Journal: Cell Death Discovery

Article Title: ACSL1-induced ferroptosis and platinum resistance in ovarian cancer by increasing FSP1 N-myristylation and stability

doi: 10.1038/s41420-023-01385-2

Figure Lengend Snippet: Expression of antioxidant-related proteins SCL7A11, FSP1 after ( A ) transient overexpression of ACSL1 in NM cells and OVCA429 cells and ( B ) knockdown of ACSL1 in HM cells. C Overexpression of ACSL1-WT and ACSL1-MT in NM and OVCA429 cells and protein levels of SLC7A11 and GPX4. D After ACSL1 was knocked out in HM cells and OVCA429 cells respectively, ACSL1, SLC7A11, and GPX4 were detected by western blotting. E The protein level of FSP1 following CHX inhibitor treatment. F After overexpressing ACSL1 and its mutants in OVCA429 ovarian cancer cells, respectively, FSP1 expression was detected by western blotting of cells in the presence or absence of the autophagy blocker 3-MA and the proteasome inhibitor MG132. G After the knockdown of ACSL1 ovarian cancer OVCA429, FSP1 expression in each group was detected by western blotting when the cells were exposed to 3-MA and MG132 or untreated cells. H FSP1 ubiquitination in ACSL1 wild type or ACSL1 mutant overexpression in HEK293T cells was detected by immunoprecipitation. I , J Gene expression and protein levels of the SREBP1 and ACSL1 in HM cells treated with the SREBP1 small molecule inhibitor fatostatin. * P < 0.05 and *** P < 0.001 indicate statistical significance.

Article Snippet: Cycloheximide (CHX, cat:HY-12320), 3-methylamine (3-MA,cat:HY-19312), deferoxamine mesylate (DFO, cat:HY-B0988) were purchased from MCE, China; Necrostatin-1(cat:A4213), ferrostatin-1(cat:A4371), erastin(cat:B1524), MG-132(cat:A2585), ZVAD-FMK(cat:A1902) were purchased from APEXBIO, USA; anti-NRF2 mAb, anti-GPX4 mAb (cat:59735) were purchased from CST, USA; anti-AIFM2 polyclonal antibody(cat:abs138156) was from Absin, China; anti-SLC7A11 polyclonal antibody(cat:26864-1-AP), anti-PTGS2 polyclonal antibody(cat: 66351-1-Ig), anti-alpha tubulin antibody(cat:11224-1-AP), and anti-β-actin antibody(cat: 81115-1-RR) were purchased from PTG, China; anti-4-HNE antibody(cat:MAB3249) was purchased from R&D Systems, USA; Protein A/G Mix magnetic beads kit (cat: 88802) was from Thermo fisher, USA; Myristic acid alkyne(cat: 82909-47-5), Biotinamine (Azide-PEG3-biotin conjugate, cat:762024) were purchased from Santa Cruz, USA; The Click-it Chemistry Kit(cat: C10276), C11 BODIPY581/591(cat:D3861), Streptavidin (DynabeadsTM, cat:65001), and anti-TfR1 polyclonal antibody (cat:14-0719-82), anti-SREBP1 (cat:MA5-11685) were purchased from Thermo Fisher, USA.

Techniques: Expressing, Over Expression, Western Blot, Mutagenesis, Immunoprecipitation

Journal: Molecular Medicine Reports

Article Title: Activating autophagy and ferroptosis of 3‑Chloropropane‑1,2‑diol induces injury of human umbilical vein endothelial cells via AMPK/mTOR/ULK1

doi: 10.3892/mmr.2023.12963

Figure Lengend Snippet: Fer-1 reverses 3-MCPD-induced ferroptosis in HUVECs. (A) Assessment of mitochondrial membrane potential in HUVECs treated with 3-MCPD (scale bar, 10 µm). (B) FerroOrange staining demonstrated that 3-MCPD markedly elevated intracellular Fe 2+ levels in HUVECs (scale bar, 10 µm). (C) C11-BODIPY581/591 staining demonstrated that 3-MCPD markedly enhanced accumulation of lipid reactive oxygen species in HUVECs (scale bar, 10 µm). (D) 3-MCPD significantly increased the accumulation of SOD and MDA in HUVECs; Fer-1 partially neutralized these effects. (E) 3-MCPD significantly decreased expression levels of ferroptosis-related proteins, including GPX4, SLC7A11 and FTH1, in HUVECs. *P<0.05, **P<0.01 and ***P<0.001 vs. Con; # P<0.05 and ## P<0.01 vs. 3-MCPD. 3-MCPD, 3-Chloropropane-1,2-diol; HUVEC, human umbilical vein endothelial cell; Fer-1, ferrostatin-1; GPX4, glutathione peroxidase; 4SLC7A11, cystine/glutamate antiporter solute carrier family 7 member 11; SOD, superoxide dismutase; MDA, malondialdehyde; Con, control.

