Elsevier

Journal of Surgical Research

Volume 178, Issue 2, December 2012, Pages 1038-1045
Journal of Surgical Research

Vascular
Differential gender- and species-specific formation of aneurysms using a novel method of inducing abdominal aortic aneurysms

https://doi.org/10.1016/j.jss.2012.04.073Get rights and content

Abstract

Background

The objective of this study was to test a novel model of inducing abdominal aortic aneurysms (AAAs) in different mouse strains and genders.

Materials and methods

Male and female C57BL/6 and B6129 mice (n = 5 per group) underwent periaortic dissection and porcine pancreatic elastase (30 μL) or inactivated elastase application (5 min) to the aorta. Aortic measurements were taken on days 0 and 14. Aortic samples were analyzed for histology and zymography for matrix metalloproteinase (MMP) activity. Comparison statistics were performed using unpaired t-test.

Results

AAA phenotype (50% aortic increase) occurred in external elastase–treated males (100%) and females (90%). No control animals developed AAAs. The aortic diameter was larger in C57BL/6 and B6129 elastase-treated versus control males (P = 0.0028 and P < 0.0001, respectively) and females (P < 0.0001 and P = 0.0458, respectively). Histology verified phenotype via disrupted internal elastic laminae. Macrophage counts in elastase-treated animals were >6-fold higher than in controls (all groups significant). MMP9 activity was greater in elastase-treated males and females in C57BL/6 (P = 0.0031, P = 0.0004) and B6129 (P = 0.025, P = 0.2) mice; MMP2 activity was greater in C57BL/6 versus B6129 male elastase-treated mice.

Conclusions

This rodent model produced AAAs in both genders and strains of mice. This model is simple, has little variability, and occurs in the infrarenal aorta, substantiating the external elastase model for future studies.

Introduction

Several experimental methods have been used to induce experimental abdominal aortic aneurysms (AAAs) in animal models. While there is no perfect model for murine development of AAAs that has universal applicability, we developed a model of topical elastase application to produce aneurysms in the infrarenal aorta that is reproducible and easy to perform. To date, the external elastase model has not been tested in mice.

Various experimental AAA models have had varying incidences of AAA development among different murine strains. For example, Thompson et al. showed AAA resistance in wild-type B6129/SvEv, SvJ, and CBA mice using the elastase perfusion model. Intermediate susceptibility to AAAs was found in Balb/c mice, among others, while C57BL/6 was highly susceptible to AAA formation [1]. Also, as many knockout strains are on different genetic backgrounds, it must be known if the control animals of the same background develop AAAs for comparisons and conclusions to be valid. Therefore, models should be tested on more than one strain of mouse.

In addition, confirmation that the female resistance to AAA formation is maintained with external elastase application is necessary for utilization of this model in further studies of gender differences. There have been several studies from our lab, utilizing the elastase perfusion model, demonstrating that females are protected from AAA formation [2], [3], [4], [5]. The mechanisms underlying this gender difference are not completely clear. Some evidence exists that macrophage infiltration is inhibited, thus decreasing the inflammatory reaction in females as compared with males. Matrix metalloproteinase 9 (MMP9), a crucial proteinase in AAA development released by macrophages, is decreased in females [3]. Cho et al. [4] and Sinha et al. [5] have demonstrated that neutrophils and certain proinflammatory mediators, such as chemokines and cytokines, are decreased in females compared with males during AAA formation. Thus, both direct and indirect mechanisms have been documented showing that estradiol exerts protection from AAA formation females.

The objective of this study was to test this novel model of AAA formation in different murine strains, in both male and female mice.

Section snippets

Animal surgery

After induction of general anesthesia with isoflurane, male and female mice, both C57BL/6 and B6129 strains, obtained from Jackson Laboratories (Bar Harbor, ME) (8 wk, 20–30 g) (n = 5 for each of eight groups) had their abdominal aorta exposed via midline laparotomy. Aortic diameter was measured using a video micrometer (Nikon Elements software; Nikon, Melville, NY) in three locations: just below the renal arteries, mid–abdominal aorta, and at the iliac bifurcation. The infrarenal aorta was

AAA phenotype is confirmed in males and females in C57BL/6 and B6129 strains

AAA phenotype, as defined by a 50% increase in aortic diameter at harvest compared to baseline, occurred in 100% of males (5 of 5 for both strains) treated with external elastase (Fig. 1). In addition, 90% of female mice (4 of 5 of the C57BL/6 and 5 of 5 of the B6129) also formed AAAs. Visually at harvest, treated animals had thin-walled, fusiform AAAs, similar to those seen in humans. None of the control animals in any group (male and female, both strains) developed AAAs (0 of 20). The aortic

Discussion

In the present study, we have documented that external elastase application to the infrarenal aorta results in the ability to generate AAAs in two strains of mice, and equally in male and female mice. As opposed to previous work, the ME B6129 mice formed larger aneurysms than their C57BL/6 counterparts. In this model the AAA phenotype (as determined by aortic diameter and disrupted elastic laminae by van Gieson stain) occurs in association with an increase in Mac-2-stained cells in both the

Conclusions

In contrast to previous studies, which have documented vast variability in AAA formation between mouse strains and genders, the external elastase model of AAA formation produced significant aortic dilatation in males and females of two strains of mice despite differences in inflammatory cells. This study further serves to highlight the variability in rodent AAA models and substantiates the external elastase model for studying AAAs in future, as it can be easily duplicated in both genders and in

Acknowledgment

This work was supported by NIH R01 HL081629-01, NIH R01 supplemental HL081629-03S1, and the University of Michigan Cardiovascular Center Aortic Program Research Fellowship 2009–2010.

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