Journal: Stem cells and development

Article Title: Characterization of superoxide dismutases in cardiac progenitor cells demonstrates a critical role for manganese superoxide dismutase.

doi: 10.1089/scd.2012.0191

Figure Lengend Snippet: FIG. 2. CPCs have higher basal SOD activity and protein levels compared to myocytes and fibroblasts. (A) CPCs have higher SOD1 (left panel) and SOD2 (right panel) activities compared to fibroblasts and myocytes. n ‡ 4, **P < 0.01 versus CPC, ANOVA followed by the Dunnett’s post-test. (B) In all the 3 cell types, 90% of total SOD activity is due to SOD1 and the remaining is due to SOD2. (C) Western blot analysis shows that CPCs have significantly higher basal SOD1 and SOD2 protein levels versus myocytes and (D) versus FB. n ‡ 4, *P < 0.05 versus CPC, t-test; **P < 0.01 versus CPC, t-test. SOD, superoxide dismutase; CM, cardiomyocytes; FB, fibroblasts.

Article Snippet: Protein-blotted PVDF membranes were blocked with 4% milk in tris-buffered saline containing 0.1% Tween and probed with antibodies for SOD1, SOD2, GAPDH, or beta-actin (Santa Cruz Biotechnology).

Techniques: Activity Assay, Western Blot

Journal: Stem cells and development

Article Title: Characterization of superoxide dismutases in cardiac progenitor cells demonstrates a critical role for manganese superoxide dismutase.

doi: 10.1089/scd.2012.0191

Figure Lengend Snippet: FIG. 4. XXO treatment increases SOD1 and SOD2 protein and activity levels in CPCs. (A) Shows a representative blot of both SOD1 and SOD2 in cells treated with XXO for the indicated time. Both (B) SOD1 and (C) SOD2 activities re- presented as fold change normalized to initial levels are significantly increased following 48 h of XXO treatment compared to its time-matched controls. n ‡ 4, 2-way ANOVA followed by the Bonferroni post-test, **P < 0.01, ***P < 0.001 versus respective time-matched controls.

Article Snippet: Protein-blotted PVDF membranes were blocked with 4% milk in tris-buffered saline containing 0.1% Tween and probed with antibodies for SOD1, SOD2, GAPDH, or beta-actin (Santa Cruz Biotechnology).

Techniques: Activity Assay

Journal: Stem cells and development

Article Title: Characterization of superoxide dismutases in cardiac progenitor cells demonstrates a critical role for manganese superoxide dismutase.

doi: 10.1089/scd.2012.0191

Figure Lengend Snippet: FIG. 3. XXO treatment increases SOD1 and SOD2 expres- sion in CPCs. (A) SOD1 and (B) SOD2 mRNA expressions normalized to 18 s mRNA expression are significantly in- creased following XXO addition compared to control cells without XXO. n ‡ 7, ANOVA followed by the Dunnett’s post- test, *P < 0.05 versus control, **P < 0.01 versus control.

Article Snippet: Protein-blotted PVDF membranes were blocked with 4% milk in tris-buffered saline containing 0.1% Tween and probed with antibodies for SOD1, SOD2, GAPDH, or beta-actin (Santa Cruz Biotechnology).

Techniques: Expressing, Control

Journal: Stem cells and development

Article Title: Characterization of superoxide dismutases in cardiac progenitor cells demonstrates a critical role for manganese superoxide dismutase.

doi: 10.1089/scd.2012.0191

Figure Lengend Snippet: FIG. 5. Silencing of SOD2 and not SOD1 sensitizes cells to XXO-induced apoptosis. (A) Representative images of CPCs in 20 · magnification; nuclei stained blue with DAPI and apoptotic cells stained red using TUNEL assay. (B, C) Grouped data following 24 h of XXO treatment after 48 h of siRNA transfection show that the percentage of TUNEL- positive cells (B) and cells in the sub-G1 phase (C) increases significantly only in SOD2 silenced CPCs. *P < 0.05, **P < 0.01 versus own control; 2-way ANOVA followed by the Bon- ferroni post-test.

Article Snippet: Protein-blotted PVDF membranes were blocked with 4% milk in tris-buffered saline containing 0.1% Tween and probed with antibodies for SOD1, SOD2, GAPDH, or beta-actin (Santa Cruz Biotechnology).

Techniques: Staining, TUNEL Assay, Transfection, Control

Journal: Redox Biology

Article Title: MG53 protein rejuvenates hUC-MSCs and facilitates their therapeutic effects in AD mice by activating Nrf2 signaling pathway

doi: 10.1016/j.redox.2022.102325

Figure Lengend Snippet: MG53 reduces oxidative stress in senescent hUC-MSCs by activating the Nrf2 signaling pathway. (A) ROS production measured by DCFH-DA kit. (B) MDA level. (C) SOD viability. (D) Protein expression of Nrf2 signaling pathway in each group. (E) Densitometric analysis of Nrf2 expression in the nucleus. (F) Densitometric analysis of Keap1, NQO1, SOD1 and Nrf2 in the cytosol. Data were presented as mean ± SEM. Data were presented as mean ± SEM. N = 3 per group. *: Compared with P5 group, P < 0.05; #: Compared with P10 group, P < 0.05; Δ: Compared with P15, P < 0.05.

