Reactive oxygen species accelerate production of vascular endothelial growth factor by advanced glycation end...

Advanced glycation end products (AGEs) are believed to play an important role in the development of angiopathy in diabetes mellitus. Previous reports suggested a correlation between accumulation of AGEs and production of vascular endothelial growth factor (VEGF) in human diabetic retina. However, the mechanisms involved were not revealed. In this study, we investigated the transcriptional regulation of the expression of vascular endothelial growth factor (VEGF) by AGEs, and possible involvement of reactive oxygen species (ROS) in the induction. We employed an AGE of bovine serum albumin (BSA) prepared by an incubation of BSA with D-glucose for 40 weeks and N(epsilon)-(carboxymethyl)lysine (CML), a major AGE. The expression of VEGF was induced by CML-BSA in RAW264.7 mouse macrophage-like cells. CML-BSA stimulated the DNA-binding activity of activator protein-1 (AP-1). Promoter assay showed that the induction of VEGF was dependent on AP-1. The activity of Ras/Raf-1/MEK/ERK1/2 was involved in the CML-BSA-stimulated signaling pathways to activate the AP-1 transcription with a peak at 1 h. AGE-BSA also induced VEGF mediated by AP-1, however, there was a difference of effect between AGE-BSA and CML-BSA in the activation of AP-1. AGE-BSA-stimulated AP-1 activity showed a peak at 5 h, which paralleled the formation of ROS. Reduction of AGE-BSA with NaBH(4) or addition of vitamin E attenuated the AGE-BSA-stimulated signaling pathways leading to the same pattern as for CML-BSA-stimulated signals. These results suggest an important role for AGEs in stimulation of the development of angiogenesis observed in diabetic complications, and that ROS accelerates the AGE-stimulated VEGF expression.