Oligodendrocytes Derived from Human Embryonic Stem Cells for Remyelination and Treatment of Spinal Cord Injury

Abstract

This invention provides populations of neural cells bearing markers of glial cells, such as oligodendrocytes and their precursors. The populations are generated by differentiating pluripotent stem cells such as human embryonic stem cells under conditions that promote enrichment of cells with the desired phenotype or functional capability. Various combinations of differentiation factors and mitogens can be used to produce cell populations bearing markers of oligodendrocyte precursor cells. Upon further differentiation form complex processes characteristic of mature oligodendrocytes. The cells are capable of forming myelin sheaths, and can be used therapeutically improve function of the central nervous system.

1 .- 34 . (canceled) 
     
     
         35 . A first cell population and a second cell population, comprising:
 a) a first cell population comprising a line of undifferentiated human pluripotent stem (PS) cells in a cell culture medium comprising a mitogen, a ligand for a thyroid hormone receptor, and a ligand for a retinoic acid receptor, wherein the mitogen and the ligands are present in an amount effective for differentiation of the line of undifferentiated human PS cells into oligodendrocyte precursor cells; and   b) a second cell population comprising the oligodendrocyte precursor cells which are the in vitro progeny of the line of undifferentiated human PS cells.   
     
     
         36 . The first cell population and the second cell population of  claim 35 , wherein the oligodendrocyte precursor cells also express A2B5. 
     
     
         37 . The first cell population and the second cell population of  claim 36 , wherein the oligodendrocyte precursor cells stain with antibody specific for NG2 proteoglycan and are negative for the neuronal marker NeuN. 
     
     
         38 . The first cell population and the second cell population of  claim 35 , wherein the oligodendrocyte precursor cells also express platelet-derived growth factor receptor-α (PDGFR α). 
     
     
         39 . The first cell population and the second cell population of  claim 35 , wherein 20% or more of the oligodendrocyte precursor cells show a bipolar morphology characteristic of oligodendrocyte precursors. 
     
     
         40 . The first cell population and the second cell population of  claim 35 , wherein implantation of the second cell population into the spinal cord of a shiverer mutant mouse causes deposition of compact myelin around neuronal axons. 
     
     
         41 . The first cell population and the second cell population of  claim 35 , wherein implantation of the second cell population in or around the spinal cord in a contusion-injured rat causes improvement in overground locomotion compared with a contusion injured rat that does not receive the implantation of the differentiated cell population. 
     
     
         42 . The first cell population and the second cell population of  claim 35 , wherein the mitogen is selected from the group consisting of basic fibroblast growth factor (bFGF) and epidermal growth factor (EGF). 
     
     
         43 . The first cell population and the second cell population of  claim 35 , wherein the ligand for the thyroid hormone receptor is thyroid hormone triiodothyronine (T3). 
     
     
         44 . The first cell population and the second cell population of  claim 35 , wherein the ligand for the retinoic acid receptor is retinoic acid (RA). 
     
     
         45 . The first cell population and the second cell population of  claim 35 , wherein the mitogen is basic fibroblast growth factor (bFGF), the ligand for the thyroid hormone receptor is thyroid hormone triiodothyronine (T3), and the ligand for the retinoic acid receptor is retinoic acid (RA). 
     
     
         46 . The first cell population and the second cell population of  claim 45 , wherein the concentration of thyroid hormone T3 is 20 ng/ml. 
     
     
         47 . The first cell population and the second cell population of  claim 45 , wherein the concentration of RA is 10 μM. 
     
     
         48 . The first cell population and the second cell population of  claim 45 , wherein the concentration of bFGF is 4 ng/ml. 
     
     
         49 . The first cell population and the second cell population of  claim 45 , wherein the concentration of thyroid hormone T3 is 20 ng/ml, the concentration of RA is 10 μM, and the concentration of bFGF is 4 ng/ml. 
     
     
         50 . The first cell population and the second cell population of  claim 35 , wherein the first cell population and the second cell population are in separate containers. 
     
     
         51 . The first cell population and the second cell population of  claim 35 , wherein the oligodendrocyte precursor cells stain positive for GalC or MBP. 
     
     
         52 . The first cell population and the second cell population of  claim 35 , wherein the oligodendrocyte precursor cells stain positive for NP2 or GalC. 
     
     
         53 . A method for producing glial cells from human pluripotent stem (PS) cells, comprising culturing human PS cells in medium comprising a growth factor, a ligand for a thyroid hormone receptor, and a ligand for a retinoic acid receptor. 
     
     
         54 . The method of  claim 53 , comprising formulating glial cells in a composition for myelination of neurons in need thereof, wherein the glial cells are positive for GalC or MBP.