Article Title: Sensitization of neuroblastoma for vincristine-induced apoptosis by Smac mimetic LCL161 is attended by G2 cell cycle arrest but is independent of NFκB, RIP1 and TNF-α
Figure Lengend Snippet: Smac mimetic LCL161 cooperates with vincristine to induce activation of extrinsic and intrinsic apoptosis and caspase-dependent cell death, G2 cell cycle arrest and reduction of migratory potential in neuroblastoma cell lines Neuroblastoma cell lines were treated with the indicated concentrations of vincristine, LCL161, and 50 μmol/L caspase inhibitors. Expression of cIAP-1, XIAP and β-actin was detected by Western Blot analysis 48 h after treatment initiation (A) (representative images of at least three independent Western Blots are shown). The proportions of apoptotic cells (B) (‡; p≤ 0.05 (NC/VCR-treated vs. NC/VCR/LCL161-treated), * ; p≤ 0.05 (NC/VCR/LCL161-treated vs. caspase inhibitor/VCR/LCL161-treated)), and cells with decreased mitochondrial membrane potential (MMP) (C) were determined by flow cytometry 48 hours after treatment initiation. Activation of caspases-8 (D) and -9 (E) was detected by Caspase-Glo assays 24 h following start of treatment. Flow cytometric cell cycle analysis was performed using propidium iodide DNA staining at 24 h time point (F) . Migrated cells 24 h post treatment start were visualized by fluorescence microscopy (G) and quantified using ImageJ and nucleus counter plugin (H) . Values represent the mean ± SD of three independent experiments. (C-H) * p ≤ 0.05; ** p≤ 0.01 (VCR vs VCR/LCL161).
Article Snippet: Apoptosis was detected by Annexin V-FITC (556419; BD Pharmingen) and propidium iodide (PI) (1 mg/ml in ddH2 O; Invitrogen) staining and flow cytometry.
Techniques: Activation Assay, Expressing, Western Blot, Flow Cytometry, Cytometry, Cell Cycle Assay, Staining, Fluorescence, Microscopy