Sodium dichloroacetate

PLC/PRF/5, human hepatoma cell line was obtained from the American Type Culture Collection (Rockville, MD). The cells were cultured in low glucose (5.5 mM) or high glucose (25 mM) Dulbecco's modified Eagle's medium (DMEM) containing 10% fetal bovine serum (FBS; Gibco) and antibiotic. Mito Tracker Green and Mito Tracker red CMXRos were obtained from Invitrogen. Sodium lactate and sodium dichloroacetate were obtained from Sigma-Aldrich. The antibodies obtained were: anti-CD133/1 (AC133)-PE and anti-CD133 (W6B3C1) pure (Miltenyi Biotec), anti-HNF4α (Santa Cruz Biotechnology), anti-HNF1α and anti-HNF3α (Millipore), anti-HNF1β (Genway), anti-PDK4 (ABGENT), and anti-β-actin (Sigma). Synthetic siRNAs against LDHA and PDK4, miR-122, U6 primers and mimic were obtained from Qiagen. Cells were transfected at a final concentration of 50 nM siRNA or 20 nM of miR-122 mimic using Oligofectamine reagent (Invitrogen) according to the manufacturer's instructions. The lentiviral particle expressing miR-122 and lentiviral negative control mimic were obtained from GenCopoiea.

Reagents used for experiments: 2-deoxy-d-glucose (D0051, TCI), l-phenylalanine (P0134, TCI), oligomycin A (75351, Sigma-Aldrich), rotenone (R8875, Sigma-Aldrich), FCCP (C2920, Sigma-Aldrich), sodium pyruvate (11360070, Gibco), sodium dichloroacetate (347795, Sigma-Aldrich), palmitic acid (P0500, Sigma-Aldrich), oleic acid (O1008, Sigma-Aldrich), l-glutamine (LS002-01, Welgene), CPI-613 (SML0404, Sigma-Aldrich), (+)-Etomoxir sodium salt hydrate (E1905, Sigma-Aldrich), BPTES (SML0601, Sigma-Aldrich), nicotinamide riboside chloride (SMB00907, Sigma-Aldrich),

The human HCC cell line HCCLM3 and mouse HCC cell line H22 were obtained from the China Center for Type Culture Collection, the mouse HCC cell line Hepa1–6 was obtained from Cell Bank of Type Culture Collection Chinese Academy of Sciences, authenticated by short tandem repeat (STR) analysis, and tested for mycoplasma contamination. HCCLM3 (abbreviated LM3) and Hepa1–6 cells were cultured in high glucose (4.5 g/L) Dulbecco’s modified Eagle’s medium (DMEM) and H22 cells were cultured in Roswell Park Memorial Institute 1640 Medium (RPMI 1640) supplemented with 10% foetal bovine serum, 2 mM l-glutamine, 100 units/mL penicillin, and 0.1 mg/mL streptomycin (all from Thermo Fisher Scientific, Gibco, Grand Island, NY, USA). All cells were maintained in a humidified incubator with an atmosphere containing 5% CO₂ at 37 °C.
The following reagents were used: sodium dichloroacetate (#347795; Sigma-Aldrich, St. Louis, MO, USA), trypan blue (#ST798; Beyotime Biotechnology, Shanghai, China), and 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) (#M2128; Sigma-Aldrich).