D galn g1639

Normal human hepatic cell line of L-02, purchased from the Cell Bank of Chinese Academy of Sciences (Shanghai, China), was cultured in Roswell Park Memorial Institute (RPMI) 1,640 medium (Gibco, Grand Island, NY, United States) containing 10% fetal bovine serum (Gibco), penicillin of 100 U/ml, and streptomycin of 100 μg/ml in a humidified incubator, with CO₂ at 5% and 37°C. The dose of the d-galactosamine (GalN)/lipopolysaccharide (LPS) model (D-GalN/LPS: 10 mM/10 ng/ml) was determined based on previous research Liu et al. (2018) (link) and our pre-experimental results. LPS (055: B5) and D-GalN (G1639) were obtained from Sigma-Aldrich Chemical Company (St. Louis MO, United States). In subsequent experiments, L-02 hepatocytes were divided into four groups: (a) Control group: Cells were cultured in RPMI 1640 medium alone. (b) D-GalN/LPS group: Cells were stimulated by D-GalN (10 mM)/LPS (10 ng/ml). (c) DEX + D-GalN/LPS groups: Cells were incubated with DEX (1, 10 μM) and D-GalN (10 mM)/LPS (10 ng/ml). (d) DEX + EX527+D-GalN/LPS groups: Cells were incubated with DEX (1, 10 μM), EX527 (10 μM), and D-GalN (10 mM)/LPS (10 ng/ml). According to the results of Cell Counting Kit-8 (CCK-8) assays (MedChemExpress, Monmouth Junction, NJ, United States), the concentration of DEX treatment in the subsequent experiments was determined as 1 μM.

Chrysophanol-8-O-glucoside (PHL84206), N-Acetyl-L-cysteine (A9165), LPS (L4391) and D-GalN (G1639) were purchased from Sigma. Primary antibodies against ATG5 (10181-2-AP), Beclin1 (11306-1-AP), β-actin (66,009-1-lg), LC3B (18725-1-AP), and ERK (16443-1-AP) were purchased from Proteintech. Antibody against p-ERK (sc-101761) was purchased from Santa Cruz. Antibody against p62 was purchased from OriGene (TA502127). NF-κB Pathway Sampler Kit (#9936) was purchased from Cell signaling technology.