Mtesr1 medium

hESC lines (H1 hESCs, WiCell Research Institute) were cultured as we previously reported [13 (link), 14 (link)]. Briefly, the cells were seeded on 12-well plate (Corning) coated with matrigel (BD Bioscience) in mTeSR1 medium (Stem Cell Technologies) at 5% CO₂ and 37 °C. For MSC induction, hESCs were split into single cells with Accutase (Gibco), then seeded in mTeSR1 medium with Y27632 (10 nM) (Sigma) addition on a growth factor reduced gel (GFR, Stem Cell Technologies)-coated 6-well plate at a density of 1.3 × 10⁴ cells/mL. After 2 days, the spent medium was replaced by DMEM/F12 basal medium (Hyclone) containing 5% fetal bovine serum (Gibco), 1% penicillin-streptomycin (Gibco), 1% l-glutamine (Gibco), and 10 nM small molecule (Targetmol) from day 0 to day 7; then, cells were transferred into an adherent culture plate in DMEM/F-12 medium supplemented with 10% FBS, 1% penicillin-streptomycin (Gibco), 1% l-glutamine (Gibco), and 5 nM Y27632 (Sigma). Cells were cultured for another 7 days, and media were changed every 2 days in the entire process.

DMEM-F12 medium (cat. no. 11320082), StemPro® NSC SFM—Serum-Free Human Neural Stem Cell Culture Medium (cat. No. A1050901), knockout serum replacement (KOSR, cat. no. 10828–028), MEM nonessential amino acids (MEM-NEAA, cat. No. 11140050). N₂ supplement (cat no.17502048), B-27 supplement without vitamin A (cat. No. 12587010), B-27 supplement with vitamin A (cat. No. 17504044), glutamine/glutaMAX (200 mM, cat. No. 35050061, A2916801), Dispase (cat no. 17105–0412q), and Collagenase IV (cat. No. 17104019) were obtained from Invitrogen (USA). Fetal bovine serum (FBS, cat. No. 10438–018) and bovine serum albumin (BSA, BP9703–100) obtained from Fischer Scientific (Gibco, Invitrogen, USA). Heparin (cat. No. H3149), basic fibroblast growth factor (bFGF, F0291), and epidermal growth factor (EGF, E9644) were procured from Sigma-Aldrich (USA) while mTeSR1 medium (Cat. No. 05850) was procured from Stem cell Technologies (USA).

Human-induced pluripotent stem cell (hiPSC) lines UC and WD were obtained from Guangzhou Institutes of Biomedicine and Health, Chinese Academy of Science. The passage numbers of the cell lines ranged from 41 to 47. Human ESC line H1 was obtained from Wisconsin Cell Research Institute. The cells were cultured on plates coated with Matrigel (BD Biosciences) and maintained in mTeSR™1 medium (STEMCELL Technologies). Before the initiation of hepatocyte-like cell differentiation, hiPSCs were dissociated into clumps using Accutase® solution (Sigma-Aldrich) and were plated onto Matrigel-coated plates and maintained in mTeSR™1 medium. The primary hepatocytes (Lonza) were cultured in HCM.