Figure 6.

Pre-incubation with the phytonutrients or estradiol is required to reduce H₂O₂-generated ROS. Cells were seeded in 6-well plates (3 × 10⁵ cells/well) and were pre-incubated for 24 h with (a,c) or without (b,d,e) estradiol (10 nM), rosemary extract (5 µM carnosic acid), or tomato extract (10 µM lycopene). Then, H₂O₂ (50 μM) was added together with the treatment compounds, and ROS levels (after 90 min, a–d) or cell number (after 24 h, e) were determined as described in Section 2.5 and Section 2.3, respectively. (a,b) Typical flow cytometric histograms of DCF fluorescence obtained in a representative experiment with (a) or without (b) pre-incubation. (c,d) Averaged geometric means of DCF fluorescence intensities (MFI) with (c) or without (d) pre-incubation. (e) Cell numbers without pre-incubation, 100% of the cell number was 37,858 ± 676 cells/well. Values are the means ± SEM of 2–4 experiments, each performed in duplicate. ### p < 0.001, #### p < 0.0001, significant difference between the vehicle with and without H₂O₂. ****, p < 0.0001, significant difference between the vehicle and other treatments in the presence of H₂O₂.