| US 7,588,937 B2 | ||
| Method of in vitro differentiation of neural stem cells, motor neurons and dopamine neurons from primate embryonic stem cells | ||
| Su-Chun Zhang, Middleton, Wis. (US); and Xue-jun Li, Madison, Wis. (US) | ||
| Assigned to Wisconsin Alumni Research Foundation, Madison, Wis. (US) | ||
| Filed on Aug. 27, 2004, as Appl. No. 10/928,805. | ||
| Application 10/928805 is a continuation in part of application No. 09/970382, filed on Oct. 03, 2001, granted, now 6,887,706. | ||
| Claims priority of provisional application 60/498831, filed on Aug. 29, 2003. | ||
| Claims priority of provisional application 60/499570, filed on Sep. 02, 2003. | ||
| Prior Publication US 2005/0095706 A1, May 05, 2005 | ||
| Int. Cl. C12N 5/00 (2006.01); C12N 5/02 (2006.01); A01N 63/00 (2006.01) | ||
| U.S. Cl. 435—377 [435/363; 435/366] | 2 Claims |
| 1. A method of obtaining a population of midbrain dopamine neurons, comprising:
(a) obtaining a synchronous population of cells that are cultured from embryonic stem cells, and which are characterized by
an early rosette morphology and are Sox1−, Pax6+;
(b) culturing the cells obtained in (a) in a neural basal medium with fibroblast growth factor-8 (FGF8), wherein resulting
cells are Pax6+, Sox1+ and engrailed 1 positive (EN-1+) neuroepithelial cells;
(c) expanding the cells from step (b) in a neural basal medium with sonic hedgehog (SHH) and FGF8, wherein resulting cells
are dopaminergic neural precursors; and
(d) culturing the cells from step (c) in a neural basal differentiation medium without SHH and FGF8 until resulting cells
express tyrosine hydroxylase (TH), aromatic acid decarboxylase (AADC), EN-1, vesicular monoamine transporter 2 (VMAT2) and
dopamine transporter (DAT) but do not express dopamine beta hydroxylase (DbH) and phenylethanolamine N-methyltransferase (PNMT),
and produce dopamine.
|