Small molecules supporting pluripotent cell growth and methods thereof
The present invention relates to compositions and methods for maintaining undifferentiated pluripotent stem cell cultures.
1. An in vitro composition comprising single human pluripotent cells whereby said pluripotent cells are plated as single cells at a low cell density in a medium comprising an effective amount of a neurotransmitter or a structural analog, derivative or physiologically acceptable salt thereof sufficient to achieve low density survival and expansion of the pluripotent cells.
2. The composition of claim 1 , wherein the neurotransmitter is a neurotransmitter receptor agonist.
3. The composition of claim 2 , wherein the neurotransmitter is norepinephrine.
4. The composition of claim 1 , wherein the human pluripotent cells can expand in the absence of differentiation.
5. The composition of claim 1 , further comprising at least one growth factor.
6. The composition of claim 5 , wherein the at least one growth factor is selected from the group consisting of: an ErbB3 ligand or a functional fragment thereof; a TGF-β family member or a functional fragment thereof; an activator of insulin-like growth factor receptor (IGF-1R) or a functional fragment thereof; and an activator of a fibroblast growth factor (FGF) receptor or a functional fragment thereof.
7. The composition of claim 6 , wherein the ErbB3 ligand is heregulin.
8. The composition of claim 1 , wherein the pluripotent cells are plated at about 1000 cells per ml of the medium.
9. The composition of claim 1 , wherein the pluripotent cells can be passaged at least 2 times.
10. The composition of claim 9 , wherein the pluripotent cells retain expression of pluripotency markers upon passaging.
11. The composition of claim 1 , wherein the composition is serum free.
12. The composition of claim 1 , wherein the composition is Activin A free.
13. The composition of claim 1 , further comprising at least one growth factor.
14. An in vitro suspension culture comprising single human pluripotent cells whereby said pluripotent cells are in suspension as single cells and an effective amount of a neurotransmitter or a structural analog, derivative or physiologically acceptable salt thereof.
15. The composition of claim 14 , wherein the composition is Activin A free.
16. A method of expanding human pluripotent cells in vitro comprising:
a. obtaining single human pluripotent cells; and
b. plating said single human pluripotent cells as single cells at low cell density in a medium comprising an effective amount of a neurotransmitter or a structural analog, derivative or physiologically acceptable salt thereof sufficient to achieve low density survival and expansion of the pluripotent cells.
17. The method of claim 16 , wherein the neurotransmitter is a neurotransmitter receptor agonist.
18. The method of claim 17 , wherein the neurotransmitter receptor agonist is norepinepherine.
19. The method of claim 16 , wherein the medium does not comprise Activin A.
20. The composition of claim 1 , said composition being essentially free of serum and serum replacement.
21. The composition of claim 1 , wherein the composition is free of a feeder cell layer.
22. The composition of claim 1 , wherein the effective amount of the neurotransmitter or the structural analog, derivative or physiologically acceptable salt thereof is sufficient to achieve improved survival and expansion of the pluripotent cells as compared to a control composition without the neurotransmitter or the structural analog, derivative or physiologically acceptable salt thereof.
23. The composition of claim 14 , wherein the neurotransmitter is norepinephrine.