Fig. 7: Trehalose and rapamycin differently counteract docetaxel-induced apoptosis in PC3 cells.

a Mitochondrial localization of cytochrome c was evaluated by IF. Cells were treated with 20 nM docetaxel and/or 100 mM trehalose. After 48 h, the cells were incubated for 30 min with 250 nM MitoTracker Orange, fixed and stained with cytochrome c antibody followed by FITC secondary antibody. Images were acquired with ×63/1.4 objective lens linked to a Coolsnap Es CCD camera (Ropper Scientific-Trenton, NJ, USA). Scale bar 20 μm. b WB shows caspase-9, caspase-3 and PARP and their cleaved active form after 48 h of treatment with 20 nM docetaxel and/or 100 mM trehalose. Each experiment was repeated three times and representative blots are shown. c Cells were treated with 20 nM docetaxel and/or 100 nM rapamycin. The mitochondrial localization of cytochrome c was evaluated by IF analysis as described in a. d The cells were treated with 20 nM docetaxel and/or 100 nM rapamycin for 48 h. The analysis of apoptotic-related proteins was conducted as described in b. e PC3 cells were treated with 20 nM docetaxel, 100 mM trehalose and/or 100 nM rapamycin. After 24 h of the treatment the cells were labelled with Annexin V-FITC and PI. Dot-plots represent flow cytometry analysis of 10,000 events. Experiment was repeated three times and representative plots are shown