Granted Patent B2

US 9,125,743 · App. 11/826,523 · Granted Sep 8, 2015

Abstract

Claims (39)

1. A process of recellularizing a devitalized cartilage graft with viable cells to render the tissue vital, the process comprising

seeding recellularizable cells on the devitalized cartilage graft,

introducing a mechanical force of at least about 2 MPa on the graft after the recellularizable cells have been seeded onto the graft and are adherent to the graft and

applying at least one chemical stimulus to the graft by applying microparticles comprising the chemical stimulus to the graft, wherein said microparticles do not include enzyme linked microparticles.

2. A process for repairing a cartilage defect and implanting a cartilage graft into a human or animal, comprising

a) crafting a cartilage matrix into individual grafts;

b) cleaning and disinfecting said cartilage graft;

c) applying a pretreatment solution to said cartilage graft;

d) removing cellular debris using an extracting solution to produce a devitalized cartilage graft;

e) recellularizing said devitalized cartilage graft according to the process of claim 1

f) implanting said cartilage graft into said cartilage defect with or without an insertion device; and

g) sealing the implanted cartilage graft with the recipient tissue; and

wherein said devitalized cartilage graft is optionally stored between said removing cellular debris and said recellularizing steps.

3. The process of claim 2 , wherein said recellularizable cells are mixed with a carrier.

4. The process of claim 2 , wherein said devitalized cartilage graft is hyaline cartilage, elastic cartilage, or fibrocartilage.

5. The process of claim 4 , wherein said devitalized cartilage graft is in the form of whole condyles, whole plateaus, hemicondyles, hemiplateaus, femoral heads, phalanges, talus, tibia, fibula, rib, intervertebral discs, menisci, nose, ear, osteochondral plugs, cartilage discs, cartilage slices, cartilage curls, or cartilage flakes.

6. The process of claim 5 , wherein the cartilage portion and the bone portion of said devitalized osteochondral plug are recellularized with same or different cell type(s).

7. The process of claim 5 , wherein the circumferential surface and/or the superficial surface of said cartilage discs or the cartilage portion of said osteochondral plug is microperforated to facilitate the integration of graft tissue to a recipient tissue.

8. The process of claim 7 , wherein said microperforation comprises using enzyme-linked microparticles, micromachining, or a laser.

9. The process of claim 5 , wherein said devitalized cartilage slice or disc is recellularized with one or more than one type of recellularizable cells comprising autologous or allograft chondrocytes isolated from articular cartilage, fibrocartilage, or elastic cartilage; bone marrow aspirate; stromal cells from bone marrow, synovium, periostieum, perichondrium, muscle, dermis, umbilical cord blood, adipose tissue, or Warton's jelly; or pericytes.

10. The process of claim 2 , wherein said microparticles are microbeads, wherein said microbeads are conjugated with cytokines or bioactive growth supplements.

11. The process of claim 2 , wherein said recellularizable cells are a single cell type or mixed cell types.

12. The process of claim 2 , wherein the chemical stimulus is applied to the devitalized cartilage graft prior to cell seeding.

13. The process claim 12 , wherein said chemical stimulus is applied by spraying or blasting said microparticles comprising the chemical stimulus onto the devitalized cartilage graft.

14. The process of claim 2 , wherein the chemical stimulus is applied to the devitalized cartilage graft after cell seeding.

15. The process of claim 14 , further comprising coating the recellularized devitalized cartilage graft with a group of bioactive growth supplements.

16. The process of claim 2 , further comprising implanting the recellularized devitalized cartilage graft into a cartilage defect site in a recipient.

17. The process of claim 2 , wherein the mechanical force is at least one of centrifugal force, compression or positive pressure.

18. The process of claim 2 , wherein the mechanical force is applied at a frequency between about 0.001 and about 5 Hz.

19. The process of claim 2 , wherein the mechanical force is cycled between 0 and at least about 2 MPa.

20. The process of claim 2 , further comprising immobilizing a bonding agent to the graft.

21. The process of claim 20 , wherein the immobilizing the bonding agent comprises activating a carboxylic group of the bonding agent and contacting the activated bonding agent with the graft.

