Journal: Animals : an Open Access Journal from MDPI

Article Title: Effect of Zearalenone-Induced Ferroptosis on Mice Spermatogenesis

doi: 10.3390/ani12213026

Figure Lengend Snippet: List of primers employed for q-RT-PCR.

Article Snippet: The proteins were then transferred onto PVDF membranes (GE Bioscience, Newark, NJ, USA) and then blocked with 5% BSA (dissolved in TBST) for 1 h and the membrane was incubated with anti-SYCP3 (ab97672; Abcam, Cambridge, UK), anti-DDX4 (bs-22896R; Bioss Biotech, Beijing, China), anti-SOX9 (ab185966; Abcam, Cambridge, UK), anti-SLC7A11 (bs-6883R; Bioss Biotech, Beijing, China), anti-GPX4 (bs-3884R; Bioss Biotech, Beijing, China), anti-4-HNE (ab48506; Abcam), and anti-β-actin (bs-0061R; Bioss Biotech, Beijing, China) overnight at 4 °C.

Techniques: Sequencing

Journal: Animals : an Open Access Journal from MDPI

Article Title: Effect of Zearalenone-Induced Ferroptosis on Mice Spermatogenesis

doi: 10.3390/ani12213026

Figure Lengend Snippet: Expressions of genes, proteins involved in ferroptosis in testis of mice in each group. ( A ) Nrf2 mRNA expression. ( B ) SLC7A11 mRNA expression. ( C ) GPX4 mRNA expression. ( D ) Western blot detection of SLC7A11, GPX4, and 4-HNE protein. ( E ) Image J analysis showing the grey value of SLC7A11. ( F ) Image J analysis showing the grey value of GPX4. ( G ) Image J analysis showing the grey value of 4-HNE. The data are expressed as means ± SEM, with n ≥ 3; Different lowercase letters (a, b, c, d) indicate significant differences ( p < 0.05).

Article Snippet: The proteins were then transferred onto PVDF membranes (GE Bioscience, Newark, NJ, USA) and then blocked with 5% BSA (dissolved in TBST) for 1 h and the membrane was incubated with anti-SYCP3 (ab97672; Abcam, Cambridge, UK), anti-DDX4 (bs-22896R; Bioss Biotech, Beijing, China), anti-SOX9 (ab185966; Abcam, Cambridge, UK), anti-SLC7A11 (bs-6883R; Bioss Biotech, Beijing, China), anti-GPX4 (bs-3884R; Bioss Biotech, Beijing, China), anti-4-HNE (ab48506; Abcam), and anti-β-actin (bs-0061R; Bioss Biotech, Beijing, China) overnight at 4 °C.

Techniques: Expressing, Western Blot

Journal: Experimental and therapeutic medicine

Article Title: Novel function of fluvastatin in attenuating oxidized low-density lipoprotein-induced endothelial cell ferroptosis in a glutathione peroxidase4- and cystine-glutamate antiporter-dependent manner.

doi: 10.3892/etm.2021.10710

Figure Lengend Snippet: Figure 1. ox‑LDL promotes endothelial cell dysfunction via induction of ferroptosis. (A) Immunoblots of TNF‑α, IL‑α, 4‑HNE, GPx4 and xCT in the normal (control), ox‑LDL and ox‑LDL + DFOM groups. (B) Representative Transwell migration assay images of HUVECs treated with ox‑LDL and ox‑LDL + DFOM (magnification, x10). (C) Representative tube formation assay images of HUVECs treated with PBS, 100 µg/ml ox‑LDL and 125 µM DFOM (magnification, x10). (D) Analysis of HUVEC viability using the MTT assay after ox‑LDL treatment. Columns, means of three experiments; bars, SD. Histograms of (E) relative TNF‑α/β‑actin levels, (F) relative IL‑α/β‑actin levels, (G) relative 4‑HNE/β‑actin levels, (H) relative GPx4/β‑actin levels and (I) relative xCT/β‑actin levels. *P<0.05 vs. control group. ox‑LDL, oxidized low‑density lipoprotein; GPx4, glutathione peroxidase 4; xCT, cystine‑glutamate antiporter; 4‑HNE, 4‑hydroxynonenal; DFOM, deferoxamine mesylate; HUVEC, human umbilical vein cell.

