Micro and ultrastructural changes monitoring during decellularization for the generation of a biocompatible liver.

ABSTRACT

Decellularization of a whole organ is an attractive process that has been used to create 3D scaffolds structurally and micro-architecturally similar to the native one. Currently used decellularization protocols exhibit disrupted extracellular matrix (ECM) structure and denatured ECM proteins. Therefore, maintaining a balance between ECM preservation and cellular removal is a major challenge. The aim of this study was to optimize a multistep Triton X-100 based protocol (either using Triton X-100/ammonium hydroxide mixture alone or after its modification with DNase, sodium dodecyl sulfate or trypsin) that could achieve maximum decellularization with minimal liver ECM destruction suitable for subsequent organ implantation without immune rejection. Based on our findings, Triton X-100 multistep protocol was insufficient for whole liver decellularization and needed to be modified with other detergents. Among all Triton X-100 modified protocols, a Triton X-100/DNase-based one was considered the most suitable. It maintains a gradual but sufficient removal of cells to generate decellularized biocompatible liver scaffolds without any significant alteration to ECM micro- and ultra-structure.