Pluripotent cells

Abstract

Pluripotent cells that are immunopositive for both the neural progenitor marker nestin and a pluripotent cell marker are provided. The cells exhibit rapid doubling times and can be maintained in vitro for extended periods. Also provided are cell cultures containing the pluripotent cells, a method of transplanting human pluripotent cells to a host, and a method of reducing seizure activity in a subject. These pluripotent cells, when transplanted into the ventricle of a host animal, migrate to the site of damage and adopt a suitably corrective phenotype, resulting in both structural and functional restoration.

1 . A plurality of isolated mammalian cells that are immunopositive for nestin and for a marker of pluripotent cells.  
   
   
       2 . The cells of  claim 1 , wherein the marker of pluripotent cells is selected from the group consisting of TRA-1-60, TRA-1-81 and SSEA-4.  
   
   
       3 . The cells of  claim 1 , wherein the cells have a doubling rate of less than 12 days.  
   
   
       4 . The cells of  claim 1 , wherein the cells have a doubling rate of about 5 days.  
   
   
       5 . The cells of  claim 1 , wherein the cells continue to proliferate for at least 2 years in vitro.  
   
   
       6 . The cells of  claim 1 , wherein the cells are obtained from fetal forebrain.  
   
   
       7 . A composition comprising the cells of  claim 1  and a pharmaceutically acceptable carrier.  
   
   
       8 . A cell culture comprising: 
 (a) a culture medium, wherein the calcium concentration of the medium is not greater than 0.15 mM;    (b) about 15-100 ng/μl epidermal growth factor (EGF);    (c) about 10-150 ng/μl basic fibroblast growth factor (bFGF);    (d) about 10-75 ng/μl transforming growth factor-alpha (TGFα);    (e) the cells of  claim 1  suspended in the medium.    
   
   
       9 . The cell culture of  claim 8 , further comprising: 
 (f) about 0.05 to about 0.12 mM calcium chloride, wherein the medium is brought to full volume in a calcium-free minimum essential medium and has a total calcium concentration of less than 0.1 mM.    
   
   
       10 . The cell culture of  claim 8 , further comprising; 
 (g) about 25-150 ng/μl leukemia inhibiting factor (LIF).    
   
   
       11 . The cell culture of  claim 9 , wherein the total calcium concentration is about 0.05 mM.  
   
   
       12 . The cell culture of  claim 8 , wherein the EGF is about 20 ng/μl.  
   
   
       13 . The cell culture of  claim 8 , wherein the bFGF is about 10 ng/μl.  
   
   
       14 . The cell culture of  claim 8 , wherein the TGFα is about 10 ng/μl.  
   
   
       15 . The cell culture of  claim 10 , wherein the LIF is about 25 ng/μl.  
   
   
       16 . The cell culture of  claim 8 , wherein the culture medium is serum-free.  
   
   
       17 . The cell culture of  claim 8 , further comprising 0.5-2.5% B27 supplement.  
   
   
       18 . The cell culture of  claim 8 , wherein the growth factors EGF, bFGF and TGFα are recombinant growth factors.  
   
   
       19 . The cell culture of  claim 8 , wherein the cells and the growth factors are human.  
   
   
       20 . The cell culture of  claim 8 , further comprising about 0.11 mg/ml sodium pyruvate.  
   
   
       21 . A method of propagating pluripotent cells, comprising culturing the cell culture of  claim 8 .  
   
   
       22 . A method of transplanting human pluripotent cells to a mammalian host, comprising: 
 (a) obtaining the cells of  claim 1;  and    (b) transplanting the cells to the host.    
   
   
       23 . The method of  claim 22 , wherein glutamine (to a concentration 0.5 mM) and LIF (25-150 ng/μl) are added to the culture medium prior to the transplanting.  
   
   
       24 . A method of reducing seizure activity in a mammalian subject comprising: 
 (a) identifying a mammalian subject who has had a seizure within the preceding week; and    (b) implanting the cells of  claim 1  into a cerebral ventricle of the host.    
   
   
       25 . The method of  claim 24 , wherein the subject has had a seizure within the preceding 48 hours.