Efficient Differentiation of iPSCs into Hepatocytes

The differentiation of iPSCs into hepatocytes was carried out as described previously35 (link) with slight modification. Briefly, iPSCs were cultured to 70% confluence before switching to a basal differentiation medium containing 100 ng/ml activin A (R&D systems) and 50 ng/ml WNT3a (R&D systems) for 6 days (step 1), followed by 10 ng/ml FGF2 (R&D systems) and 50 ng/ml BMP4 (R&D systems) in differentiation medium (half basal differentiation medium with half STEMdiff™ APEL™ medium (Stemcell Technologies)) for 4 days (step 2). On the following 4 days, 50 ng/ml FGF1 (R&D systems), 10 ng/ml FGF4 (R&D systems) and 25 ng/ml FGF8 (R&D systems) was applied in the differentiation medium (step 3). Finally, the cells were incubated with 20 ng/ml HGF (R&D systems) and 100 ng/ml Follistatin-288 (R&D systems) for 4 days with additional 20 ng/ml Oncostatin (R&D systems) for another 2 days (step 4). The medium was changed every other day during the 20-days differentiation period.