Influence of apolipoprotein E, age and aortic site on calcium phosphate induced abdominal aortic aneurysm in mice

Highlights

• ApoE deficiency, but not age, predisposes to AAA induced within the calcium phosphate model.

• AAA induction is related to aortic inflammation, calcification, apoptosis and increased concentrations of LDL/VLDL cholesterol.

• Local application of calcium phosphate also promotes dilation of distant aortic segments.

Abstract

Objective

To assess relevant features of abdominal aortic aneurysms (AAA) induced by calcium phosphate within a mouse model. Specifically we investigated: (1) whether apolipoprotein E deficiency and older age promoted AAA formation, and (2) whether the local application of calcium phosphate affected the size of distant aortic segments.

Methods

AAA was induced by application of calcium phosphate to the infra-renal aortas of 3 and 7 month old male mice. AAA induction was assessed by calculating expansion of the infra-renal aortic diameter over 1–4 weeks. Aortic samples were assessed to quantify calcification, macrophages infiltration, elastic lamellar degradation and apoptosis. Blood pressure was measured by the tail cuff method, and plasma concentrations of total cholesterol, low density lipoprotein and very low density lipoprotein cholesterol, and pro-inflammatory cytokines were measured using commercially available kits. The maximum diameters of the aortic arch, thoracic and supra-renal aorta at sacrifice were measured by morphometry and the mean maximal diameter of these three aortic segments was calculated.

Results

The median expansion of the infra-renal aorta 2 weeks after AAA induction was significantly greater in apolipoprotein E deficient (ApoE^−/−) mice than in age- and gender-matched wild type controls [275.8% (IQR 193.8%–348.5%) versus 94.7% (IQR 47.8%–163.4%), P = 0.02]. The greater aortic expansion in ApoE^−/− mice was associated with aortic calcification, macrophage infiltration, elastic lamellar degradation and apoptosis of cells in the media and adventitia. The plasma low density lipoprotein/very low density lipoprotein cholesterol concentrations 2 weeks after AAA induction were positively correlated with the expansion of the infra-renal aorta induced by calcium phosphate. The median expansion of the infra-renal aorta 2 weeks after AAA induction was similar in 3 and 7 month old wild type mice. The local administration of calcium phosphate was associated with an increase in the mean maximal diameter of distant aortic segments, but not associated with changes in the concentrations of pro-inflammatory markers in either the plasma or the spleen.

Conclusion

This study suggests that apolipoprotein E deficiency, but not age, predisposes to AAA induced within the calcium phosphate model. Increased AAA expansion in ApoE^−/− mice was associated with calcification, macrophage infiltration, elastic lamellar degradation, and cell apoptosis. Local application of calcium phosphate also promoted dilation of distant aortic segments.

Introduction

Abdominal aortic aneurysm (AAA) affects 1–5% men aged >65 years and is an important cause of mortality in the elderly due to aortic rupture and associated cardiovascular events [1]. There is a current deficiency in therapies for AAA and there is a significant interest in identifying drug therapies which can limit progression of AAA [2], [3], [4]. The study of AAA pathogenesis in humans is limited by a number of problems including the difficulty in obtaining aortic samples, the problems of adjusting for other co-morbidities and the complexity of performing interventional trials in a slowly progressive disease [5]. The use of appropriate animal models could potentially have an important role in furthering the understanding of the pathogenesis of human AAA and in targeting the development of new therapies for the management of AAA [6], [7], [8]. Recently, a new animal model was reported in which AAA was induced by peri-adventitial application of calcium phosphate to the infra-renal aorta [9]. This model mimics some key features of human AAA, including aortic inflammation, calcification and elastic lamellar degradation [9].

Athero-thrombosis and older age are important risk factors for AAA [1], [10], [11]. Apolipoprotein E (ApoE) plays an important role in cholesterol metabolism by serving as a cholesterol transporter and promoting cholesterol efflux and degradation [12], [13]. Deficiency or dysfunction in ApoE has been critically implicated in the development and progression of atherosclerosis, a known risk factor for AAA [14]. Allelic variation of ApoE has been reported to be associated with differential expansion rates of small AAAs [15]. The prevalence of AAA has been reported to be ∼5% in men aged 65–74 years [16] and ∼10% in men aged ≥75 years [17], [18].

AAA is regarded as a local manifestation of a systemic disease [19]. Most patients with AAA have other enlarged arteries including dilation of other aortic sites and large distal arteries [20]. The ideal animal model of AAA would mimic all of the features of the human disease. It has not previously been investigated whether ApoE or age influence AAA induction in the calcium phosphate model. It is also not clear whether sites distant to the local application of calcium phosphate dilate in response to AAA induction. The aims of this study were to: (1) investigate the effect of ApoE deficiency (ApoE^−/−) on the AAA formation in the calcium phosphate-induced AAA model; (2) assess whether older age promotes AAA formation in this model; and (3) to examine if local application of calcium phosphate to the infra-renal aorta stimulates expansion of distant aortic segments.