Compositions and Methods for Enrichment of Neural Stem Cells Using Ceramide Analogs

Abstract

The present invention provides compositions and methods for human neural cell production. More particularly, the present invention provides cellular differentiation methods employing amphiphilic lipid compounds, preferably ceramide analogs of the β-hydroxyalkylamine type and optionally employing an essentially serum free MEDII conditioned medium for the generation of human neural cells from pluripotent human cells. The methods alternatively comprise modulating apoptosis by modifying the levels of PAR-4, with or without the presence of amphiphilic lipid compounds and optionally employing MEDII conditioned medium. The methods alternatively encompass modulating apoptosis by modulating the intracellular concentration of endogenous lipid second messengers, such as ceramide.

1 - 23 . (canceled) 
     
     
         24 : A method of producing a human neural cell comprising,
 a) providing a pluripotent human cell,   b) forming an embryoid body comprising the pluripotent human cell by culturing the pluripotent human cell in a medium that is optionally essentially serum free; and   c) culturing cells from the embryoid body with a composition comprising a sphingosine compound of the general formula   
       
         
           
           
               
               
           
         
         
           wherein 
         
         R=a saturated or mono- or polyunsaturated (cis or trans) alkyl group having greater than 2 carbon atoms, and 
         R1, R2, R3 and R4 may be the same or different and are saturated or mono- or polyunsaturated hydroxylated alkyl groups, aryl groups, or hydrogen. 
       
     
     
         25 : The method of  claim 24 , wherein the medium is an essentially serum free medium and comprises a MEDII conditioned medium. 
     
     
         26 : A method of producing a human neural cell comprising,
 a) providing a pluripotent human cell;   b) forming an embryoid body comprising the pluripotent human cell by culturing the pluripotent human cell in a medium that is optionally essentially serum free; and   c) culturing cells from the embryoid body with a composition comprising a hydroxyalkyl ester compound of the general formula   
       
         
           
           
               
               
           
         
         
           wherein 
         
         R=a saturated or mono- or polyunsaturated (cis or trans) alkyl group having greater than 2 carbon atoms, and 
         R1 is a saturated or mono- or polyunsaturated hydroxylated alkyl group, aryl group, or hydrogen. 
       
     
     
         27 : The method of  claim 26 , wherein the medium is an essentially serum free medium and comprises a MEDII conditioned medium. 
     
     
         28 : The method of  claim 27 , wherein the composition comprises an hydroxyalkyl ester compound of the structure 
       
         
           
           
               
               
           
         
       
     
     
         29 . A human neural cell produced by the method of  claim 24 . 
     
     
         30 : A method for treating a patient, comprising a step of administering to the patient having a neural disease a therapeutically effective amount of the neural cell of  claim 29 . 
     
     
         31 : The method of  claim 30 , wherein the neural disease is Parkinson's disease. 
     
     
         32 - 71 . (canceled) 
     
     
         72 : A human neural cell produced by the method of  claim 26 . 
     
     
         73 : A method for treating a patient, comprising a step of administering to the patient having a neural disease a therapeutically effective amount of the neural cell of  claim 72 . 
     
     
         74 : The method of  claim 73 , wherein the neural disease is Parkinson's disease.