RSL3, FER‐1, DFO, liproxstatin‐1 (Lip‐1) and Z‐VAD‐FMK were purchased from Selleck (Chemicals, Houston, TX). Cisplatin and N‐acetyl‐L‐cysteine (NAC) were purchased from Sigma‐Aldrich (Saint Louis, USA). RSL3, FER‐1, DFO, Lip‐1, Z‐VAD‐FMK and NAC were initially dissolved in dimethylsulfoxide (DMSO) and applied at final concentrations (1, 2, 3 and 5 μmol/L with RSL3; 0.5, 1, 2, 5, 10, 20, 30 and 40 μmol/L with FER‐1; 5, 10, 20, 40, 60 and 80 μmol/L with DFO; 0.5, 1, 2, 5, 10 and 20 μmol/L with Lip‐1; 1, 2, 5, 10, 20 and 40 μmol/L with Z‐VAD‐FMK; 5 mmol/L with NAC). Cisplatin was supplied as a 1 mmol/L stock solution in PBS and diluted in culture medium. Final Cisplatin concentrations ranged from 10 to 40 μmol/L.
Liproxstatin 1 lip 1
Manufactured by Selleck Chemicals
Sourced in United States
Liproxstatin-1 (Lip-1) is a small molecule that functions as a potent and selective inhibitor of ferroptosis, a form of regulated cell death. It serves as a valuable tool for research and investigation into the mechanisms and pathways involved in ferroptosis.
Automatically generated - may contain errors
Lab products found in correlation
Ferrostatin 1 fer 1, by Merck Group (2 mentions)
Erastin, by Selleck Chemicals (2 mentions)
Cisplatin, by Merck Group (1 mentions)
N acetyl l cysteine, by Merck Group (1 mentions)
Fer 1, by Selleck Chemicals (1 mentions)
Z vad fmk, by Selleck Chemicals (1 mentions)
Compound c hy 13418a, by MedChemExpress (1 mentions)
Kyse150, by Shanghai Institute of Biochemistry and Cell Biology (1 mentions)
Celecoxib, by Selleck Chemicals (1 mentions)
Bodipy 581 591 c11, by Thermo Fisher Scientific (1 mentions)
Imidazole ketone erastin ike, by Selleck Chemicals (1 mentions)
Sulfasalazine, by Selleck Chemicals (1 mentions)
5 protocols using liproxstatin 1 lip 1
1
Ferroptosis Induction and Inhibition Assay
2
Cytotoxicity Screening of Ferroptosis Inhibitors
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Ferrostatin-1 (Fer-1; SML0583) was obtained from Sigma-Aldrich, Liproxstatin-1 (Lip-1; S7699), and RLS3 (S8155) and Erastin (S7242) were obtained from Selleck. cn. MDA-MB-231 cells (5 × 104/mL) were seeded in 96-well plates with 100 μL per well for 24 h, then 0.099 μM, 0.197 μM, 0.375 μM, 0.625 μM, 1.25 μM, 2.5 μM, 5 μM, 10 μM, and 20 μM RLS3, Sulfasalazine and Erastin were added into plates. After 16 h, 24 h and 48 h, 10 μL of the Cell Counting Kit-8 (CCK-8) reagent was added into each well and incubated at 37 °C, 5% CO2 for 2 h. Then, cell growth was detected, and the inhibition rate was calculated.
3
Cyanidin-3-Glucoside Modulates Ferroptosis
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Cyanidin-3-glucoside (C3G, #C832095, with the purity ≥ 98%) was purchased from Macklin (Shanghai, China). Erastin (Era) (HY-15763) and Compound C (CC) (HY-13418A) were purchased from Med Chem Express (Shanghai, China). Liproxstatin-1 (Lip-1), ferroptosis inhibitor, was bought from Selleck (S7699, Selleck, TX, USA). In vitro experiments, the following reagents were used at the appropriate concentrations: Era (1 μM) and CC (10 μM) for HK-2 cells. C3G (50 μM) or Lip-1(0.5 μM) were added, respectively, and incubated with Era. According to the manufacturer and refer to published literatures (Shan et al. 2021 (link); Krama et al. 2022 (link); Li et al. 2020 (link); Qin et al. 2018b (link)), we finally determined a relatively low dosage and timing of C3G and final concentration and timing of CC. For in vivo experiments, the dosage of the drugs was C3G (10 mg/kg) and CC (10 mg/kg). C3G was intraperitoneal injection daily for a week with or without intravenous injection of CC once 30 min before I/R. C3G was dissolved in dimethyl sulfoxide (DMSO) and then diluted with saline to make sure DMSO concentration was less than 0.1% (v/v), just like previously described (Shan et al. 2021 (link)).
4
Ferroptosis induction in esophageal cancer
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The ESCC cell lines ECA9706 and KYSE150 were purchased from Shanghai Institute of Cell Biology (Shanghai, China) and were cultured in RPMI 1640 supplemented with 10% fetal bovine serum and 1% penicillin/streptomycin. The cells were maintained in an atmosphere containing 5% CO2 at 37°C. Imidazole ketone erastin (IKE, #S8877), liproxstatin-1 (Lip-1, #S7699), KX2-391 (Tirbanibulin, #S2700) and celecoxib (#S1261) were obtained from Selleck Chemicals (United States). BODIPY-C11 (581/591) (#D3861) was obtained from Invitrogen (United States).
5
Cytotoxicity Screening of Small Molecules
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Ferrostatin-1 (Fer-1; SML0583) was obtained from Sigma-Aldrich, Liproxstatin-1 (Lip-1; S7699), RLS3 (S8155), Sulfasalazine (SULF; S1576) and Erastin (S7242) was obtained from Selleck. cn. MDA-MB-231 cells (5×10 4 /mL) were seeded in 96-wells plates with 100 µL per well for 24 hs, then 0.099 µM, 0.197 µM, 0.375 µM, 0.625 µM, 1.25 µM, 2.5 µM, 5 µM, 10 µM and 20 µM RLS3, Sulfasalazine and Erastin were added in to plates. After 16 h, 24 h and 48 h, 10 µL of the Cell Counting Kit-8 (CCK-8) reagent was added into each well and incubated in 37°C, 5% CO 2 for 2 hours. Then cell growth was detected and calculated the inhibition rate.
Cell culture and lentivirus-mediated transduction of shRNA Human breast cancer cell lines MDA-MB-231 and MDA-MB-468 cell lines were cultured in L-15 medium with 10% fetal bovine serum (FBS), in 37°C incubator without CO 2 .
The procedure of lentivirus infection is as follows: the plate containing cells was added with appropriate amount of lentivirus in concentration gradient, followed by adding 1/1000 polybrene to enhance infection. The sequence of the HCP5-132aa shRNA was showed in Table S1. Lentivirus vector LV5 containing full-length HCP5-132aa or empty vector were purchased from GenePharma Co., Ltd. (Shanghai, China).
Cell culture and lentivirus-mediated transduction of shRNA Human breast cancer cell lines MDA-MB-231 and MDA-MB-468 cell lines were cultured in L-15 medium with 10% fetal bovine serum (FBS), in 37°C incubator without CO 2 .
The procedure of lentivirus infection is as follows: the plate containing cells was added with appropriate amount of lentivirus in concentration gradient, followed by adding 1/1000 polybrene to enhance infection. The sequence of the HCP5-132aa shRNA was showed in Table S1. Lentivirus vector LV5 containing full-length HCP5-132aa or empty vector were purchased from GenePharma Co., Ltd. (Shanghai, China).
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