Abstract 705: Blockade of RAGE Signaling in Endothelial Cells Reduces Vascular Inflammation and Atherosclerosis in ApoE Null Mice
We previously demonstrated that homozygous deletion of RAGE in the apoE null background resulted in significant suppression of atherosclerosis. The striking expression of RAGE in endothelial cells (EC) in apoE null aorta led us to test the premise that RAGE signal transduction in EC mediated, at least in part, exacerbation of vascular inflammation and plaque formation in these mice. Transgenic mice expressing cytosolic domain deleted RAGE (dominant negative DN-RAGE) in EC were prepared using the preproendothelin-1 promoter. When bred into apoE null, mean atherosclerotic lesion area at the aortic root of DN-RAGE/apoE null mice at age 14 and 24 weeks was significantly reduced compared to apoE null mice: 19,779 ± 2,261 vs. 68,300 ± 10,523 μm2 (p = 0.012) at 14 weeks; and 61,913 ± 15,653 vs. 221,370 ± 12,856 μm2 (p = 0.0002) at 24 weeks (N ≥ 3/group). Lesion complexity was also reduced in DN-RAGE/apoE null vs. apoE null mice at 14 and 24 weeks of age (p < 0.05). There were no differences in plasma lipids. Consistent with key roles for EC RAGE signaling in vascular inflammation, Western blots of aortic lysates from DN-RAGE/apoE null mice revealed significantly reduced expression of VCAM-1, IL-10, MCP-1, MMP-2 and -9 vs. apoE nulls at 14 weeks (p < 0.001). Aortic EC were isolated from Wild Type (WT) or DN-RAGE expressing mice; upon incubation with S100B (prototypic RAGE ligand; 10 μg/ml), a 5–7 fold increase in Erk1/2 phosphorylation and VCAM-1 expression was observed in WT vs 2-fold in DN-RAGE EC (p < 0.0001), compared to untreated EC. MMP-2 activity was increased 8-fold over baseline in WT vs. 2-fold in DN-RAGE EC (p < 0.0001). Finally, to test these pathways in human vascular cells, aortic EC and smooth muscle cells were transduced with lentivirus expressing full-length RAGE (FL) or DN-RAGE. Upon incubation with S100B, assays performed with HAEC displayed a significant 30% inhibition in permeability in DN-RAGE-expressing EC (p < 0.0001), and a 5-fold suppression of migration of AoSMC expressing DN-RAGE vs. FL-RAGE cells (p < 0.0001). These findings demonstrate that EC RAGE signaling pivotally impacts on vascular inflammation and mechanisms linked to atherogenesis. Strategies to locally modulate RAGE signaling in the vasculature may be key tools in prevention of vascular injury.