Journal: Cell Reports Medicine

Article Title: Dual ferroptosis induction in N2-TANs and TNBC cells via FTH1 targeting: A therapeutic strategy for triple-negative breast cancer

doi: 10.1016/j.xcrm.2024.101915

Figure Lengend Snippet:

Article Snippet: NCOA4 antibody , Cell Signaling Technology , Cat66849S; RRID: AB_3064842.

Techniques: Recombinant, Lysis, Staining, Saline, Transfection, Isolation, GSSG Assay, SYBR Green Assay, Mass Spectrometry, Software

Journal: Neural Regeneration Research

Article Title: Inhibition of autophagy rescues HT22 hippocampal neurons from erastin-induced ferroptosis

doi: 10.4103/1673-5374.360246

Figure Lengend Snippet: Western blot assay and densitometric analysis of immunoblots of GPX4, x-CT protein, NCOA4, ACSL4, FTH and, mtFt 6, 16 and 24 hours after incubation with 0.5 μM erastin in HT22 neurons. Each experiment was performed in triplicate. * P < 0.05, ** P < 0.01, vs . control (Ctr) group (one-way analysis of variance with Tukey’s post hoc test). ACSL4: Acyl-CoA synthetase long-chain family member 4; FTH: ferritin; GPX4: glutathione peroxidase 4; mtFT: mitochondrial ferritin; NCOA4: nuclear receptor coactivator 4; x-CT: sodium-independent cystine-glutamate antiporter.

Article Snippet: Formalin-fixed cells were incubated overnight at 4°C with primary antibodies: GPX4 (rabbit, mAb; 1:200, Cat# 52455, Cell Signaling, Frankfurt, Germany), cystine-glutamate antiporter (x-CT; rabbit mAb; 1:200, Cat# 12691, Cell Signaling), FTH (rabbit, mAb; 1:200, Cat# 4393 Cell Signaling), mtFt (rabbit, mAb, 1:200, Cat# 31718, Abcam), NCOA4 (rabbit, mAb; 1:200, Cat# 66849, Cell Signaling), ACSL4 (rabbit, polyclonal, 1:200, Cat# PA5-27137, Thermo Fisher Scientific).

Techniques: Western Blot, Incubation

Journal: Neural Regeneration Research

Article Title: Inhibition of autophagy rescues HT22 hippocampal neurons from erastin-induced ferroptosis

doi: 10.4103/1673-5374.360246

Figure Lengend Snippet: The x-CT- (A), GPX4 (B), FTH (C), mtFt (D), NCOA4 (E) and ACSL4 (F) immunoreactivities (IR) in HT 22 neurons collected from controls and at 6, 16, and 24 hours after incubation with erastin. Red and green colors represent the immunoreactivities of the corresponding antibodies. Blue (DAPI) corresponds to nuclei. Scale bar: 50 µm. Each experiment was performed in triplicate. * P < 0.05 and ** P < 0.01, vs . control (Ctr) group (one-way analysis of variance with Tukey’s post hoc test). ACSL4: Acyl-CoA synthetase long-chain family member 4; FTH: ferritin; GPX4: glutathione peroxidase 4; mtFT: mitochondrial ferritin; NCOA4: nuclear receptor coactivator 4; x-CT: sodium-independent cystine-glutamate antiporter.

Article Snippet: Formalin-fixed cells were incubated overnight at 4°C with primary antibodies: GPX4 (rabbit, mAb; 1:200, Cat# 52455, Cell Signaling, Frankfurt, Germany), cystine-glutamate antiporter (x-CT; rabbit mAb; 1:200, Cat# 12691, Cell Signaling), FTH (rabbit, mAb; 1:200, Cat# 4393 Cell Signaling), mtFt (rabbit, mAb, 1:200, Cat# 31718, Abcam), NCOA4 (rabbit, mAb; 1:200, Cat# 66849, Cell Signaling), ACSL4 (rabbit, polyclonal, 1:200, Cat# PA5-27137, Thermo Fisher Scientific).

