Journal: Oncotarget

Article Title: A novel pan-Nox inhibitor, APX-115, protects kidney injury in streptozotocin-induced diabetic mice: possible role of peroxisomal and mitochondrial biogenesis

doi: 10.18632/oncotarget.18540

Figure Lengend Snippet: Diabetes was induced in mice by intraperitoneal injection of STZ (50 mg/kg). Then APX-115 (60 mg/kg/day) or losartan (1.5 mg/kg/day) was administered orally for 12 weeks to diabetic mice. After 12 weeks, urine and blood samples were collected for analysis of ( A ) urinary albumin excretion, ( B ) albumin/creatinine ratio, ( C ) creatinine clearance rate, and ( D ) plasma cystatin C. ( E ) Kidneys were fixed in paraffin and cut into 3 μm sections that were subsequently stained with PAS reagent. Scale bar: 10 μm; original magnification: 630×. After PAS staining, ( F ) glomerular volume, ( G ) mesangial area, and ( H ) tuft area were analyzed using Image-Pro Plus 4.5.1. DM, STZ-induced diabetic mice. Data are presented as means ± SE of 10–12 mice/group; * p < 0.05 vs. control, † p < 0.05 vs. DM.

Article Snippet: A murine mesangial cell line (MES-13, cloned from mice transgenic for the early region of SV-40 virus, passage 25) was obtained from American Type Culture Collection (ATCC, Rockville, MD, USA).

Techniques: Injection, Clinical Proteomics, Staining, Control

Journal: Oncotarget

Article Title: A novel pan-Nox inhibitor, APX-115, protects kidney injury in streptozotocin-induced diabetic mice: possible role of peroxisomal and mitochondrial biogenesis

doi: 10.18632/oncotarget.18540

Figure Lengend Snippet: ( A ) Plasma LPO, ( B ) urinary LPO, ( C ) kidney tissue LPO, ( D ) Nox1, ( E ) Nox2, and ( F ) Nox4 mRNA expression levels in kidneys were measured using real-time PCR. ( G and H ) Frozen kidney sections were stained with DHE at 5 µM (original magnification: 400×; scale bar: 20 μm). (A–H) Data are presented as means ± SE of 10–12 mice/group; * p < 0.05 vs. control, † p < 0.05 vs. DM. ( I ) Mesangial cells were incubated with or without APX-115 (1 µM) for 30 min and stimulated with or without 30 mM high glucose (HG) for 24 h followed by angII for 30 min. After that cells were incubated with 10 µM DCF-DA for 10 min and the fluorescence intensity was measured with a Zeiss vision system. Data are presented as means ± SE of at least 2 independent experiments; * p < 0.05 vs. control, † p < 0.05 vs. angII or angII+HG in DMSO.

Article Snippet: A murine mesangial cell line (MES-13, cloned from mice transgenic for the early region of SV-40 virus, passage 25) was obtained from American Type Culture Collection (ATCC, Rockville, MD, USA).

Techniques: Clinical Proteomics, Expressing, Real-time Polymerase Chain Reaction, Staining, Control, Incubation, Fluorescence

Journal:

Article Title: Poisoning of human DNA topoisomerase I by ecteinascidin 743, an anticancer drug that selectively alkylates DNA in the minor groove

doi:

Figure Lengend Snippet: Et743 induces top1-mediated DNA single-strand breaks. SV40 DNA was reacted with top1 in the absence or presence of drugs, as indicated. Reactions were stopped with 0.5% SDS followed by proteinase K incubation. Samples were run in 1% agarose gels containing ethidium bromide. Percent nicked DNA was computed after FluorImager analysis (Molecular Dynamics) by using the imagequant software and is indicated below each lane. (A) top1-mediated nicking assay by using supercoiled negative SV40 DNA. Lane 1, DNA alone; lane 2, + top1; and lanes 3–6, + top1 and the indicated concentrations of Et743 or CPT. (B) Et743 induces more top1-mediated DNA nicking in supercoiled than in linear SV40 DNA. Lanes 1 and 5, DNA alone; lanes 2 and 6, DNA + top1; lanes 3 and 7, DNA + top1 + Et743 (0.1 μM); lanes 4 and 8, Et743 without top1. (C) Et743 does not induce detectable top2 cleavage complexes. Lane 1, DNA alone; lane 2, + top2; and lanes 3–4, + top2 and the indicated drug concentrations.

Article Snippet: Each reaction (10 μl final volume) in 10 mM Tris⋅HCl, pH 7.5/50 mM KCl/5 mM MgCl 2 /0.1 mM EDTA/1 mM ATP/15 μg/ml BSA contained 0.3 μg supercoiled simian virus 40 (SV40) DNA (GIBCO/BRL).

Techniques: Incubation, Software

Journal:

Article Title: Poisoning of human DNA topoisomerase I by ecteinascidin 743, an anticancer drug that selectively alkylates DNA in the minor groove

doi:

Figure Lengend Snippet: DNA alkylation by Et743 induces salt-reversible top1-mediated DNA breaks at different sites from CPT. (A) Differences in sequence selectivity of top1-mediated DNA breaks for Et743 and CPT. The long BanI-HpaII fragments of SV40 DNA 3′-end-labeled at the BanI Site was used. Lane 1, formic acid (FA); lane 2, DNA alone; lane 3, + top1 without drug; lane 4, top1 + Et743 (10 μM); lane 5, top1 + CPT (10 μM). (B) Salt reversibility of the top1-mediated DNA breaks induced by Et743. Drug-induced DNA cleavage was reversed by adding 0.35 M NaCl (final concentration) (at time 0 corresponding to a 30-min incubation of top1 with Et743 or CPT; lanes 3 and 8, respectively) and by further incubation at 25°C for the indicated times. (C) Et743-DNA adducts induce the top1 cleavage complexes. 3′-end-labeled PvuII/HindIII fragment of pSK(−) phagemid DNA was used for these reactions. In lanes 2–9, reactions were performed in two consecutive steps. First, the DNA was treated overnight (O/N ≈16 hr) with or without drug, as indicated above pairs of lanes, then DNA was ethanol precipitated to remove free drug. Secondly, the DNA was reacted ± top1 in the absence of added drug (lanes 2–10) or in the presence of 10 μM CPT (lane 11). Lane 1, FA-sequencing lane.

Article Snippet: Each reaction (10 μl final volume) in 10 mM Tris⋅HCl, pH 7.5/50 mM KCl/5 mM MgCl 2 /0.1 mM EDTA/1 mM ATP/15 μg/ml BSA contained 0.3 μg supercoiled simian virus 40 (SV40) DNA (GIBCO/BRL).

Techniques: Sequencing, Labeling, Concentration Assay, Incubation