Article Snippet: The primary antibodies were MG53 (1:1000), P16 (1:1000, C610021, Sangon, China), P21 (1:1000, D220091, Sangon, China), P53(1:1000, 10442-1-AP, Proteintech, China), Tau (1:1000, D121297, Sangon, China), p-Tau (Ser396) (1:1000, D155045, Sangon, China), p-Tau (Ser231) (1:1000, D155301, Sangon, China) and p-Tau (Ser235) (1:1000, D155045, Sangon, China), Nrf2 (1:1000, WL02135, Wanlei, China), SOD1 (1:1000, WL01846, Wanlei, China), NQO1 (1:1000, WL04860, Wanlei, China) and Keap-1 (1:1000, WL03285, Wanlei, China), pCNA (1:1000, 10205-2-AP, Proteintech, China), Sirt1 (1:2000, 13161-1-AP, Proteintech, China), β-actin (1:2000, 20536-1-AP, Proteintech, China) and Histone-3 (1:2000, WL0984a, Wanlei, China). β-Actin and Histone-3 were used as internal reference of protein expression in the cytosol and nucleus, respectively.

Techniques: Expressing

Journal: Frontiers in Pharmacology

Article Title: Enhancing Fatty Acids Oxidation via L-Carnitine Attenuates Obesity-Related Atrial Fibrillation and Structural Remodeling by Activating AMPK Signaling and Alleviating Cardiac Lipotoxicity

doi: 10.3389/fphar.2021.771940

Figure Lengend Snippet: LCA inhibits cardiac oxidative stress and mitigates inflammation. Representative images and analysis of the subcellular localization of the oxidation products of (A,B) MitoSOX and (C,D) DHE. Red: oxidation products-staining. Scar: 10 μm n = 4. (E,F) Comparisons of serum MDA and SOD using commercially available kits among the groups. (G,H) Levels of atrial MDA and SOD normalized to tissue protein concentration. (I) Schematic diagram illustrating NRF2-related signals. (J,K) Localization of NRF2 in atria by confocal immune-cyto-chemical analysis: Blue: nucleus (DAPI); Red: NRF2-staining; Pink: merge of blue and red indicated nuclear localization of NRF2 (Arrow). Scar: 30 μm n = 4. (L,M) Representative images and quantitative analysis of anti-oxidative system involved protein expressions (NRF2 and SOD2) using Western blot. (N,O) Representative images and analysis of oxidative DNA damage by 8-OH-dG staining. Green: 8-OH-dG -staining; Blue: DAPI. Scar: 50 μm n = 4. (P,Q) Representative images and analysis of DNA damage by TUNEL staining. Red: TUNEL-staining; Blue: DAPI; Pink: merge. Scar: 50 μm n = 4. Comparisons of (R) serum LDH and serum (S) CK-MB using commercially available kits. n = 8. (T) Quantitative analysis of the expression of inflammation-related genes in the atria using RT-qPCR. (U) Representative images and quantitative analysis of expression and phosphorylation of NFκB using Western blot. One-way ANOVA with Bonferroni post-hoc test was used to compare data among groups. Data are expressed as mean ± SEM. * p < 0.05, ** p < 0.01. STD, standard diet; HFD, high-fat diet; LCA, l -carnitine; DHE, dihydroethidium; MDA, molondialdehyde; SOD, superoxide dismutase; NRF2, nuclear erythroid 2 p45-related factor 2; p-, phoso-; SOD2, manganese superoxide dismutase, superoxide dismutase 2; 8-OH-Dg, 8-hydroxydeoxyguanosine; IL, interleukin; TNF-α, tumor necrosis factor-α; MCP-1, monocyte chemoattractant protein-1; NFκB, the nuclear factor kappa B; LDH, lactate dehydrogenase; CK-MB, creatine kinase-MB.

Article Snippet: The membranes were blocked with 5% skim milk and incubated with antibodies against total-AMPK (1:1,000; Cell Signaling Technology/CST, Massachusetts, United States), CD36 (1:1,000; Abcam), collagen I (1:1,000; Abcam), collagen III (1:1,000; Abcam), connexin-43 (1:200; Invitrogen), CPT1B (1:1,000; Proteintech), GLUT4 (1:500; Proteintech), NFκB (1:1,000; Proteintech), NRF2 (1:1,000; Proteintech), phoso-AMPK (Thr172; 1:1,000; CST), phoso-Akt (Ser473; 1:1,000; CST), phoso-NFκB (1:1,000; CST), PGC1α (1:1,000; Proteintech), SOD2 (1:1,000; Proteintech), TGF-β (1:1,000; Abcam), α-SMA (1:1,000; CST), β-actin (1:5,000; Proteintech).

