Covalent linkage of recombinant hirudin to poly(ethylene terephthalate) (Dacron): creation of a novel antithrombin surface

摘要: Abstract Thrombus formation and intimal hyperplasia on the surface of implantable biomaterials such as poly(ethylene terepthalate) (Dacron) vascular grafts are major concerns when utilizing these materials in clinical setting. Thrombin, a pivotal enzyme blood coagulation cascade primarily responsible for thrombus smooth muscle cell activation, has been target numerous strategies to prevent this phenomenon from occurring. The purpose study was covalently immobilize potent, specific antithrombin agent recombinant hirudin (rHir) modified Dacron characterize vitro efficacy thrombin inhibition by novel biomaterial surface. Bovine serum albumin (BSA), which selected ‘basecoat’ protein, reacted with various molar ratios cross-linker sulphosuccinimidyl 4-( N -maleimidomethyl) cyclohexane-1-carboxylate (sulpho-SMCC; 1:5-1:50). These BSA-SMCC complexes were then linked sodium hydroxide-hydrolysed (HD) segments via 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride (EDC). Covalent linkage HD (HD-BSA-SMCC) not affected any sulpho-SMCC assayed. rHir, initially 2-iminothiolane (Traut's reagent) order create sulphydryl groups, bound HD-BSA-SMCC surfaces (HD-BSA-SMCC-S-rHir). 1:50 (BSA: sulpho-SMCC) HD-BSA-SMCC-S-rHir 22-fold more rHir (111 ng per mg Dacron) compared control also possessed greatest evaluated using chromogenic substrate assay thrombin. Further characterization demonstrated that maximum 20.43 NIHU, 14.6-fold greater than (1.4 NIHU). Thrombin results confirmed 125 L-thrombin binding experiments, had significantly adhesion segments. Non-specific I-thrombin release less Thus, demonstrate can be clinically utilized while still maintaining its ability bind inhibit