Hepg2

The human LIHC cell lines HepG-2 and Huh-7 were obtained from Procell Life Science&Technology Co., Ltd. (Wuhan, China). The human LIHC cell lines MHCC97-H and HCCLM-3 were obtained from Zhong Qiao Xin Zhou Biotechnology Co., Ltd. (Shanghai, China). The human normal liver cell line HL-7702 (L02) were obtained from Guan&Dao Biological Engineering Co., Ltd. (Shanghai, China). HepG-2, Huh-7, MHCC97-H and HCCLM-3 cells were cultured in DMEM (Cytiva, Utah, USA) with 10% FBS (Gibco, NY, USA) plus 1% penicillin and streptomycin (Sangon, Shanghai, China). L02 cell line was cultured in RIPA 1640 medium (Cytiva, Utah, USA) with 10% FBS plus 1% penicillin and streptomycin. All of the cells were cultured under humidified condition with 5% CO₂ at 37 °C. The authentication of these cell lines was performed via comparisons with the STR database.
The lentivirus vectors LV–NC–shRNA, LV-CCDC115-shRNA and LV-CCDC115-OE were purchased from GenePharma (Shanghai, China). HepG-2 and HCCLM-3 cells were infected with lentivirus (MOI: HepG-2 cell = 80 pfu/cell; HCCLM-3 cell = 100 pfu/cell) with hexadimethrine bromide (5 μg/mL, GenePharma) for 72 h and puromycin (GenePharma) at the concentration of 2 μg/mL (HepG-2) or 2.5 μg/mL (HCCLM-3) was used to select for stably transfected cells.

The human hepatoma cell lines HepG2, Huh7, and HepG2.2.15 were purchased from the Cell Bank of the Chinese Academy of Sciences (Shanghai, China). HepAD38 was provided by Professor Zhi Li, College of Life Sciences, Shaanxi Normal University.Ethics approval for the work was granted by the Ethics Committee of The Second Xiangya Hospital.Cells were maintained in Dulbecco’s modified Eagle’s medium (DMEM, HyClone, Shanghai, China) supplemented with 10% fetal bovine serum (FBS, Gibco, New York, USA) containing 100 U/mL penicillin and streptomycin (Cat. No. ST488, Beyotime, Beijing, China) at 37°C in a humidified atmosphere with 5% CO₂. Cells were plated into 6-well plates at a density of 1×10⁶ cells/well. After 24 hours, a concentration gradient of the HBV-miR-3 agomir or of the negative control (NC) RNA (artificially synthesized by GenePharma, Shanghai, China) was transfected into HepG2 cells and into HepAD38 cells. Transfection was performed using GP-siRNA-Mate Plus (GenePharma, Shanghai, China) according to the manufacturer’s instructions. The growth medium was changed after 8 hours. Transfected cells were harvested at 48 hours, and total cellular RNA and protein were isolated for RT-qPCR and Western blot analyses. All transfections were performed in triplicate. The blank group refers to cells that were not transfected with any RNA.