Abstract 532: Periadventitial Adenoviral Ang-1/2 Gene Transfer Interferes with Atherosclerotic Plaque Progression and Angiogenesis in the Carotid Artery of LDLr ^-/- ApoB100/100 Mice

Accumulation of macrophages, red blood cells and lipids in atherosclerotic plaques is associated with plaque rupture. It has been shown that macrophages and erythrocytes can enter the plaque also via plaque microvasculature originating from the adventitia. Furthermore, it has been reported that in advanced or ruptured human coronary lesions the microvessel density is increased and these microvessels are leaky. The leaky phenotype is characterized by poor pericyte coverage and dysfunctional inter-endothelial junctions. As angiopoietins are involved in angiogenic growth and maturation, we hypothesized that periadventitial Ang-1 or Ang-2 gene transfer would reduce atherogenesis and affect microvasculature in mouse carotid artery. To study this, atherosclerosis was induced by placing a perivascular silastic collar around the carotid artery of LDLr -/- ApoB100/100 mice fed a high cholesterol (0,15 %) diet for 3 weeks. Simultaneous to surgery adenoviral gene transfer (5μl, 5 x 107 pfu/ml) of either Ang-1, Ang-2 or LacZ as a control was applied to the carotid artery (20 mice per group). After 1 or 5 weeks follow-up on high cholesterol diet mice were sacrificed. Plaque size and microvessel density were analyzed with morphometry software (Leica Qwin) using cross-sections stained with hematoxylin and eosin or CD31 respectively. Microvessel ultrastructure was studied using electron microscopy. Ang-2 gene transfer led to significantly smaller plaque area and plaque volume 5 weeks after collar placement compared to LacZ group (2,7 fold, p<0,001 and 3,9 fold, p<0,01 respectively). Also the microvessel density in intima, media and adventitia was lower in the Ang-2 group compared to LacZ (4,5 fold, p<0,05; 7,5 fold p<0,05 and 11,8 fold p<0,05 respectively). Ang-1 gene transfer led to a non-significant trend towards smaller lesions and lower microvessel density compared to LacZ gene transfer. Plaque phenotype was quite fibrous in all groups at 5 weeks, although lesions in Ang-2 group were most human atheroma-like. Electron microscopy showed intact pericyte coverage and endothelial junctions of microvessels in all groups. Endothelial cell morphology after Ang-1 gene transfer showed less filopodia and cytoplasmic vacuoles compared to Ang-2 and LacZ, suggestive of a quiescent phenotype. In conclusion: Ang-2 over expression in mouse carotid arteries led to smaller plaque size and lower microvessel density, suggesting the involvement of angiopoietin signaling in angiogenesis of the atherosclerotic vessel wall.