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Fig. 5 | Journal of Experimental & Clinical Cancer Research

Fig. 5

From: ROS-p53-cyclophilin-D signaling mediates salinomycin-induced glioma cell necrosis

Fig. 5

ROS is required for salinomycin-induced necrosis in cultured glioma cells. U87MG glioma cells were pre-treated with anti-oxidants NAC (50 μM) or PDTC (25 μM) for 1 h, followed by salinomycin (Sali, 5 μM) stimulation, ROS production was analyzed 3 h after stimulation (a), mitochondrial expression and association of p53 and Cyp-D were tested at 6 h after stimulation (b), JC-10 green fluorescence intensity was examined after 6 h (c); cell necrosis was analyzed similarly 72 h after salinomycin stimulation (d). The effect of NAC (50 μM, 1 h pretreatment) on salinomycin (Sali, 5 μM)-induced viability loss was tested by MTT assay in U251MG and EFC-2 cells (e). Non-transfected control U87MG cells, Cyp-D siRNA-transfected U87MG cells or Cyp-D-over-expressing U87MG cells were treated with salinomycin (Sali, 5 μM) for 3 h, ROS production was analyzed (f). (g) U251 cells (1 × 106) were injected subcutaneously into nude mice as described in Materials and Methods, and treatment was started when the tumors reached 200 mm3. Salinomycin (5.0 mg/kg) and/or CsA (5.0 mg/kg) were administered intraperitoneally once daily for two weeks. Control mice received vehicle only, according to the same schedule. Tumor volume was measured by caliper with the formula: π/6 × width 2× length. N = 10 for each group. (h) The proposed signaling pathway of this study. Experiments were repeated three times in this figure, and similar results were obtained. Error bars indicate SD. * p < 0.05 vs. Sali only group (a, b, d, f and g). # p < 0.05 vs. vehicle group (g)

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