Glyceraldehyde-derived pyridinium (GLAP) evokes oxidative stress and inflammatory and thrombogenic reactions in endothelial cells via the interaction with RAGE

Abstract

Background: We have previously shown that serum levels of glyceraldehyde-derived advanced glycation end products (Gly-AGEs) are elevated under oxidative stress and/or diabetic conditions and associated with insulin resistance, endothelial dysfunction and vascular inflammation in humans. Further, Gly-AGEs not only evoke oxidative and inflammatory reactions in endothelial cells (ECs) through the interaction with a receptor for AGEs (RAGE), but also mimic vasopermeability effects of AGE-rich serum purified from diabetic patients on hemodialysis. These observations suggest that Gly-AGE-RAGE system might be a therapeutic target for vascular complications in diabetes. However, since incubation of glyceraldehyde with proteins will generate a large number of structurally distinct AGEs, it remains unclear what type of AGE structures could mediate the deleterious effects of Gly-AGEs on ECs. Aims and Methods: Therefore, in this study, we examined (1) whether glyceraldehyde-derived pyridinium (GLAP), one of the Gly-AGEs generated by the incubation of lysine with glyceraldehyde, elicited reactive oxygen species (ROS) generation and inflammatory and thrombogenic gene expression in human umbilical vein ECs (HUVECs) via the interaction with RAGE and (2) if DNA aptamers raised against Gly-AGEs or GLAP (AGE-aptamer or GLAP-aptamer) inhibited the binding of GLAP to RAGE and subsequently suppressed the harmful effects of GLAP on HUVECs. Results: GLAP stimulated ROS generation in a bell-shaped manner; GLAP at 10 μg/ml increased ROS generation in HUVECs by 40%, which was blocked by the treatment with RAGE-antibody (RAGE-Ab). Ten μg/ml GLAP significantly up-regulated mRNA levels of RAGE, monocyte chemoattractant protein-1, intercellular adhesion molecule-1, vascular cell adhesion molecule-1 and plasminogen activator inhibitor-1 in HUVECs, which were also suppressed by RAGE-Ab. AGE-aptamer or GLAP-aptamer significantly blocked these deleterious effects of GLAP on HUVECs. Moreover, quartz crystal microbalance analyses revealed that GLAP actually bound to RAGE and that AGE-aptamer or GLAP-aptamer inhibited the binding of GLAP to RAGE. Conclusions: The present study suggests that GLAP might be a main glyceraldehyde-related AGE structure in Gly-AGEs that bound to RAGE and subsequently elicited ROS generation and inflammatory and thrombogenic reactions in HUVECs. Blockade of the GLAP-RAGE interaction by AGE-aptamer or GLAP-aptamer might be a novel therapeutic strategy for preventing vascular injury in diabetes.