Article Snippet: The following primary antibodies were used: 4‐hydroxynonenal (4‐HNE; 1:2,000; ab46544; Abcam), TNF‐α (1:500; WL01581; Wanleibio; Co., Ltd.), IL‐1α (1:2,000; ab134908; Abcam), GPx4 (1:1,000; 14432‐1‐AP; ProteinTech Group, Inc.), xCT (1:1,000; 26864‐1‐AP; ProteinTech Group, Inc.); Caspase‐3/cleaved‐caspase3 (1:500; WL02117; Wanleibio; Co., Ltd.), Bax (1:1,000; WL01637; Wanleibio; Co., Ltd.), Bcl‐2 (1:1,000; WL01556; Wanleibio; Co., Ltd.), β‐actin (1:2,000; WL01845 Wanleibio; Co., Ltd.).

Techniques: Western Blot, Control, Transwell Migration Assay, Tube Formation Assay, MTT Assay

Journal: Experimental and therapeutic medicine

Article Title: Novel function of fluvastatin in attenuating oxidized low-density lipoprotein-induced endothelial cell ferroptosis in a glutathione peroxidase4- and cystine-glutamate antiporter-dependent manner.

doi: 10.3892/etm.2021.10710

Figure Lengend Snippet: Figure 2. RNAi‑mediated GPx4 and xCT knockdown promotes ox‑LDL‑induced ferroptosis in human endothelial cells. (A) Immunoblots of TNF‑α, IL‑α, 4‑HNE, GPx4 and xCT in the normal (control), ox‑LDL, si‑GPx4 + ox‑LDL, si‑GPx4 + ox‑LDL + DFOM, si‑xCT + ox‑LDL and si‑xCT + ox‑LDL + DFOM treatment groups. (B) Representative Transwell migration assay images of HUVECs treated with ox‑LDL, si‑GPx4 + ox‑LDL, si‑GPx4 + ox‑LDL + DFOM, si‑xCT + ox‑LDL and si‑xCT + ox‑LDL + DFOM (magnification, x10). #P<0.05 vs. control group; *P<0.05 vs. si‑GPx4 + ox‑LDL; -P<0.05 vs. si‑xCT + ox‑LDL. (C) Representative images showing HUVEC tube formation in the presence of PBS (control) or 100 µg/ml ox‑LDL,si‑GPx4 + 100 µg/ml ox‑LDL, si‑GPx4 + 100 µg/ml ox‑LDL + 125 µM DFOM, si‑xCT + 100 µg/ml ox‑LDL or si‑xCT + 100 µg/ml ox‑LDL + 125 µM DFOM (all magnifications, x10). #P<0.05 vs. control group, *P<0.05 vs. si‑GPx4 + ox‑LDL, ‑P<0.05 vs. si‑xCT + ox‑LDL. (D) Analysis of cell viability using the MTT assay in HUVECs after ox‑LDL, si‑GPx4 + ox‑LDL, si‑GPx4 + ox‑LDL + DFOM, si‑xCT + ox‑LDL and si‑xCT + ox‑LDL + DFOM treatments. Columns, mean of three experiments; bars, SD. #P<0.05 vs. control group, *P<0.05 vs. si‑GPx4 + ox‑LDL, ‑P<0.05 vs. si‑xCT + ox‑LDL. (E) Histogram of relative TNF‑α/β‑actin levels. #P<0.05 vs. control group, *P<0.05 vs. si‑GPx4 + ox‑LDL. (F) Histogram of relative IL‑α/β‑actin levels. #P<0.05 vs. control group, *P<0.05 vs. si‑GPx4 + ox‑LDL. (G) Histogram of relative 4‑HNE/β‑actin levels. #P<0.05 vs. control group, *P<0.05 vs. si‑GPx4 + ox‑LDL. (H) Histogram of relative xCT/β‑actin levels. #P<0.05 vs. control group, *P<0.05 vs. si‑GPx4 + ox‑LDL. (I) Histogram of relative GPx4/β‑actin levels. #P<0.05 vs. control group, *P<0.05 vs. si‑GPx4 + ox‑LDL. (J) Histogram of relative TNF‑α/β‑actin levels. #P<0.05 vs. control group, *P<0.05 vs. si‑xCT + ox‑LDL. (K) Histogram of relative IL‑α/β‑actin levels. #P<0.05 vs. control group, *P<0.05 vs. si‑xCT + ox‑LDL. (L) Histogram of relative 4‑HNE/β‑actin levels. #P<0.05 vs. control group, *P<0.05 vs. si‑xCT + ox‑LDL. (M) Histogram of relative GPx4/β‑actin levels. #P<0.05 vs. control group, *P<0.05 vs. si‑xCT + ox‑LDL. (N) Histogram of relative xCT/β‑actin levels. #P<0.05 vs. control group, *P<0.05 vs. si‑xCT + ox‑LDL. ox‑LDL, oxidized low‑density lipoprotein; GPx4, glutathione peroxidase 4; xCT, cystine‑glutamate antiporter; 4‑HNE, 4‑hydroxynonenal; DFOM, deferoxamine mesylate; HUVEC, human umbilical vein cell.

