Figure 4. Effect of curcumin on intracellular reactive oxygen species (ROS) levels. Retinal pigment epithelium (RPE) cells were separately pretreated with 15 µM curcumin and 10 µM CoPP known as heme oxygenase-1 (HO-1) stimulator for 3 h. About 10 µM ZnPP, a known HO-1 inhibitor, was added to cells preprocessed for 1 h after 2 h pretreatment with 15 µM curcumin. Cells were then treated with 1000 µM H₂O₂ for 3 h. Intracellular ROS levels in RPE cells were measured with flow cytometry (see Materials and Methods). In contrast, ROS levels were increased (shift in relative fluorescence intensity) in cells cotreated with curcumin and ZnPP. A: (a) dimethyl sulfoxide (DMSO), (b) DMSO + H₂O₂, (c) curcumin (15 µM) + H₂O₂, (d) curcumin (15 µM) + H₂O₂ +ZnPP (10 µM), (e) CoPP (10 µM) + H₂O₂. B: Columns, percentage of DCF-CA-stained cells for indicated conditions. Each bar represents the mean±SD of three independent experiments (**p<0.01, ***p<0.001)