Article Snippet: Membranes were incubated with primary antibodies against phosphorylated (p)-AMPK (1:1,000; cat. no. 2535; Cell Signaling Technology, Inc.), AMPK (1:1,000; cat. no. 5831; Cell Signaling Technology, Inc.), p-unc-51 like autophagy activating kinase (1:1,000; ULK1; cat. no. 37762; Cell Signaling Technology, Inc.), ULK1 (1:1,000; cat. no. 8054; Cell Signaling Technology, Inc.), p-mTOR (1:1,000; cat. no. 5536; Cell Signaling Technology, Inc.), mTOR (1:1,000; cat. no. 2983; Cell Signaling Technology, Inc.), p53 (1:1,000; cat. no. 2527; Cell Signaling Technology, Inc.), p16 (1:1,000; cat. no. 18769; Cell Signaling Technology, Inc.), p21 (1:1,000; cat. no. 2947; Cell Signaling Technology, Inc.), light chain (LC)3II/I (1:1,000; cat. no. 3868; Cell Signaling Technology, Inc.), p62 (1:1,000; cat. no. 88588; Cell Signaling Technology, Inc.), GPX4 (1:1,000; cat. no. 59735; Cell Signaling Technology, Inc.), SLC7A11 (1:1,000; cat. no. 12691; Cell Signaling Technology, Inc.), FTH1 (1:1,000; cat. no. 4393; Cell Signaling Technology, Inc.) and GAPDH (1:4,000; cat. no. 5174; Cell Signaling Technology, Inc.) at 4°C overnight.

Techniques: Staining, Expressing

Journal: Frontiers in Pharmacology

Article Title: Role of ferroptosis in hypoxic preconditioning to reduce propofol neurotoxicity

doi: 10.3389/fphar.2023.1121280

Figure Lengend Snippet: Protein expression levels of TFR1, FPN1, DMT1, SLC7A11 and GPX4 in primary hippocampal neurons exposed to propofol with or without hypoxic preconditioning. (A) TFR1, FPN1, DMT1, SLC7A11 and GPX4 protein expression was detected by Western blot analysis. (B–F) Quantitative analysis of the ratio of TFR1, FPN1, DMT1, SLC7A11 and GPX4 to β-actin (one-way ANOVA, * p < 0.05).

Article Snippet: Microtubule-associated protein 2 (MAP2) (Abcam, Cambridge, England, UK, Cat#ab183830), Transferrin receptor protein 1 (TFR1) (Beijing Bioss Biotechnology Co., Ltd., Beijing, China, Cat#bsm-54633R), Divalent metal transporter 1 (DMT1) (Proteintech, Chicago, IL, United States, Cat#20507-1-AP), Glutathione peroxidase 4 (GPX4) (Cell Signaling Technology, United States, Cat#59735), Solute carrier family seven member 11 (SLC7A11) (Abcam, United States, Cat#ab175186), Ferroportin 1 (FPN1) (Proteintech, Chicago, IL, United States, Cat#26601-1-AP), β-actin (Abcam, United States, Cat#ab8227), LI-COR fluorescent secondary antibody (LI-COR, Lincoln, NE, United States).

Techniques: Expressing, Western Blot

Journal: Frontiers in Pharmacology

Article Title: Role of ferroptosis in hypoxic preconditioning to reduce propofol neurotoxicity

doi: 10.3389/fphar.2023.1121280

Figure Lengend Snippet: Protein expression levels of TFR1, FPN1, DMT1, SLC7A11 and GPX4 in primary hippocampal neurons exposed to propofol with or without hypoxic preconditioning, inhibitor or agonist. (A) TFR1, FPN1, DMT1, SLC7A11 and GPX4 protein expression was detected by Western blot analysis. (B–F) Quantitative analysis of the ratio of TFR1, FPN1, DMT1, SLC7A11 and GPX4 to β-actin (one-way ANOVA, * p < 0.05).

Article Snippet: Microtubule-associated protein 2 (MAP2) (Abcam, Cambridge, England, UK, Cat#ab183830), Transferrin receptor protein 1 (TFR1) (Beijing Bioss Biotechnology Co., Ltd., Beijing, China, Cat#bsm-54633R), Divalent metal transporter 1 (DMT1) (Proteintech, Chicago, IL, United States, Cat#20507-1-AP), Glutathione peroxidase 4 (GPX4) (Cell Signaling Technology, United States, Cat#59735), Solute carrier family seven member 11 (SLC7A11) (Abcam, United States, Cat#ab175186), Ferroportin 1 (FPN1) (Proteintech, Chicago, IL, United States, Cat#26601-1-AP), β-actin (Abcam, United States, Cat#ab8227), LI-COR fluorescent secondary antibody (LI-COR, Lincoln, NE, United States).

Techniques: Expressing, Western Blot