Journal: Experimental and therapeutic medicine

Article Title: 1,25(OH) 2 D 3 ameliorates doxorubicin‑induced cardiomyopathy by inhibiting the NLRP3 inflammasome and oxidative stress.

doi: 10.3892/etm.2023.12112

Figure Lengend Snippet: Figure 3. Increased oxidative stress and NLRP3 inflammasome pathways are ameliorated by 1,25(OH)2D3 treatment in DOX‑induced cardiomyopathy in mice. (A) AC16 cells treated with different concentrations of 1,25(OH)2D3 (0, 25, 50, 100, 200 and 400 nM) for 24 h. Cell viability was measured using CCK‑8 assay (n=3). (B) AC16 cells were treated with 1 µM DOX and different concentrations of 1,25(OH)2D3 (0, 25, 50, 100 and 200 nM) for 24 h. Cell viability was measured using CCK‑8 assay (n=3). ****P<0.0001 vs. Control group; #P<0.05 and ####P<0.0001 vs. DOX‑only group; &&&P<0.001 vs. DOX + 25 nM 1,25(OH)2D3‑treated group; @@P<0.01 vs. DOX + 50 nM 1,25(OH)2D3‑treated group. (C) Levels of reactive oxygen species and (D) densitometric analysis in AC16 cells after treatment with DOX or DOX + 1,25(OH)2D3 via DCFH‑DA staining (n=5). *P<0.05 and ****P<0.0001 vs. Control; ###P<0.001 vs. DOX group. Levels of (E) MDA, (F) T‑SOD and (G) GSH‑Px in cardiac tissue 4 weeks after the first injection of DOX (n=5). (H) Western blotting images of cardiac tissue extracts analyzing protein expression of SOD1, SOD2, Nrf2 and KEAP1. GAPDH was used as the loading control. (I) Semi‑quantification of protein expression levels of SOD1, SOD2, Nrf2 and KEAP1. Protein expression levels relative to the Sham group were assessed by densitometric analysis (n=3). (J) Western blotting images of cardiac tissue extracts analyzing protein expression of NLRP3, ASC and Caspase‑1. GAPDH was used as the loading control. (K) Semi‑quantification of protein expression levels NLRP3, ASC and Caspase‑1. Protein expression levels relative to the Sham group were assessed by densi‑ tometric analysis (n=3). *P<0.05, **P<0.01, ***P<0.001 and ****P<0.0001 vs. Sham; #P<0.05, ##P<0.01, ###P<0.001 and ####P<0.001 vs. DOX. DOX, doxorubicin hydrochloride; CCK‑8, Cell Counting Kit‑8; ns, no significant difference; DCFH‑DA, 2',7'‑dichlorofluorescin diacetate; MDA, malondialdehyde; T, total; GSH‑Px, glutathione peroxidase; SOD, superoxide dismutase; Nrf2, nuclear erythroid 2‑related factor 2; KEAP1, Kelch‑like ECH‑associated protein 1; ASC, caspase recruitment domain; NLRP3, nod‑like receptor family pyrin domain‑containing 3 inflammasome.

Article Snippet: The dilutions of primary antibodies used were as follows: Collagen I (cat. No. GTX26308; 1:1,000; GeneTex, Inc.), α‐SMA (α‐smooth muscle actin; cat. No. ab5694; 1:1,000; Abcam), IL‐1β (cat. no. ab9722; 0.25 μg/ml; Abcam), IL‐6 (cat. No. A0286; 1:1,000; Abclonal Biotech Co., Ltd.), TNF‐α (cat. No. GTX110520; 1:500; GeneTex, Inc.), SOD1 (cat. No. ab16831; 1:2,000; Abcam), SOD2 (cat. No. NB100‐1992; 1:2,000; Novus Biologicals, LLC), Nrf2 (cat. No. GTX135165; 1:1,000; GeneTex, Inc.), KEAP1 (Kelch‐like ECH‐associated protein 1; cat. No. ab119403; 1:1,000; Abcam), Caspase‐1 (cat. No. 83383; 1:500; Cell Signaling Technology, Inc.), NLRP3 (cat. No. ab214185; 1:1,000; Abcam), ASC (caspase recruitment domain; cat. No. GTX55818; 1:1,000; GeneTex, Inc.) and GAPDH (cat. No. 2118; 1:1,000; Cell Signaling Technology, Inc.).

