Figure 1. Osmolarity-dependent production of VEGF in RPE cells. The mRNA expression (A, C, E, F) was determined with real-time RT–PCR analysis and is expressed as a fold of isoosmotic unstimulated control. The level of VEGF-A₁₆₅ protein in the cultured media (B, D) was determined with ELISA and is expressed as a percentage of isoosmotic unstimulated control (100%, corresponding to 169.1±38.9 [B, 6 h], 230.6±55.1 [B, 24 h], and 525.0±90.6 pg/ml VEGF[D], respectively). A. Relative expression level of VEGFA in cells cultured for 2, 6, and 24 h in isoosmotic (- NaCl) and hyperosmotic (+ 100 mM NaCl) media in the absence (- Triam) and presence (+ Triam) of triamcinolone acetonide (50 µM). B. Release of VEGF protein from RPE cells measured under hyperosmotic conditions. The cells were stimulated for 6 and 24 h, respectively, with hyperosmotic medium (+ 100 mM NaCl) in the absence (- Triam) and presence (+ Triam) of triamcinolone acetonide (50 µM). The data are expressed as a percentage of isoosmotic untreated control (100%). C. Effect of hyperosmotic medium (+ 100 mM sucrose) on the expression of VEGFA. D. Effect of hyperosmotic medium (+ 100 mM sucrose) on the release of VEGF protein from RPE cells. E. Dose-dependent effect of high extracellular NaCl on the cellular level of VEGF mRNA. The cells were cultured for 6 h in media that were made hyperosmotic by the addition of 10 to 100 mM NaCl. F. Effect of hypoosmotic medium (60% osmolarity) on the expression of VEGFA. Data are means ± SEM of n=4 (C, D, E), 5 (F), 7 (B), and 10 (A) independent experiments performed in triplicate. Significant difference versus isoosmotic unstimulated control: *p<0.05. Significant difference versus NaCl control: ●p<0.05.