Huh7 cell line

The human HCC cell line SMMC-7721 was obtained from the Cell Bank of the Institute of Biochemistry and Cell Biology of the Chinese Academy of Sciences (Shanghai, China). The PLC/PRF/5, HepG2, SK-HEP-1, and Hep3B2.1-7 cell lines were purchased from the American Type Culture Collection (Manassas, VA, USA). The Huh7 cell line was obtained from the Riken Cell Bank (Tsukuba, Japan). The MHCC-97L and MHCC-LM3 cell lines were kindly provided by the Liver Cancer Institute, Zhongshan Hospital of Fudan University (Shanghai, China). All cell lines were maintained in Dulbecco’s modified Eagle’s medium (DMEM) (Sigma-Aldrich, St. Louis, MO, USA) supplemented with 10% fetal bovine serum (Hyclone, Logan, UT, USA) at 37 °C in a 5% CO₂ incubator. Standard transient transfections for all cell lines were conducted using Lipofectamine 2000 (Life Technologies, Carlsbad, CA, USA) according to the manufacturer’s instructions.

Cancer stem cells, miPS-BT549cmP cells [12 (link)] and miPS-Huh7cmP cells [14 (link)], were obtained by the conversion of miPSCs (iPS-MEF-Ng-20D-17, Lot No. 012, Riken Cell Bank, Tokyo, Japan), in which the puromycin (puro) resistant gene and green fluorescent protein (GFP) gene were cloned under the control of the Nanog promoter, in the presence of CM from human breast cancer cell line BT549 cells (ATCC HTB-122) and the human liver cancer cell line Huh7 cell line (Riken Cell Bank). All cell lines were with documents confirming their STR profiling.
CSCs were cultured using miPS medium (DMEM media (Wako, Tokyo, Japan) supplemented with 15% fetal bovine serum (FBS), 0.1 mM MEM non-essential amino acids (NEAA) (Gibco, Waltham, MA, USA), 2 mM L-glutamine (Nacalai Tesque, Kyoto, Japan), 50 U/mL penicillin/streptomycin, and 0.1 mM 2-mercaptoethanol (Millipore, MA, USA)) and CM from BT549 cells or Huh7 cell lines, according to the protocol designed by Said et al. [13 (link)]. The ratio of the media was 1:1 of miPS medium to CM. CSCs was maintained on 1% gelatin-coated 60 mm-dishes.

The PLC/PRF/5, Hep3B, and SK-Hep-1 HCC cell lines were obtained from the American Type Culture Collection (Manassas, VA, USA). The Huh7 cell line was purchased from the Riken Cell Bank (Tsukuba, Japan). The SMMC-7721 cell line was purchased from the cell bank of the Institute of Biochemistry and Cell Biology of the Chinese Academy of Sciences (Shanghai, China). The MHCC-97L, MHCC-97H and MHCC-LM3 cell lines were kindly provided by the Liver Cancer Institute, Zhongshan Hospital of Fudan University (Shanghai, China). These cell lines were cultured in Dulbecco's modified Eagle's medium (DMEM; Sigma-Aldrich, St. Louis, MO, USA) with 10% fetal bovine serum (FBS; Hyclone, Logan, UT, USA) at 37°C in 5% CO₂. The HCC-LY5 cell line was established in our laboratory.