Article Snippet: The following primary antibodies were used: 4‐hydroxynonenal (4‐HNE; 1:2,000; ab46544; Abcam), TNF‐α (1:500; WL01581; Wanleibio; Co., Ltd.), IL‐1α (1:2,000; ab134908; Abcam), GPx4 (1:1,000; 14432‐1‐AP; ProteinTech Group, Inc.), xCT (1:1,000; 26864‐1‐AP; ProteinTech Group, Inc.); Caspase‐3/cleaved‐caspase3 (1:500; WL02117; Wanleibio; Co., Ltd.), Bax (1:1,000; WL01637; Wanleibio; Co., Ltd.), Bcl‐2 (1:1,000; WL01556; Wanleibio; Co., Ltd.), β‐actin (1:2,000; WL01845 Wanleibio; Co., Ltd.).

Techniques: Knockdown, Western Blot, Control, Transwell Migration Assay, MTT Assay

Journal: Experimental and therapeutic medicine

Article Title: Novel function of fluvastatin in attenuating oxidized low-density lipoprotein-induced endothelial cell ferroptosis in a glutathione peroxidase4- and cystine-glutamate antiporter-dependent manner.

doi: 10.3892/etm.2021.10710

Figure Lengend Snippet: Figure 3. Fluvastatin attenuates ox‑LDL‑induced endothelial cell dysfunction and ferroptosis. (A) Immunoblots of TNF‑α, IL‑α, 4‑HNE, GPx4 and xCT in the normal, ox‑LDL, ox‑LDL + 2.5 µM fluvastatin, ox‑LDL + 5 µM fluvastatin, ox‑LDL + 10 µM fluvastatin treatment groups. (B) Analysis of cell viability using the MTT assay in HUVECs after ox‑LDL, ox‑LDL + 2.5 µM fluvastatin, ox‑LDL + 5 µM fluvastatin and ox‑LDL + 10 µM fluvastatin treatments. Columns, means of three experiments; bars, SD. *P<0.05 vs. control group, ΔP<0.05 vs. ox‑LDL group. (C) Histogram of relative TNF‑α/β‑actin levels. *P<0.05 vs. control group, ΔP<0.05 vs. ox‑LDL group. (D) Histogram of relative IL‑α/β‑actin levels. *P<0.05 vs. control group, ΔP<0.05 vs. ox‑LDL group. (E) Histogram of relative 4‑HNE/β‑actin levels. *P<0.05 vs. control group, ΔP<0.05 vs. ox‑LDL group. (F) Histogram of relative GPx4/β‑actin levels. *P<0.05 vs. control group, ΔP<0.05 vs. ox‑LDL group. (G) Histogram of relative xCT/β‑actin levels.*P<0.05 vs. control group, ΔP<0.05 vs. ox‑LDL group. (H) Representative Transwell migration assay images of HUVECs treated with ox‑LDL, ox‑LDL + 2.5 µM fluvastatin, ox‑LDL + 5 µM fluvastatin, or ox‑LDL + 10 µM fluvastatin. *P<0.05 vs. control group, ΔP<0.05 vs. ox‑LDL group (magnification, 100x). (I) Representative tube formation assay images of HUVEC in the presence of PBS (control) or with 100 µg/ml ox‑LDL, 100 µg/ml ox‑LDL + 2.5 µM fluvastatin, 100 µg/ml ox‑LDL + 5 µM fluvastatin, or 100 µg/ml ox‑LDL + 10 µM fluvastatin (magnification, 100x). *P<0.05 vs. control group, ΔP<0.05 vs. ox‑LDL group. ox‑LDL, oxidized low‑density lipoprotein; GPx4, glutathione peroxidase 4; xCT, cystine‑glutamate antiporter; 4‑HNE, 4‑hydroxynonenal; HUVEC, human umbilical vein cell.

