IP Library Granted Patent US 9,938,502
Granted Patent B2
US 9,938,502 · App. 13/882,429 · Granted Apr 10, 2018

Tissue-specific extracellular matrix with or without tissue protein components for cell culture

Inventors: Yuanyuan Zhang (Winston Salem, NC); Shay Soker (Greensboro, NC); Anthony Atala (Winston Salem, NC); Aleksander Skardal (Clemmons, NC)
Assignee: Wake Forest University Health Sciences
C12N5/0671C12N5/0068C12N2513/00C12N2533/54C12N2533/80C12N2533/90
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Quick Facts
Patent No.
US 9,938,502
App. No.
13/882,429
Granted
Apr 10, 2018
Kind
B2
Abstract

The present invention relates to compositions and methods for mimicking an in vivo environment for culturing cells in vitro. The in vivo mimicking environment is based on the generation of a tissue-specific extracellular matrix wherein the matrix provides a substrate for which the cultured cell originated from. The tissue-specific extracellular matrix can further comprise a component of a whole tissue-specific homogenate.

Claims (91)

1. A tissue-specific in vitro cell culturing system for expanding a cell while maintaining cellular function, the cell culture system comprising:

a cell;

a culture medium; and

a tissue-specific component;

wherein said tissue-specific component comprises material isolated from a tissue that normally supports the growth and maturation of the cell in vivo by lyophilizing said tissue, grinding said lyophilized tissue and sterilizing said ground lyophilized tissue;

wherein said tissue is a whole tissue; and

wherein said isolated material is in the form of a ground lyophilized particle.

2. The cell culturing system of claim 1 , wherein said whole tissue is a homogenate.

3. The cell culturing system of claim 1 , wherein said cell is selected from the group consisting of an endothelial cell, a muscle cells, a smooth muscle cell, a fibroblast, an osteoblast, a myoblast, a neuroblast, a glioblast, a germ cell, a hepatocyte, a chondrocyte, a keratinocyte, a cardiac muscle cell, a connective tissue cell, an epithelial cell, a hormone-secreting cell, a cell of the immune system, a neuron, a stem cell, and any combination thereof.

4. The cell culturing system of claim 1 , wherein said tissue is selected from the group consisting of heart, kidney, liver, lung, pancreas, spleen, bladder, cartilage, bone, brain, spine cord, peripheral nerve, ureter, urethra, and any combination thereof.

5. The cell culturing system of claim 1 , further comprising a secreted factor selected from the group consisting of a growth factor, cytokine, and any combination thereof.

6. The cell culturing system of claim 1 , wherein said tissue-specific component is in the form of a sterilized fine particle, wherein the particle size is less than about 40 μm.

7. The cell culturing system of claim 1 , wherein said tissue-specific component is incorporated into a 2.5 D tissue particle coating.

8. The cell culturing system of claim 1 , wherein said tissue-specific component is incorporated into a 3-D culture gel.

9. The cell culturing system of claim 1 , wherein said tissue-specific component is incorporated into a 3-D porous tissue disc.

10. The cell culturing system of claim 1 , wherein the tissue specific component further comprises a whole tissue extract supernatant.

11. The cell culturing system of claim 1 , wherein said tissue-specific component comprises at least one selected from the group consisting of a tissue-specific whole tissue particle, a tissue-specific whole tissue particulate, and any combination thereof.

12. The cell culturing system of claim 11 , wherein said tissue-specific component is incorporated with a biopolymer selected from the group consisting of type I collagen, hyaluronic acid, heparin-conjugated hyaluronic acid, and any combination thereof.

13. A method of maintaining cellular function of a cell cultured in vitro, the method comprising culturing a cell in an in vitro cell culturing system comprising a tissue-specific component isolated from a tissue of a mammal, wherein said tissue-specific component comprises material isolated from a tissue that normally supports the growth and maturation of said cell in vivo by lyophilizing said tissue, grinding said lyophilized tissue and sterilizing said ground lyophilized tissue;

wherein said tissue is a whole tissue; and

wherein said isolated material is in the form of a ground lyophilized particle.

14. The method of claim 13 , wherein said whole tissue is a homogenate.

15. The method of claim 13 , further comprising a cell selected from the group consisting of an endothelial cell, a muscle cells, a smooth muscle cell, a fibroblast, an osteoblast, a myoblast, a neuroblast, a glioblast, a germ cell, a hepatocyte, a chondrocyte, a keratinocyte, a cardiac muscle cell, a connective tissue cell, an epithelial cell, a hormone-secreting cell, a cell of the immune system, a neuron, a stem cell, and any combination thereof.

16. The method of claim 13 , wherein said tissue is selected from the group consisting of heart, kidney, liver, lung, pancreas, spleen, bladder, cartilage, bone, brain, spine cord, peripheral nerve, ureter, urethra, and any combination thereof.

17. The method of claim 13 , wherein said tissue-specific component further comprises a secreted factor selected from the group consisting of a growth factor, cytokine, and any combination thereof.

