Tenocyte containing bioscaffolds and treatment using the same
The present invention relates to methods for preparing bio scaffolds useful in the repair of tears. More specifically, the invention relates to a method of treating rotator cuff tear in a mammalian subject in need thereof comprising the steps of: (i) selectively expanding tenocytes in vitro in culture medium comprising insulin or a functional derivative and a glucocorticoid or a glucocorticoid-like molecule to produce a culture of expanded tenocytes; (ii) seeding a bioscaffold with said expanded tenocytes to produce a tenocyte seeded bioscaffold; and (iii) implanting said tenocyte seeded bioscaffold proximal to the rotator cuff tear. The present invention also relates to a bioscaffold comprising cells, wherein more than 80% of said cells are tenocytes.
1. A method of treating rotator cuff tear in a mammalian subject in need thereof comprising the steps of:
(i) selectively expanding tenocytes in vitro in a culture medium comprising:
(a) at least 10% v/v serum;
(b) insulin; and
(c) betamethasone
to produce a culture of expanded tenocytes;
(ii) seeding a bioscaffold with said expanded tenocytes to produce a tenocyte seeded bioscaffold; and
(iii) implanting said tenocyte seeded bioscaffold proximal to the rotator cuff tear.
2. The method of claim 1 , wherein said rotator cuff tear is a massive rotator cuff tear.
3. The method of claim 1 , wherein said tenocytes are isolated from a tenocyte-containing tissue before selective expansion.
4. The method of claim 3 , wherein said tenocyte-containing tissue is a tendon.
5. The method of claim 1 , wherein said tenocytes are from a mammalian animal selected from the group consisting of a sheep, a cow, a pig and a human.
6. The method of claim 5 , wherein said mammalian animal is a human.
7. The method of claim 1 , wherein said tenocytes are autologous.
8. The method of claim 1 , wherein said culture medium comprises about 0.0006% w/v insulin and about 0.0002% w/v betamethasone.
9. The method of claim 1 , wherein said culture of expanded tenocytes comprises at least 80% tenocytes.
10. The method of claim 1 , wherein said culture of expanded tenocytes comprises at least 80% tenocyte cells that express one or more genes encoding the following: type I collagen, type III collagen, EphA4, scleraxis, Six1, COMP and Cbfa1.
11. The method of claim 1 , wherein said tenocyte-seeded bioscaffold is cultured in vitro for a sufficient time to establish the tenocytes before implantation.
12. The method of claim 1 , wherein said mammalian subject is a human.
13. The method of claim 1 , wherein said bioscaffold comprises a matrix, a membrane, a microbead, a fleece, a thread, a gel, or mixtures thereof.
14. The method of claim 1 , wherein said bioscaffold comprises a type I/III collagen matrix or a small intestinal submucosa.
15. The method of claim 1 , wherein the step of seeding a bioscaffold with said expanded tenocytes to produce a tenocyte seeded bioscaffold is carried out at a cell density of 0.05 to 5 times the physiological cell density of native tendon.
16. The method of claim 15 , wherein the cell density is at least 1×10 6 to 1×10 7 cells per cm 2 .
17. The method of claim 15 , wherein the cell density is 4×10 6 cells per cm 2 .
18. The method of claim 1 , wherein said culture medium comprises at least 15% v/v serum.
19. A method of treating massive rotator cuff tear in a human subject in need thereof comprising the steps of:
(i) isolating tenocytes from tendon tissue isolated from said subject;
(ii) selectively expanding the isolated tenocytes in vitro in a culture medium comprising:
(a) at least 10% v/v serum;
(b) about 0.0006% w/v insulin; and
(c) about 0.0002% w/v betamethasone
to produce a culture of expanded tenocytes comprising at least 80% tenocytes;
(iii) seeding a bioscaffold with said expanded tenocytes and culturing said bioscaffold and tenocytes for no more than five days to produce a tenocyte-seeded bioscaffold; and
(iv) implanting said tenocyte-seeded bioscaffold into the rotator cuff tear and securing with sutures.
20. The method of claim 19 , wherein said bioscaffold comprises a matrix, a membrane, a microbead, a fleece, a thread, a gel, or mixtures thereof.
21. The method of claim 19 , wherein said bioscaffold comprises a type I/III collagen matrix or a small intestinal submucosa.
22. The method of claim 19 , wherein the step of seeding a bioscaffold with said expanded tenocytes to produce a tenocyte seeded bioscaffold is carried out at a cell density of 0.05 to 5 times the physiological cell density of native tendon.
23. The method of claim 22 , wherein the cell density is at least 1×10 6 to 1×10 7 cells per cm 2 .
24. The method of claim 22 , wherein the cell density is 4×10 6 cells per cm 2 .
25. The method of claim 19 , wherein said culture medium comprises at least 15% v/v serum.
26. A method of treating a massive rotator cuff tear in a human subject in need thereof comprising the steps of:
(i) isolating tenocytes from tendon tissue isolated from said subject;
(ii) selectively expanding the isolated tenocytes in vitro in a culture medium comprising:
(a) at least 15% v/v serum;
(b) about 0.0006% w/v insulin; and
(c) about 0.0002% w/v betamethasone
to produce a culture of expanded tenocytes comprising at least 80% tenocytes;
(iii) seeding a bioscaffold with said expanded tenocytes to a cell density of 4×10 6 cells per cm 2 and culturing said bioscaffold and tenocytes for no more than five days in said culture medium to produce a tenocyte-seeded bioscaffold; and
(iv) implanting said tenocyte-seeded bioscaffold into the rotator cuff tear and securing with sutures.