Cell-free tissue replacement for tissue engineering
View Patent ↗The present invention is a natural, cell-free tissue replacement that does not require difficult or extensive preparation made by washing tissue replacement in a solution including one or more sulfobetaines and an anionic surface-active detergent and washing the tissue replacement in serial solutions of the buffered salt to remove excess detergent. The natural, cell-free tissue replacement may be a nerve graft that supports axonal regeneration, guides the axons toward the distal nerve end and/or is immunologically tolerated. Other forms of the invention are a composition and kit prepared by the method of making a native, cell-free tissue replacement. The present invention may be modified for use in diagnostic, therapeutic, and prophylactic applications.
1. A method for preparing a native, acellular nerve tissue replacement comprising the steps of:
obtaining a nerve tissue;
soaking the nerve tissue for at least six hours in a solution comprising one or more sulfobetaines;
treating the nerve tissue in a mixture of one or more sulfobetaines and an anionic sulfonate detergent; and
washing the nerve tissue in one or more solutions of a buffered salt to remove the excess anionic sulfonate detergent to form the native, acellular nerve tissue replacement that is immunologically tolerated when implanted.
2. The method of claim 1 , further comprising the step of storing the native, acellular nerve tissue replacement in a buffered salt solution until needed.
3. The method of claim 1 , wherein the sulfobetaines have hydrophilic tails of 10 to 16 carbons.
4. The method of claim 1 , further comprising the step of adhering one or more bioactive agents to the nerve tissue replacement.
5. The method of claim 1 , wherein the native, acellular nerve tissue comprises mammalian tissue.
6. The method of claim 4 , wherein the one or more bioactive agents comprises a drug.
7. The method of claim 1 , wherein the native, acellular nerve tissue replacement further comprises a structure selected from the group consisting of a tube, a sheet, a film, a scaffold, and a tissue transplant for delivery into the body.
8. The method of claim 1 , wherein the sulfobetaine comprises SB-16.
9. The method of claim 1 , wherein the anionic sulfonate detergent is Triton X-200.
10. The method of claim 1 , wherein the step of washing the nerve tissue comprises one or more washes in a buffered salt solution comprising 100 mM sodium and 50 mM phosphate for at least 15 minutes each.
11. The method of claim 5 , wherein the nerve tissue is harvested from a mammalian cadaver.
12. The method of claim 11 , wherein the nerve tissue is cleaned of fat and blood after harvesting and rinsed two or more times in deionized distilled water.
13. The method of claim 1 wherein basal laminae and endoneurium layer of the native, acellular nerve tissue replacement substantially retain the native extracellular matrix structure.
14. The method of claim 1 , wherein the native, acellular nerve tissue replacement, when implanted, elicits a T-cell mediated immune response that is less than an immune response triggered by an allogeneic implant.
15. The method of claim 1 , wherein the native, acellular nerve tissue replacement allows for higher axon density when implanted relative to an untreated nerve tissue graft.
16. The method of claim 1 further comprising a step of: implanting the native, acellular nerve tissue into a mammal.
17. A method for preparing a native, acellular nerve tissue replacement comprising the steps of:
obtaining a nerve tissue;
soaking the nerve tissue for at least six hours in a solution comprising one or more sulfobetaines;
treating the nerve tissue in a mixture of one or more sulfobetaines and a detergent selected from the group consisting of polyether sulfonate sold under the trademark TRITON X-200, sulfosuccinate, polyether phosphate ester, alkoxylate sulfate, sodium deoxycholate, and low foam; and
washing the nerve tissue in one or more solutions of a buffered salt to remove the excess detergent to form the native, acellular nerve tissue replacement that is immunologically tolerated when implanted.