Human pluripotent stem cell-based screening for smooth muscle cell differentiation and disease
AssigneeWisconsin Alumni Research Foundation (WARF)
InventorsJames A. Thomson, Jue Zhang
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Abstract
Methods of using a small molecule MYH11 agonist to inhibit intimal hyperplasia and to maintain a contractile phenotype in vitro and in vivo are described. Also described herein are methods for generating human contractile smooth muscle cells from human pluripotent stem cells under defined conditions in the presence of the small molecule MYH11 agonist.
Claims17
A tissue-engineered blood vessel comprising an isolated population of contractile smooth muscle cells that does not undergo contractile-to-synthetic phenotype switching, wherein the isolated population of contractile smooth muscle cells is obtained by a method comprising:
- (1) obtaining smooth muscle cell (SMC) progenitor cells;
- and
- (2) culturing the SMC progenitor cells in a culture medium comprising a myosin heavy chain 11 (MYH11) agonist, wherein the MYH11 agonist is RepSox, until a cell population comprising MYH11+, alpha-actin 2 positive (ACTA2+), smoothelin positive (SMTN+), calponin1 positive (CNN1+), and transgelin positive (TAGLN+) contractile smooth muscle cells is obtained, and
- wherein the SMC progenitor cells are obtained by a method selected from the group consisting of:
- (a) a first method comprising:
- (i) culturing mesoderm cells under conditions and for a time sufficient to obtain a population of cells expressing MEOX1;
- (ii) culturing the population of cells expressing MEOX1 under conditions and for a time sufficient to suppress MEOX1 expression;
- and
- (iii) culturing the population of cells from step (ii) under conditions and for a time sufficient to obtain a population of SMC progenitor cells, and
- (b) a second method comprising:
- (i) culturing mesoderm cells in chemically defined medium that comprises a fibroblast growth factor (FGF) or a vascular endothelial growth factor (VEGF) for a period of time sufficient to induce SMC progenitor cells;
- and
- (ii) culturing the population of cells from step (i) under conditions and for a time sufficient to obtain a population of SMC progenitor cells.
A tissue-engineered construct comprising an isolated population of contractile smooth muscle cells that does not undergo contractile-to-synthetic phenotype switching, wherein the isolated population of contractile smooth muscle cells is obtained by a method comprising:
- (1) obtaining smooth muscle cell (SMC) progenitor cells;
- and
- (2) culturing the SMC progenitor cells in a culture medium comprising a myosin heavy chain 11 (MYH11) agonist, wherein the MYH11 agonist is RepSox, until a cell population comprising MYH11+, alpha-actin 2 positive (ACTA2+), smoothelin positive (SMTN+), calponin1 positive (CNN1+), and transgelin positive (TAGLN+) contractile smooth muscle cells is obtained, and
- wherein the SMC progenitor cells are obtained by a method selected from the group consisting of:
- (a) a first method comprising:
- (i) culturing mesoderm cells under conditions and for a time sufficient to obtain a population of cells expressing MEOX1;
- (ii) culturing the population of cells expressing MEOX1 under conditions and for a time sufficient to suppress MEOX1 expression;
- and
- (iii) culturing the population of cells from step (ii) under conditions and for a time sufficient to obtain a population of SMC progenitor cells, and
- (b) a second method comprising:
- (i) culturing mesoderm cells in chemically defined medium that comprises a fibroblast growth factor (FGF) or a vascular endothelial growth factor (VEGF) for a period of time sufficient to induce SMC progenitor cells;
- and
- (ii) culturing the population of cells from step (i) under conditions and for a time sufficient to obtain a population of SMC progenitor cells.
Dependencies are inferred from the claim text.