Rapid hepatic perfusion decellularization: technique and critique.

Abstract

Organ shortage facing the increasing success of liver transplantation has provoked research into the utilization of animal organs for clinical transplantation. The technique of whole-organ decellularization aims at the removal of the antigenic cellular content, thus evading the immune rejection cascade and the production of complex three-dimensional extracellular matrices of the entire organs with preservation of their intrinsic vascular networks rendering them transplantable. The aim of this study was the production of decellularized rabbit liver matrices by applying a simple, rapid perfusion decellularization technique and their characterization (both qualitatively and quantitatively). Decellularization of the caudate hepatic lobes of New Zealand white rabbits (n=22) was achieved through sequential perfusion of the portal venous system with deionized water, 0.8% Triton X-100 and 0.8% sodium dodecyl sulphate (SDS). Decellularized specimens were characterized both qualitatively (histology, fluoroscopy, corrosion casting and scanning electron microscopy) and quantitatively (total collagen assay [colorimetric] and total DNA assay [Hoechst 33258]). A Student's t-test was used to compare quantitative laboratory results before and after decellularization. A probability (P) value of <0.05 was considered significant. Effective decellularization was achieved as proven by histology and quantitative assessment (DNA remnants <1.5%, P=0.0009), while preserving 68% of the total collagen content (P=0.003). Portal vascular network integrity was confirmed by fluoroscopy and corrosion casting. Scanning electron microscopy also confirmed the preservation of the three-dimensional architecture. Liver perfusion decellularization technique using both 0.8% Triton X-100 and 0.8% SDS is a simple and rapid technique, yielding efficiently decellularized liver matrices preserving their vascular integrity, 3D architecture and 68% of total collagen content.