Cryopreservation of Hepatocyte Organoid-Derived Cells Generated from Human iPS Cells

Hepatic organoids (iHOs) generated from human-induced pluripotent stem cell-derived hepatocyte-like cells (HLCs) have attracted attention as a new source of hepatocytes to replace primary human hepatocytes (PHHs) in drug discovery research. Recently, we established iHOs from HLCs, which proliferated for more than 10 passages, and developed a two-dimensional (2D) culture protocol for iHOs, by which the cells showed much higher hepatic functions. In this study, we examined the cryopreservation of iHOs for the purpose of improving their versatility. Cryopreserved iHOs had comparable cell proliferative capacity to fresh iHOs. Hepatocyte marker gene expression levels in the cryopreserved iHOs were comparable to those without cryopreservation. When cryopreserved iHOs were directly matured by our 2D culture protocol, they showed higher hepatocyte marker gene expression levels and higher CYP3A4 activity than HLCs. Some of the hepatic functions in 2D-cultured cryopreserved iHOs were at levels comparable to those in PHHs. These results suggest that cryopreserved iHOs also differentiated into mature hepatocytes by 2D culture. We expect cryopreserved iHOs to be used as a user-friendly cell source for drug discovery research in many laboratories.