Journal: Experimental neurology

Article Title: KU-596 Decreases Mitochondrial Superoxide and Improves Bioenergetics Following Downregulation of Manganese Superoxide Dismutase in Diabetic Sensory Neurons

doi: 10.1016/j.expneurol.2018.12.006

Figure Lengend Snippet: A) MnSOD expression in sensory neurons after infection with MnSOD shRNA adenovirus. (B & C) EPR assessment of the effect of MnSOD downregulation and KU-596 treatment on the superoxide signal in (B) non-diabetic and (C) diabetic neurons from WT mice. D) Quantification of the area under the curve of the first superoxide signal peak, normalized to total protein and expressed as the fold of the non-diabetic plus scrambled shRNA control. *, p < 0.05 versus non-diabetic plus scrambled shRNA; **, p < 0.05 versus diabetic neurons plus scrambled shRNA; ^, p < 0.05, vs diabetic neurons plus MnSOD shRNA. Results are from two separate experiments in which 3-4 wells of neurons per group were obtained from 6 non-diabetic or diabetic mice.

Article Snippet: Recombinant adenoviruses expressing shRNA targeting mouse MnSOD (Ad-GFP-U6-mSOD2-shRNA, #shADV-272843) or scrambled shRNA with GFP (Ad-GFP-U6-scrambled-shRNA, #1122) were purchased from Vector BioLabs (Malvern, PA).

Techniques: Expressing, Infection, shRNA

Journal: Experimental neurology

Article Title: KU-596 Decreases Mitochondrial Superoxide and Improves Bioenergetics Following Downregulation of Manganese Superoxide Dismutase in Diabetic Sensory Neurons

doi: 10.1016/j.expneurol.2018.12.006

Figure Lengend Snippet: A) Quantification of MRC in diabetic WT diabetic neurons in the presence and absence of KU-596 following MnSOD downregulation. *, p < 0.05 versus non-diabetic neurons treated with scrambled shRNA; ^, p < 0.05, versus diabetic neurons treated with scrambled shRNA; B) Quantification of MRC in diabetic neurons in the presence and absence of mito-TEMPO-H and KU-596 following MnSOD downregulation. *, p < 0.05, versus non-diabetic neurons treated with scrambled shRNA. ^, p < 0.05, versus diabetic neurons treated with scrambled shRNA; #, p < 0.05, versus diabetic neurons treated with MnSOD shRNA. For each figure, the results are from 4 - 5 wells of neurons per group obtained from 6 non-diabetic or diabetic mice.

Article Snippet: Recombinant adenoviruses expressing shRNA targeting mouse MnSOD (Ad-GFP-U6-mSOD2-shRNA, #shADV-272843) or scrambled shRNA with GFP (Ad-GFP-U6-scrambled-shRNA, #1122) were purchased from Vector BioLabs (Malvern, PA).

Techniques: shRNA

Journal: Scientific Reports

Article Title: Aging disrupts cell subpopulation dynamics and diminishes the function of mesenchymal stem cells

doi: 10.1038/srep07144

Figure Lengend Snippet: (A) Hierarchical clustering of cells from young (left) and aged (right) mice. Gene expression is presented as fold change from median on a color scale from yellow (high expression, 32-fold above median) to blue (low expression, 32-fold below median). (B) Whisker plots presenting raw qPCR cycle threshold values for each gene across all young and aged ASCs. Individual dots represent single gene/cell qPCR reactions, with increased cycle threshold values corresponding to decreased mRNA content. Cycle threshold values of 40 were assigned to all reactions that failed to achieve detectable levels of amplification within 40 qPCR cycles. (C) Median-centered Gaussian fit curves of selected genes relating to cell stemness and vasculogenesis displaying grossly differential expression profiles between young and aged cells. Non-parametric two-sample Kolmogorov-Smirnov analysis confirmed the differential expression of the anti-oxidative enzyme Sod-2 in aged versus young ASCs (p < 0.01). The left bar for each panel represents the fraction of qPCR reactions that failed to amplify in each group. (D,E) Partitional clustering of young and aged cells based on the expression patterns of all 71 genes (k = 2). (F) Pie charts representing the fraction of ASCs comprising each cluster (Young [red], Aged [green]), with selected cluster 1-defining genes listed below.