Article Snippet: The following primary antibodies were used: 4‐hydroxynonenal (4‐HNE; 1:2,000; ab46544; Abcam), TNF‐α (1:500; WL01581; Wanleibio; Co., Ltd.), IL‐1α (1:2,000; ab134908; Abcam), GPx4 (1:1,000; 14432‐1‐AP; ProteinTech Group, Inc.), xCT (1:1,000; 26864‐1‐AP; ProteinTech Group, Inc.); Caspase‐3/cleaved‐caspase3 (1:500; WL02117; Wanleibio; Co., Ltd.), Bax (1:1,000; WL01637; Wanleibio; Co., Ltd.), Bcl‐2 (1:1,000; WL01556; Wanleibio; Co., Ltd.), β‐actin (1:2,000; WL01845 Wanleibio; Co., Ltd.).

Techniques: Western Blot, MTT Assay, Control, Transwell Migration Assay, Tube Formation Assay

Journal: Experimental and therapeutic medicine

Article Title: Novel function of fluvastatin in attenuating oxidized low-density lipoprotein-induced endothelial cell ferroptosis in a glutathione peroxidase4- and cystine-glutamate antiporter-dependent manner.

doi: 10.3892/etm.2021.10710

Figure Lengend Snippet: Figure 4. GPx4 and xCT are required for the fluvastatin‑mediated attenuation of ferroptosis in human endothelial cells. (A) Immunoblots of TNF‑α, IL‑α, 4‑HNE, GPx4 and xCT in the normal, ox‑LDL, ox‑LDL + 10 µM fluvastatin, si‑GPx4 + ox‑LDL, si‑GPx4 + ox‑LDL + 10 µM fluvastatin, si‑xCT + ox‑LDL and si‑xCT + ox‑LDL + 10 µM fluvastatin treatment groups. (B) Analysis of cell viability using the MTT assay in HUVECs after ox‑LDL, ox‑LDL + 10 µM fluv astatin, si‑GPx4 + ox‑LDL, si‑GPx4 + ox‑LDL + 10 µM fluvastatin, si‑xCT + ox‑LDL or si‑xCT + ox‑LDL + 10 µM fluvastatin treatments. Columns, means of three experiments; bars, SD. *P<0.05 vs. control group, &P<0.05 vs. si‑GPx4 + ox‑LDL, +P<0.05 vs. si‑xCT + ox‑LDL. (C) Histogram of relative TNF‑α/β‑actin levels. *P<0.05 vs. control group, &P<0.05 vs. si‑GPx4 + ox‑LDL, +P<0.05 vs. si‑xCT + ox‑LDL. (D) Histogram of relative IL‑α/β‑actin levels. *P<0.05 vs. control group, &P<0.05 vs. si‑GPx4 + ox‑LDL, +P<0.05 vs. si‑xCT + ox‑LDL. (E) Histogram of relative 4‑HNE/β‑actin levels. *P<0.05 vs. control group, &P<0.05 vs. si‑GPx4 + ox‑LDL, +P<0.05 vs. si‑xCT + ox‑LDL. (F) Histogram of relative GPx4/β‑actin levels. *P<0.05 vs. control group, &P<0.05 vs. si‑GPx4 + ox‑LDL, +P<0.05 vs. si‑xCT + ox‑LDL. (G) Histogram of relative xCT/β‑actin levels. *P<0.05 vs. control group, &P<0.05 vs. si‑GPx4 + ox‑LDL, +P<0.05 vs. si‑xCT + ox‑LDL. (H) Representative Transwell migration assay images of HUVECs treated with ox‑LDL, ox‑LDL + 10 µM fluvastatin, si‑GPx4 + ox‑LDL, si‑GPx4 + ox‑LDL + 10 µM fluvastatin, si‑xCT + ox‑LDL and si‑xCT + ox‑LDL + 10 µM fluvastatin. *P<0.05 vs. control group, &P<0.05 vs. si‑GPx4 + ox‑LDL, +P<0.05 vs. si‑xCT + ox‑LDL (magnification, 100x). (I) Representative tube formation assay images of HUVECs in the presence of PBS or with 100 µg/ml ox‑LDL, 100 µg/ml ox‑LDL + 10 µM fluvastatin, si‑GPx4 + 100 µg/ml ox‑LDL, si‑GPx4 + 100 µg/ml ox‑LDL + 10 µM fluvastatin, si‑xCT + 100 µg/ml ox‑LDL or si‑xCT + 100 µg/ml ox‑LDL + 10 µM fluvastatin (magnification, 100x). *P<0.05 vs. control group, &P<0.05 vs. si‑GPx4 + ox‑LDL, +P<0.05 vs. si‑xCT + ox‑LDL. ox‑LDL, oxidized low‑density lipoprotein; GPx4, glutathione peroxidase 4; xCT, cystine‑glutamate antiporter; 4‑HNE, 4‑hydroxynonenal; HUVEC, human umbilical vein cell.

