A New Type of Human iPS Cells: Human Induced Hepatic Lineage-Oriented Stem Cells

Human induced hepatic lineage-oriented stem cells (hiHSCs) can be established as a new type of human induced pluripotent stem cells (hiPSCs) with only some modifications of their culture conditions in both generation and self-renewal. hiHSCs were generated and expanded at a very high density under non-typical coculture with feeder cells in a chemically defined hiPSC medium . In detail, hiHSCs were generated from human non-hepatic cells by gene transfer with pantropic retrovirus vectors carrying the three transcription factor genes OCT3/4, SOX2, and KLF4. They were expanded at a very high density under non-typical coculture with mouse embryonic fibroblasts (MEF) in a chemically defined mTeSR1 medium on gelatin-coated dishes. Self-renewing hiHSCs expressed markers of both human embryonic stem cells (hESCs) and hepatocytes. The potential hepatic specification of hiHSCs was defined by their expression profiles of hepatic markers under a self-renewal culture. Using the AllPrep DNA/RNA Mini kit (Qiagen), total RNA was prepared from a hepatocellular carcinoma cell line (HuH-7) and human primary adult hepatocytes (obtained from the Health Science Research Resources Bank) as well as clones AFB1-1, NGC1-1, and NGC1-2 that were cultured on the MEFs (5 X 10^4 cells/cm^2) with mTeSR1 (StemCell Technologies) medium in gelatin-coated 100-mm dishes before long-term serial passage. The microarray study was carried out using a Whole Human Genome Oligo microarray 4X44K (Agilent). The analysis was performed by Bio Matrix Research according to Agilent technical protocols. Data from these experiments and the GEO database for hESCs (hES_ES01, GSM194392; hES_BG03, GSM194391; hES_H9, GSM194390) and hiPSCs (201B7, GSM241846) established from human adult fibroblasts (fibroblast, GSM242095) were analyzed with GeneSpring GX 11.5 software (Agilent). The 50th percentile of fluorescent intensity distribution was normalized across arrays.