Journal: Mediators of Inflammation

Article Title: Thiopurines Induce Oxidative Stress in T-Lymphocytes: A Proteomic Approach

doi: 10.1155/2015/434825

Figure Lengend Snippet: Schematic representation of experimental system. (a) Silver stained 2-DE gel (Jurkat wt DMSO), (b) phosphor stained 2-DE gel (Jurkat wt DMSO), and (c) Jurkat wt and kd cells were treated with IC 60 doses of 6-MP, 6-TG, and vehicle (DMSO) for 48 h. Total protein lysates from DMSO, 6-MP, and 6-TG treated Jurkat wt and kd cells were separated by 2-DE, followed by staining with phosphor specific and silver stain. Consistently regulated (circled) spots were identified by Q-TOF MS/MS analysis. Densitometric analysis of Jurkat wt and kd groups (a); DMSO treated, silver stained, and phosphor stained representative gels of Jurkat wt ((b) and (c)) cell lysates.

Article Snippet: The wild-type Jurkat cell line (wt) was obtained from the German Collection of Microorganisms and Cell Cultures (DSMZ), Braunschweig, Germany.

Techniques: Staining, Silver Staining, Tandem Mass Spectroscopy

Journal: Mediators of Inflammation

Article Title: Thiopurines Induce Oxidative Stress in T-Lymphocytes: A Proteomic Approach

doi: 10.1155/2015/434825

Figure Lengend Snippet: ROS assay after 6-MP or 6-TG treatment (a) and differential expression of ROS related SOD1, SOD2, and CAT proteins (b). (a) Cells were treated with vehicle DMSO or IC 60 doses of 6-MP or 6-TG for 48 h. After treatment media were removed and the cells were resuspended in PBS containing 10 μ mol/L DCFDA. Fluorescence intensity was measured after 1 h incubation at 37°C. The error bars represent mean ± SD of four independent experiments (in triplicate format each). * P ≤ 0.05; *** P ≤ 0.0005. For expressional regulation of ROS related proteins, total protein lysates from DMSO or 6-MP or 6-TG treated Jurkat wt and kd cells were separated by 1D gel electrophoresis and immunoblotted with antibody against SOD1, SOD2, and CAT (b). Beta-actin was used as a loading control.

Article Snippet: The wild-type Jurkat cell line (wt) was obtained from the German Collection of Microorganisms and Cell Cultures (DSMZ), Braunschweig, Germany.

Techniques: ROS Assay, Quantitative Proteomics, Fluorescence, Incubation, Nucleic Acid Electrophoresis, Control

Journal: Mediators of Inflammation

Article Title: Thiopurines Induce Oxidative Stress in T-Lymphocytes: A Proteomic Approach

doi: 10.1155/2015/434825

Figure Lengend Snippet: Differential expression of STMN1 as shown by silver stained gels (a) and Western blot analysis (b). (a) STMN1 spot in all six groups: significant downregulation in wt and kd 6-MP treated groups. Graphical representation of spot density (% volume) with mean ± SD of five independent experiments ( * P ≤ 0.05). (b) Total protein lysates from DMSO or 6-MP or 6-TG treated Jurkat wt and kd cells were separated by 1D gel electrophoresis and immunoblotted with antibody against STMN1. Densitometric analyses were performed using LabImage 2.71 software. Beta-actin was used as a loading control. The error bars represent mean ± SD of four independent experiments ( * P ≤ 0.05).

Article Snippet: The wild-type Jurkat cell line (wt) was obtained from the German Collection of Microorganisms and Cell Cultures (DSMZ), Braunschweig, Germany.

Techniques: Quantitative Proteomics, Staining, Western Blot, Nucleic Acid Electrophoresis, Software, Control