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Whole organ engineering for liver replacement: a regenerative medicine approach

Ken Fukumitsu, Alejandro Soto-Gutierrez

University of Pittsburgh, Pittsburgh, PA, United States

Liver transplantation is the only effective treatment for end-stage liver disease and the treatment is limited by the critical donor organ shortage. A promising regenerative medicine approach to address this issue is the construction of organs in vitro for transplantation. In this study, we aimed to regenerate liver tissue using adult hepatocytes, endothelial cells and biliary cells using decellularized rat livers as scaffolds in an organ perfusion system. Rat livers were decellularized using a new, minimally disruptive method based on trypsin/triton X. Native liver matrices were recellularized with adult primary rat hepatocytes for the parenchymal space, rat biliary cells for the bile duct and microvascular endothelial cells for the vascular system of the liver. The grafts were perfusion cultured for 5 d in vitro. The perfusate was sampled daily to assess hepatic secretory and synthesis function. Decellularized livers were examined by morphologic, biochemical, and immunolabeling techniques for preservation of the native matrix architecture and composition. The decellularization process preserves the three-dimensional macrostructure, the ultrastructure, the composition of the extracellular matrix components, the native microvascular network of the liver, and the bile drainage system, and up to 50% of growth factor content (HGF, bFGF). The three-dimensional liver matrix reseeded with the multistep infusion of hepatocytes generated ~90% of cell engraftment and supported liver-specific functional capacities of the engrafted cells, including albumin production, urea metabolism, and cytochrome P450 induction. More than 60% of the bile duct was adequately recellularized and the endothelial cells preferably aligned approximately 50% of vascular spaces. Whole-organ liver decellularization is possible with maintenance of structure and composition suitable to support hepatic function.