Conditioned medium for culturing Schwann cells

• US 20050142660A1
• Filed: 11/18/2004
• Published: 06/30/2005
• Est. Priority Date: 12/30/2003
• Status: Abandoned Application

• Current Assignee: Industrial Technology Research Institute
• Original Assignee: Industrial Technology Research Institute
• Inventors: Shen, Hsin-Hsin, Chang, Pei-Ching, Chen, Yu-Hua, Lou, Wan-Shiun
• Application Number:
  US10/990,465
• Publication Number:
  US 20050142660A1
• Time in Patent Office: Days
• US Class Current: 435/368
• CPC Class Codes: C12N 2502/08 cells of the nervous system
• CPC Class Codes: C12N 5/0622 Glial cells, e.g. astrocytes, oligodendrocytes; Schwann cells

Abstract

The invention provides a conditioned medium that enhances the survival and/or proliferation of Schwann cells in cell culture, and a cell line useful for production of such medium. The cell line used in the invention is a differentiated NTera2/D1 (NT2) cell line.

Claims

1. A method for preparing a conditioned medium for culturing Schwann cells, comprising the steps of:

a) providing a culture medium, and cells;

b) culturing the cells in the culture medium, thereby conditioning the medium; and

c) harvesting the conditioned medium from the culture;

wherein the cells used to condition the medium are differentiated Ntera2/D1 (NT2)cells.

2. The method of claim 1, wherein the culture medium is DMEM/F12.

3. The method of claim 1, wherein the conditioned medium was produced after differentiated Ntera2/D1 (NT2) cells were cultured for at least 10 days.

4. The method of claim 1,wherein the conditioned medium is filtrated after harvesting or before use.

5. The method of claim 1,wherein the filtration is performed with a 0.22 μ

m pore sized filter.

6. The method of claim 1, wherein further comprises a step a1), differentiating NTera2/D1 (NT2) cells before step a).

7. The method of claim 6, wherein the NTera2/D1 (NT2) cells are differentiated through incubating undifferentiated NTera2/D1 (NT2) cells in a culture medium containing a differentiation agent in step a).

8. The method of claim 7, wherein the differentiation agent is hexamethylene bisacetamide (HMBA), retinoic acid(RA) or bromodeoxyuridine (BrdUrd).

9. The method of claim 8, wherein the differentiation agent is retinoic acid(RA).

10. The method of claim 8, wherein the differentiation agent is All-trans-Retinoic acid.

13. The method of claim 1, wherein the conditioned medium was harvested after differentiated Ntera2/D1 (NT2)cells were cultured for at least 10 days.

14. The method of claim 1, wherein the conditioned medium is filtrated after harvesting or before use.

11. A method for culturing Schwann cells which comprises stpes of:

culturing the Schwann cells in a growth environment that contains a conditioned medium produced by a method comprising steps of;

a) providing a culture medium, and cells;

b) culturing the cells in the culture medium, thereby conditioning the medium; and

c) harvesting the conditioned medium from the culture;

wherein the cells used for the medium conditioning are differentiated Ntera2/D1 (NT2) cells.

12. The method of claim 11, wherein the culture medium is DMEM/F12.

15. The method of claim 11, further comprises a diluting method of the conditioned medium.

16. The method of claim 15, wherein the diluting method comprises steps of:

a) providing a conditioned medium and a diluent medium; and

b) mixing the conditioned medium with the diluent medium.

17. The method of claim 16, wherein the conditioned medium and the diluent medium are mixed in a ratio of 1:

1.

18. The method of claim 16, wherein the diluent medium is SC medium.

19. A conditioned medium for culturing Schwann cells, produced by a method comprising:

a) providing a culture medium, and cells

b) culturing the cells in the culture medium, thereby conditioning the medium; and

c) harvesting the conditioned medium from the culture;

wherein the cells used to condition the medium are differentiated Ntera2/D1 (NT2)cells.

20. The conditioned medium of claim 19,wherein the culture medium is DMEM/F12.

21. The conditioned medium of claim 19, wherein the conditioned medium was harvested after differentiated Ntera2/D1 (NT2)cells were cultured for at least 10 days.

22. The conditioned medium of claim 19, wherein the conditioned medium is filtrated after harvesting or before use.