Generation of human induced pluripotent stem cell-derived liver buds with chemically defined and animal origin-free media

• Published in: Scientific reports 2020-10, Vol.10 (1), p.17937-17937, Article 17937
• Main Authors: Sekine, Keisuke, Ogawa, Shimpei, Tsuzuki, Syusaku, Kobayashi, Tatsuya, Ikeda, Kazuki, Nakanishi, Noriko, Takeuchi, Kenta, Kanai, Eriko, Otake, Yugo, Okamoto, Satoshi, Kobayashi, Tsuyoshi, Takebe, Takanori, Taniguchi, Hideki
• Format: Article
• Language: English
• Subjects: 639/301/1023/1025; 639/301/357/1016; 639/301/357/537; 639/301/357/995; Animals; Cell culture; Cell Culture Techniques - methods; Cell Differentiation; Cells, Cultured; Contamination; Culture Media; Dietary supplements; Endothelial Cells; Hepatocytes; Humanities and Social Sciences; Humans; Induced Pluripotent Stem Cells - physiology; Liver; Liver - cytology; Liver diseases; Mesenchymal Stem Cells; Mesenchyme; Mimicry; multidisciplinary; Organoids; Pluripotency; Regenerative medicine; Regenerative Medicine - methods; Science; Science (multidisciplinary); Septum; Stem cells; Toxins
• ISSN: 2045-2322; 2045-2322
• DOI: 10.1038/s41598-020-73908-1

Advances in organoid technology have broadened the number of target diseases and conditions in which human induced pluripotent stem cell (iPSC)-based regenerative medicine can be applied; however, mass production of organoids and the development of chemically defined, animal origin-free (CD-AOF) media and supplements are unresolved issues that hamper the clinical applicability of these approaches. CD-AOF media and supplements ensure the quality and reproducibility of culture systems by lowering lot-to-lot variations and the risk of contamination with viruses or toxins. We previously generated liver organoids from iPSCs, namely iPSC-liver buds (iPSC-LBs), by mimicking the organogenic interactions among hepatocytes, endothelial cells (ECs), and mesenchymal cells (MCs) and recently reported the mass production of iPSC-LBs derived entirely from iPSCs (all iPSC-LBs), which should facilitate their large-scale production for the treatment of liver failure. However, in previous studies we used media originating from animals for differentiation except for the maintenance of undifferentiated iPSCs. Therefore, we developed a CD-AOF medium to generate all iPSC-LBs. We first developed a CD-AOF medium for hepatocytes, ECs, and stage-matched MCs, i.e., septum transversum mesenchyme (STM), in 2D cultures. We next generated all iPSC-LBs by incubating individual cell types in ultra-low attachment micro-dimple plates. The hepatic functions of all iPSC-LBs generated using the CD-AOF medium were equivalent to those of all iPSC-LBs generated using the conventional medium both in vitro and in vivo. Furthermore, we found that this CD-AOF medium could be used in several cell culture settings. Taken together, these results demonstrate the successful development of a CD-AOF medium suitable for all iPSC-LBs. The protocol developed in this study will facilitate the clinical applicability of all iPSC-LBs in the treatment of liver diseases.