Induction of a mature hepatocyte phenotype in adult liver derived progenitor cells by ectopic expression of transcription factors

Background/aims: By ectopic expression of a distinct combination of transcription factors we aimed to induce a mature hepatocyte phenotype in an adult liver derived progenitor cell population (ALDPC).

Methods: The open reading frames encoding murine Foxa2, Hnf4α and C/ebpα were cloned into lentivirus vectors and sequentially expressed in target cells. After seven days of culture, cells were analysed for expression of liver specific genes, and functional assays were performed. Fresh primary hepatocytes, twenty four hours in culture, served as positive controls.

Results: Untransduced ALDPC under established differentiation conditions exhibited moderate signs of maturation, in particular in comparison with fresh hepatocyte controls. In transcription factor transduced cells, fifteen mRNA´s coding for secreted proteins, cytochrome p450 isoenzymes, liver metabolic enzymes were detected by RT-qPCR at levels close to controls. Albumin secretion increased incrementally in single (Foxa2), double (Foxa2, Hnf4α) and triple-transduced cells (Foxa2, Hnf4α, C/ebpα) and reached levels observed in primary hepatocytes. Glycogen storage as determined by PAS staining was detectable in double and triple transduced cells, comparable to controls. Ureagenesis was also induced in triple transduced cells, but at lower levels compared to primary hepatocytes.

Conclusions: Sequential expression of Foxa2, Hnf4α and C/ebpα induces a mature hepatocyte phenotype in an expandable liver derived progenitor cell line.