Method of reducing cell differentiation

• US 20050106725A1
• Filed: 11/19/2003
• Published: 05/19/2005
• Est. Priority Date: 11/19/2003
• Status: Abandoned Application

• Current Assignee: Wisconsin Alumni Research Foundation (University of Wisconsin System)
• Original Assignee: Wisconsin Alumni Research Foundation (University of Wisconsin System)
• Inventors: Ji, Lin, Palecek, Sean P., de Pablo, Juan J.
• Application Number:
  US10/717,677
• Publication Number:
  US 20050106725A1
• Time in Patent Office: Days
• US Class Current: 435/366
• CPC Class Codes: C12M 35/04 Mechanical means, e.g. sonic waves, stretching forces, pressure or shear stimuli
• CPC Class Codes: C12N 2533/90 Substrates of biological origin, e.g. extracellular matrix, decellularised tissue
• CPC Class Codes: C12N 5/0606 Pluripotent embryonic cells, e.g. embryonic stem cells [ES] embryonic germ cells C12N5/0611, induced pluripotent stem cells C12N5/0696

Abstract

The present invention relates to methods and compositions for culturing embryonic stem (ES) cells. The methods relate to growing the cells in culture on a flexible solid porous matrix, suitably without conditioned media or fibroblast feeder cells and applying an effective amount of periodic strain to stretch the flexible matrix, such that the cells proliferate and exhibit reduced differentiation. These methods in part facilitate a reduction in cross-species contamination for ES cells used in medical applications.

Claims

1. A method of culturing embryonic stem (ES) cells with reduced differentiation comprising:

a) growing the cells in culture on a flexible solid porous matrix without conditioned media and in the absence of fibroblast feeder cells; and

b) applying an effective amount of periodic strain to stretch the flexible matrix, such that the cells proliferate and exhibit reduced differentiation.

2. The method of claim 1 wherein the cells are mammalian embryonic stem cells.

3. The method of claim 1 wherein the cells are human embryonic stem cells.

4. The method of claim 1 wherein the cell differentiation is eliminated.

5. The method of claim 1 wherein the cells are grown on Matrigel™

using BioFlex®

untreated culture plates.

6. The method of claim 1 wherein the cells are grown without the presence of cross-species biological material.

7. The method of claim 1 wherein the flexible matrix is Matrigel™

.

8. The method of claim 1 wherein the strain is mechanically produced.

9. The method of claim 1 wherein the flexible matrix is stretched using vacuum pressure.

10. The method of claim 1 wherein the strain exerted on the flexible matrix is at least about 5%.

11. The method of claim 1 wherein the flexible matrix undergoes at least about 6 stretches per minute.

12. The method of claim 1 wherein the mechanical strain is from oscillatory stretching of the flexible matrix surface.

13. A culture comprising embryonic stem (ES) cells grown on a flexible solid porous matrix without conditioned media or fibroblast feeder cells, wherein an effective amount of periodic strain is applied to stretch the flexible matrix, such that the cells proliferate and exhibit reduced differentiation.

14. The method of claim 13 wherein the cells are mammalian embryonic stem cells.

15. The method of claim 13 wherein the cells are human embryonic stem cells.

16. The method of claim 13 wherein the cell differentiation is eliminated.

17. The method of claim 13 wherein the cells are grown on Matrigel™

using BioFlex®

untreated culture plates.

18. The method of claim 13 wherein the cells are grown without the presence of cross-species biological material.

19. The method of claim 13 wherein the flexible solid porous matrix is Matrigel™

.

20. The method of claim 13 wherein the strain is mechanically produced.

21. The method of claim 13 wherein the flexible matrix is stretched using vacuum pressure.

22. The method of claim 13 wherein the mechanical strain is from oscillatory stretching of the flexible matrix surface.

23. The method of claim 13 wherein the strain exerted on the flexible matrix is at least about 5%.

24. The method of claim 13 wherein the flexible matrix undergoes at least about 6 stretches per minute.