Three-dimensional cell culture system

• US 20060134781A1
• Filed: 12/07/2005
• Published: 06/22/2006
• Est. Priority Date: 12/07/2004
• Status: Abandoned Application

• Current Assignee: Bacterin International Incorporated (OrbiMed Advisors LLC (China))
• Original Assignee: Bacterin International Incorporated (OrbiMed Advisors LLC (China))
• Inventors: Shelby, Nancy Jane, Yang, Young Il
• Application Number:
  US11/296,802
• Publication Number:
  US 20060134781A1
• Time in Patent Office: Days
• US Class Current: 435/366
• CPC Class Codes: C12N 2533/54 Collagen; Gelatin
• CPC Class Codes: C12N 2533/56 Fibrin; Thrombin
• CPC Class Codes: C12N 2533/90 Substrates of biological origin, e.g. extracellular matrix, decellularised tissue
• CPC Class Codes: C12N 5/0667 Adipose-derived stem cells [ADSC]; Adipose stromal stem cells

Abstract

A three-dimensional culture system provides an efficient mechanism for producing stems cells derived from adipose tissue. A three-dimensional matrix is formed in the presence of adipose tissue to incorporate the adipose tissue into the three-dimensional matrix. After incubation, the 3-D matrix may be degraded to liberate stem cells, or progeny cells arising from the stem cells, from the 3-D matrix.

Claims

1. A method for culturing adipose-derived stem cells, the method comprising:

forming a three-dimensional (3-D) matrix in the presence of adipose tissue to incorporate the adipose-tissue into the 3-D matrix; and

incubating the 3-D matrix in a nutrient medium to produce a population of adipose-derived stem cells within the 3-D matrix.

2. The method of claim 1, wherein the adipose tissue fragment includes a native extracellular matrix.

3. The method of claim 1, and further comprising:

removing an adipose-derived stem cell from the 3-D matrix.

4. The method of claim 3, wherein removing the adipose-derived stem cell comprises digesting the 3-D matrix under conditions suitable for preserving viability of the adipose-derived stem cell.

5. The method of claim 3, and further comprising:

implanting the adipose-derived stem cell in a host.

6. The method of claim 5, and further comprising:

combining the adipose-derived stem cell with a carrier prior to implanting the adipose-derived stem cell in the host.

7. The method of claim 6, wherein the carrier comprises demineralized bone matrix.

8. The method of claim 3, and further comprising:

exposing the adipose-derived stem cell to a differentiation factor.

9. The method of claim 8, wherein the adipose-derived stem cell and differentiation factor are combined and implanted into a host.

10. The method of claim 8, wherein the differentiation factor comprises demineralized bone matrix.

11. The method of claim 8, wherein the differentiation factor comprises adipose tissue.

12. The method of claim 3, and further comprising:

cryopreserving the adipose-derived stem cell removed from the 3-D matrix.

13. The adipose-derived stem cell obtained from the method of claim 3.

14. The method of claim 1, wherein the adipose tissue comprises an adipose tissue fragment.

15. The method of claim 14, wherein a maximum dimension of the adipose tissue fragment ranges from as low as about 0.1 mm to as high as about 10 mm.

16. The method of claim 1, wherein the adipose tissue is not subjected to enzymatic digestion prior to forming the 3-D matrix.

17. The method of claim 1, and further comprising:

implanting the 3-D matrix in a host.

18. The method of claim 1, and further comprising:

removing an adipose tissue sample from a human and processing the adipose tissue sample to produce the adipose tissue.

19. The three-dimensional matrix obtained by the method of claim 1 and including the population of adipose-derived stem cells.

20. A method for culturing adipose-derived stem cells, the method comprising:

exposing an adipose tissue fragment to a fluid containing a substance;

crosslinking the substance to form a 3-D matrix, wherein the adipose tissue fragment is incorporated within the 3-D matrix; and

providing nutrients to the 3-D matrix.

21. The method of claim 20, and further comprising:

mincing adipose tissue to produce the adipose tissue fragment.

22. The method of claim 20, and further comprising:

removing one or more adipose-derived stem cells from the 3-D matrix.

23. The method of claim 22, wherein the 3-D matrix is disassociated to remove the one or more adipose-derived stem cells from the 3-D matrix.

24. The method of claim 22, and further comprising:

cryopreserving the one or more adipose-derived stem cells removed from the 3-D matrix.

25. The method of claim 22, and further comprising:

differentiating the one or more adipose-derived stem cells removed from the 3-D matrix.

26. The method of claim 20, and further comprising:

incubating the 3-D matrix to increase a number of the adipose-derived stem cells.

27. The method of claim 20, wherein the substance is selected from the group consisting of fibrin, collagen, gelatin, hyaluronan, chondroitin sulfate, alginate, nitrocellulose, carboxymethycellulose, polyglycolic acid (PGA), polyethylene glycol (PEG), poly(lactic-co-glycolic acid) (PLGA), poly-L-lysine, Matrigel®

compositions, poly(lactic acid) (PLA), and combinations thereof.

28. A composition prepared in vitro comprising a 3-D matrix and adipose tissue embedded in the 3-D matrix.

29. The composition of claim 28, wherein the adipose tissue includes a native extracellular matrix.

30. The composition of claim 28, wherein the adipose tissue comprises an adipose tissue fragment embedded in the 3-D matrix having a maximum dimension ranging from as low as about 0.1 mm to as high as about 10 mm.

31. The composition of claim 28, wherein the 3-D matrix comprises a fibrin matrix.

32. The composition of claim 28, wherein the 3-D matrix comprises a collagen matrix.

33. The composition of claim 28, wherein the 3-D matrix comprises a substance selected from the group consisting of fibrin, collagen, gelatin, hyaluronan, chondroitin sulfate, alginate, nitrocellulose, carboxymethylcellulose, polyglycolic acid (PGA), polyethylene glycol (PEG), poly(lactic-co-glycolic acid) (PLGA), poly-L-lysine, Matrigel®

compositions, poly(lactic acid) (PLA), and combinations thereof.