Figure 2. High extracellular osmolality activates human RPE cells in vitro and stimulates gene expression of angiogenic factors. A: Western blot analysis shows that the addition of 100 mM salt (NaCl) to the culture medium for 20 min induces increased phosphorylation levels of p38 mitogen-activated protein kinase (MAPK) and extracellular signal-regulated kinase (ERK) 1/2 proteins. Phosphorylation of p38 MAPK and ERK1/2 is not stimulated by chemical hypoxia (induced by addition of 150 µM CoCl₂) for 20 min. Amounts of total proteins are shown above; amounts of phosphorylated proteins are shown below. B, C: Relative mRNA levels of growth factors in cells cultured in media with high osmolality for 2, 6, and 24 h. High osmolality was induced by the addition of 100 mM sucrose (B) and 100 mM salt (C), respectively, to the culture medium. EGF = epidermal growth factor; HGF = hepatocyte growth factor. *p<0.05 vs. isoosmotic control. Modified after Hollborn et al. [57,138] and Veltmann et al. [139].