Multiple lines of evidence suggest that intraplaque (IP) neovascularization promotes atherosclerotic plaque growth, destabilization, and rupture. However, pharmacological inhibition of IP... Show moreMultiple lines of evidence suggest that intraplaque (IP) neovascularization promotes atherosclerotic plaque growth, destabilization, and rupture. However, pharmacological inhibition of IP neovascularization remains largely unexplored due to the limited number of animal models that develop IP neovessels and the lack of reliable methods for visualizing IP angiogenesis. Here, we applied 3D confocal microscopy with an optimized tissue-clearing process, immunolabeling-enabled three-dimensional imaging of solvent-cleared organs, to visualize IP neovessels in apolipoprotein E-deficient (ApoE(-/-)) mice carrying a heterozygous mutation (C1039+/-) in the fibrillin-1 gene. Unlike regular ApoE(-/-) mice, this mouse model is characterized by the presence of advanced plaques with evident IP neovascularization. Plaques were stained with antibodies against endothelial marker CD31 for 3 days, followed by incubation with fluorescently labeled secondary antibodies. Subsequent tissue clearing with dichloromethane (DCM)/methanol, DCM, and dibenzyl ether allowed easy visualization and 3D reconstruction of the IP vascular network while plaque morphology remained intact. Show less
During my PhD we have investigated different approaches to block intraplaque angiogenesis in atherosclerosis. Intraplaque angiogenesis is a physiological response to the increased oxygen demand in... Show moreDuring my PhD we have investigated different approaches to block intraplaque angiogenesis in atherosclerosis. Intraplaque angiogenesis is a physiological response to the increased oxygen demand in the plaque but also has adverse effects by facilitating intraplaque hemorrhage and influx of inflammatory mediators, resulting in plaque instability and consequent rupture. To study this phenomenon we used in vitro assays as well as the accelerated atherosclerosis vein graft model in ApoE3*Leiden mice, a unique model in which the formed plaque shows characteristics that highly resemble human atherosclerotic lesions, including intraplaque angiogenesis and hemorrhage and a high inflammatory cell content. We focused on different approaches to restore plaque stability via improving intraplaque oxygen levels as well as via blocking different growth factors signaling. Moreover we studied the effects of our treatments on the interaction between angiogenesis and inflammation both in vitro and in vivo. Show less
