Figure 9. Effects of the knockdown of AQP8 gene expression with siRNA (siAQP8; 10 nM) on the expression and secretion of VEGF from cultured RPE cells. As negative control, scrambled nontargeted siRNA (siNon; 10 nM) was used. siRNA transfection was performed 24 h before the addition of 100 mM NaCl or CoCl₂ (150 µM), and the culture in a 0.2% O₂ atmosphere, respectively. A: Effects of siAQP8 and siNon on the level of VEGFA mRNA in cells cultured for 6 h in a high-NaCl medium and in the presence of CoCl₂, respectively. The mRNA level is shown as folds of unstimulated control. B: Effects of siAQP8 and siNon on the secretion of VEGF. The level of the VEGF-A protein was determined with enzyme-linked immunosorbent assay (ELISA) in the media of cells cultured for 24 h in the presence of high NaCl and CoCl₂, and in a 0.2% O₂ atmosphere, respectively. The protein level is expressed as percent of unstimulated control (100%). Each bar represents mean ± standard error of the mean (SEM) obtained in four to nine independent experiments using cell lines from different donors. Significant difference vs. unstimulated control: *p<0.05. Significant difference vs. siNon and vs. NaCl, CoCl₂, and low-O₂ controls, respectively: ●p<0.05.