Hepatic Differentiation of Murine Disease-Specific Induced Pluripotent Stem Cells Allows Disease Modelling In Vitro

Figure 4

FAH^-/--iPS cells. (a)–(c) iPS generated from fibroblasts isolated from FAH^−/−-mice using retroviral vectors expressing human OCT4, KLF4, SOX2, and c-MYC were stained for murine Nanog and Oct4. (d) Karyotype analysis of DAPI stained metaphase spreads. (e) Expression of endogenous Oct4, Nanog, and Sox2 determined by qRT PCR compared to ES cells. (f)–(h) FAH^−/−-iPSCs were able to form tissues from all three germ layers when transplanted subcutaneously into NOD/SCID mice. (i) qRT PCR analysis for hepatic marker genes Afp, Alb, Ck18, Abcc2, Ttr, and Hnf4α. Undifferentiated cells served as negative control, in which no hepatic gene expression was determined until cycle 45 (↓). (j)-(k) FAH^−/−-iPSCs differentiated using the hanging drop method and transduced with lentiviral Alb-eGFP expression vector. (l)-(m) FAH^−/−-iPSCs differentiated with the same method but with NTBC treatment. The NTBC-treatment rescued the lethal phenotype of hepatic cells, and more Alb-eGFP positive cells were detectable.