Figure 3.
Interference with GCS augments intracellular ROS production and improves chemotherapy-induced cell death. Cells were exposed to doxorubicin (DOXO, 8.6 µM) ±2 h pretreatment with the GCS inhibitor PDMP (10 µM) or the catalase inhibitor 3-amino-1,2,4-triazole (3AT, 1 mM). Alternatively, cells were transfected with 200 nM siRNA directed against GCS (siGCS) or non-targeted siRNA (siSCR), 48 hours prior to treatment. The production of intracellular ROS was examined after 90 min treatment in human SH-SY5Y neuroblastoma, U-87 MG glioblastoma or LN-18 glioblastoma cells, using the redox-sensitive indicator H2DCFDA, while cellular viability was determined after 48 hour treatment by XTT assay. Fluorescence, corresponding to ROS, was normalized to the average fluorescence of the control (relative DCF-fluorescence). (A) ROS in SH-SY5Y cells. (B) ROS in U-87 MG cells. (C) ROS in LN-18 cells. (D) Viability of SH-SY5Y cells. (E) Viability of U-87 MG cells. (F) Viability of LN-18 cells. Data represent the mean ± SEM of four independent experiments; *p < 0.05 compared to control (CON), **p < 0.05 compared to DOXO, ***p < 0.05 compared to DOXO + PDMP, as determined by 1-way ANOVA.