Novel genipin-crosslinked acellular biogenic conduits for tissue engineering applications

Abstract

Background: Collagen-based conduits have been generated in-vivo stimulating a fibrotic response through the implantation of a non-resorbable material in animal models, creating biogenic substitutes. However, they often exhibit clinical limitations due to prolonged generation times, exclusive autologous use and insufficient mechanical strength. Consequently, decellularization and cross-linking could solve the aforementioned drawbacks, providing a non-immunogenic and ready-to-use natural substitute with enhanced biomechanical properties. Nevertheless, these processes may alter microarchitecture and biocompatibility. Hence, this is the first study to characterize ex-vivo the biogenic conduits of 1-and 2-months maturation time which were subjected to decellularization and genipin (GP) cross-linking procedures performing histological, structural, biomechanical, biocompatibility, and immunological analyses to identify the most suitable option for peripheral nerve regeneration. Results: Histological examination indicated consistent uniformity of the biogenic conduits at both timepoints post-implantation, maintaining their overall structural integrity and collagen pattern following decellularization and GP crosslinking treatments. Furthermore, no evidence of nuclear debris was observed in the decellularized groups at either stage of maturation, confirming the decellularization protocol’s efficiency. The substitutes with longer maturation time presented a generally higher preservation of ECM key components. In addition, the GP crosslinking significantly increased the resistance values of decellularized biogenic conduits, without drastically affecting the ex-vivo cell biocompatibility nor macrophage polarization rate phenotype. Conclusions: These findings indicate the suitability of our decellularization protocol for biogenic conduits, and subsequent crosslinking with GP improves their biomechanical properties without altering their biocompatibility or immunological profile, suggesting their potential as a ready-to-use tubular substitute for nerve and other tissue engineering applications.