18. The method of claim 13 , wherein said tissue-specific component is in the form of a sterilized fine particle, wherein the particle size is less than about 40 μm.

19. The method of claim 13 , wherein said tissue-specific component is incorporated into a 2.5 D tissue particle coating.

20. The method of claim 13 , wherein said tissue-specific component is incorporated into a 3-D culture gel.

21. The method of claim 13 , wherein said tissue-specific component is incorporated into a 3-D porous tissue disc.

22. The method of claim 13 , wherein the tissue specific component further comprises a whole tissue extract supernatant.

23. The method of claim 13 , wherein said tissue-specific component comprises at least one selected from the group consisting of a tissue-specific whole tissue particle, a tissue-specific whole tissue particulate, and any combination thereof.

24. The method of claim 13 , wherein said tissue-specific component is incorporated with a biopolymer selected from the group consisting of type I collagen, hyaluronic acid, heparin-conjugated hyaluronic acid, and any combination thereof.

25. A method of making tissue-specific extract for expanding a cell while maintaining cellular function, the method comprising:

obtaining a tissue from a mammal;

lyophilizing the tissue;

powderizing the lyophilized tissue thereby generating a tissue-specific particle;

forming a suspension comprising said tissue-specific particle by mixing the tissue-specific particle with a culture medium, hydrochloric acid and pepsin thereby generating a tissue-specific particle suspension;

separating the tissue-specific particle suspension thereby generating a supernatant and a particulate, wherein the supernatant is a tissue-specific extract and wherein the particulate is a tissue-specific particulate; and

isolating the tissue-specific extract from the particulate, and collecting the tissue-specific extract;

wherein said tissue is a tissue selected from a whole tissue and a decellularized tissue.

26. A method of making a culture system for expanding a cell while maintaining cellular function, the method comprising:

applying a tissue-specific component onto a surface, wherein said tissue-specific component comprises material isolated from a tissue that normally supports the growth and maturation of said cell in vivo by lyophilizing said tissue, grinding said lyophilized tissue and sterilizing said ground lyophilized tissue;

wherein said tissue is a whole tissue; and

wherein said tissue-specific component comprises at least one selected from the group consisting of a tissue-specific whole tissue particle, a tissue-specific whole tissue particulate, and any combination thereof.

27. A method of making a culture system for expanding a cell while maintaining cellular function, the method comprising:

mixing a first solution comprising a tissue-specific component with a second solution comprising a biopolymer, thereby generating a hydrogel;

wherein said tissue-specific component comprises material isolated from a tissue that normally supports the growth and maturation of said cell in vivo by lyophilizing said tissue, grinding said lyophilized tissue and sterilizing said ground lyophilized tissue;

wherein said tissue is a whole tissue; and

wherein said tissue-specific component comprises at least one selected from the group consisting of a tissue-specific whole tissue particle, a tissue-specific whole tissue particulate, and any combination thereof.

28. A tissue-specific in vitro cell culturing system for expanding a cell while maintaining cellular function, the cell culture system comprising:

a cell;

a culture medium; and

a tissue-specific component;

wherein said tissue-specific component comprises material isolated from a tissue that normally supports the growth and maturation of the cell in vivo by lyophilizing said tissue, grinding said lyophilized tissue and sterilizing said ground lyophilized tissue;

wherein said tissue is selected from the group consisting of a whole tissue and a decellularized tissue;

wherein said isolated material is in the form of a ground lyophilized particle; and

wherein said tissue-specific component is incorporated with a biopolymer selected from the group consisting of hyaluronic acid, heparin-conjugated hyaluronic acid, and any combination thereof.

29. The cell culturing system of claim 28 , wherein said tissue-specific component comprises material isolated from a whole tissue.

30. The cell culturing system of claim 29 , wherein said whole tissue is a homogenate.

31. The cell culturing system of claim 28 , wherein said tissue-specific component comprises material isolated from a decellularized tissue.

32. The cell culturing system of claim 31 , wherein said decellularized tissue is a homogenate.

33. The cell culturing system of claim 28 , wherein said cell is selected from the group consisting of an endothelial cell, a muscle cells, a smooth muscle cell, a fibroblast, an osteoblast, a myoblast, a neuroblast, a glioblast, a germ cell, a hepatocyte, a chondrocyte, a keratinocyte, a cardiac muscle cell, a connective tissue cell, an epithelial cell, a hormone-secreting cell, a cell of the immune system, a neuron, a stem cell, and any combination thereof.

34. The cell culturing system of claim 28 , wherein said tissue is selected from the group consisting of heart, kidney, liver, lung, pancreas, spleen, bladder, cartilage, bone, brain, spine cord, peripheral nerve, ureter, urethra, and any combination thereof.

35. The cell culturing system of claim 28 , further comprising a secreted factor selected from the group consisting of a growth factor, cytokine, and any combination thereof.

36. The cell culturing system of claim 28 , wherein said tissue-specific component is in the form of a sterilized fine particle, wherein the particle size is less than about 40 μm.

