title: Engineered aprotinin for improved stability of fibrin biomaterials.
abstract: Fibrin has been long used clinically for hemostasis and sealing, yet extension of use in other applications has been limited due to its relatively rapid resorption in vivo, even with addition of aprotinin or other protease inhibitors. We report an engineered aprotinin variant that can be immobilized within fibrin and thus provide extended longevity. When recombinantly fused to a transglutaminase substrate domain from α(2)-plasmin inhibitor (α(2)PI(1-8)), the resulting variant, aprotinin-α(2)PI(1-8), was covalently crosslinked into fibrin matrices during normal thrombin/factor XIIIa-mediated polymerization. Challenge with physiological plasmin concentrations revealed that aprotinin-α(2)PI(1-8)-containing matrices retained 78% of their mass after 3 wk, whereas matrices containing wild type (WT) aprotinin degraded completely within 1 wk. Plasmin challenge of commercial sealants Omrixil and Tisseel, supplemented with aprotinin-α(2)PI(1-8) or WT aprotinin, showed extended longevity as well. When seeded with human dermal fibroblasts, aprotinin-α(2)PI(1-8)-supplemented matrices supported cell growth for at least 33% longer than those containing WT aprotinin. Subcutaneously implanted matrices containing aprotinin-α(2)PI(1-8) were detectable in mice for more than twice as long as those containing WT aprotinin. We conclude that our engineered recombinant aprotinin variant can confer extended longevity to fibrin matrices more effectively than WT aprotinin in vitro and in vivo.
authors.affiliation: Institute of Bioengineering, Ecole Polytechnique Fédérale de Lausanne (EPFL), Lausanne, Switzerland.
authors.forename: Kristen M
authors.initials: KM
authors.lastname: Lorentz
authors.forename: Stephan
authors.initials: S
authors.lastname: Kontos
authors.forename: Peter
authors.initials: P
authors.lastname: Frey
authors.forename: Jeffrey A
authors.initials: JA
authors.lastname: Hubbell
mesh_terms: D000818:Animals; D007611:Aprotinin / Q000737:chemistry*; D001672:Biocompatible Materials / Q000737:chemistry* / Q000378:metabolism; D002478:Cells, Cultured; D005260:Female; D005337:Fibrin / Q000737:chemistry*; D015718:Fibrin Tissue Adhesive / Q000737:chemistry; D005341:Fibrinolysin / Q000737:chemistry; D006801:Humans; D051379:Mice; D008810:Mice, Inbred C57BL; D014945:Wound Healing / Q000502:physiology
chemical_list: D001672:Biocompatible Materials; D015718:Fibrin Tissue Adhesive; D005337:Fibrin; D007611:Aprotinin; D005341:Fibrinolysin