Effect of methylglyoxal on eNOS phosphorylation in EA. hy926 endothelial cells. A and B. Original Western blots (A) and the respective densitometric analysis (B) of the relative abundance of total eNOS protein (white bars; relative to β-actin) and Ser-1177 phospho-eNOS (black bars; relative to β-actin) in EA.hy296 endothelial cells incubated in the absence (-) or in the presence of MG (100 μM) for 8 h alone (+) or with 100 μM BH4, 100 μM NH4, 20 μM sepiapterin or 50 μM L-NAME in the last 4 h of MG treatment or with 24 h pre-treatment with 300 μM Tempol and 50 μM L-NAME (ONOO- suppression). Data are arithmetic means ± SEM (A, representative of three experiments; B, n = 3). * indicates significant difference (p < 0.05) from the absence of MG (ANOVA). # indicates significant difference (p < 0.05) from MG treatment alone (ANOVA).