Augmented angiogenesis in adventitia promotes growth of atherosclerotic plaque in apolipoprotein E-deficient mice

Abstract

Objective

Accumulating evidence suggests that exaggerated formation of vasa vasorum (VV) plays an important role in the pathogenesis of atherosclerosis. However, it remains unclear whether augmented angiogenesis in the adventitia could promote hyperlipidemia-induced atherosclerotic lesion formation.

Methods and results

First, we analyzed the time course of VV development in apolipoprotein E-deficient (ApoE−/−) mice. VV proliferation was observed only after atherosclerotic lesion formation. Next, we investigated whether forced perivascular angiogenesis could promote plaque progression. Basic fibroblast growth factor (bFGF) (100 μg/body) incorporated in acid gelatin hydrogel microspheres (AGHM) (bFGF + AGHM group, n = 10), AGHM alone (AGHM group, n = 7), or PBS (control group, n = 8) was administered into the periaortic area of the retroperitoneal space in 10- to 11-week-old male ApoE−/− mice. At 13 weeks after the operation, lesions were significantly larger in the bFGF + AGHM group than in others (bFGF + AGHM: 3.4 ± 0.7 × 10⁴ μm²; AGHM: 0.1 ± 0.1 × 10⁴ μm²; control: 0 μm²; p < 0.0001), which was associated with increased neovascularization in the adventitia. The number of adventitial capillaries correlated with plaque size (r = 0.69, p < 0.0001). In the bFGF + AGHM group, an increase in the number of VV and accumulation of Mac3-positive macrophages were observed prior to atherosclerotic lesion formation.

Conclusions

Our findings demonstrated that local administration of bFGF in the adventitia induced development of VV and accelerated plaque progression in ApoE−/− mice, supporting the notion that VV formation plays a crucial role in the pathogenesis of atherosclerosis.

Introduction

The vasa vasorum (VV) are the microvasculature present in the adventitial layer of the vessel wall, presumably supplying nutrients to the vessel wall of large arteries [1]. VV play a significant role in maintaining vessel integrity. Previous postmortem studies showed that adventitial VV developed in human atherosclerotic coronary arteries [2], [3]. Chronic inflammation of large arteries is associated with proliferation of VV which function to perfuse the vessel wall beyond the limit of diffusion from the luminal side [2]. It has been proposed that VV could function as a conduit supplying inflammatory cells and lipid into atherosclerotic plaques [4]. VV may also cause intraplaque hemorrhage, which contributes to lesion progression and destabilization [3], [4]. Neovascularization was reported to increase in ruptured plaques in the human aorta [5].

Recent advances in imaging techniques, particularly micro CT technique, have enabled us to visualize VV [6]. Micro CT has made it possible to perform quantitative analysis of the number of VV, spatial density, vascular area fraction, and endothelial surface fraction [7]. Recently, Kampschulte et al. investigated the spatio-temporal distribution of vasa vasorum (VV) relative to advanced atherosclerotic lesions of mice using high-resolution nano-CT [7]. The authors convincingly demonstrated that atherosclerotic lesion type is correlated to the number and cross-sectional area of VV. Although these studies suggested that VV formation plays a crucial role in the pathogenesis of atherosclerosis, it has been unclear whether proliferation of adventitial microvessels is a cause or a result of atherosclerotic lesion progression. Precise role of VV in progression and destabilization of atherosclerosis remains to be clarified.

Histological analysis can provide high resolution images of capillary plexuses in close proximity to the vascular wall, particularly at sites where atherosclerosis develops, with information on inflammatory cells, cytokine expression, connective tissues and surrounding tissues [8]. Here, we carefully analyzed the time course of atherosclerotic lesion formation and VV progression in ApoE−/− mice. We also tested the hypothesis that forced angiogenesis in the adventitia could promote atherosclerotic lesion formation in ApoE−/− mice.