Journal: European Journal of Medical Research

Article Title: Ferrostatin-1 facilitated neurological functional rehabilitation of spinal cord injury mice by inhibiting ferroptosis

doi: 10.1186/s40001-023-01264-7

Figure Lengend Snippet: Ferrostatin-1 promotes GPX4 to disturbe Nrf2/HO-1 signaling pathway during ferroptosis. A Representative results of western blot of Nrf2/HO-1 signaling pathway change with Ferrostatin-1 and Ferrostatin-1 + Spinosin administration. B The quantification of Fe 2+ concentration in injured spinal cord

Article Snippet: After conventional electrophoretic separation, membrane transfer, and blocking, primary antibodies against Glutathione Peroxidase 4 (GPX4) (ab252833, abcam), HO-1 (ab305290, abcam), Nrf2 (ab92946, abcam) and secondary antibodies were sequentially applied.

Techniques: Western Blot, Concentration Assay

Journal: Journal of Orthopaedic Translation

Article Title: Paeonol inhibits ACSL4 to protect chondrocytes from ferroptosis and ameliorates osteoarthritis progression

doi: 10.1016/j.jot.2024.10.005

Figure Lengend Snippet: PAE mitigates IL-1β/FAC-induced ferroptosis in chondrocytes: (A) Western Blot analysis of ACSL4, SLC7A11, and GPx4 protein levels. (B) Quantification of protein expression by band densitometry, normalized to GAPDH (n = 3 per group). (C) MDA content measured using a lipid peroxidation MDA assay kit (n = 3 per group). (D) Fe 2+ levels in chondrocytes detected using a Fe 2+ content assay kit (n = 4 per group). (E) 4-HNE levels in chondrocytes determined by ELISA (n = 3 per group). (F) GPx4 protein levels detected by ELISA (n = 3 per group). (G) Measurement of intracellular SOD activity (n = 4 per group). (H) Intracellular GSH, GSSG content, and GSH/GSSG (glutathione redox ratio) (n = 4 per group). (I) Glutamate and glutamine levels (n = 3 per group). (J) Intracellular Fe 2+ visualized by FerroOrange using fluorescence microscopy (scale bar = 100 μm). (K) Quantification of FerroOrange fluorescence intensity (n = 3 per group). (L) Prussian blue staining of chondrocytes with red arrows indicating iron deposition (scale bar = 100 μm). Two-tailed P values were calculated with statistical significance defined as P < 0.05, denoted as 'ns' for no statistical difference, and indicated as ∗ P < 0.05, ∗∗ P < 0.01, ∗∗∗ P < 0.001. All experiments and images shown are representative. (For interpretation of the references to colour in this figure legend, the reader is referred to the Web version of this article.)

Article Snippet: The commercial enzyme-linked immunosorbent assay (ELISA) kits used were: Mouse 4-Hydroxynonenal (4-HNE) ELISA Kit (EM1583, FineTest), Mouse Nitric Oxide Synthase 2 (iNOS) ELISA Kit (EM0272, FineTest), Mouse Phospholipid Hydroperoxide Glutathione peroxidase (GPx4) ELISA Kit (EM1964, FineTest), Mouse Interleukin-6 (IL-6) ELISA Kit (RK00008, ABclonal), Mouse Cyclooxygenase-2 (COX-2) ELISA Kit (RK03142, ABclonal), and Mouse Tumor Necrosis Factor-alpha (TNF-α) ELISA Kit (RK00027, ABclonal).

Techniques: Western Blot, Expressing, Multiple Displacement Amplification, Enzyme-linked Immunosorbent Assay, Activity Assay, Fluorescence, Microscopy, Staining, Two Tailed Test

Journal: Sensors (Basel, Switzerland)

Article Title: Simultaneous Down-Regulation of Intracellular hTERT and GPX4 mRNA Using MnO 2 -Nanosheet Probes to Induce Cancer Cell Death

doi: 10.3390/s26030836

Figure Lengend Snippet: Schematic illustration of designation of MnO 2 -NS probes, mechanism of inducing cancer cell death by the synergism of GSH depletion and down-regulation of hTERT and GPX4 simultaneously through gene silencing.

Article Snippet: The GPX4 activity in the cell lysates was determined using a commercial GPX4 Fluorogenic Assay Kit (Amyjet Scientific, Wuhan, China).

Techniques:

Journal: Sensors (Basel, Switzerland)

Article Title: Simultaneous Down-Regulation of Intracellular hTERT and GPX4 mRNA Using MnO 2 -Nanosheet Probes to Induce Cancer Cell Death

doi: 10.3390/s26030836

Figure Lengend Snippet: ( A ) TEM, ( B ) AFM images, ( C , D ) XPS characterization, ( E ) UV−vis absorption spectrum, and ( F ) size distribution of MnO 2 -NSs. ( G ) Zeta potential of MnO 2 -NSs and Anti-hTERT/GPX4-MnO 2 -NS probes.

Article Snippet: The GPX4 activity in the cell lysates was determined using a commercial GPX4 Fluorogenic Assay Kit (Amyjet Scientific, Wuhan, China).

