Advanced glycation end products quench nitric oxide in vitro.
BACKGROUND
Nitric oxide (NO) is known as an important mediator of endothelial function. The aim of this investigation was to evaluate the influence of mediators of retinal pathology - vascular endothelial growth factor (VEGF) and advanced glycation end products (AGEs) - on NO release from choroidal endothelial cells (CEC) and retinal pigment epithelial (RPE) cells to elucidate the complex role of NO.


METHODS
Bovine CEC were stimulated using VEGF (1, 10, and 100 ng/ml), and RPE cells were exposed to interferon-gamma (IFN-gamma 100 U/ml) and lipopolysaccharide (LPS 1 micro g/ml). NO release into the media was assessed by an amperometric NO sensor. The influence of AGEs (10 and 100 micro g/ml) on NO release from CEC and RPE cells was investigated. The competitive NO synthase inhibitor L(omega)-nitro-L-arginine methyl ester (L-NAME 2 nmol) was used to pretreat the cells 2 h before NO measurement.


RESULTS
Unstimulated CEC produced low basal levels of NO in vitro (39.1+/-13.9 nmol), and unstimulated RPE cells produced minimal basal levels of NO (15.7+/-7.0 nmol). Exposure of CEC to VEGF for 30 min resulted in a dose-dependent rise of NO in the medium, which was significantly inhibited by L-NAME. Stimulation of RPE cells with IFN-gamma and LPS resulted in a rise of NO in the bath to 125.9+/-18.5% of basal values. Basal NO release from CEC, and stimulated NO release from RPE cells, was significantly reduced by AGE treatment and L-NAME.


CONCLUSIONS
These data demonstrate that AGEs formed from the nonenzymatic glycation of proteins with reducing sugars quench NO activities in vitro. The results implicate AGEs as important modulators of NO activity and may be relevant to the impairment of endothelial functions observed in diabetes and aging.
Uhlmann
Susann
S
Department of Ophthalmology, University of Leipzig, Liebigstr. 10-14, 04103 Leipzig, Germany. Scos@medizin.uni-leipzig.de
Rezzoug
Karen
K
Friedrichs
Ulrike
U
Hoffmann
Stephan
S
Wiedemann
Peter
P
D000818:Animals; D002417:Cattle; D002478:Cells, Cultured; D002829:Choroid / Q000166:cytology / Q000378:metabolism; D016228:Endothelial Growth Factors / Q000494:pharmacology; D004727:Endothelium / Q000166:cytology / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology; D017127:Glycation End Products, Advanced / Q000494:pharmacology*; D036341:Intercellular Signaling Peptides and Proteins / Q000494:pharmacology; D007371:Interferon-gamma / Q000494:pharmacology; D008070:Lipopolysaccharides / Q000494:pharmacology; D008222:Lymphokines / Q000494:pharmacology; D019331:NG-Nitroarginine Methyl Ester / Q000494:pharmacology; D009569:Nitric Oxide / Q000037:antagonists & inhibitors* / Q000378:metabolism; D010857:Pigment Epithelium of Eye / Q000166:cytology / Q000378:metabolism; D042461:Vascular Endothelial Growth Factor A; D042442:Vascular Endothelial Growth Factors
D016228:Endothelial Growth Factors; D004791:Enzyme Inhibitors; D017127:Glycation End Products, Advanced; D036341:Intercellular Signaling Peptides and Proteins; D008070:Lipopolysaccharides; D008222:Lymphokines; D042461:Vascular Endothelial Growth Factor A; D042442:Vascular Endothelial Growth Factors; D009569:Nitric Oxide; D007371:Interferon-gamma; D019331:NG-Nitroarginine Methyl Ester