Fig. 2

Oxidative stress induces NRF2 activation in CuONPs-treated HUVECs. A Representative FACS results of HUVECs cells stained with DHE. HUVECs cells were pretreated with NAC (10 mM) for 1 h and then treated with 20 μg/ml CuONPs for 12 h. Unstained, non-labeled cells. Control, normal culture media. MFI, mean fluorescence intensity. B Quantification analysis of DHE intensity in A. C Representative confocal images of HUVECs cells treated with NAC (10 mM) for 1 h, followed by treatment with 20 μg/mL CuONPs for 12 h. Scale bar, 20 μm. Nuclei, DAPI. D, E Immunoblotting analysis and quantification of protein levels of HMOX1, GCLM, SLC7A11 and β-Actin (loading control) in HUVECs cells treated with NAC (10 mM) for 1 h and then CuONPs (20 μg/mL) for 6 h or 9 h. F, G 7-AAD fluorescence intensity analysis of HUVECs cells treated with NAC and CuONPs for 12 h. All data are representative of three independent experiments. Statistical significance was evaluated using one-way ANOVA followed by a Tukey multiple comparison test. The values are expressed in mean ± S.D. “***”, “P ≤ 0.001”