Markers for differentiation of stem cells into differentiated cell populations

Abstract

Provided herein are systems and methods for identifying cell-specific differentiation markers. In particular, provided herein are systems and methods for generating induced pluripotent cells (IPS) from human cells, differentiating the IPS into differentiated cells, and identifying differentiation specific markers.

1 . A method of identifying differentiation markers, comprising:
 a) culturing a primary cell under conditions such that said primary cell de-differentiates to generate an induced pluripotent stem cell (iPSC);   b) treating said iPSC under conditions such that said iPSC differentiates into a differentiated cell; and   c) identifying molecular markers that are indicative of differentiation into said differentiated cell.   
     
     
         2 . The method of  claim 1 , wherein said primary cell is a pancreatic beta cell. 
     
     
         3 . The method of  claim 1 , wherein said differentiated cell is a pancreatic beta cell. 
     
     
         4 . The method of  claim 1 , wherein said culturing comprises expanding said cell. 
     
     
         5 . The method of  claim 1 , wherein said culturing comprises inducing expression of one or more genes selected from the group consisting of Oct4 (Pou5f1), Sox2, cMyc, nanog, LIN28, Glis1, and Klf4. 
     
     
         6 . The method of  claim 1 , wherein said treating comprises the steps of
 (a) culturing the iPSCs in a chemically defined medium, fibroblast growth factor, Activin A, and bone morphogenetic protein;   (b) culturing the cells from step (a) in the presence of chemically defined medium comprising insulin, transferrin, and selenium, a fibroblast growth factor and nicotinamide;   (c) culturing the cells from step (b) in the presence of a chemically defined medium comprising insulin, transferrin, and selenium; retinoic acid; a bone morphogenetic protein inhibitor; and nicotinamide; and   (d) culturing the cells from step (c) in the presence of a serum-free medium, an insulin-like growth factor, insulin, nicotinamide, exendin-4 and/or GLP-1, a TGF-beta inhibitor, and an agent that increase cAMP.   
     
     
         7 . The method of  claim 6 , further comprising:
 (e) maintaining the cells of the pancreatic lineage for 1-50 days by culturing the cells from step (d) in the presence of a serum-free medium, an insulin-like growth factor, fibroblast growth factor, insulin, nicotinamide, exendin-4 and/or GLP-1, and an agent that increase cAMP.   
     
     
         8 . The method of  claim 6 , wherein the fibroblast growth factor of step (a) is basic fibroblast growth factor (bFGF). 
     
     
         9 . The method of  claim 6 , wherein the bone morphogenetic protein of step (a) is BMP4. 
     
     
         10 . The method of  claim 6 , wherein the chemically defined medium comprising insulin, transferrin, and selenium of steps (b) and/or step (c) is ITS medium. 
     
     
         11 . The method of  claim 6 , wherein the fibroblast growth factor of step (b) and/or step (d) is FGF7. 
     
     
         12 . The method of  claim 6 , wherein the bone morphogenetic protein inhibitor of step (c) is a BMP4 inhibitor. 
     
     
         13 . The method of  claim 6 , wherein the serum-free medium of step (d) is B27. 
     
     
         14 . The method of  claim 6 , wherein the primary and differentiated cells are the same or different cell types. 
     
     
         15 . The method of  claim 1 , wherein step (a) has a duration of 2-10 days. 
     
     
         16 . The method of  claim 1 , wherein said identifying comprises sequencing expressed genes in said primary cell, said iPSC, and said differentiated cell and comparing said expressed genes to identify gene expression markers indicative of differentiation into said differentiated cell. 
     
     
         17 . The method of  claim 16 , wherein said comparing comprises the use of bioinformatics methods. 
     
     
         18 . One or more differentiation markers identified by the method of  claim 1 . 
     
     
         19 . A method of transplanting cells into a subject, comprising:
 a) identifying therapeutically suitable differentiated cells that express one or more of the markers of  claim 18 ; and   b) transplanting said cells into a subject in need thereof.   
     
     
         20 . The method of  claim 19 , wherein said transplanting results in reduction or elimination of one or more symptoms of a disease or condition.