Multipotent neural cells

Abstract

The inventions disclosed herein are based on the identification of novel cell populations derived from human embryonic stem cells and other pluripotent cells. The inventive cell populations may be used for cell therapies for the treatment of various neurological diseases and as substrates in pharmacological assays.

1 . A substantially homogenous population of cells, wherein the cells of said population are characterized as CD24 HI  and express at least one marker selected from the group consisting of TuJ1, MAP2, and doublecortin. 
     
     
         2 . The population of  claim 1 , wherein said cells are further characterized as CD29 LO . 
     
     
         3 . The population of  claim 2 , wherein said cells are further characterized as CD15 − . 
     
     
         4 . The population of  claim 3 , wherein said cells further express tyrosine hydroxylase. 
     
     
         5 . The population of  claim 4 , wherein said cells adopt a neuronal morphology when cultured in the presence of at least one growth factor. 
     
     
         6 . The population of  claim 5 , wherein said growth factor is selected from the group consisting of fibroblast growth factor 8 (FGF8), brain-derived neurotrophic factor (BDNF), basic fibroblast growth factor (bFGF), transforming growth factor type beta-3 (TGF-β3), and glial cell line-derived neurotrophic factor (GDNF). 
     
     
         7 . The population of  claim 5 , wherein said cells are characterized as CD15 − /CD24 HI /CD29 LO . 
     
     
         8 . A substantially homogenous population of cells, wherein the cells of said population are characterized as CD24 LO  and CD29 HI . 
     
     
         9 . The population of any one of  claim 8 , wherein said cells are further characterized as CD15 − . 
     
     
         10 . The population of any one of  claim 8 , wherein said cells are further characterized as CD15 + . 
     
     
         11 . A therapeutic composition comprising a population of  claim 1 . 
     
     
         12 . The therapeutic composition of  claim 11 , wherein said population of cells is suspended in a physiologically compatible solution. 
     
     
         13 . The therapeutic composition of  claim 11 , wherein said population of cells is encapsulated. 
     
     
         14 . A method for treating a neurodegenerative disease in a patient, comprising administering to the brain of said patient a therapeutic composition of  claim 11 . 
     
     
         15 . A method for isolating early neurons and neuroblasts from in vitro culture comprising:
 (i) culturing pluripotent cells in the presence of a growth factor that induces at least some of said pluripotent cells to increase expression of TuJ1 relative to pluripotent cells cultured in the absence of said growth factor; and   (ii) selecting the cells obtained from step (i) that express relatively high levels of CD24, wherein said subpopulation is identified as early neurons and neuroblasts.   
     
     
         16 . The method of  claim 15 , wherein said growth factor is selected from the group consisting of sonic hedgehog (SHH), fibroblast growth factor-8 (FGF-8), basic fibroblast growth factor (bFGF), and brain-derived neurotrophic factor (BDNF). 
     
     
         17 . The method of  claim 16 , wherein CD15 +  cells are removed from said subpopulation. 
     
     
         18 . The method of  claim 17 , wherein cells expressing relatively high levels of CD29 are removed. 
     
     
         19 . The method of  claim 15 , wherein the cells obtained in step (ii) represent not more 75% of the CD15 −  cells obtained from step (i). 
     
     
         20 . The method of  claim 15 , wherein the cells obtained from step (ii) represent not more 25% of the CD15 −  cells from the cells obtained from step (i). 
     
     
         21 . A method for isolating neural crest cells from in vitro culture comprising:
 (i) culturing pluripotent cells in the presence of a growth factor that induces at least some of said pluripotent cells to increase expression of TuJ1 relative to pluripotent cells cultured in the absence of said growth factor;   (ii) removing CD15 +  cells from the cells obtained from step (i); and   (iii) selecting cells that express relatively high levels of CD29 from the cells obtained from step (i),   
       wherein the cells obtained following steps (ii) and (iii) are identified as neural crest cells. 
     
     
         22 . The method of  claim 21 , wherein said growth factor is selected from the group consisting of sonic hedgehog (SHH), fibroblast growth factor-8 (FGF-8), basic fibroblast growth factor (bFGF), and brain-derived neurotrophic factor (BDNF). 
     
     
         23 . The method of  claim 21 , wherein the cells obtained from step (iii) represent not more than 75% of the CD15 −  cells obtained from step (i). 
     
     
         24 . The method of  claim 21 , wherein the cells obtained from step (iii) represent not more than 25% of the CD15 −  cells obtained from step (i). 
     
     
         25 . The method of  claim 21 , wherein said method further comprises selecting cells that express relatively low levels of CD24. 
     
     
         26 . A method for isolating neural precursor cells from in vitro culture comprising:
 (i) culturing pluripotent cells in the presence of a growth factor that induces at least some of said pluripotent cells to increase expression of TuJ1 relative to pluripotent cells cultured in the absence of said growth factor;   (ii) removing CD15 −  cells from obtained from step (i); and   (iii) selecting cells that express relatively high levels of CD29 from the cells obtained from step (i),   
       wherein the cells obtained following steps (ii) and (iii) are identified as early neurons and neuroblasts. 
     
     
         27 . The method of  claim 26 , wherein said growth factor is selected from the group consisting of sonic hedgehog (SHH), fibroblast growth factor-8 (FGF-8), basic fibroblast growth factor (bFGF), and brain-derived neurotrophic factor (BDNF). 
     
     
         28 . The method of  claim 26 , wherein the cells obtained from step (iii) represent not more than 75% of the CD15 +  cells from said population cultured in step (i). 
     
     
         29 . The method of  claim 26 , wherein the cells obtained from step (iii) represent not more than 25% of the CD15 +  cells from said population cultured in step (i). 
     
     
         30 . The method of  claim 26 , wherein said method further comprises selecting cells that express relatively low levels of CD24. 
     
     
         31 . A method for preparing substantially homogenous populations of cells having a stellate morphology, comprising:
 (i) culturing in vitro cells having a stellate morphology;   (ii) preparing a fluid composition containing detached cells obtained from the culture of step (i); and   (iii) selecting said cells by flow cytometry to generate substantially homogenous cell populations, wherein the flow cytometer fluid pressure is less than about 50 psi and the fluid ejection nozzle has a diameter of greater than about 75 μm.   
     
     
         32 . The method of  claim 31 , wherein said fluid pressure is between about 20 psi and about 50 psi. 
     
     
         33 . The method of  claim 31 , wherein said ejection nozzle has a diameter of between about 75 μm and about 125 μm. 
     
     
         34 . The method of  claim 31 , wherein said cells having a stellate morphology are neurons or neuronal precursor cells.