Impact of heart valve decellularization on 3-D ultrastructure, immunogenicity and thrombogenicity

Decellularized xenogeneic tissue represents an interesting material for heart valve tissue engineering. The prospect objective is removal of all viable cells while preserving extracellular matrix (ECM) integrity. The major concerns of all decellularization protocols remain ECM disruption, immunogenicity and thrombogenicity. Accordingly the aim of this study was visualization of ultrastructural ECM disruption and human immune response and thrombogenicity using different decellularization protocols of porcine heart valves. Porcine pulmonary leaflets were decellularized with four different protocols: sodium deoxycholate, sodium dedecylsulfate, trypsin/EDTA, and trypsin-detergent-nuclease. Then the tissues were processed for histology and two-photon laser scanning microscopy (LSM). For thrombogenicity and immunogenicity testing tissues were incubated with human blood. The histological examination revealed no remaining cells and no significant differences in the ECM histoarchitecture in any group. LSM detected significant ECM alterations in all groups except sodium deoxycholate group with an almost completely preserved ECM. There was no increased immunogenicity between fresh and decellularized tissue. Compared to GA-fixed tissue however significantly increased immune responses and thrombogenicity was observed in all protocols. From our experiment, sodium deoxycholate enables cell removal with almost complete preservation of ECM structures. And all of these four decellularization protocols affected human immunological response and increased thrombogenicity.