Techniques: Incubation

Journal: Oxidative Medicine and Cellular Longevity

Article Title: Modulation of the p38 MAPK Pathway by Anisomycin Promotes Ferroptosis of Hepatocellular Carcinoma through Phosphorylation of H3S10

doi: 10.1155/2022/6986445

Figure Lengend Snippet: Anisomycin-induced phosphorylation of H3S10 was associated with transcriptional activation of ferroptosis-related genes. (a) Representative graphs of chromatin immunoprecipitation- (ChIP-) sequencing results. (b) Gene Ontology (GO) analyses of differentially enriched genes. BP: biological processes; CC: cell components; MF: molecular functions. (c) Kyoto Encyclopedia of Genes and Genomes (KEGG) results ranked by degree of enrichment. (d) Protein-protein interactions of ferroptosis-related genes enriched in the term of ferroptosis in (c) (NCOA4, SLC3A2, LPCAT3, PRNP, FTL, FTH1, SLC40A1, GCLC, and GPX4).

Article Snippet: The anti-tubulin antibody was obtained from InTech (Shanghai, China), while anti-c-Myc (#18583), CD133 (#64326), Nanog (#4903), EpCAM (#2929), caspase-3 (#14220), Bcl-2 (#15071), Bax (#5023), N-cadherin (#13116), E-cadherin (#3195), vimentin (#5741), α -smooth muscle actin ( α -SMA, #19245), p38 MAPK (#8690), phospho-p38 MAPK (p-p38 MAPK, #9216), histone H3 (#4499), phospho-histone H3 (Ser10) (p-H3S10, #53348), phospho-histone H3 (Ser28) (p-H3S28, #9713), NRF2 (#12721), SLC7A11 (#12691), FTH1 (#4393), and NCOA4 (#66849) antibodies were purchased from Cell Signaling Technology (Boston, USA).

Techniques: Activation Assay, Chromatin Immunoprecipitation, ChIP-sequencing

Journal: Oxidative Medicine and Cellular Longevity

Article Title: Modulation of the p38 MAPK Pathway by Anisomycin Promotes Ferroptosis of Hepatocellular Carcinoma through Phosphorylation of H3S10

doi: 10.1155/2022/6986445

Figure Lengend Snippet: NCOA4 was upregulated in anisomycin- (AN-) induced ferroptosis. (a) The heatmap based on qRT-PCR indicated changes in ferroptosis-related genes in HCCLM3 after treatment with AN for 12 h. In the inhibitor group, cells were pretreated with SB203580 (SB) for 1 h before being treated with anisomycin (2.5 μ M for Hep3B or 5 μ M for HCCLM3) for 12 h. (b) The qRT-PCR results showed the mRNA expression levels of five ferroptosis-related genes (SLC40A1, FTH1, FTL, NCOA4, and SLC3A2) in different groups of Hep3B and HCCLM3 cells. N = 3; ∗ P < 0.05, ∗∗ P < 0.01, and ∗∗∗ P < 0.001. (c) The protein level of NCOA4 was detected in HCC cells after AN treatment by immunofluorescence. (d) The accumulation of lipid-ROS was detected in NOCA4 knockdown or negative control (NC) HCCLM3 after 12 h AN treatment (left). The accumulation of lipid-ROS was detected in NOCA4 cDNA or pEX-6 carrier transfected HCCLM3 after 12 h AN treatment (right).

Article Snippet: The anti-tubulin antibody was obtained from InTech (Shanghai, China), while anti-c-Myc (#18583), CD133 (#64326), Nanog (#4903), EpCAM (#2929), caspase-3 (#14220), Bcl-2 (#15071), Bax (#5023), N-cadherin (#13116), E-cadherin (#3195), vimentin (#5741), α -smooth muscle actin ( α -SMA, #19245), p38 MAPK (#8690), phospho-p38 MAPK (p-p38 MAPK, #9216), histone H3 (#4499), phospho-histone H3 (Ser10) (p-H3S10, #53348), phospho-histone H3 (Ser28) (p-H3S28, #9713), NRF2 (#12721), SLC7A11 (#12691), FTH1 (#4393), and NCOA4 (#66849) antibodies were purchased from Cell Signaling Technology (Boston, USA).

Techniques: Quantitative RT-PCR, Expressing, Immunofluorescence, Negative Control, Transfection