Culturing different cell populations on a decellularized natural biostructure for organ reconstruction

• US 6,479,064 B1
• Filed: 12/29/1999
• Issued: 11/12/2002
• Est. Priority Date: 12/29/1999
• Status: Expired due to Term

• Current Assignee: Children's Medical Center Corporation
• Original Assignee: Children's Medical Center Corporation
• Inventors: Atala, Anthony
• Primary Examiner(s): Naff, David M.
• Application Number:
  US09/474,525
• Time in Patent Office: 1,049 Days
• Field of Search: 424/93.7, 424/423, 435/177, 435/180, 435/395
• US Class Current: 424/423
• 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: A61L 27/3604 characterised by the human or animal origin of the biological material, e.g. hair, fascia, fish scales, silk, shellac, pericardium, pleura, renal tissue, amniotic membrane, parenchymal tissue, fetal tissue, muscle tissue, fat tissue, enamel
• CPC Class Codes: A61L 27/3808 Endothelial cells
• CPC Class Codes: A61L 27/3839 characterised by the site of application in the body materials for artificial blood vessels A61L27/507; materials for use in artificial skin A61L27/60
• CPC Class Codes: A61L 27/3891 as distinct cell layers
• 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: A61P 13/02 of urine or of the urinary tract, e.g. urine acidifiers
• CPC Class Codes: A61P 13/10 of the bladder
• CPC Class Codes: A61P 13/12 of the kidneys
• CPC Class Codes: A61P 9/00 Drugs for disorders of the cardiovascular system
• CPC Class Codes: C12N 2502/28 Vascular endothelial cells
• CPC Class Codes: C12N 2503/04 Screening or testing on artificial tissues
• CPC Class Codes: C12N 2533/52 Fibronectin; Laminin
• CPC Class Codes: C12N 2533/54 Collagen; Gelatin
• CPC Class Codes: C12N 2533/90 Substrates of biological origin, e.g. extracellular matrix, decellularised tissue
• CPC Class Codes: C12N 5/0068 General culture methods using substrates for specific animal cell type C12N5/06
• CPC Class Codes: C12N 5/0686 Kidney cells

Abstract

artificial organs are reconstructed using a three-dimensional scaffold produced by decellularizing biostructures from a donor organ. The three-dimensional scaffold is perfused with isolated endothelial cells that develop to produce an endothelial tissue layer with a primitive vascular system that sustains the growth and development of a second cultured cell population. When grown in the three-dimensional scaffold containing the endothelial tissue layer, with the primitive vascular system, the cells of the second cell population proliferate, mature and differentiate into neomorphic organ structures that are analogous to their in vivo counterparts.

Claims

1. An artificial organ construct comprising:

a three-dimensional scaffold of connective tissue formed by decellularizing a natural biostructure to remove cellular content, which has been perfused with a first population of isolated and cultured endothelial cells, such that the endothelial cells attach to the three-dimensional scaffold to produce an endothelial tissue layer comprising a vascular system, and at least one second population of cultured cells that is different from the population of cultured endothelial cells, such that the second cell population attaches to the endothelial tissue layer comprising a vascular system and differentiates into a neomorphic organ structure.

2. The artificial organ of claim 1, wherein the natural biostructure is an organ selected from the group consisting of heart, kidney, liver, pancreas, spleen, bladder, ureter and urethra.

3. The artificial organ of claim 1, wherein the natural biostructure is a part of an organ selected from the group consisting of heart, kidney, liver, pancreas, spleen, bladder, ureter and urethra.

4. The artificial organ of claim 1, wherein the artificial organ construct is an artificial kidney construct.

5. The artificial organ of claim 1, wherein the scaffold is derived from a decellularized mammalian kidney.

6. The artificial organ construct of claim 1, wherein the endothelial cells are human endothelial cells.

7. The artificial organ construct of claim 1, wherein the second cell population comprises human kidney cells.

8. An artificial kidney construct comprising:

a three-dimensional scaffold of connective tissue formed by decellularizing a mammalian kidney to remove cellular content, which has been perfused with a population of isolated and cultured endothelial cells, such that the endothelial cells attach to the three-dimensional scaffold to produce an endothelial tissue layer comprising a vascular system, and a population of cultured kidney cells, such that the kidney cell population attaches to the endothelial tissue layer comprising the vascular system and differentiates into nephron structures.

9. The artificial kidney of claim 8, wherein the endothelial cells are human endothelial cells.

10. The artificial kidney of claim 8, wherein the kidney cells are human kidney cells.

11. A method of treating a subject with an organ disorder comprising:

implanting a three-dimensional scaffold of connective tissue formed by decellularizing a natural biostructure to remove cellular content, which has been perfused with a first population of isolated and cultured endothelial cells, such that the endothelial cells attach to the three-dimensional scaffold to produce an endothelial tissue layer comprising a vascular system, and at least one second population of cultured cells that is different from the population of cultured endothelial cells, such that the second cell population attaches to an endothelial tissue layer comprising a vascular system and differentiates into a neomorphic organ structure; and

monitoring the subject for a modulation in the organ disorder.