Techniques: Staining, Protein Concentration, Western Blot, TUNEL Assay, Expressing, Quantitative RT-PCR, Phospho-proteomics

Journal: Frontiers in Pharmacology

Article Title: Enhancing Fatty Acids Oxidation via L-Carnitine Attenuates Obesity-Related Atrial Fibrillation and Structural Remodeling by Activating AMPK Signaling and Alleviating Cardiac Lipotoxicity

doi: 10.3389/fphar.2021.771940

Figure Lengend Snippet: Pharmacological inhibition of AMPK via CC attenuated LCA-conferred beneficial effects in palmitate-treated primary atrial cardiomyocytes. (A) Schematic diagram illustrating the cell isolation, culture and treatment. (B) FAO rate in different treatment groups. The measure of substrate utilization after 18°C unsaturated fatty acid (Oleate, 100 μM) addition was normalized with maximal O 2 consumption in Control cells. (C) Representative images and (D) quantitative analysis of the expression of FAO-related proteins using Western blot. (E) Cellular MDA concentrations among the groups. (F) Representative images and (G) quantitative analysis of the expression of oxidative stress-related proteins (NRFS and SOD2) and inflammation-related protein (NFκB) using Western blot. n = 3 or 4 each group. Two-way ANOVA with Bonferroni post-hoc test was used to compare data among groups. Data are expressed as mean ± SEM. * p < 0.05, ** p < 0.01. CC, Compound C; LCA, l -carnitine; PA, palmitate; FAO, fatty acids oxidation; MDA, molondialdehyde; NRF2, nuclear erythroid 2 p45-related factor 2; AMPK, AMP-activated protein kinase; CD36, FAT; CPT1B, carnitine palmitoyltransferase-1B; p-, phoso-; PGC1α, peroxisome proliferator-activated receptor γ coactivator1α; NRF2, Nuclear factor erythroid 2-related factor 2; NFκB, the nuclear factor kappa B; SOD2, manganese superoxide dismutase, superoxide dismutase 2.

Article Snippet: The membranes were blocked with 5% skim milk and incubated with antibodies against total-AMPK (1:1,000; Cell Signaling Technology/CST, Massachusetts, United States), CD36 (1:1,000; Abcam), collagen I (1:1,000; Abcam), collagen III (1:1,000; Abcam), connexin-43 (1:200; Invitrogen), CPT1B (1:1,000; Proteintech), GLUT4 (1:500; Proteintech), NFκB (1:1,000; Proteintech), NRF2 (1:1,000; Proteintech), phoso-AMPK (Thr172; 1:1,000; CST), phoso-Akt (Ser473; 1:1,000; CST), phoso-NFκB (1:1,000; CST), PGC1α (1:1,000; Proteintech), SOD2 (1:1,000; Proteintech), TGF-β (1:1,000; Abcam), α-SMA (1:1,000; CST), β-actin (1:5,000; Proteintech).

Techniques: Inhibition, Cell Isolation, Control, Expressing, Western Blot

Journal: Redox Biology

Article Title: The loss of pancreatic islet NADPH oxidase (NOX)2 improves islet transplantation

doi: 10.1016/j.redox.2022.102419

Figure Lengend Snippet: The loss of NOX2 increases the expression of insulin and antioxidative proteins in hypoxic islets. ( A ) Representative Western blot analysis of (from top to bottom) Nrf2, β-actin, HO-1, SOD2, SOD1 and insulin from whole cell extracts of isolated hypoxic WT and Nox2 −/− islets. ( B ) Quantitative analysis of insulin expression (Fold change) (n = 5 each). Mean ± SEM. *P < 0.05 vs. WT. ( C ) Quantitative analysis of SOD1 expression (Fold change) (n = 3 each). Mean ± SEM. *P < 0.05 vs. WT. ( D ) Quantitative analysis of SOD2 expression (Fold change) (n = 3 each). Mean ± SEM. *P < 0.05 vs. WT. ( E ) Quantitative analysis of HO-1 expression (Fold change) (n = 5 each). Mean ± SEM. *P < 0.05 vs. WT. ( F ) Quantitative analysis of Nrf2 expression (Fold change) (n = 5 each). Mean ± SEM. *P < 0.05 vs. WT.

Article Snippet: The anti-SOD1 (TA321133) and anti-SOD2 (TA321189) antibodies were purchased from OriGene (Rockville, USA).

Techniques: Expressing, Western Blot, Isolation