Concept for Organ Engineering: A Reconstruction Method of Rat Liver for in Vitro Culture

Abstract

In the past decade, there have been remarkable advances in tissue engineering technology toward the goal of creating organoids in vitro from cells and cellular scaffolding. Indeed, tissue-engineered organoids such as skin and cartilage, each with comparatively simple architectures, are presently at the clinical stage. However, conventional tissue engineering techniques have not allowed for the reconstruction of an organoid that mimics an organ of complex architecture of abundant vascular networks. We established a method for organ engineering that can remodel a rat liver into a reconstructed organoid without separating the majority of liver cells by a continuous three-step perfusion. The liver was perfused through its vascular system with a buffered balanced salt solution to cleanse blood from the organ, with a collagenase/dispase medium to deconstruct cellular scaffolds, and with a culture medium containing collagen type I to reorganize the multicellular architecture. The reconstructed organoid was then prepared by excising the perfused liver from the rat and culturing it at 37°C for 2 h. Histologically healthy parenchymal hepatocytes expressing albumin were observed in the excised organoid even after culture for 3 weeks. Furthermore, a fibroblast-implanted organoid was prepared by using a culture medium containing suspended fibroblasts in the third step of the perfusion procedure, demonstrating the efficacy of heterogeneous cells for the reconstruction of an organoid. This method may be applicable to the formation of organoids from other organs, such as kidney and spleen, each of which have abundant capillaries, and therefore the method provides a novel concept for the development of lab-grown organs, i.e., organ engineering.