Techniques: CCK-8 Assay, Control, Staining, Injection, Western Blot, Expressing

Journal: Oxidative Medicine and Cellular Longevity

Article Title: Superoxide Anion Production and Bioenergetic Profile in Young and Elderly Human Primary Myoblasts

doi: 10.1155/2018/2615372

Figure Lengend Snippet: SOD1 and SOD2 protein expression. Western blotting analysis of superoxide enzymes cytosolic type 1 and mitochondrial type 2 was performed on young and elderly myoblasts and myotubes. β -Actin content was used for normalization. Panels (a), (b), (c), and (d) show SOD1 and SOD2 densitometric analysis of Western blots performed on three young and three old samples expressed as mean ± SEM. Representative patterns of SOD1, SOD2, and β -actin (as loading control) expression in young and elderly myoblasts and myotubes are shown in panels (e) and (f). The bands were taken from two nonadjacent lanes originating from exactly the same gel and blot with exactly the same exposure time, but spliced together indicated with double-dotted lines. Moreover, we performed no change in contrast ( ∗ p ≤ 0.05).

Article Snippet: Western blotting (WB) analysis was performed on 40 μ g lysates from young and old myoblasts and myotubes, using SOD1 (71G8) mouse mAb (number 4266, Cell Signalling Technology, Danvers, MA, USA) at 1 : 1000, SOD2 (D9V9C) rabbit mAb (number 13194, Cell Signalling Technology) at 1 : 1000, and β -actin (8H10D10) mouse mAb (number 3700, Cell Signalling Technology) at 1 : 1000, as primary antibody, and secondary HRP-conjugated antibodies (Cell Signalling Technology) at 1 : 5000.

Techniques: Expressing, Western Blot, Control

Journal: Cell metabolism

Article Title: SELECTIVE PERSULFIDE DETECTION REVEALS EVOLUTIONARILY CONSERVED ANTI-AGING EFFECTS OF S -SULFHYDRATION

doi: 10.1016/j.cmet.2019.10.007

Figure Lengend Snippet: KEY RESOURCES TABLE

Article Snippet: MnSOD , Creative BioMart , Cat# SOD2-1039H.

Techniques: Ab Array, Recombinant, Protease Inhibitor, Magnetic Beads, Sequencing, Mutagenesis, Plasmid Preparation, Software

Journal: Scientific Reports

Article Title: AOH induces oxidative stress and DNA damage in ovarian cancer cells via modulation of GPER1 and HIF1α/PI3K/CLDNs signaling pathway

doi: 10.1038/s41598-025-26042-9

Figure Lengend Snippet: AOH induces oxidative stress in OVCAR3 cells. ( a ) Oxidative stress: The analysis of the generation of ROS with flow cytometry. ( b ) Representative results of flow cytometry. ( c ) SOD2 protein expression: Expression of SOD2 was evaluated with Western Blot, the representative results. ( d ) SOD1, SOD2, PARP1 and XRCC1 gene expression: Relative expression of SOD1, SOD2 , PARP1 and XRCC1 . ( e ) DNA damage: evaluated with flow cytometry (ATM and H2A.X activation). ( f ) Representative results of flow cytometry. ( g ) p-H2A.X, PARP1 and cleaved-PARP1 protein expression: Expression of phospho-H2A.X, PARP1 and cleaved—PARP1 evaluated with Western Blot, the representative results. ( h ) Confocal microscopy: Bar graph showing the difference in normalized mean fluorescence intensity signal. ( i ) Immunofluorescent staining of phospho-H2A.X (green) and nucleus (orange, NucSpot 555 / 570). All the results are expressed as mean ± SE. One-way ANOVA with Bonferroni correction was used for statistical analysis. P < 0.05 was considered statistically significant, * p < 0.05, ** p < 0.01, *** p < 0.001. AOH alternariol, G15 selective GPER1 receptor antagonist, G1 selective GPER1 agonist, SOD1 Superoxide dismutase 1, SOD2 Superoxide dismutase 2, PARP1 Poly [ADP-ribose] polymerase 1, XRCC1 X-ray repair cross-complementing protein 1, GAPDH anti-glyceraldehyde 3-phosphate dehydrogenase, pH2AX phospho-H2A histone family member X. Cnt control (non-treated cells).

Article Snippet: Akt (#9272), phospho-Akt (Ser473) (#4060), PARP1 (#46,011), SOD1 (#4266), SOD2 (#13,141), pH2AX (#9718), CLDN1 (#13,995), CLDN3 (#83,609), PI3K (#11,889), p44/42 (#4695), phospho-p44/42 (#4370), CLDN4 (STJ114778), GPER1 (STJ192629) (St John’s Laboratory Ltd, University Way, London) antibodies were used according to the manufacturer’s recommendations.

Techniques: Flow Cytometry, Expressing, Western Blot, Gene Expression, Activation Assay, Confocal Microscopy, Fluorescence, Staining, Control