37. The cell culturing system of claim 28 , wherein said tissue-specific component is incorporated into a 3-D porous tissue disc.

38. The cell culturing system of claim 28 , wherein said tissue-specific component is incorporated into a 3-D culture gel.

39. The cell culturing system of claim 28 , wherein the tissue specific component further comprises a whole tissue extract supernatant.

40. The cell culturing system of claim 28 , wherein said tissue-specific component comprises at least one selected from the group consisting of a tissue-specific whole tissue particle, a tissue-specific decellularized tissue particle, a tissue-specific whole tissue particulate, a tissue-specific decellularized tissue particulate, and any combination thereof.

41. A method of making a culture system for expanding a cell while maintaining cellular function, the method comprising:

mixing a first solution comprising a tissue-specific component with a second solution comprising a biopolymer, thereby generating a hydrogel;

wherein the biopolymer is selected from the group consisting of hyaluronic acid, heparin-conjugated hyaluronic acid, and any combination thereof;

wherein said tissue-specific component comprises material isolated from a tissue that normally supports the growth and maturation of said cell in vivo by lyophilizing said tissue, grinding said lyophilized tissue and sterilizing said ground lyophilized tissue;

wherein said tissue is selected from the group consisting of a whole tissue and a decellularized tissue; and

wherein said tissue-specific component comprises at least one selected from the group consisting of a tissue-specific whole tissue particle, a tissue-specific decellularized tissue particle, a tissue-specific whole tissue particulate, a tissue-specific decellularized tissue particulate, and any combination thereof.

42. A method of maintaining cellular function of a cell cultured in vitro, the method comprising culturing a cell in an in vitro cell culturing system comprising a tissue-specific component isolated from a tissue of a mammal, wherein said tissue-specific component comprises material isolated from a tissue that normally supports the growth and maturation of said cell in vivo by lyophilizing said tissue, grinding said lyophilized tissue and sterilizing said ground lyophilized tissue;

wherein said tissue is selected from the group consisting of a whole tissue and a decellularized tissue;

wherein said isolated material is in the form of a ground lyophilized particle; and

wherein said tissue-specific component is incorporated with a biopolymer selected from the group consisting of hyaluronic acid, heparin-conjugated hyaluronic acid, and any combination thereof.

43. The method of claim 42 , wherein said tissue-specific component comprises material isolated from a whole tissue.

44. The method of claim 43 , wherein said whole tissue is a homogenate.

45. The method of claim 42 , wherein said tissue-specific component comprises material isolated from a decellularized tissue.

46. The method of claim 45 , wherein said decellularized tissue is a homogenate.

47. The method of claim 42 , further comprising a cell selected from the group consisting of an endothelial cell, a muscle cells, a smooth muscle cell, a fibroblast, an osteoblast, a myoblast, a neuroblast, a glioblast, a germ cell, a hepatocyte, a chondrocyte, a keratinocyte, a cardiac muscle cell, a connective tissue cell, an epithelial cell, a hormone-secreting cell, a cell of the immune system, a neuron, a stem cell, and any combination thereof.

48. The method of claim 42 , wherein said tissue is selected from the group consisting of heart, kidney, liver, lung, pancreas, spleen, bladder, cartilage, bone, brain, spine cord, peripheral nerve, ureter, urethra, and any combination thereof.

49. The method of claim 42 , wherein said tissue-specific component further comprises a secreted factor selected from the group consisting of a growth factor, cytokine, and any combination thereof.

50. The method of claim 42 , wherein said tissue-specific component is in the form of a sterilized fine particle, wherein the particle size is less than about 40 μm.

51. The method of claim 42 , wherein said tissue-specific component is incorporated into a 3-D culture gel.

52. The method of claim 42 , wherein said tissue-specific component is incorporated into a 3-D porous tissue disc.

53. The method of claim 42 , wherein the tissue specific component further comprises a whole tissue extract supernatant.

54. The method of claim 42 , wherein said tissue-specific component comprises at least one selected from the group consisting of a tissue-specific whole tissue particle, a tissue-specific decellularized tissue particle, a tissue-specific whole tissue particulate, a tissue-specific decellularized tissue particulate, and any combination thereof.

Assignments (2)
ASSIGNMENT OF ASSIGNOR'S INTEREST Recorded Jan 23, 2018
From: SKARDAL, ALEKSANDER
To: WAKE FOREST UNIVERSITY HEALTH SCIENCES
Reel/Frame 044698/0244 →
ASSIGNMENT OF ASSIGNOR'S INTEREST Recorded Jul 17, 2013
From: ZHANG, YUANYUAN; SOKER, SHAY; ATALA, ANTHONY
To: WAKE FOREST UNIVERSITY HEALTH SCIENCES
Reel/Frame 030818/0687 →
Continuity (2)
Provisional Application 61412193 · Nov 10, 2010
Related Publication 20130288375A1 · Oct 31, 2013