Vemurafenib (cat No. S1267), sorafenib (cat No. S1040), ferroptosis inhibitor Ferrostatin-1 (Fer-1) (cat no. S7243), caspase inhibitor Z-VAD-FMK (cat No. S7023) and MLKL inhibitor Necrosulfonamide (cat No. S8251) were purchased from Selleck (Shanghai, China).
Ferrostatin 1 fer 1
Manufactured by Selleck Chemicals
Sourced in United States, China
Ferrostatin-1 (Fer-1) is a chemical compound used in laboratory research. It functions as a potent and selective inhibitor of ferroptosis, a form of regulated cell death. Fer-1 has been widely utilized in scientific studies to investigate the mechanisms and significance of ferroptosis in various biological systems.
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Lab products found in correlation
Erastin, by Selleck Chemicals (4 mentions)
Ab125066, by Abcam (3 mentions)
Fetal bovine serum (fbs), by Thermo Fisher Scientific (3 mentions)
Sorafenib, by Selleck Chemicals (2 mentions)
Z vad fmk, by Selleck Chemicals (2 mentions)
Dulbecco modified eagle medium (dmem), by Thermo Fisher Scientific (2 mentions)
β actin ab8226, by Abcam (2 mentions)
Ferric ammonium citrate, by Merck Group (2 mentions)
A1933, by ABclonal (2 mentions)
Ab175186, by Abcam (2 mentions)
4 6 diamidino 2 phenylindole (dapi), by Thermo Fisher Scientific (2 mentions)
Necrosulfonamide, by Selleck Chemicals (1 mentions)
Vemurafenib, by Selleck Chemicals (1 mentions)
Ab125068, by Abcam (1 mentions)
Ab252833, by Abcam (1 mentions)
Ab51052, by Abcam (1 mentions)
Nvp auy922, by Selleck Chemicals (1 mentions)
Sa00001 1, by Proteintech (1 mentions)
Sa00001 2, by Proteintech (1 mentions)
Ab46545, by Abcam (1 mentions)
23 protocols using ferrostatin 1 fer 1
1
Ferroptosis and Necroptosis Inhibition
2
Antibody-based Analysis of Cellular Stress Pathways
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Glyceraldehyde 3-phosphate dehydrogenase (60004-1-Ig) and horseradish peroxidase-conjugated secondary antibodies (SA00001-1 and SA00001-2) were purchased from Proteintech (Wuhan, China). Antibodies against Lamp-2a (ab125068), HSP90 (ab32568), GPX4 (ab252833), HSC70(ab51052), and 4-HNE (ab46545) were obtained from Abcam (Cambridge, UK). NVP-AUY922 and ferrostatin-1 (Fer-1) were purchased from Selleck Chemicals (Houston, TX, USA).
3
Metformin and Ferroptosis in Lung Cancer
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A549 and H1299 cells (Cell Bank of the Typical Culture Preservation Committee, Chinese Academy of Sciences) were cultured in RPMI-1640 medium (Gibco, China ) supplemented with 10% fetal bovine serum (FBS) (Biological Industries, Israel) and 100 µg/mL PenStrep. The cells cultured at 37 °C in 5% CO2 humidified chamber were passaged when they grew to 80% confluence. Cells were inoculated into six-well plates with 1 × 106 cells per well and cultured under normal conditions for 48 h. Subsequently, the cells were treated with 0.5 mM, 1.0 mM or 2.0 mM metformin [24 (link)] (Met, S1950, Selleck, China) for 48 h with or without the supplement of 10 µM Ferrostatin-1 (Fer-1, S7243, Selleck, China).
4
Targeting FTO and Ferroptosis in NPC
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FB23-2 (Selleck, China) is an inhibitor that directly binds to FTO and selectively inhibits the m6A demethylase activity of FTO in mammalian cells [16] (link). Ferrostatin-1 (fer-1, Selleck, China) is an effective and selective ferroptosis inhibitor, while erastin (Selleck, China) is a ferroptosis activator that acts on mitochondrial VDAC in cancer cells [17] (link). The three drugs were dissolved in dimethyl sulfoxide (DMSO) and stored at -80 °C. The NPC cells were treated with 10 μmol of FB23-2, 1 μmol of Fer-1, or 10 μmol of erastin in a medium containing 10% of FBS for 24–72 h, and the corresponding concentration of DMSO was used as a control.
5
CX3CL1 Modulation in 786-O Cells
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The human ccRCC cell line 786-O was obtained from the Chinese Academy of Sciences. 786-O cells were incubated with Ferrostatin-1 (Fer-1, S7243, Selleckchem, Houston, TX, USA), Necrostatin-1 (MCE, HY-15760), and Z-VAD-FMK (MCE, HY-16658B). 786-O cells were infected with small interfering RNA (Si-RNA) (Ubi-MCS-CBh-gcGFP-IRES-Puro-CX3CL1) against CX3CL1 and infected with a lentiviral vector (Ubi-MCS-3FLAG-SV40-mcherry-IRES-Puro-CX3CL1) (Shanghai Gene Chem Co., Ltd.) to induce CX3CL1 overexpression.
6
Establishing High-Glucose Cell Model
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SC line RSC96 was purchased from the cell bank of the China Academy of Sciences (Shanghai, China). The cell line was cultured in DMEM (Gibco, Grand Island, USA) supplemented with 10% fetal bovine serum (Gibco) and 1% pen/strep (Gibco), and incubated at 5% CO2 and 37°C.
