HUMAN LIVER SCAFFOLDS AS BASIS FOR RECELLULARIZATION AND ORGAN RECOVERY

Abstract

Background In Brazil, the donation waiting list for a liver is growing and the mortality rate in this list can reach 23%. Therefore, the investigation for new therapeutic proposals, such as the production of bio-artificial organs, is extremely relevant. From this perspective, hepatocyte-like cells, derived from iPS generated from blood cells collected from end-stage liver diseased patient, can be expanded ex vivo and repopulate liver scaffolds to be transplanted in patients who might end up dying on the waiting list. In this study we have evaluated the feasibility of using controlled perfusion of detergents to obtain decellularized human whole liver scaffolds (DHLS) from diseased liver. Methods One amyloidotic liver discarded for transplantation purpose by State Transplant Program from Rio de Janeiro (Brazil) was decellularized by freezing and thawing followed by detergent perfusion (Triton X-100, SDC and SDS) through the portal vein (PV) and inferior vena cava (IVC) in a fourteen-day process. Histological processing (HE, PAS, Picrosirius red, Congo red staining) and DNA content analysis were made to evaluate our new protocol of decellularization of diseased liver. Results Prior decellularization, the liver, weighing 2100g, presented harden, important hepatomegaly, blunted edge and yellowish. At the end of the decellularization process, DHLS retained an opaque and pale appearance. The weight of the liver was reduced by 40% (1333g) indicating loss of cellular content. In consonance, DNA content analysis indicated significant reduction from 404,8±136.9 [average ± standard deviation (SD)] before decellularization to 40±3,2 ng DNA/mg wet weight in the DHLS. H&E staining analysis showed that the decellularization process removed cells and preserved vascular structures and components of the DHLS. PAS staining for glycogen after process showed that hepatocytes were removed when compared to liver sample before decellularization. Picrosirius red staining revealed that collagen content was preserved in the acellular scaffold. Congo red staining confirmed intense protein deposit in the liver, such content was partially removed during decellularization. Conclusion Production of whole human liver scaffolds from discarded organ is feasible and may provide a natural structure for organ recovery through recellularization. These organs may offer an alternative to patients who have no life expectancy in donation waiting list.