Stem cells and method of separating the same

• US 20050003529A1
• Filed: 10/27/2003
• Published: 01/06/2005
• Est. Priority Date: 04/24/2001
• Status: Abandoned Application

• Current Assignee: Ajinomoto Co., Inc.
• Original Assignee: Ajinomoto Co., Inc.
• Inventors: Taniguchi, Hideki, Suzuki, Atsushi
• Application Number:
  US10/693,712
• Publication Number:
  US 20050003529A1
• Time in Patent Office: Days
• US Class Current: 435/366
• CPC Class Codes: A61K 35/12 Materials from mammals; Compositions comprising non-specified tissues or cells; Compositions comprising non-embryonic stem cells; Genetically modified cells vaccines or medicinal preparations containing antigens or antibodies A61K39/00
• CPC Class Codes: A61P 1/00 Drugs for disorders of the alimentary tract or the digestive system
• CPC Class Codes: A61P 1/16 for liver or gallbladder disorders, e.g. hepatoprotective agents, cholagogues, litholytics
• CPC Class Codes: A61P 1/18 for pancreatic disorders, e.g. pancreatic enzymes
• CPC Class Codes: C12N 2503/02 Drug screening
• CPC Class Codes: C12N 5/0678 Stem cells; Progenitor cells; Precursor cells
• CPC Class Codes: C12Q 1/6881 for tissue or cell typing, e.g. human leukocyte antigen [HLA] probes
• CPC Class Codes: C12Q 2600/158 Expression markers
• CPC Class Codes: G01N 33/5005 involving human or animal cells immunoassay G01N33/56966; immunoassays of protozoa G01N33/56905; protozoa in screening assays C12Q1/025

Abstract

The present invention provides a method of separating a pancreatic stem cell from the pancreas of a mammal, a method of identifying a pancreatic stem cell of a mammal, and use of a pancreatic stem cell that can be separated or identified by this method. More particularly, the present invention provides a method of separating or identifying a pancreatic stem cell of a mammal, which includes analyzing the state of expression of 2 or more marker proteins selected from the group consisting of c-Met, c-Kit, CD45, TER119 and Flk-1, or a gene encoding the marker protein.

Claims

1. A method of separating a pancreatic stem cell from the pancreas of a mammal using two or more kinds of substances having specific affinity for a marker protein selected from the group consisting of c-Met, c-Kit, CD45 and TER119, or a gene encoding the same.

3. The method of claim 1, wherein the substance having specific affinity is an antibody against the marker protein.

11. A pancreatic stem cell that can be separated from the pancreas of a mammal by the method described in claim 1.

12. The pancreatic stem cell of claim 11, which shows 4 markers of c-Met, c-Kit, CD45 and TER119 in a pattern of c-Met⁺, c-Kit⁻

, CD45⁻

, TER119⁻

.

13. The pancreatic stem cell of claim 11, which shows 5 markers of c-Met, c-Kit, CD45, TER119 and Flk-1 in a pattern of c-Met⁺, c-Kit⁻

, CD45⁻

, TER119⁻

, Flk-1⁻

.

15. A method of screening a substance that induces differentiation into liver•

bile duct or stomach•

intestine of a mammal, which comprises the following steps;

(i) a step of reacting a pancreatic stem cell of claim 12 with a test substance, and

(ii) a step of determining the expression of a liver•

bile duct or stomach•

intestine marker in the cell after the reaction.

17. A method of screening a substance that regulates the function of liver•

bile duct or stomach•

intestine of a mammal, which comprises the following steps;

(i) a step of reacting a pancreatic stem cell of claim 12 or a cell differentiated from the stem cell with a test substance, and

(ii) a step of determining the expression of a liver•

bile duct or stomach•

intestine marker in the cell after the reaction.

32. An agent for the prophylaxis or treatment of a pancreatic hypofunctional disease, which comprises the pancreatic stem cell of claim 12, or a cell differentiated from the stem cell.

33. The agent of claim 32, wherein the pancreatic hypofunctional disease is a disease selected from the group consisting of diabetes, chronic pancreatitis, autoimmune pancreatitis and pancreatic functional disorder from surgical removal of all or part of a pancreas.

34. An agent for the prophylaxis or treatment of a hypofunctional disease of the liver•

bile duct, which comprises the pancreatic stem cell of claim 12, or a cell differentiated from the stem cell.

