Fig. 3.
Effects of antioxidants on cisplatin-induced apoptosis and ROS generation. a, b HFDPC and HaCaT cells were pretreated for 30 min with CAT (7,500 U/ml), MnTBAP (50 µM), or sodium formate (NaFM, 2.5 mM) followed by cisplatin treatment (100 and 25 µM in HFDPC and HaCaT cells, respectively) for 24 h and analyzed for apoptosis by Hoechst 33342 assay. Increasing the dose of CAT or MnTBAP had no significant effect on the apoptotic signals (not shown). c, d HFDPC and HaCaT cells were similarly treated with the antioxidants and cisplatin as described above and analyzed for ROS generation at the maximum response time of 2 h by flow cytometry using H2DCF-DA as a fluorescent probe. Increasing the dose of antioxidants had no further effect on the ROS generation (not shown). e, f HFDPC and HaCaT cells were treated with various concentration of cisplatin (0–250 and 0–50 µM in HFDPC and HaCaT cells, respectively) in the presence or absence of sodium formate (2.5 mM) for 30 min and cellular hROS generation was determined by fluorescence plate reader using HPF as a fluorescent probe as described in “Material and methods” section. Plots are mean ± SD (n = 4). *P < 0.05 versus non-treated control. #P < 0.05 versus cisplatin-treated control