Comparison of commercially available media for hepatic differentiation and hepatocyte maintenance

Abstract

Human hepatocytes are essential materials in pharmaceutical researches. Not only primary human hepatocytes (PHH) but also human iPS cell-derived hepatocyte-like cells (human iPS-HLCs) are expected to be applied as materials for pharmaceutical researches. To date, several culture media have been developed for culturing human hepatocytes. However, there have been no reports comparing these media to determine which is most suitable for culturing human hepatocytes. In this study, we compared five commercial media (Hepatocyte Culture Medium (HCM), HepatoZYME-SFM, Cellartis Power Primary HEP Medium, DMEM/F12, and William's E Medium (WEM)) to determine which is most suitable for culturing PHH and human iPS-HLCs. In hepatic differentiation of human iPS cells (day 14-25 of differentiation), albumin (ALB) and urea secretion abilities and CYP2C9, CYP2C19, and CYP3A4 activities were the highest when using HCM or WEM. During maintenance of human iPS-HLCs, ALB and urea producing abilities and CYP2C9, CYP2C19, and CYP3A4 activities were the highest when using HCM. Importantly, we found that human iPS-HLCs cultured in HCM were maintained for 3 weeks or more without impairment of their hepatic functions. These results suggest that it is necessary to select an optimal medium for hepatic differentiation and maintenance of human iPS-HLCs. In the case of PHH culture, there was little difference in hepatic functions among the five media. However, the CYP2C9, CYP2C19, and CYP3A4 activities were the highest when using HCM and WEM. In conclusion, it is important to select the optimal medium for specific application when carrying out pharmaceutical researches using human hepatocytes.

Conditions for maintenance of hepatocyte differentiation and function in 3D cultures

“…The average albumin secretion was within the normal range observed in vivo ( Figure 1 B) and comparable to, or greater than, that observed in other PHH models ( Davidson et al., 2016 ; Hu et al., 2018 ; Leite et al., 2016 ; Ong et al., 2017 ; Ramaiahgari et al., 2014 ; Toba et al., 2020 ; Török et al., 2011 ; Tostões et al., 2012 ). Albumin secretion was stable (defined here as within +/− 35% of first week) over time for the two commercial and the three WE-media, but lower from the spheroids cultured in the DF media ( Figure 1 C).…”

Enhancement and Maintenance of Hepatic Metabolic Functions by Controlling Three-Dimensional Aggregation of Cryo-Preserved Human iPS Cells Derived Hepatocyte-Like Cells

“…However. previous report shows the functions are quite lower than primary human hepatocytes 3,4, and protocol which facilitated further maturation is still under development 13,14 . Spheroid is thought to be a good strategy to enhance functionality due to compaction of the cells with high cell density leading to enhancement of cell-cell contact to mimic an in vivo-like microtissue.…”

“…Spheroid formation by our original method is previously 13 . In this experiment, 0.5 or 1 µl of the cell suspension was serially injected into the MC medium, using an electronic micropipette or automated 3D cell injection system (Mito Kogyo Company Limited, Japan), which is utilized for large-scale production of the spheroids (more than 800 spheroids).…”

iPSC-derived hepatocytes generated from NASH donors provide a valuable platform for disease modeling and drug discovery

“…There are numerous published protocols for hepatic differentiation of human embryonic and pluripotent stem cells ( Fig. 1 A and Toba et al, 2020 ), but these lack robustness for performance across donor lines and for the most part yield boutique quantities of cells at the end of the process. The protocol described here routinely produces tens of millions of cryopreserved cells across multiple lines from AHN and NASH donors.…”