12. The method of claim 11, wherein the natural biostructure is an organ selected from the group consisting of heart, kidney, liver, pancreas, spleen, bladder, ureter and urethra.

13. The method of claim 11, wherein the natural biostructure is a part of an organ selected from the group consisting of heart kidney, liver, pancreas, spleen, bladder, ureter and urethra.

14. The method of claim 11, wherein the artificial organ construct is an artificial kidney construct.

15. The method of claim 11, wherein the scaffold is derived from a decellularized mammalian kidney.

16. The method of claim 11, wherein the endothelial cells are human endothelial cells.

17. The method of claim 11, wherein the second cell population comprises human kidney cells.

18. A method of treating a subject with a kidney disorder comprising;

implanting a three-dimensional scaffold of connective tissue formed by decellularizing a mammalian kidney to remove cellular content, which has been perfused with a population of isolated and cultured endothelial cells, such that the endothelial cells attach to the three-dimensional scaffold to produce an endothelial tissue layer comprising a vascular system, and a population of cultured kidney cells, such that the kidney cell population attaches to the endothelial tissue layer comprising the vascular system and differentiates into nephron structures; and

monitoring the subject for a modulation in the kidney disorder.

19. The method of claim 18, wherein the endothelial cells are human endothelial cells.

20. The method of claim 18, wherein the kidney cells are human kidney cells.

21. A method for screening a compound that modulates kidney cells comprising:

providing an artificial kidney construct with a three-dimensional scaffold of connective tissue formed by decellularizing a mammalian kidney to remove cellular content, which has been perfused with a population of isolated and cultured endothelial cells, such that the endothelial cells attach to the three-dimensional scaffold to produce an endothelial tissue layer comprising vascular system, and a population of cultured kidney cells, such that the kidney cell population attaches to the endothelial tissue layer comprising the vascular system and differentiates into nephron structures;

contacting the artificial kidney construct with a library of test compounds;

selecting from the library of test compounds a compound of interest that modulates kidney cells.

22. The method of claim 21, wherein the modulator is cytotoxic to the kidney cells.

23. The method of claim 21, wherein the modulator is therapeutic to the kidney cells.

24. The method of claim 21, wherein the compound is a chemical agent.

25. The method of claim 21, wherein the compound is a pharmaceutical agent.

26. A method for processing an aqueous solution comprising:

providing an artificial kidney construct having a three-dimensional scaffold of connective tissue formed by decellularizing a mammalian kidney to remove cellular content, which has been perfused with a population of isolated and cultured endothelial cells, such that the endothelial cells attach to the three-dimensional kidney scaffold to produce an endothelial tissue layer comprising a vascular system, and a population of cultured kidney cells, such that the kidney cell population attaches to the endothelial tissue layer comprising the vascular system and differentiates into nephron structures;

delivering the aqueous solution to a luminal side of the artificial kidney construct;

collecting a processed aqueous solution from an abluminal side of the artificial kidney construct.

27. The method of claim 26, wherein the aqueous solution is unfiltered blood.

28. The method of claim 26, wherein the processed aqueous solution is filtered blood.

29. A method of reconstructing an artificial organ construct comprising:

decellularizing a natural biostructure by removing cellular content to produce a three-dimensional scaffold of connective tissue;

perfusing a first population of isolated and cultured endothelial cells into the three-dimensional scaffold formed by decellularizing a natural biostructure, such that endothelial cells attach to the three dimensional scaffold;

culturing the endothelial cells in the scaffold until the endothelial cells produce an endothelial tissue layer comprising a vascular system;

seeding at least one second population of cultured cells that is different from the population of cultured endothelial cells into the three-dimensional scaffold such that the second cell population attaches to the endothelial tissue layer comprising the vascular system and differentiates into a neomorphic organ structure.

30. The method of claim 29, wherein the natural biostructure is an organ selected from the group consisting of heart, kidney, liver, pancreas, spleen, bladder, ureter and urethra.

31. The method of claim 29, wherein the natural biostructure is a part of an organ selected from the group consisting of heart, kidney, liver, pancreas, spleen, bladder, ureter and urethra.

32. The method of claim 29, wherein the artificial organ construct is an artificial kidney construct.

33. The method of claim 29, wherein the scaffold is derived from a decellularized mammalian kidney.

34. The method of claim 29, wherein the endothelial cells are human endothelial cells.

35. The method of claim 29, wherein the second cell population comprises human kidney cells.

36. A method of reconstructing an artificial kidney construct comprising:

decellularizing a mammalian kidney by removing cellular content to produce a three-dimensional scaffold of connective tissue;

perfusing a population of isolated and cultured endothelial cells into the three-dimensional scaffold formed by decellularizing a mammalian kidney, such that endothelial cells attach to the three-dimensional scaffold;

culturing the endothelial cells in the three-dimensional scaffold until the endothelial cells produce an endothelial tissue layer comprising a vascular system;

seeding a population of cultured kidney cells into the three-dimensional scaffold such that the kidney cell population attaches to the endothelial tissue layer comprising the vascular system and differentiates into nephron structures.

37. The method of claim 36, wherein the endothelial cells are human endothelial cells.

38. The method of claim 36, wherein the kidney cells are human kidney cells.