Figure 5.

Effect of different media compositions and/or co-culture with human umbilical vein endothelial cells (HUVEC) on mRNA expression of stage-specific markers after hepatic differentiation of human induced pluripotent stem cells (hiPSC). Differentiation of definitive endodermal cells was performed over 14 days using hepatocyte culture medium (HCM-I/II), a 1:1 mixture of hepatocyte culture medium and endothelial cell growth medium, consisting of basal medium and supplements (HCM-I/II + EGM complete) or HCM enriched with endothelial cell growth supplements (HCM-I/II + EGM supplements) with or without HUVEC addition. In addition mRNA expression analysis was performed with HUVEC mono-cultures cultured in EGM complete as control. Graphs show POU class 5 homeobox 1 (POU5F1, A), α-fetoprotein (AFP, B), albumin (ALB, C), cytokeratin 18 (KRT18, D), hepatocyte nuclear factor 4 α (HNF4A, E) and platelet endothelial cell adhesion molecule 1 (PECAM1, F). Fold changes of mRNA expression were calculated relative to undifferentiated hiPSC with normalization to glyceraldehyde-3-phosphate‎ dehydrogenase (GAPDH) expression by the ∆∆Ct method. Differences between HCM and all other groups and differences between test media and their corresponding co-cultures were detected with the unpaired, two-tailed Student’s t-test, n = 8; Co-cultures: n = 3; mean ± standard error of the mean.