Et1604 45

All prepared cells were homogenized in protein lysate buffer for 30 min at 4 ℃, and debris was removed by centrifugation at 12,000 g for 15 min at 4 ℃. The protein concentration was ascertained by a BCA protein assay kit (KeyGEN BioTECH, Jiangsu, China). Following the loading buffer addition, 30 µg of each protein sample was electrophoresed, transferred to PVDF membranes (0.2 μm; Millipore, Bedford, MA), and blocked for 2 h in silk milk. Subsequently, the anti-human primary antibody was applied to the membranes for immunoblotting overnight at 4 ℃. Following three times TBST washing, the membranes were incubated for 1 h at room temperature with horseradish peroxidase (HRP)-conjugated secondary antibodies. The HRP signal was then detected by an enhanced chemiluminescence kit (Fdbio Science, Zhejiang, China). Antibodies for GAPDH (10494-1-AP), Tubulin (66031-1-Ig), LILRB3 (18260-1-AP), BAX (50599-2-Ig), BCL2 (12789-1-AP), Caspase 3/p17/p19 (19677-1-AP), P21 (10355-1-AP), Cyclin D1 (60186-1-Ig), and CDK4 (11026-1-AP) were obtained from Proteintech (Wuhan, China). Antibodies for NRF2 (R1312-8), HO-1 (ET1604-45), SOD2 (ET1701-54), and NQO1 (JF440-1) were provided by HUABIO (Hangzhou, China).

Reagents: ferrostatin‐1 (CAS:347174–05‐4) and erastin (CAS:571203–78‐6) were all purchased from MedChemExpress (shanghai; China). SBFI26 (CAS:1541207–06‐0) was purchased from GLPBIO Technology (American). Antibodies: Primary antibodies were used as following: anti‐β‐actin antibody (1:1000, mAbcam 8226, Abcam), anti‐ALOX5 (1:500–1:1000, R1512‐14, HuaBio), anti‐ALOX12 (1:2000, AP8877B Abcepta Biotech), anti‐NFE2L2 (1:1000–1:2000, R1312‐8, HuaBio), anti‐GPX4 (1:500–1:2000, ET1706‐45, HuaBio), anti‐HO‐1 (1:1000, ET1604‐45, HuaBio), anti‐ATF4 (1:500–1:2000, ET1612‐37, HuaBio), secondary antibody was purchased from Beyotime (Shanghai, China).