Fig. 4. Cyclosporin A (CsA) enhances reactive oxygen species (ROS) production induced by H2O2 in human kidney proximal tubule epithelial cells without altering expression level of antioxidant enzymes. HK-2 cells were treated with either 10 nM CsA or vehicle for 60 minutes and then exposed to 1 mM H2O2 or control for 60 minutes (n=3 experiments per condition). (A) ROS production was measured using oxidative sensitive dye 2′,7′-dichlorodihydrofluorescein diacetate. (B) Expression levels of manganese superoxide dismutase (MnSOD; 25 kDa), copper/zinc superoxide dismutase (CuZnSOD; 23 kDa), and catalase (64 kDa) were measured by western blot analysis. Antibody of β-actin (43 kDa) as a loading control was used for normalization. (C–E) Intensities of MnSOD, CuZnSOD, and catalase protein bands were quantified using the AzureSpot software. N.S., not significant. *P<0.05 vs. control, #P<0.05 vs. vehicle.
