Erk inhibitor pd98059

The human HCC HepG2 cell line was purchased from the Type Culture Collection of the Chinese Academy of Sciences (Shanghai, China). HepG2 cells were grown as sub-confluent monolayer cultures in Dulbecco's modified Eagle's medium (DMEM; cat. no. SH30022.01; Hyclone; GE Healthcare, Logan, UT, USA) supplemented with 10% fetal bovine serum (cat. no. 11011-8611; Zhejiang Tianhang Biological Technology Co., Ltd., Hangzhou, China). Cells were incubated at 37°C with 5% CO₂. The experiment was performed at the log phase of growth after the cells had been plated for 24 h. HepG2 cells were starved overnight in a humidified 5% CO₂ incubator at 37°C in serum-free DMEM, in the absence or presence of 10 µM ERK inhibitor (PD98059), JNK inhibitor (SP600125) or p38 inhibitor (SB203580) (all EMD Millipore, Billerica, MA, USA) for 5 h in a humidified 5% CO₂ incubator at 37°C; subsequently cells were treated with 9 pM TGF-β1 (R&D Systems, Inc., Minneapolis, MN, USA) for 1 h in a humidified 5% CO₂ incubator at 37°C. The cells in the control groups were added to an equal volume of serum-free medium.