Fig 2: H2O2 production and release by cultured KGN cells and effects of exogenous H2O2. (A) NOX4 RT-PCR analysis and Western blot of cultured KGN cells show single bands of 160 bp and 68 kDa, respectively. Controls using RNA (-RT) or H2O instead of cDNA (H2O) were negative. (B) Hydrogen peroxide assay of untreated KGN cells showed increasing H2O2 levels in the supernatant over a time period of 2 h (n = 6). Signal intensities were normalized to start point values. Bars indicate SEM. (C) Exogenously added H2O2 reduced cell viability in a dose dependent manner. Cell counts after treatment of KGN cells with different concentrations of H2O2 for 24 h (n = 2–5 for each concentration) are shown with an interpolated sigmoidal standard curve (r2 = 0.9361). Bars indicate SEM. (D) Images of KGN cells treated with H2O2 (1 mM) for 24 h compared to untreated control cells. Scale bars indicate 200 µm.