22. The process of claim 21 , wherein the activating comprises activating with EDC and NHS.

23. The process of claim 20 , wherein the bonding agent is selected from the group consisting of photoactive dyes, hyaluronidase, chondroitinase, collagenase, trypsin, superoxide dismutase (SOD), catalase, bioactive growth supplements, dihydroxyphenylalanine (DOPA) based adhesive, glucose, concentrated albumin, cyanoacrylate adhesive, gelatin-resorcin-formalin adhesive, chondroitin sulfate aldehyde N-acetylglucosamine (GlcNAc), mussel-based adhesive, poly(amino acid)-based adhesive, cellulose-based adhesive, synthetic acrylate-based adhesives, platelet rich plasma (PRP), monostearoyl glycerol co-Succinate (MGSA), monostearoyl glycerol co-succinate/polyethylene glycol (MGSAPEG) copolymers, collagen, gelatin, agarose, modified hyaluronic acid, fibrin, chitin, biotin, avidin, native or crosslinked chitosan, alginate, demineralized bone matrix, MATRIGEL, HUMAN EXTRACELLULAR MATRIX, homogenized connective tissue, proteoglycans, fibronectin, laminin, fibronectin, elastin, heparin, glycerol, polymethylmethacrylate, polyurethane, acryloilmorpholine, N,N-dimethyl acrylamide, N-vinyl pyrrolidone and tetrahydrofurfuryl methacrylate, hydroxyapatite, cross-linkage or functionalization of hyaluronan-based collagen and alginate, polyurethane, polylactic acid, sodium nitroprusside, cartilage matrix glycoprotein (CMGP), vitamins C, vitamin E, selenium, N-Acetylcysteine (NAC) estradiol, glutathione, melatonin, resveratrol, flavonoid, carotene, aminoguanidine, lycopene, glutaraldehyde, glyceraldehydes, genipin, glucose or ribose, poly(ethylene glycol) diepoxide crosslinker, poly(ethylene glycol) diglycidyl ether, EDC and NHS, transglutaminase, lysyl oxidase family, hexamethylene diisocyanate (HMDIC), dimethyl suberimidate (DMS), dimethyl-3-3′-dithiobispropionimidate (DTBP), acryl azide, or a combination thereof.

24. The process of claim 20 , wherein the bonding agent is selected from the group consisting of collagen, gelatin, agarose, modified hyaluronic acid, fibrin, chitin, biotin, avidin, native or crosslinked chitosan, alginate, demineralized bone matrix, MATRIGEL, human extracellular matrix, homogenized connective tissue, proteoglycans, fibronectin, laminin, fibronectin, elastin, heparin, glycerol, or a combination thereof.

25. The process of claim 2 , wherein the chemical stimulus comprises a bioactive growth supplement selected from the group consisting of a natural or recombinant FGF-family, TGF-family, IGF-1, PDGF, EGF, VEGF, HGF, PTHrP, Ihh, dexamethasone, insulin, transferrin, selenium, ITS, ascorbate, extractions of demineralized bone matrix, basement membrane, submucosa matrix, an IL-1αR antibody, TNF-α receptor antagonist, cyclooxygenase-2 specific inhibitors, MAP kinase inhibitors, NO synthase inhibitors, NE-κB inhibitors, inhibitors of MMP, and a combination thereof.

26. The process of claim 2 , wherein said microparticles are conjugated with a bioactive growth supplement selected from the group consisting of a natural or recombinant FGF-family, TGF-family, IGF-1, PDGF, EGF, VEGF, HGF, PTHrP, Ihh, dexamethasone, insulin, transferrin, selenium, ITS, ascorbate, extractions of demineralized bone matrix, basement membrane, submucosa matrix, an IL-1αR antibody, TNF-α receptor antagonist, cyclooxygenase-2 specific inhibitors, MAP kinase inhibitors, NO synthase inhibitors, NE-κB inhibitors, inhibitors of MMP, and a combination thereof.

27. The process of claim 2 , further comprising differentiating the recellularizable cells, wherein the chemical stimulus is a growth factor of TGF-family.

28. The process of claim 2 , further comprising proliferating the recellularizable cells.

Assignments (8)