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6 protocols using deferoxamine mesylate

1

Culturing A549 and Small Airway Cells

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A549 cells (American Type Culture Collection, #CCL-185) were cultured in DMEM supplemented with 10% heat-inactivated fetal bovine serum (FBS). The cells were passaged <20 times. Small airway epithelial cells (American Type Culture Collection, #PCS-301-010) were cultured in small airway epithelial growth media (Lonza #CC-3118) and passaged < 7 times. For studies using cell death inhibitors, deferoxamine mesylate (Selleckchem #S5742), necrosulfonamide (Selleckchem #S8251), and Z-VAD-FMK (Selleckchem #S7023) were dissolved in DMSO, and cells were treated at indicated concentrations
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2

Ferroptosis Regulation by Molecular Inhibitors

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Myrislignan (S3261), Z-VAD-FMK (S7023), necrosulfonamide (S8251), ferrostatin-1 (S7243), deferoxamine mesylate (S5742), and RSL3 (S8155) were purchased from Selleck (USA). 3-MA (HY-19312) and erastin (HY-15763) were purchased from MedChemExpress (USA). Dimethyl sulfoxide (DMSO) was obtained from Servicebio (G5051, China). The following antibodies were used: anti-GAPDH (60004-1-Ig, Proteintech), anti-E-cadherin (14472, Cell Signaling Technology), anti-Snail-1 (13099-1-AP, Proteintech), anti-Slug (GTX128796, GeneTex), anti-p65 (sc-8008, Santa Cruz), anti-p-p65 (sc-136548, Santa Cruz), anti-phospho-IκB-α (2859, Cell Signaling Technology), anti-IκB-α (CSB-RA015761A0HU, Cusabio Technology), anti-SLC7A11 (NB300-318, Novus), anti-β-catenin (phospho Y142, ab27798, Abcam), anti-β-catenin (ab32572, Abcam), anti-Cyclin-D1 (60186-1-Ig, Proteintech), anti-Bad (9292, Cell Signaling Technology), anti-Bcl2 (15017, Cell Signaling Technology), anti-Vimentin (GB111308, Servicebio), anti-Twist (25465-1-AP, Proteintech), anti-ZEB1 (21544-1-AP, Proteintech), anti-Nrf2 (M200-3, Medical and Biological Laboratories), anti-TFR1 (ab214039, Abcam), and anti-GPX4 (67763-1-Ig, Proteintech).
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Comprehensive Toolkit for Cancer Research

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RSL3 (S8155), neratinib (S2150), lapatinib (S2111), gefitinib (S1025), ML210 (S0788), afatinib (S1011), dacomitinib (S2727), sapitinib (S2192), Z-VAD-FMK (S7023), necrostatin-1 (S8037), 3-Methyladenine (3-MA, S2767), tucatinib (S8362), liproxstatin-1 (S7699), erastin (S7242), deferoxamine mesylate (DFO, S5742), deferiprone (S4067), ferrostatin-1 (S7243), cobimetinib (S8041), Trastuzumab (A2007) and AZD6738 (S7693) were obtained from Selleck Chemicals. L-glutathione (G6013) and N-Acetyl-l-cysteine (A9165) were purchased from Sigma-Aldrich. T-DM1 (HY-P9921) was obtained from MedChemExpress.
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Cell Culture of GIST Cancer Lines

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The human GIST882 cell line [31 (link)] was kindly provided by Dr. Jonathan A. Fletcher, Harvard Medical School, Boston, MA, USA. Cells were cultured at 37 °C in DMEM (Gibco, CA, USA, Cat# 41965-062), supplemented with 10% (v/v) FBS, 2% (v/v) penicillin–streptomycin (Gibco, CA, USA, Cat# 15140-122). The human GIST48 cell line [32 (link)] was kindly provided by Dr. Ronald DeMatteo, Perelman School of Medicine, Philadelphia, PA, USA. Cells were cultured at 37 °C in RPMI-1640 (Gibco, CA, USA, Cat# 31870-074), supplemented with 10% (v/v) FBS, 1% (v/v) L-glutamine (Gibco, CA, USA, Cat# 35050-038), 1% (v/v) HEPES (Gibco, CA, USA, Cat# 15630-056), 0.1% (v/v) 2-mercaptoethanol (Gibco, CA, USA, Cat# 31350-010), and 0.5% (v/v) penicillin–streptomycin. Verteporfin and cumene hydroperoxide were purchased from Sigma, St. Louis, MO, USA (Cat# C0737, SML0534, and 247502, respectively). Erastin, liproxstatin-1, deferoxamine mesylate, RSL3, CA3, and staurosporine were purchased from Selleckchem, Houston, TX, USA (Cat# S7242, S7699, S5742, S8155, S8661, and S1421, respectively).
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5

Cardiomyocyte Isolation and Inhibitor Pretreatment

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We isolated primary cardiomyocytes from Sprague‒Dawley rat heart tissue using previously described procedures [22 (link)]. H9c2 cardiomyocytes were acquired from the National Collection of Authenticated Cell Cultures (Shanghai, China). All cells were cultured with DMEM containing glucose (Gibco, USA, 4.5 g/L), and 10 % fetal bovine serum (Gibco, USA) and 1 % penicillin and streptomycin solution were added at 37 °C in a 5 % CO2 incubator. Cardiomyocytes were pretreated with A-438079 (an inhibitor of P2X7R) from Selleck (#S7705, 10 μM), deferoxamine mesylate from Selleck (#S5742, 50 or 100 μM), or ferrostatin-1 from Selleck (#S7243, 1 or 5 μM) for 2 h before stimulation with Ang II (1 μM).
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6

Pharmacological Compounds for Cell Death Pathways

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Doxorubicin (#T1020) was purchased from TargetMol (Wellesley Hills, MA, USA). 17-PT-PGE2 (#14810) and SC-51322 (#10010744) were obtained from Cayman Chemical Company (Ann Arbor, MI, USA). U73122 (#U6756), and staurosporine (#539648) was obtained from Sigma Company (Sigma-Aldrich, St. Louis, MO, USA). Erastin (#S7242), RSL3 (#S8155), Fer-1 (#S7243), Nec-1s (#S8641), Bafilomycin A1 (#S1413), Z-VAD-FMK (#S7023), Trolox (#S3665), Deferoxamine mesylate (#S5742) and ML-385 (#S8790) were purchased from Selleck Chemicals (Houston, TX, USA). Wortmannin (#HY-10197), SP600125 (#HY-12041), and Ravoxertinib (#HY-15947) were purchased from MedChemExpress (MCE) Company (Shanghai, China).
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