Development and Characterization of a Porcine Liver Scaffold

Abstract

The growing number of patients requiring liver transplantation for chronic liver disease cannot be currently met due to a shortage in donor tissue. As such, alternative tissue engineering approaches combining the use of acellular biological scaffolds and different cell populations (hepatic or progenitor) are being explored to augment the demand for functional organs. Our goal was to produce a clinically relevant sized scaffold from a sustainable source within 24 hours, whilst preserving the extra cellular matrix (ECM) to facilitate cell repopulation at a later stage. Whole porcine livers underwent perfusion de-cellularisation via the hepatic artery and hepatic portal vein using a combination of saponin, sodium deoxycholate (SOC) and deionised water washes resulting in an acellular scaffold with an intact vasculature and preserved ECM. Molecular and immuno-histochemical analysis (collagen I and IV and laminin) showed complete removal of any DNA material, together with excellent retention of glycosaminoglycans and collagen. FTIR analysis showed both absence of nuclear material and removal of any detergent residue, which was successfully achieved after additional ethanol gradient washes.Samples of the de-cellularised scaffold were assessed for cytotoxicity by seeding with porcine adipose derived mesenchymal stem cells in vitro, these cells over a 10 day period showed attachment and proliferation. Perfusion of the vascular tree with contrast media followed by CT imaging showed an intact vascular network. In vivo implantation of whole intact non-seeded livers, into a porcine model (as auxiliary graft) showed uniform perfusion macroscopically and histologically. Using this method, it is possible to create an acellular, clinically sized, liver scaffold with intact vasculature in less than 24 hours.

A decellularized human corneal scaffold for anterior corneal surface reconstruction

“…In this study, we established and validated a rapid alternative method (~ 1 day) for efficient decellularization of the human cornea using SD and DNAse. Our data indicate that 1% or 4% SD with DNAse and 4% dextran allow for efficient decellularization by removal of cellular components (HLA-ABC and nuclear debris) within 24hrs, in line with earlier reports on other tissues 11,17,18 .…”

“…Moreover, protocols using SDS or hypertonic salt with nucleases are very time-consuming (~ 5-8 days) 6,7,10 and failure to fully elute SDS can have cytotoxic effects 6,7 . Sodium deoxycholate (SD), a naturally occurring bile salt metabolite, has been used to decellularize various organs or tissues such as liver 11 , peripheral nerves 12 , ovary 13 , uterus 14 , conchal cartilage 15 , trachea 16 , lungs and heart valves 17 . The combination of SD/ deoxyribonuclease I (DNAse) was also used to generate decellularized human corneal limbus, however, a long elution time for removal of cellular components was a drawback of the protocol suggested 18 .…”

A Decellularized Human Corneal Scaffold for Anterior Corneal Surface Reconstruction

“…In this study, we established and validated a rapid alternative method for efficient decellularization of the human cornea using SD and DNAse. Our data indicate that 1 % or 4 % SD with DNAse and 4% dextran allow for efficient decellularization by removal of cellular components (HLA-ABC and nuclear debris) within 24hrs, in line with earlier reports on other tissues 11,17,18 .…”

Tissue-Specific Microparticles Improve Organoid Microenvironment for Efficient Maturation of Pluripotent Stem-Cell-Derived Hepatocytes

“…In this study, we successfully developed a simple and efficient method to produce liver-ECM-derived MPs with a round shape and homogenous size in the range of the typical size of single cells (~8 µm). Moreover, FTIR assessment of the LEM gel, MPs, and liver ECM verified the preservation of the key components of liver tissue after processing into the liver ECM solution and hydrogel MPs, which was consistent with previously reported data [1,23].…”

“…End-stage liver diseases account for almost two million deaths per year worldwide [1]. Moreover, drug-induced liver injury, as a public health concern, remains a potential health challenge [2].…”