hedgehog signalling to obtain cells expressing Pdx-1; and (e) culturing the cells obtained from step (d) in at least one fourth medium comprising at least one inhibitor of notch signalling to differentiate cells expressing Pdx-1 to C-peptide producing cells. - View Dependent Claims (2, 3, 4, 5, 6, 7, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18)
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8. The method according to claim, wherein the inhibitor of hedgehog signalling is SANT-1.
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