Multi-Step Connective Tissue Stabilization Method and Stabilized Tissue Formed Thereby

Abstract

A multi-step stabilization method for connective tissue is described. Stabilized tissues can exhibit increased resistance to degradation due to enzyme activity, fatigue and storage. The multi-step method includes a first step during which the tissue can be incubated with a glycosaminoglycanase inhibitor such as a sulfated oligosaccharide, one example of which being neomycin, a second step during which the tissue can be incubated with a crosslink activator such as a carbodiimide crosslink activator and/or a crosslinking agent such as a heterobifunctional crosslinking agent and/or a phenolic compound such as a tannin, examples of which include tannic acid and pentagalloylglucose, and a third step during which the tissue can be incubated with a second crosslink activator that can be the same or different as the first crosslink activator.

What is claimed is: 
     
         1 . A method for stabilizing ex vivo or in vitro connective tissue comprising:
 a first step including contacting the ex vivo or in vitro connective tissue with a glycosaminoglycanase inhibitor, the connective tissue including collagen, elastin, and one or more glycosaminoglycans;   a second step carried out subsequent to the first step, the second step including contacting the ex vivo or in vitro connective tissue with a solution comprising a first crosslinking reagent and a phenolic compound comprising multiple phenolic groups and a hydrophobic core;   a third step carried out subsequent to the second step, the third step including contacting the connective tissue with a second crosslinking reagent.   
     
     
         2 . The method of  claim 1 , wherein the ex vivo or in vitro connective tissue is a component of dura mater, a tendon, a ligament, a dermal structure, a blood vessel, umbilical material, pericardium, fascia, or submucosa. 
     
     
         3 . The method of  claim 2 , wherein the ex vivo or in vitro connective tissue is a component of a blood vessel. 
     
     
         4 . The method of  claim 3 , wherein the blood vessel is a vein. 
     
     
         5 . The method of  claim 1 , wherein the glycosaminoglycanase inhibitor comprises neomycin or a salt of neomycin. 
     
     
         6 . The method of  claim 5 , wherein the glycosaminoglycanase inhibitor comprises neomycin trisulfate salt. 
     
     
         7 . The method of  claim 1 , wherein the first crosslinking reagent comprises a carbodiimide crosslinking reagent. 
     
     
         8 . The method of  claim 7 , wherein the first crosslinking reagent comprises 1-ethyl-3-(3 dimethyl-aminopropyl)carbodiimide. 
     
     
         9 . The method of  claim 1 , wherein the phenolic compound comprises a tannic acid or a derivative thereof. 
     
     
         10 . The method of  claim 1 , wherein the phenolic compound comprises pentagalloylglucose. 
     
     
         11 . The method of  claim 1 , wherein the second crosslinking reagent is the same as the first crosslinking reagent. 
     
     
         12 . A stabilized ex vivo or in vitro blood vessel comprising a glycosaminoglycanase inhibitor and a phenolic compound, the phenolic compound comprising multiple phenolic groups and a hydrophobic core. 
     
     
         13 . The stabilized ex vivo or in vitro blood vessel of  claim 12 , wherein the glycosaminoglycanase inhibitor is neomycin or a neomycin salt. 
     
     
         14 . The stabilized ex vivo or in vitro blood vessel of  claim 13 , wherein the glycosaminoglycanase inhibitor is neomycin trisulfate salt. 
     
     
         15 . The stabilized ex vivo or in vitro blood vessel of  claim 12 , wherein the phenolic compound comprises tannic acid or a derivative of tannic acid. 
     
     
         16 . The stabilized ex vivo or in vitro blood vessel of  claim 15 , wherein the phenolic compound comprises pentagalloylglucose. 
     
     
         17 . The stabilized ex vivo or in vitro blood vessel of  claim 12 , wherein the tissue is a component of a bioprosthetic implant. 
     
     
         18 . A bioprosthesis comprising a stabilized ex vivo or in vitro tissue, the stabilized tissue comprising a glycosaminoglycanase inhibitor and a phenolic compound, the phenolic compound comprising multiple phenolic groups and a hydrophobic core. 
     
     
         19 . The bioprosthesis of  claim 18 , wherein the glycosaminoglycanase inhibitor comprises neomycin or a neomycin salt. 
     
     
         20 . The bioprosthesis of  claim 18 , wherein the phenolic compound comprises pentagalloylglucose. 
     
     
         21 . The bioprosthesis of  claim 18 , wherein the bioprosthesis comprises a dermal graft, a vascular graft, a stent, a vascular or cardiovascular shunt, a dura mater graft, a cartilage graft, a cartilage implant, a pericardium graft, a ligament prosthesis, a tendon prosthesis, a urinary bladder prosthesis, a pledget, a suture, an artificial joint, an artificial limb, a bionic construct, or a surgical patch. 
     
     
         22 . The bioprosthesis of  claim 18 , further comprising an implantable support material in conjunction with the stabilized ex vivo or in vitro tissue. 
     
     
         23 . The bioprosthesis of  claim 18 , wherein the bioprosthesis is a transcatheter bioprosthesis.