Preparation of endodermal stem cells

• US 20060281136A1
• Filed: 05/20/2004
• Published: 12/14/2006
• Est. Priority Date: 05/20/2003
• Status: Abandoned Application

• Current Assignee: Riken
• Original Assignee: Riken
• Inventors: Era, Takumi, Nishikawa, Shinichi
• Application Number:
  US10/556,886
• Publication Number:
  US 20060281136A1
• Time in Patent Office: Days
• US Class Current: 435/7.200
• CPC Class Codes: C12N 2506/02 from embryonic cells
• CPC Class Codes: C12N 5/0602 Vertebrate cells

Abstract

It is intended to prepare endodermal stem cells by differentiation from multipotential stem cells followed by isolation. This object can be achieved by separating various endodermal stem cells having been differentiated from multipotential stem cells with the use of, as indications, a specific cell surface marker, an organizer-specific marker and the expression manner of E-cadherin.

Claims

1. An endodermal stem cell which has been differentiated in vitro from a multipotential stem cell, wherein the cell is organizer-specific-marker positive and E-cadherin positive.

2. The cell according to claim 1, wherein the multipotential stem cell is an embryonic stem cell.

3. The cell according to claim 1 or 2, wherein the organizer-specific marker is goosecoid.

4. The cell according to claim 3, wherein a label protein is fused to the organizer-specific marker.

5. The cell according to claim 4, wherein the label protein is green fluorescence protein (GFP).

9. A method for differentiating an endodermal stem cell according to any of claims 1 to 5 to prepare an endodermal cell of interest.

10. The method according to claim 9, wherein the endodermal stem cells are cultured in the presence of activin in a serum-free medium on a culture plate coated with collagen IV, thereby differentiating the endodermal stem cells into endodermal cells.

11. An endodermal cells obtained by the preparation method according to claim 9 or 10.

13. The method according to claim 11, wherein the multipotential stem cell is an embryonic stem cell and the label protein is green fluorescence protein.

6. A method for preparing an endodermal stem cell, which comprises the steps of:

a) culturing multipotential stem cells, and

b) selecting and separating cells expressing an organizer-specific marker and E-cadherin.

7. The method according to claim 6, wherein a cell sorter is employed at the selection step.

8. The method according to claim 6 or 7, wherein the multipotential stem cells are cultured in the presence of activin in a serum-free medium on a culture plate coated with collagen IV, thereby differentiating the multipotential stem cells into endodermal stem cells.

12. A method for preparing an intermediate stem cell of interest from a multipotential stem cell, which comprises the steps of:

a) introducing a gene of a label protein in the genome of a multipotential stem cell so as to be adapted to the expression system of a predetermined marker gene, such that said label protein is expressed instead of or together with the predetermined marker gene, and

b) selecting and separating intermediate stem cells of interest using the label of said label protein as an indicator.

14. The method according to claim 12 or 13, wherein the intermediate stem cells are endodermal stem cells.