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 17beta-estradiol through Sp-1 in human vascular endothelial cells.
The binding of advanced glycation end products (AGE) to the receptor for AGE (RAGE) is known to deteriorate various cell functions and is implicated in the pathogenesis of diabetic vascular complications. Here we show that AGE, tumor necrosis factor-alpha (TNF-alpha), and 17beta-estradiol (E(2)) up-regulated RAGE mRNA and protein levels in human microvascular endothelial cells and ECV304 cells, with the mRNA stability being essentially invariant. Transient transfection experiments with human RAGE promoter-luciferase chimeras 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 an nuclear factor-kappaB (NF-kappaB) site at -671 or of Sp-1 sites at -189 and -172 residing in those regions resulted in an abrogation of the AGE/TNF-alpha- or E(2)-mediated transcriptional activation. Electrophoretic mobility shift assays revealed that ECV304 cell nuclear extracts contained factors which retarded the NF-kappaB and Sp-1 elements, and that the DNA-protein complexes were supershifted by anti-p65/p50 NF-kappaB and anti-Sp-1/estrogen receptor alpha antibodies, respectively. These results suggest that AGE, TNF-alpha, and E(2) can activate the RAGE gene through NF-kappaB and Sp-1, causing enhanced AGE-RAGE interactions, which would lead to an exacerbation of diabetic microvasculopathy.
Tanaka
N
N
Department of Biochemistry and the Department of Ophthalmology, Kanazawa University School of Medicine, Japan.
Yonekura
H
H
Yamagishi
S
S
Fujimori
H
H
Yamamoto
Y
Y
Yamamoto
H
H
D015153:Blotting, Western; D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism; D002478:Cells, Cultured; D004305:Dose-Response Relationship, Drug; D004730:Endothelium, Vascular / Q000378:metabolism; D004926:Escherichia coli / Q000378:metabolism; D004958:Estradiol / Q000378:metabolism*; D017127:Glycation End Products, Advanced / Q000378:metabolism*; D006801:Humans; D008833:Microcirculation / Q000378:metabolism; D016297:Mutagenesis, Site-Directed; D016328:NF-kappa B / Q000378:metabolism*; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D012333:RNA, Messenger / Q000378:metabolism; D000067759:Receptor for Advanced Glycation End Products; D011971:Receptors, Immunologic / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D016329:Sp1 Transcription Factor / Q000378:metabolism*; D013997:Time Factors; D014158:Transcription, Genetic; D015533:Transcriptional Activation; D014162:Transfection; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism*; D015854:Up-Regulation
D017127:Glycation End Products, Advanced; D016328:NF-kappa B; D012333:RNA, Messenger; D000067759:Receptor for Advanced Glycation End Products; D011971:Receptors, Immunologic; D011993:Recombinant Fusion Proteins; D016329:Sp1 Transcription Factor; D014409:Tumor Necrosis Factor-alpha; D004958:Estradiol