Article Snippet: Real-time qPCR reactions were performed using Taqman gene expression assays (Applied Biosystems, Foster City, CA) for murine Angpt1 (Angiopoietin 1, Mm00456503_m1), Vegfa (Vascular endothelial growth factor-A, Mm01281447_m1), Sod3 (Superoxide dismutase 3, Mm01213380_s1), Sod2 (Superoxide dismutase 2, Mm00449726_m1), Angpt2 (Angiopoitin 1, Mm00545822), Fgf2 (Fibroblast growth factor 2, Mm00433287_m1) , Fgfr2 (Fibroblast growth factor receptor 2, Mm01269930_m1), Pdgfa (Platelet derived growth factor-A, Mm01205760_m1) , and Pdfgra (Platelet derived growth factor receptor-A, Mm01205760_m1) using a Prism 7900HT Sequence Detection System (Applied Biosystems).

Techniques: Gene Expression, Expressing, Whisker Assay, Amplification, Quantitative Proteomics

Journal: Neurotoxicity Research

Article Title: Prolonged Exposure to Silver Nanoparticles Results in Oxidative Stress in Cerebral Myelin

doi: 10.1007/s12640-018-9977-0

Figure Lengend Snippet: Protein expression of SOD1 and SOD2 in myelin fractions isolated from brain of control (saline-treated) and silver-exposed rats. Representative immunoblots and the graphs illustrating the results of densitometric measurements, calculated against β-actin as an internal standard, of five different immunoblots performed using four distinct animals. Bars represent means ± SD expressed as a percentage of control; * p < 0.05 or ** p < 0.01 vs. control; # p < 0.05 vs. Ag citrate

Article Snippet: The rat superoxide dismutase specific primers for Sod1 (Rn00566938_m1) and Sod2 (Rn00690588_m1) were obtained from Life Technologies.

Techniques: Expressing, Isolation, Control, Saline, Western Blot

Journal: Neurotoxicity Research

Article Title: Prolonged Exposure to Silver Nanoparticles Results in Oxidative Stress in Cerebral Myelin

doi: 10.1007/s12640-018-9977-0

Figure Lengend Snippet: Expression of mRNA of superoxide dismutases ( Sod1 and Sod2 ) in myelin isolated from control (saline-treated) and silver-exposed rat brains. The mRNA levels were determined by quantitative real-time PCR and normalized against ACTB as a reference gene. Graphs indicate the results expressed as a percentage of control. The values represent the means ± SD from four distinct brain samples, each performed in duplicate; * p < 0.05 vs. control

Article Snippet: The rat superoxide dismutase specific primers for Sod1 (Rn00566938_m1) and Sod2 (Rn00690588_m1) were obtained from Life Technologies.

Techniques: Expressing, Isolation, Control, Saline, Real-time Polymerase Chain Reaction

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: Journal of cell science

Article Title: A new activating role for CO in cardiac mitochondrial biogenesis.

doi: 10.1242/jcs.03318

Figure Lengend Snippet: Fig. 7. Schematic of the mitochondrial respiratory chain showing the effect of CO binding (x) at the terminal oxidase on hydrogen peroxide (H2O2) production. CO slows the rate of electron transport (solid arrows), enabling electrons to accumulate, including at complex III. Complex III and the proton motive Q cycle have been expanded to show sites of ubi-semiquinone (Q·-) formation from which electrons can be donated to produce superoxide (·O2-) (dotted arrows). Complex III O2 availability is also increased by CO, which promotes ·O2-production and its conversion to H2O2 by SOD2 (MnSOD). SOD2 induction may increase the extra-mitochondrial H2O2 leak rate. Cyt, cytochrome; ISP, iron sulfur protein; Q, oxidized coenzyme; QH2, reduced coenzyme Q. The details of the Q cycle, including the transmembrane proton flux, have been omitted for clarity.

Article Snippet: Membranes were incubated with validated polyclonal rabbit Abs against mouse PGC-1 , NRF1, TFAM, and Pol , or Akt, pAkt, p38, p-p38 and SOD2 (Santa Cruz Biotechnology, Santa Cruz, CA), or anti-tubulin or -actin (1:1000; Sigma).

Techniques: Binding Assay