Definitive Endoderm Formation from Plucked Human Hair-Derived Induced Pluripotent Stem Cells and SK Channel Regulation
Figure 2
Differentiation of hiPSCs into DE (definitive endoderm) cells. (a) Scheme of monolayer differentiation of hiPSCs into DE cells. (b) Immunocytochemistry shows that hiPSCs-derived DE cells express the early endodermal markers SOX 17 (red) or FOXA2 (green). Scale bars as indicated. (c) FACS analysis reveals that after 3 days of differentiation approximately 45% of the cells have become SOX17 positive. After 6 days of differentiation about 80% of the cells express SOX17. This data was consistent within two different hiPSC lines. (d) (e) Number of CXCR4/cKIT double-positive cells after 3 days and 5 days of differentiation, respectively. After 5 to 6 days of differentiation both hiPSCs lines express the highest amount of CXCR4/cKIT double-positive cells in the region of 75–80%. (f) Virus-containing hiPSCs (hiPSC_1/hiPSC_2) did not show differences in the number of CXCR4/cKIT double-positive cells compared to virus-free iPSCs (hiPSC_Cre). (g) Continuous loss of OCT4 mRNA levels during DE differentiation. In contrast, transcript levels of early endodermal genes such as SOX17 and FOXA2 steadily increased and reached highest levels after 5 days of differentiation. Expression levels are shown relative to the housekeeping gene HMBS (, two different hiPSCs lines).