35. The agent of claim 34, wherein the hypofunctional disease of the liver•

bile duct is a disease selected from the group consisting of acute hepatitis, chronic hepatitis, metabolic liver disease and hepatic functional disorder from surgical removal of all or part of a liver.

36. An agent for the prophylaxis or treatment of a hypofunctional disease of the stomach•

intestine, which comprises the pancreatic stem cell of claim 12, or a cell differentiated from the stem cell.

37. The agent of claim 36, wherein the hypofunctional disease of the stomach•

intestine is a disease selected from the group consisting of short bowel syndrome, inflammatory bowel disease, and stomach functional disorder from surgical removal of all or part of a stomach.

49. A method of screening a substance that induces differentiation into liver•

bile duct or stomach•

intestine of a mammal, which comprises the following steps;

(i) a step of reacting a pancreatic stem cell of claim 13 with a test substance, and

(ii) a step of determining the expression of a liver•

bile duct or stomach•

intestine marker in the cell after the reaction.

50. A method of screening a substance that regulates the function of liver•

bile duct or stomach•

intestine of a mammal, which comprises the following steps;

(i) a step of reacting a pancreatic stem cell of claim 13 or a cell differentiated from the stem cell with a test substance, and

(ii) a step of determining the expression of a liver•

bile duct or stomach•

intestine marker in the cell after the reaction.

57. An agent for the prophylaxis or treatment of a pancreatic hypofunctional disease, which comprises the pancreatic stem cell of claim 13, or a cell differentiated from the stem cell.

60. The agent of claim 57, wherein the pancreatic hypofunctional disease is a disease selected from the group consisting of diabetes, chronic pancreatitis, autoimmune pancreatitis and pancreatic functional disorder from surgical removal of all or part of a pancreas.

63. An agent for the prophylaxis or treatment of a hypofunctional disease of the liver•

bile duct, which comprises the pancreatic stem cell of claim 13, or a cell differentiated from the stem cell.

66. The agent of claim 63, wherein the hypofunctional disease of the liver•

bile duct is a disease selected from the group consisting of acute hepatitis, chronic hepatitis, metabolic liver disease and hepatic functional disorder from surgical removal of all or part of a liver.

69. An agent for the prophylaxis or treatment of a hypofunctional disease of the stomach•

intestine, which comprises the pancreatic stem cell of claim 13, or a cell differentiated from the stem cell.

72. The agent of claim 69, wherein the hypofunctional disease of the stomach•

intestine is a disease selected from the group consisting of short bowel syndrome, inflammatory bowel disease, and stomach functional disorder from surgical removal of all or part of a stomach.

2. A method of separating a pancreatic stem cell from the pancreas of a mammal using two or more kinds of substances having specific affinity for a marker protein selected from the group consisting of c-Met, c-Kit, CD45, TER119 and Flk-1, or a gene encoding the same.

38. The method of claim 2, wherein the substance having specific affinity is an antibody against the marker protein.

40. A pancreatic stem cell that can be separated from the pancreas of a mammal by the method described in claim 2.

43. The pancreatic stem cell of claim 40, which shows 4 markers of c-Met, c-Kit, CD45 and TER119 in a pattern of c-Met⁺, c-Kit⁻

, CD45⁻

, TER119⁻

.

46. The pancreatic stem cell of claim 40, which shows 5 markers of c-Met, c-Kit, CD45, TER119 and Flk-1 in a pattern of c-Met⁺, c-Kit⁻

, CD45⁻

, TER119⁻

, Flk-1⁻

.

4. A method of identifying a pancreatic stem cell of a mammal using two or more kinds of substances having specific affinity for a marker protein selected from the group consisting of c-Met, c-Kit, CD45 and TER119, or a gene encoding the same.

6. The method of claim 4, wherein the substance having specific affinity is an antibody against the marker protein.

5. A method of identifying a pancreatic stem cell of a mammal using two or more kinds of substances having specific affinity for a marker protein selected from the group consisting of c-Met, c-Kit, CD45, TER119 and Flk-1, or a gene encoding the same.