Article Snippet: The following primary antibodies were used: 4‐hydroxynonenal (4‐HNE; 1:2,000; ab46544; Abcam), TNF‐α (1:500; WL01581; Wanleibio; Co., Ltd.), IL‐1α (1:2,000; ab134908; Abcam), GPx4 (1:1,000; 14432‐1‐AP; ProteinTech Group, Inc.), xCT (1:1,000; 26864‐1‐AP; ProteinTech Group, Inc.); Caspase‐3/cleaved‐caspase3 (1:500; WL02117; Wanleibio; Co., Ltd.), Bax (1:1,000; WL01637; Wanleibio; Co., Ltd.), Bcl‐2 (1:1,000; WL01556; Wanleibio; Co., Ltd.), β‐actin (1:2,000; WL01845 Wanleibio; Co., Ltd.).

Techniques: Western Blot, MTT Assay, Control, Transwell Migration Assay, Tube Formation Assay

Journal: PeerJ

Article Title: 3,4-dihydroxyphenylethyl alcohol glycoside reduces acetaminophen-induced acute liver failure in mice by inhibiting hepatocyte ferroptosis and pyroptosis

doi: 10.7717/peerj.13082

Figure Lengend Snippet: MDA levels of DAG are decreased compared with APAP. (** p < 0.0001 APAP compared with control, # p = 0.0012 compared with DAG). Proteins extracted from the liver were identified using western blot. DAG decreased the levels of p-ERK and HO-1, whereas it increased the levels of GPX4 (B).

Article Snippet: At room temperature, the membrane was sealed with 5% (w/v) skimmed milk for 2 h. Next, the membrane was incubated with the primary antibody overnight at 4 °C: ERK, p-ERK, HO-1, GPX4, NLRP3, Gasdermin-D (Cell Signaling Technology, MA, USA), Caspase 1 (p20) (Santa Cruz, CA, USA), which would combine HRP.

