Abstract-We examined whether albumin-derived advanced glycosylation end products (AGEs) downregulate the expression of endothelial nitric oxide synthase (NOS). Significant reductions in NOS activity and cGMP levels in bovine aortic endothelial cells were observed when exposed to different concentrations of albumin-derived AGEs. Western and Northern blot analyses showed significant decreases at the protein and transcript levels. Both reductions became evident after 24 hours of exposure. Nuclear run-on assays showed that AGE-BSA did not modify the transcription rate of the NOS III gene; however, AGE-BSA treatment markedly reduced the half-life of NOS III mRNA. In addition, AGE-treated endothelial cells displayed significant reduction on their antiplatelet properties. These results indicate that NOS expression is reduced by AGEs by increasing the rate of mRNA degradation and may be relevant to the impairment of some endothelial functions observed in diabetes and aging. Key Words: advanced glycosylation end products Ⅲ nitric oxide synthase Ⅲ diabetes E ndothelial cell-generated nitric oxide (NO) accounts for the biological activity of endothelium-derived relaxing factor. NO is derived from the guanidino nitrogen atoms(s) of the amino acid L-arginine through a reaction catalyzed by NO synthase. In endothelial cells, NO synthase (NOS III) has been characterized as a constitutively expressed calcium-and calmodulin-dependent enzyme. However, the level of NOS III expression can be altered by a growing number of different stimuli. Tumor necrosis factor-␣ 1 and lipopolysaccharide 2 decrease expression of NOS III. In contrast, estrogens 3 and shear stress 4 can increase NOS III expression. The effect of hypoxia is less clear; some authors have reported either an increase 5 or decrease 6 in NOS III mRNA.Vascular diseases account for the majority of the clinical complications of diabetes mellitus. 7 Although the molecular basis of the mechanisms involved for the development of vascular disease in diabetes mellitus is poorly understood, several lines of evidences demonstrate impairment in endothelium-dependent relaxation of diabetic blood vessels. 8 Reducing sugar such as glucose can react nonenzymatically with the amino groups of proteins to form complex structures called advanced glycosylation end products (AGEs). In normal aging processes and in some pathological conditions such as diabetes, the excessive accumulation of AGEs could lead to tissue dysfunction. 9 Recently, AGEs have been shown to quench NO and, therefore, may play a role in the defective endothelium-dependent vasodilatation in experimental diabetes. 10 To our knowledge, the effect of AGEs on NOS III expression is unknown. We have undertaken the present study to test the hypothesis that AGEs reduce the expression level of NOS III. We show a significant decrease in the amount of NOS III protein and mRNA in AGE-exposed bovine endothelial cells.