Investigating Cell Signaling Pathways

Phosphate-buffered saline (PBS), fetal bovine serum (FBS), Dulbecco’s modified Eagle’s medium (DMEM), and trypsin were all bought from Gibco (Grand Island, NY, USA). HepG2 and LO2 cell lines were purchased from the American Type Culture Collection (Manassas, VA, USA). Cells were grown in RPMI-1640 medium including 1% penicillin-streptomycin and 10% FBS in a 37 °C incubator supplied with 5% CO₂. cytochrome C, Bax, Bcl-2, p53, p21, cleaved PARP, PARP, cleaved caspase-3, caspase-3, cleaved caspase-9, caspase-9, CDK2, cyclin A, cyclin B, cyclin E, p-JNK, JNK, p-Erk, Erk, p-p38, p38, p-Akt, Akt, p-mTOR, mTOR, p-FAK, FAK, p-PI3K, PI3K, LC3A/B, p62, GAPDH, β-actin, and all secondary antibodies were purchased from Cell Signaling Technology (Beverly, MA, USA). DAPI, cyclosporin A (CsA, an inhibitor of permeability transition), N-acetyl-L-cysteine (NAC), ROS assay kit, LDH-release assay kit, cell cycle and apoptosis analysis kit, and mitochondrial membrane potential assay kit for JC-1 were bought from Beyotime Institute of Biotechnology (Shanghai, China). Smp24 and FITC-labeled Smp24 were obtained as previously described by us [9 (link)].