Techniques: Control, Western Blot

Journal: PeerJ

Article Title: 3,4-dihydroxyphenylethyl alcohol glycoside reduces acetaminophen-induced acute liver failure in mice by inhibiting hepatocyte ferroptosis and pyroptosis

doi: 10.7717/peerj.13082

Figure Lengend Snippet: (A) GSH content in AML12 cells. (**** p < 0.0001 control compared with APAP and APAP compared with adding 150 µM DAG. ** p < 0.05 APAP compared with adding 100 µM DAG) ( n = 3 per group df = 2). (B) ROS in AML12 cells. (C) The expression of IL-1β, IL-18 and NLRP3 in AML12 cells. # p < 0.05 APAP compared with control, ** p < 0.005 APAP compared with adding 100 µM DAG, ** p < 0.005 APAP compared with adding 150 µM DAG (IL-1β). # p < 0.005 APAP compared with control, ** p < 0.05 APAP compared with adding 50 µM DAG, **** p < 0.0001 APAP compared with adding 100 µM DAG, **** p < 0.0001 APAP compared with adding 150 µM DAG (IL-18). # p < 0.005 APAP compared with control, ** p < 0.005 APAP compared with adding 100 µM DAG, ** p < 0.005 APAP compared with adding 150 µM DAG (NLRP3) ( n = 3 per group df = 2). (D) Proteins extracted from AML12 cells were identified using western blot. DAG decreased the levels of p-ERK, HO-1, NLRP3, GSDMD and Caspase1 (p20), while at the same time it increased the levels of GPX4.

Article Snippet: At room temperature, the membrane was sealed with 5% (w/v) skimmed milk for 2 h. Next, the membrane was incubated with the primary antibody overnight at 4 °C: ERK, p-ERK, HO-1, GPX4, NLRP3, Gasdermin-D (Cell Signaling Technology, MA, USA), Caspase 1 (p20) (Santa Cruz, CA, USA), which would combine HRP.

Techniques: Control, Expressing, Western Blot

Journal: Oxidative medicine and cellular longevity

Article Title: Melatonin Inhibits the Ferroptosis Pathway in Rat Bone Marrow Mesenchymal Stem Cells by Activating the PI3K/AKT/mTOR Signaling Axis to Attenuate Steroid-Induced Osteoporosis.

doi: 10.1155/2022/8223737

Figure Lengend Snippet: Figure 1: DEX activates the ferroptotic pathway of BMSCs. (a) ROS staining was performed to test the correlation between different concentrations of DEX and the level of oxidative stress. (b) Quantitative analysis of the number of ROS-positive cells per field in (a). (c) Annexin V-mCherry/SYTOX Green detection kit was used to detect cell death. (d) Quantitative analysis of the percentage of SYTOX green-positive cells in (c). (e–i) BMSCs were treated with various concentrations of DEX for 24 h, and the expressions of system xc-, ACSL4, GPX4, and FSP1were analyzed by western blot and qRT-PCR. (j) Images of immunofluorescence staining of GPX4 in BMSCs after treatment with DEX (10-3 M) for 72 h. (k) Quantification of the fluorescence intensity of GPX4 immunofluorescence positively stained cells. These studies were performed at least 3 biological replicates. Data represent mean ± SD (n = 3). ∗P < 0:05, ∗∗P < 0:01, ∗∗∗P < 0:005 compared with control group.

Article Snippet: To more fully locate and quantitatively examine antigenic substances in tissues, sections were first stained with primary antibodies against GPX4 (1 : 200, orb340797, Biorbyt), CD90 (1 : 500,RT1615, 0 20 40 60 80 G PX 4 po sit iv e c el l nu m be r ( m m –2 ) ⁎⁎⁎ Co nt ro l M od el (i) 0 20 40 60 FS P1 p os iti ve ce ll nu m be r ( m m –2 ) ⁎⁎⁎ Co nt ro l M od el (j) 0 20 40 60 M T1 p os iti ve ce ll nu m be r ( m m –2 ) ⁎⁎⁎ Co nt ro l M od el (k) Figure 2: DEX activates the ferroptotic pathway in SIOP. (a) Quantification of the protein level of MT by ELISA. (b) Images of micro-CT. (c) BMD(g/cm3). (d) BV/TV (%). (e) Images of immunofluorescence double staining of CD90 and GPX4 in bone tissues. (f) Quantitative analysis of the area of GPX4/CD90-positive stains. (g) IHC staining of system xc-, GPX4, FSP1, and MT1, and the IHC-positive cells were marked with black arrows. (h–k) Quantitative analysis of the number of the IHC-positive cells in (g).

