The receptor for advanced glycation end products is induced by the glycation products themselves and tumor necrosis factor-alpha through nuclear Factor-kappa B, and by 17 beta-estradiol through Sp-1 in human vascular endothelial cells

The binding of advanced glycation end products (AGE) to the receptor for AG E (RAGE) is known to deteriorate various cell functions and is implicated i n the pathogenesis of diabetic vascular complications. Here we show that AG E, tumor necrosis factor-alpha (TNF-alpha), and 17 beta-estradiol (E-2) up- regulated RAGE mRNA and protein levels in human microvascular endothelial c ells and ECV304 cells, with the mRNA stability being essentially invariant. Transient transfection experiments with human RAGE promoter-luciferase chi meras revealed that the region from nucleotide number -751 to -629 and the region from -239 to -89 in the RAGE 5'-flanking sequence exhibited the AGE/ TNF-alpha and E-2 responsiveness, respectively. Site-directed mutation of a n nuclear factor-kappa B (NF-kappa B) site at -671 or of Sp-1 sites at -189 and -172 residing in those regions resulted in an abrogation of the AGE/TN F-alpha- or E-2-mediated transcriptional activation. Electrophoretic mobili ty shift assays revealed that ECV304 cell nuclear extracts contained factor s which retarded the NF-kappa B and Sp-1 elements, and that the DNA-protein complexes were supershifted by anti-p65/p50 NF-kappa B and anti-Sp-1/estro gen receptor or antibodies, respectively. These results suggest that AGE, T NF-alpha, and E-2 can activate the RAGE gene through NF-kappa B and Sp-1, c ausing enhanced AGE-RAGE interactions, which would lead to an exacerbation of diabetic microvasculopathy.