Figure 1. Effect of hydrogen peroxide on the activation of H-Ras and its signaling pathway

Bovine retinal endothelial cells were incubated with 250 μM H₂O₂ for 1 h. At the end of the incubation, the cells were rinsed with DMEM, and incubated in 5 mM glucose and 20 mM glucose media for 96 h. The activation of H-Ras was determined by Raf-1 binding assay, Raf-1 by measuring the expression of Raf-1 by Western blot, and that of MAP kinase by quantifying the expression of phospho-p38 of MAP kinase. Each sample was analyzed in duplicate, and the Western blots presented are representative of at least 3-4 experiments.