Techniques: Staining, Western Blot, Quantitative RT-PCR, Control

Journal: Biomedicine & pharmacotherapy = Biomedecine & pharmacotherapie

Article Title: Dexmedetomidine attenuates myocardial ischemia/reperfusion-induced ferroptosis via AMPK/GSK-3β/Nrf2 axis.

doi: 10.1016/j.biopha.2022.113572

Figure Lengend Snippet: Fig. 4. Dexmedetomidine produced antioxidant effect and inhibited HR-induced accumulation of intracellular ferrous iron and lipid peroxidation in H9c2 cells. (A-B) Intracellular ferrous iron was detected by Phen Green SK staining. (C-D) Lipid ROS level was assessed by C11-BODIPY staining. (E-F) Levels of MDA and 4-HNE, markers of lipid peroxidation were investigated by Elisa. (G) Intracellular ROS production was measured with H2DCFDA. (H-I) levels of GSH and SOD activity were assessed by Elisa. Data were expressed as mean ± SD, (n = 3 for each group). *P < 0.05; * *P < 0.01; * **P < 0.001; * ** *P < 0.0001; ns, not significant.

Article Snippet: The glutathione peroxidase 4 (GPX4) activity was determined using a GPX4 Elisa kit (CUSABIO, China).

Techniques: Produced, Staining, Enzyme-linked Immunosorbent Assay, Activity Assay

Journal: Biomedicine & pharmacotherapy = Biomedecine & pharmacotherapie

Article Title: Dexmedetomidine attenuates myocardial ischemia/reperfusion-induced ferroptosis via AMPK/GSK-3β/Nrf2 axis.

doi: 10.1016/j.biopha.2022.113572

Figure Lengend Snippet: Fig. 6. Effects of dexmedetomidine on the expressions of Nrf2, HO-1, SLC7A11, and GPX4 in H9c2 cells under HR. (A) Protein bands of cytoplasmic Nrf2 of H9c2 cells among four groups were assayed by Western blot, and GAPDH served as an internal control for sample loading. (B) Intensities of protein bands of cytoplasmic Nrf2. (C) Protein bands of nuclear Nrf2 of H9c2 cells among four groups were assayed by Western blot, and Lamin B served as an internal control of Nrf2 for sample loading. (D) Intensities of protein bands of nuclear Nrf2. (E) Protein bands of HO-1, SLC7A11 and GPX4 of H9c2 cells among four groups were assayed by Western blot, and GAPDH served as an internal control for sample loading. (F-G) Intensities of protein bands of HO-1, SLC7A11 and GPX4 of H9c2 cells among four groups were normalized to GAPDH. Results are expressed as mean ± SD, (n = 3 for each group). *P < 0.05; * *P < 0.01; * **P < 0.001; ns, not significant.

Article Snippet: The glutathione peroxidase 4 (GPX4) activity was determined using a GPX4 Elisa kit (CUSABIO, China).

Techniques: Western Blot, Control

Journal: Biomedicine & pharmacotherapy = Biomedecine & pharmacotherapie

Article Title: Dexmedetomidine attenuates myocardial ischemia/reperfusion-induced ferroptosis via AMPK/GSK-3β/Nrf2 axis.

doi: 10.1016/j.biopha.2022.113572

Figure Lengend Snippet: Fig. 7. Dexmedetomidine increased the levels of SLC7A11 and GPX4 in HR-treated H9c2 cells. (A) SLC7A11 expression was measured by immunofluorescent staining. Cells were immunostained using antibody specific for SLC7A11 (red). Nuclei were stained with DAPI (blue). Scale bars represent 20 µm. Images were captured at × 400 magnification. (B) Quantitative analysis of SLC7A11 expression. (C) GPX4 expression was measured by immunofluorescent staining. Cells were immunostained using antibody specific for GPX4 (red). Nuclei were stained with DAPI (blue). Scale bars represent 20 µm. Images were captured at × 400 magnification. (D) Quantitative analysis of GPX4 expression. (D) GPX4 activity was determined by Elisa. Results were expressed as mean ± SD, (n = 3 for each group). *P < 0.05; * *P < 0.01; * ** *P < 0.0001.