To establish an HG cell model RSC96 was treated with DMEM containing 100 mM glucose for 48 h as previously described [15 (link)]. Cells were cultured to logarithmic phase and grouped as follows: normal control group (DMEM containing 25 mM glucose), high-glucose group (DMEM containing 100 mM glucose), Erastin (Selleckchem, Houston, USA) group (DMEM containing 25 mM glucose and 2 μM Erastin), and the ferroptosis inhibitor ferrostatin-1 (Fer-1) (Selleckchem) group (DMEM containing 100 mM glucose and 10 μM Fer-1). Erastin, as an inducer of cell ferroptosis, was used as a positive control.
To establish an HG cell model RSC96 was treated with DMEM containing 100 mM glucose for 48 h as previously described [15 (link)]. Cells were cultured to logarithmic phase and grouped as follows: normal control group (DMEM containing 25 mM glucose), high-glucose group (DMEM containing 100 mM glucose), Erastin (Selleckchem, Houston, USA) group (DMEM containing 25 mM glucose and 2 μM Erastin), and the ferroptosis inhibitor ferrostatin-1 (Fer-1) (Selleckchem) group (DMEM containing 100 mM glucose and 10 μM Fer-1). Erastin, as an inducer of cell ferroptosis, was used as a positive control.
7
Keap1-Nrf2 Signaling Modulation
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Plasmids encoding FLAG-tagged Keap1, HA-tagged Ub, and His-tagged Nrf2 were purchased from GeneChem (Shanghai, China). Plasmids were transfected into cells using Lipofectamine 2000 reagent (Life Technologies). The small interfering RNAs (siRNAs) (nontargeting control siRNA and siRNA targeting Keap1) used in this study were synthesized by RiboBio (Guangzhou, China). The siRNAs were transfected into cells using riboFECT CP (RiboBio) according to the manufacturer’s instructions. Ferrostatin-1 (Fer-1) (catalog number S7243), RSL3 (catalog number S8155), MG132 (catalog number S2619), chloroquine (NSC-187208) (catalog number S6999), and indomethacin (IND) (NSC-77541) (catalog numberS1723) were purchased from Selleck Chemicals. Reduced l -glutathione (GSH) (catalog number HY-D0187) and staurosporine (STS) (catalog number HY-15141) were purchased from MedChemExpress (MCE). Deferoxamine (DFO), N-acetylcysteine (NAC), and TNF-α were purchased from Sigma-Aldrich.
8
Comprehensive Assays for Cell Viability and Oxidative Stress
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Assay kits for the detection of cell survival (CCK8 kit), GSH, ATP and MDA were purchased from Beyotime (Shanghai, China). Sorafenib, Ferrostatin-1 (Fer-1), Z-VAD-FMK, Necrosuifonamide (Necro), SB203580, SCH772984, SP600125 and ML385 were obtained from Selleck Chemicals (Houston, TX). DCF-DA MitoTracker, C11-BODIPY (581/591) and DAPI were obtained from Thermo Fisher Scientific (Waltham, MA). Rhodamine B-[(1,10- phenanthrolin-5-yl amino-carbonyl] benzyl ester (RPA) was obtained from Enzo Life Sciences (NY, USA). Deferoxamine (DFO), N-acetylcysteine (NAC), Cycloheximide (CHX), MG132, Disulfiram (DSF), CuCl2 and Trigonelline hydrochloride were purchased from Sigma-Aldrich (St. Louis, MO, USA). The antibodies to NRF2 (ab62352), NQO1 (ab80588), P53 (ab179477), p21 (ab109520), p-p62 (ab211324), GAPDH (ab181602), γ-H2AX (ab81299), p62 (ab109012), MDA (ab6463) and β-actin (ab8226) were obtained from Abcam (Cambridge, MA). The antibody to 4-HNE (MAB3249) was obtained from Novus Biologicals (Littleton, CO, USA). The antibodies to p38 (9212S), p-p38 (9215S), ERK (4695S), p-ERK (4370S), JNK (9252s), p-JNK (9251s), HistoneH3 (3638), β-catenin (19807s), p-GSK3β (5558p), N-cadherin (4061p), Snail (3879p) and Vimentin (5741p) were purchased from Cell Signaling Technologies (Boston, MA, USA).
9
Ferroptosis Pathway Modulation
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Dulbecco’s modified Eagle’s medium (DMEM) and fetal bovine serum (FBS) were purchased from GIBCO (Grand Island, NY, USA). Ferric ammonium citrate (FAC) and iron chloride (FeCl3) were obtained from Sigma-Aldrich (St. Louis, MO, USA). Baicalein, erastin, ferrostatin-1 (Fer-1), and liproxstatin-1 (Lipo-1) were purchased form Selleck Chemicals (Houston, TX, USA).
10
Cell Culture Conditions and Reagents
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HepG2, MHCC97H, Hep3B, and HEK293T cell lines were stocked in our laboratory. Cells are maintained in Dulbecco's Modified Eagle Medium (DMEM, Gibco, NY, USA) with 10% fetal bovine serum (FBS, Gibco, CA, USA) and 1% penicillin-streptomycin (Gibco, CA, USA) at 37 °C in a 5% CO2 humidified incubator (Thermo, CO, USA). Dimethyl sulfoxide (DMSO) was purchased from Sigma (MO, USA). Sorafenib (Sora) and Ferrostatin-1 (Fer-1) were purchased from Selleck (TX, USA).
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