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Background Mature liver organoids are potential cellular sources for research to understand the pathology of several liver-related conditions, including end-stage chronic liver disease. Although several methods exist for the differentiation of mature hepatic organoids from human induced pluripotent stem cells (hiPSCs), organoid generation have limitations due to various experimental culture conditions. Methods We differentiated homozygous hiPSCs into definitive endoderm, hepatic endoderm, and hepatic maturation in a two-dimensional culture condition. Expandable hepatic organoids (EHO) and mature hepatic organoids (MHO) were cultured in a three-dimensional culture condition. Results We successfully established hepatic organoids that were reproductively and expandably cultured for at least three months. MHO showed significant increases in hepatic-specific markers, drug-metabolizing enzymes and transporter genes, and human albumin secretion. Conclusion Therefore, we established a standard operating protocol for gen¬erating mature and expandable hepatic organoids derived from hiPSCs, and made the start¬ing materials available to promote widespread use of the protocol.