Article Snippet: The glutathione peroxidase 4 (GPX4) activity was determined using a GPX4 Elisa kit (CUSABIO, China).

Techniques: Expressing, Staining, Activity Assay, Enzyme-linked Immunosorbent Assay

Journal: The Journal of Biological Chemistry

Article Title: Cell clustering mediated by the adhesion protein PVRL4 is necessary for α6β4 integrin–promoted ferroptosis resistance in matrix-detached cells

doi: 10.1074/jbc.RA118.003017

Figure Lengend Snippet: Clustering of matrix-detached cells influences lipid peroxidation, GPX4 expression, and Src activity. A, lipid peroxidation was quantified using the MDA assay in control and β4-depleted MCF10-A and SUM-159 cells under either adherent, detached clustered (no additive), or detached single (0.5% methylcellulose) conditions for 4 h. Similar results were obtained using 2 mm EDTA. B, GPX4 mRNA expression was quantified by qPCR in control and β4-depleted MCF10-A and SUM-159 cells under either adherent, detached clustered (no additive) or detached single (0.5% methylcellulose) conditions for 2 h. C, GPX4 protein expression was assessed by immunoblotting in control MCF10-A, SUM-159, and Hs578t cells under detached clustered (no additive) or detached single (0.5% methylcellulose) conditions for 4 h. Immunoblots were quantified by densitometry, and the ratio of the intensity of the GPX4/actin bands relative to adherent cells is shown under the blots. D, SUM-159 β4-depleted cells were detached for 24 h with either no additive (clustered) or 2 mm EDTA (single) in the presence of either DMSO or 500 mm α-tocopherol, and the number of viable cells was quantified. E, control and β4-depleted cells SUM-159 cells that had been transfected with either a vector control or a GPX4 expression vector were detached for 24 h with no additive (Control) or 0.5% methylcellulose, and the number of viable cells was quantified. F, SUM-159 and Hs578t control cells were assessed for phosphorylated (Tyr-418) Src by immunoblotting under adherent conditions (Adh) as well as following 2 h of matrix detachment with no additive (NA), 2 mm EDTA (EDTA), or 0.5% methylcellulose (MC). NS, not significant. **, p < 0.01, ***, p < 0.005.

Article Snippet: To express GPX4, a plasmid construct for GPX4 was purchased from Origene (RC208065).

Techniques: Expressing, Activity Assay, Multiple Displacement Amplification, Western Blot, Transfection, Plasmid Preparation

Journal: Poultry Science

Article Title: New insights into the spleen injury by mitochondrial dysfunction of chicken under polystyrene microplastics stress

doi: 10.1016/j.psj.2024.103674

Figure Lengend Snippet: The primers involved in this study.

Article Snippet: GPX4 , 1:500 , Boster Biotechnology, China , A02059-1.

Techniques:

Journal: Poultry Science

Article Title: New insights into the spleen injury by mitochondrial dysfunction of chicken under polystyrene microplastics stress

doi: 10.1016/j.psj.2024.103674

Figure Lengend Snippet: The antibodies used in this study.

Article Snippet: GPX4 , 1:500 , Boster Biotechnology, China , A02059-1.

Techniques:

Journal: Poultry Science

Article Title: New insights into the spleen injury by mitochondrial dysfunction of chicken under polystyrene microplastics stress

doi: 10.1016/j.psj.2024.103674

Figure Lengend Snippet: MPs induced ferroptosis in spleen. (A) Western Blot results in ferroptosis-related proteins. (B) Quantitative analysis of GPX4, FTH1, and SLC7A11 protein expression. (C) Ferroptosis-related factors of mRNA expression changes. Data were expressed as mean ± SD, n = 3. *, **, ***, denotes: p < 0.05, 0.01, and 0.001, respectively.

Article Snippet: GPX4 , 1:500 , Boster Biotechnology, China , A02059-1.

Techniques: Western Blot, Expressing