39. The method of claim 5, wherein the substance having specific affinity is an antibody against the marker protein.

7. A method of separating a pancreatic stem cell from the pancreas of a mammal, which comprises a step of analyzing the expression state of two or more marker proteins selected from the group consisting of c-Met, c-Kit, CD45 and TER119, or a gene encoding the same.

41. A pancreatic stem cell that can be separated from the pancreas of a mammal by the method described in claim 7.

44. The pancreatic stem cell of claim 41, which shows 4 markers of c-Met, c-Kit, CD45 and TER119 in a pattern of c-Met⁺, c-Kit⁻

, CD45⁻

, TER119⁻

.

47. The pancreatic stem cell of claim 41, which shows 5 markers of c-Met, c-Kit, CD45, TER119 and Flk-1 in a pattern of c-Met⁺, c-Kit⁻

, CD45⁻

, TER119⁻

, Flk-1⁻

.

8. A method of separating a pancreatic stem cell from the pancreas of a mammal, which comprises a step of analyzing the expression state of two or more marker proteins selected from the group consisting of c-Met, c-Kit, CD45, TER119 and Flk-1, or a gene encoding the same.

42. A pancreatic stem cell that can be separated from the pancreas of a mammal by the method described claim 8.

45. The pancreatic stem cell of claim 42, which shows 4 markers of c-Met, c-Kit, CD45 and TER119 in a pattern of c-Met⁺, c-Kit⁻

, CD45⁻

, TER119⁻

.

48. The pancreatic stem cell of claim 42, which shows 5 markers of c-Met, c-Kit, CD45, TER119 and Flk-1 in a pattern of c-Met⁺, c-Kit⁻

, CD45⁻

, TER119⁻

, Flk-1⁻

.

9. A method of identifying a pancreatic stem cell from the pancreas of a mammal, which comprises a step of analyzing the expression state of two or more marker proteins selected from the group consisting of c-Met, c-Kit, CD45 and TER119, or a gene encoding the same.

10. A method of identifying a pancreatic stem cell from the pancreas of a mammal, which comprises a step of analyzing the expression state of two or more marker proteins selected from the group consisting of c-Met, c-Kit, CD45, TER119 and Flk-1, or a gene encoding the same.

14. A method of screening a substance that induces differentiation of a pancreatic stem cell of a mammal, which comprises the following steps:

(i) a step of reacting a pancreatic stem cell with a test substance, and

(ii) a step of determining the expression of a pancreatic marker in the cell after the reaction.

16. A method of screening a substance that regulates a pancreatic function of a mammal, which comprises the following steps:

(i) a step of reacting a pancreatic stem cell or a cell differentiated from the stem cell with a test substance, and

(ii) a step of determining the expression of a pancreatic marker in the cell after the reaction.

18. A cloned pluripotent pancreatic stem cell, which satisfies at least 2 of the characteristics selected from the group consisting of c-Met⁺, c-Kit⁻

, CD45⁻

and TER119⁻

.

20. A pharmaceutical composition, which comprises:

i) a cloned pluripotent pancreatic stem cell of claim 18 and

ii) a pharmaceutically acceptable carrier.

21. A purified composition, which comprises:

a cloned pluripotent pancreatic stem cell of claim 18.

22. Tissue regenerated from a cloned pluripotent pancreatic stem cell of claim 18.

23. An organ regenerated from a cloned pluripotent pancreatic stem cell of claim 18.

24. A method of transplanting a cloned pluripotent pancreatic stem cell into a host, which comprises:

i) obtaining the cloned pluripotent pancreatic stem cell of claim 18;

and

ii) transplanting said stem cell into the host.

58. An agent for the prophylaxis or treatment of a pancreatic hypofunctional disease, which comprises the pancreatic stem cell of claim 18, or a cell differentiated from the stem cell.

61. The agent of claim 58, wherein the pancreatic hypofunctional disease is a disease selected from the group consisting of diabetes, chronic pancreatitis, autoimmune pancreatitis and pancreatic functional disorder from surgical removal of all or part of a pancreas.

64. An agent for the prophylaxis or treatment of a hypofunctional disease of the liver•

bile duct, which comprises the pancreatic stem cell of claim 18, or a cell differentiated from the stem cell.

67. The agent of claim 64, wherein the hypofunctional disease of the liver•

bile duct is a disease selected from the group consisting of acute hepatitis, chronic hepatitis, metabolic liver disease and hepatic functional disorder from surgical removal of all or part of a liver.

