title: Genipin blues: an alternative non-toxic crosslinker for heart valves?
abstract: OBJECTIVE
  One approach in tissue-engineering involves the implantation of decellularized, xenogenic scaffolds, with the expectation of repopulation in vivo. However, a major limitation of this method is the propensity to induce a strong immune host response. The study aim was to mitigate this immunogenicity by employing a crosslinking treatment with genipin.
  
  
  METHODS
  Porcine matrices were prepared using a detergent-enzymatic treatment and fixed in 0.01% or 0.001% aqueous genipin. The mechanical properties of the matrices were monitored by tensile strength testing. The survival of chicken fibroblasts was used to determine cell-friendliness of the matrices. Non-fixed, decellularized biological scaffolds (n = 3) were implanted in a sheep model and compared to an equal number of genipin-fixed scaffolds (n = 6). Matrices implanted in the pulmonary position were explanted after six weeks and examined using light and transmission electron microscopy. The antibody reaction against porcine tissue in sheep serum was also determined.
  
  
  RESULTS
  Statistically significant differences were found between non-fixed leaflets, 0.001% genipin-and 0.6% glutaraldehyde (GA)-fixed leaflets for work to maximum load (non-fixed 0.00646 J; genipin-fixed 0.00509 J; GA-fixed 0.00543 J) and stiffness (non-fixed 9281 N/m; genipin-fixed 16214 N/m; GA-fixed 14401 N/m). Genipin-treated matrices were not cytotoxic. For all concentrations of genipin a high proportion of viable cells was present (79-100%). Low-dose GA (10 microg/ml) showed a distinct cytotoxicity (24.8% viability). At explant, an intense chronic inflammatory response was observed in non-fixed matrices, in contrast to genipin-fixed scaffolds. The sheep serum showed a marked decrease in IgG response in both 0.001% and 0.01% genipin-fixed matrices (IgG 30 and 20, respectively) when compared to non-fixed matrices (IgG 40).
  
  
  CONCLUSIONS
  Genipin crosslinking of the matrices attenuated, but did not eliminate, the inflammatory host reaction. Whether genipin treatment might extend the durability of xenogenic scaffolds remains to be investigated.
authors.affiliation: Heart Center, University Hospital Ghent, Ghent, Belgium.
authors.forename: Pamela
authors.initials: P
authors.lastname: Somers
authors.forename: Filip
authors.initials: F
authors.lastname: De Somer
authors.forename: Maria
authors.initials: M
authors.lastname: Cornelissen
authors.forename: Stefaan
authors.initials: S
authors.lastname: Bouchez
authors.forename: Frank
authors.initials: F
authors.lastname: Gasthuys
authors.forename: Kishan
authors.initials: K
authors.lastname: Narine
authors.forename: Eric
authors.initials: E
authors.lastname: Cox
authors.forename: Guido
authors.initials: G
authors.lastname: Van Nooten
mesh_terms: D000818:Animals; D001705:Bioprosthesis; D003432:Cross-Linking Reagents / Q000737:chemistry*; D005976:Glutaral / Q000737:chemistry; D006350:Heart Valve Prosthesis; D007074:Immunoglobulin G / Q000097:blood; D007249:Inflammation / Q000473:pathology; D057889:Iridoid Glycosides; D039823:Iridoids / Q000737:chemistry*; D046529:Microscopy, Electron, Transmission; D011664:Pulmonary Valve / Q000276:immunology / Q000473:pathology; D012756:Sheep; D013552:Swine; D013718:Tensile Strength; D023822:Tissue Engineering; D054457:Tissue Scaffolds
chemical_list: D003432:Cross-Linking Reagents; D007074:Immunoglobulin G; D057889:Iridoid Glycosides; D039823:Iridoids; C007834:genipin; D005976:Glutaral