Techniques: Zeta Potential Analyzer

Journal: Sensors (Basel, Switzerland)

Article Title: Simultaneous Down-Regulation of Intracellular hTERT and GPX4 mRNA Using MnO 2 -Nanosheet Probes to Induce Cancer Cell Death

doi: 10.3390/s26030836

Figure Lengend Snippet: Fluorescence quenching of ( A ) Anti-hTERT-DNA (100 nM) and ( B ) Anti-GPX4-DNA (100 nM) strands was assessed upon their adsorption onto MnO 2 -NSs at increasing concentrations. ( C , D ) Fluorescence recovery of the Anti-hTERT/GPX4-MnO 2 -NS probes (40 μg/mL) was subsequently evaluated with the addition of increasing concentrations of GSH. ( E , F ) GSH-triggered release kinetics of the Anti-hTERT/GPX4-MnO 2 -NS probes (40 μg/mL) when mixed with GSH (10 mM). The values are expressed as the mean ± SE ( n = 3).

Article Snippet: The GPX4 activity in the cell lysates was determined using a commercial GPX4 Fluorogenic Assay Kit (Amyjet Scientific, Wuhan, China).

Techniques: Fluorescence, Adsorption

Journal: Sensors (Basel, Switzerland)

Article Title: Simultaneous Down-Regulation of Intracellular hTERT and GPX4 mRNA Using MnO 2 -Nanosheet Probes to Induce Cancer Cell Death

doi: 10.3390/s26030836

Figure Lengend Snippet: CLSM images of A549, HeLa, HepG2, and Caco-2 cells after treated with Anti-hTERT/GPX4-MnO 2 -NS probes (30 μg/mL) for 6 h.

Article Snippet: The GPX4 activity in the cell lysates was determined using a commercial GPX4 Fluorogenic Assay Kit (Amyjet Scientific, Wuhan, China).

Techniques:

Journal: Sensors (Basel, Switzerland)

Article Title: Simultaneous Down-Regulation of Intracellular hTERT and GPX4 mRNA Using MnO 2 -Nanosheet Probes to Induce Cancer Cell Death

doi: 10.3390/s26030836

Figure Lengend Snippet: Determination of ( A ) hTERT and ( B ) GPX4 mRNA levels by RT-PCR in A549 and HeLa cells following treatment with PBS, Control-MnO 2 -NS probes (30 μg/mL), or Anti-hTERT/GPX4-MnO 2 -NS probes (30 μg/mL) for 24 or 48 h. Analysis of ( C ) hTERT activity, ( D , E ) relative GPX4 activity, and ( F ) Se-GPX activity in A549 and HeLa cells following treatment with PBS, Control-MnO 2 -NS probes (30 μg/mL), or Anti-hTERT/GPX4-MnO 2 -NS probes (30 μg/mL) for 24 or 48 h. The values are expressed as the mean ± SE ( n = 3); (*) p < 0.05, (**) p < 0.01, and (***) p < 0.001, significantly different from the control (PBS); (ns) p > 0.05, not significantly different from the control (PBS).

Article Snippet: The GPX4 activity in the cell lysates was determined using a commercial GPX4 Fluorogenic Assay Kit (Amyjet Scientific, Wuhan, China).

Techniques: Reverse Transcription Polymerase Chain Reaction, Control, Activity Assay

Journal: Sensors (Basel, Switzerland)

Article Title: Simultaneous Down-Regulation of Intracellular hTERT and GPX4 mRNA Using MnO 2 -Nanosheet Probes to Induce Cancer Cell Death

doi: 10.3390/s26030836

Figure Lengend Snippet: Analysis of intracellular ( A ) GSH levels, ( B ) relative ROS levels, and ( C ) MDA contents of HeLa cells following treatment with PBS, Control-MnO 2 -NS probes (30 μg/mL), or Anti-hTERT/GPX4-MnO 2 -NS probes (30 μg/mL) for 24, 48, or 72 h. The values are expressed as the mean ± SE ( n = 3); (*) p < 0.05, (**) p < 0.01, and (***) p < 0.001, significantly different from the control; (ns) p > 0.05, not significantly different from the control (PBS).

Article Snippet: The GPX4 activity in the cell lysates was determined using a commercial GPX4 Fluorogenic Assay Kit (Amyjet Scientific, Wuhan, China).

Techniques: Control

Journal: Redox Biology

Article Title: MCL attenuates atherosclerosis by suppressing macrophage ferroptosis via targeting KEAP1/NRF2 interaction

doi: 10.1016/j.redox.2023.102987

Figure Lengend Snippet: MCL enhances NRF2 nuclear translocation in macrophages and ApoE−/− mice. In macrophages, the nuclear lysate was extracted to analysis NRF2 expression. In macrophages and aorta, Nuclear and cytoplasmic lysate were extracted to analysis NRF2 expression. (A, B). Western blot is used to evaluate NRF2 levels. N = 3; (C, D). Confocal fluorescent staining is used to evaluate the nuclear translocation of NRF2. N = 6; (E, F). The protein levels of NRF2 in the aorta nuclear lysates of ApoE −/− mice. N = 5; (G) The enrichment ability of NRF2 on GPX4 and xCT promoter using ChIP-qPCR; ***p＜0.001, vs control or ND group. ###p < 0.001, vs ox-LDL or HD group.

Article Snippet: The binding of NRF2 to GPX4 and xCT promoter was analyzed using Pierce™ Agarose ChIP Kit (Thermo Fisher).

Techniques: Translocation Assay, Expressing, Western Blot, Staining