70. An agent for the prophylaxis or treatment of a hypofunctional disease of the stomach•

intestine, which comprises the pancreatic stem cell of claim 18, or a cell differentiated from the stem cell.

73. The agent of claim 70, wherein the hypofunctional disease of the stomach•

intestine is a disease selected from the group consisting of short bowel syndrome, inflammatory bowel disease, and stomach functional disorder from surgical removal of all or part of a stomach.

19. A cloned pluripotent pancreatic stem cell, which satisfies at least 2 of the characteristics selected from the group consisting of c-Met⁺, c-Kit⁻

, CD45⁻

, TER119⁻

and Flk-1⁻

.

51. A pharmaceutical composition, which comprises:

i) a cloned pluripotent pancreatic stem cell of claim 19;

and

ii) a pharmaceutically acceptable carrier.

52. A purified composition, which comprises:

a cloned pluripotent pancreatic stem cell of claim 19.

53. Tissue regenerated from a cloned pluripotent pancreatic stem cell of claim 19.

54. An organ regenerated from a cloned pluripotent pancreatic stem cell of claim 19.

55. A method of transplanting a cloned pluripotent pancreatic stem cell into a host, which comprises:

i) obtaining the cloned pluripotent pancreatic stem cell of claim 19;

and

ii) transplanting said stem cell into the host.

59. An agent for the prophylaxis or treatment of a pancreatic hypofunctional disease, which comprises the pancreatic stem cell of claim 19, or a cell differentiated from the stem cell.

62. The agent of claim 59, wherein the pancreatic hypofunctional disease is a disease selected from the group consisting of diabetes, chronic pancreatitis, autoimmune pancreatitis and pancreatic functional disorder from surgical removal of all or part of a pancreas.

65. An agent for the prophylaxis or treatment of a hypofunctional disease of the liver•

bile duct, which comprises the pancreatic stem cell of claim 19, or a cell differentiated from the stem cell.

68. The agent of claim 65, wherein the hypofunctional disease of the liver•

bile duct is a disease selected from the group consisting of acute hepatitis, chronic hepatitis, metabolic liver disease and hepatic functional disorder from surgical removal of all or part of a liver.

71. An agent for the prophylaxis or treatment of a hypofunctional disease of the stomach•

intestine, which comprises the pancreatic stem cell of claim 19, or a cell differentiated from the stem cell.

74. The agent of claim 71, wherein the hypofunctional disease of the stomach•

intestine is a disease selected from the group consisting of short bowel syndrome, inflammatory bowel disease, and stomach functional disorder from surgical removal of all or part of a stomach.

25. A method of producing a pancreatic stem cell, which comprises:

i) providing cells from the pancreas of a mammal; and

ii) selecting cells which satisfies at least 2 of the marker protein expression patterns selected from the group consisting of c-Met⁺, c-Kit⁻

, CD45⁻

and TER119⁻

, or a gene encoding the same.

27. The method according to claim 25, wherein the step of selecting cells further comprises selecting cells using an antibody having a specific affinity against the marker protein.

26. A method of producing a pancreatic stem cell, which comprises:

i) providing cells from the pancreas of a mammal; and

ii) selecting cells which satisfies at least 2 of the marker protein expression patterns selected from the group consisting of c-Met⁺, c-Kit⁻

, CD45⁻

, TER119⁻

and Flk-1⁻

or a gene encoding the same.

56. The method according to claim 26, wherein the step of selecting cells further comprises selecting cells using an antibody having a specific affinity against the marker protein.

28. A method of screening for a pancreatic stem cell of a mammal, which comprises;

i) containing a population of cells with an antibody having specific affinity for a marker protein selected from the group consisting of c-Met, c-Kit, CD45 and TER119, or a gene encoding the same; and

ii) fractionating the population of cells to obtain a cell having multi-differentiation ability.

29. The method of claim 28, wherein the cells are pancreatic cells.

30. The method of claim 28, wherein the cells express a marker protein pattern of c-Met⁺, c-Kit⁻

, CD45⁻

and TER119⁻

.

31. The method of claim 28, wherein the cells express a marker protein pattern of c-Met⁺, c-Kit⁻

, CD45⁻

, TER119⁻

and Flk-1⁻

.