Mir 23a inhibitor

The human embryonic hepatocytes cell line WRL68, hepatocellular carcinoma cell line HepG2 (HBV⁻) and human embryonic kidney cell line HEK293 were obtained from American Type Culture Collection (ATCC). HepG2.2.15 cells are stablely HBV‐transfected cells derived from a hepatoblastoma HepG2 cell line (Chongqing Medical University). All the cells were maintained in Dulbeco's Modified Eagle Medium (DMEM, Gibco) supplemented with 10% of heat‐inactivated fetal bovine serum (Gibco) and 1% of Penicillin‐Streptomyicin solution (Gibco) in a humidified atmosphere of 5% CO₂ at 37°C. In some experiments, HepG2 or HepG2.2.15 cells were treated with 5 μmol/L parthenolide (a p65 inhibitor, TOCRIS) or miR‐23a mimics (GenePharma) for 24 hours. For rescued study, cells were pretreated with miR‐23a inhibitor (GenePharma) following by 5 μmol/L parthenolide for 24 hours.

Small interfering RNA (siRNA) for GAS5, MITF, TFE3, TFEB, Raptor, DGKE, and its negative controls, as well as miR-23a mimics, miR-23b mimics, mimic NC, miR-23a inhibitor, miR-23b inhibitor, and inhibitor NC, were supplied by GenePharma (Shanghai, China). Overexpression plasmids for GAS5, MITF, TFE3, TFEB, and DGKE were supplied by GeneCreate Biological Engineering Co. (Wuhan, China), and the plasmid pcDNA-3.1 was used as a negative control. All transfections were carried out using Lipofectamine 3000 transfection reagent according to the manufacturer’s protocol (Thermo Fisher, Waltham, MA, USA, L3000075).