Fig. 1

hTERT-transduced cell lines are phenotypically stable without senescence traits. A Schematic representation of the different strategies employed to obtain hepatocyte-like cells (HLC) from human dermal fibroblasts (HDF), i.e., exogenous constitutive expression of HNF4A, HNF1A and FOXA3 using independent lentiviral vectors (HLC-3F), a doxycycline-inducible polycistronic lentiviral vector expressing HNF4A, HNF1A and FOXA3 (TetO-HHFG; diHLC), TetO-HHFG expression in large-T antigen immortalized HDF (diHLC-LT) and TetO-HHFG expression in hTERT-transduced HDF (diHLC-T). B Representative phase contrast images of iHDF and iHDF-T at PD^v.low (just derived; approximately PD24), PD^low (PD 30–40) and PD^high (PD 100–110). Images depict confluent cultures when possible. Black bar equals 100 μm. C Estimated doubling time for PD^low and PD^high iHDF-T (n = 6; three replicates each). D Volcano plot of all genes expressed in PD^high vs. PD^low iHDF-T (three replicates each) by RNAseq. Plotted genes were expressed in all samples of at least one group (absolute reads > 10). Red dots correspond to genes differentially expressed defined as FDR < 0.05 and fold change > 2 (0.6%). E mRNA level of CDKN1A and CDKN2A in iHDF-T at PD^v.low, PD^low and PD^high. Gray bars represent primary human dermal fibroblasts at PD16 and PD54 (early senescent). Values correspond to the average of 2 experiments with four replicates each. ***p < 0.005. FNone of the senescence-related gene sets was significantly different between PD^low and PD^high iHDF-T according to GSEA (see details in the text). Filled bars: NES > 0; Dashed bars: NES < 0. Red line marks FDR = 0.05