Differentiation and expansion of endothelial cells from pluripotent stem cells and the in vitro formation of vasculature like structures
First Claim
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1. A method for producing an in vitro cell culture comprising endothelial cells derived from in vitro differentiated pluripotent stein cells and smooth muscle cells derived from in vitro differentiated pluripotent stem cells or derived from pericytes, the method comprising the following steps:
- (a) culturing pluripotent stem cells in defined medium comprising ActivinA, BMP4, VEGF, and a canonical WNT ligand or GSK3 inhibitor, to produce a culture comprising differentiated cells;
(b) culturing the cells obtained in step (a) in defined medium containing VEGF and a TGF{beta} signaling inhibitor to produce endothelial cells; and
(c) culturing the endothelial cells obtained in step (b) with smooth muscle cells derived from in vitro differentiated pluripotent stem cells or derived from pericytes.
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Abstract
The disclosure is concerned among others with means and methods for obtaining endothelial cells and to means and methods for in vitro cell culture comprising endothelial cells and pericytes and/or smooth muscle cells derived from the pericytes. The endothelial cells or the pericytes and/or smooth muscle cells, or both, are preferably derived from in vitro differentiated pluripotent stem cells.
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Citations
17 Claims
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1. A method for producing an in vitro cell culture comprising endothelial cells derived from in vitro differentiated pluripotent stein cells and smooth muscle cells derived from in vitro differentiated pluripotent stem cells or derived from pericytes, the method comprising the following steps:
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(a) culturing pluripotent stem cells in defined medium comprising ActivinA, BMP4, VEGF, and a canonical WNT ligand or GSK3 inhibitor, to produce a culture comprising differentiated cells; (b) culturing the cells obtained in step (a) in defined medium containing VEGF and a TGF{beta} signaling inhibitor to produce endothelial cells; and (c) culturing the endothelial cells obtained in step (b) with smooth muscle cells derived from in vitro differentiated pluripotent stem cells or derived from pericytes. - View Dependent Claims (2, 3, 4, 5, 6, 8, 9, 10)
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7. A method of producing an in vitro cell culture comprising endothelial cells and smooth muscle cells, wherein the endothelial cells are able to integrate into a vascular network in vivo, the method comprising:
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(a) culturing pluripotent stem cells in a defined medium comprising ActivinA, BMP4, VEGF, and a canonical WNT ligand or GSK3 inhibitor, to produce a culture comprising differentiated cells; (b) culturing the cells of step (a) in defined medium containing VEGF and a TGF(beta) signaling inhibitor to produce endothelial cells; and (c) culturing the endothelial cells obtained in step (b) with smooth muscle cells.
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11. A method for producing an in vitro cell culture comprising purified endothelial cells and purified smooth muscle cells, wherein the cell culture comprises a capillary network comprising the endothelial cells and the smooth muscle cells, the method comprising:
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deriving endothelial cells from in vitro differentiated induced pluripotent stem cells (iPSCs) and purifying the endothelial cells, deriving smooth muscle cells from in vitro differentiated induced pluripotent stem cells or from pericytes that are derived from in vitro differentiated induced pluripotent stem cells and purifying the smooth muscle cells, and culturing the purified endothelial cells together with the purified smooth muscle cells. - View Dependent Claims (12, 13, 14, 15, 16, 17)
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Specification