P62 sqstm1

The chemicals and reagents used in this study were (1) RTA dh404, which was purchased from Cayman Chemicals; (2) PrestoBlue™ Cell Viability Reagent from ThermoFisher/Invitrogen; (3) fetal bovine serum (FBS), the antibiotics penicillin/streptomycin (P/S), and modified Eagle medium (MEM) were purchased from Gibco and Roswell Park Memorial Institute (RPMI) 1640 medium (USA); (4) phosphate-buffered saline (PBS), dimethyl sulfoxide (DMSO), trypsin-EDTA (0.25%), and Trypan Blue Solution were purchased from Sigma (St Louis, MO); (5) the polyvinylidene fluoride membrane (PVDF) (Millipore) and molecular weight markers were purchased from Bio Rad (USA); and (6) propidium iodide (PI) (USA).
Western blot antibodies were also purchased from commercial vendors and used at the indicated dilutions: Cyclin B (1:1000; Proteintech; 55004-1-AP), CDK1 (1:1000; Cell Signaling; E1Z6R), Wee1 (1:1000; Proteintech 14375-1-AP), p21 (1:1000; Cell Signaling E2R7A); Bcl-2 (1:1000), Nrf2 (1:1000; Cell Signaling; D1Z9C), Bcl-2(1;1000; Proteintech), Bax (1:1000; Proteintech), Caspase-3 (1:1000; Affinity), PARP (1:1000; Affinity), LC3B (1:1000; Invitrogen), p62/SQSTM1 (1:1000; Affinity), and β-actin (1:20000; Sigma; A5441).

Total proteins of cells and tissues were isolated by RIPA buffer (Beyotime, P0013) contained with 1% phenylmethylsulfonyl fluoride (Beyotime, ST506), followed by separation on 12% SDS-PAGE gel. The following primary antibodies were used: hSOD1 (Abcam, ab52950), MPO (Abcam, ab188211), GPX4 (Affinity, DF6701), FSP1 (Affinity, DF8636), NQO1 (Affinity, DF6437), Bax (Beyotime, AB026), Bcl-2 (Affinity, BF9103), caspase-3 (Bioss, bs-0081R), LC3A/B (Cell Signaling Technology, 4108S), P62/SQSTM1 (Affinity, AF7875), and GAPDH (Cell Signaling Technology, 2118S). Goat anti-rabbit IgG (H+L) HRP (MultiSciences, 70-GAR0072) and goat anti-mouse IgG (H+L) HRP (MultiSciences, 70-GAM0072) were used as secondary antibodies. The densitometry of bands was quantified by ImageJ, and the bands of GAPDH served as the loading control.