An abdominal aortic aneurysm (AAA) is an abnormal dilation of the abdominal aorta and is associated with a mortality rate of >50% in the event it ruptures. AAAs are characterized by an increase of ³50% in aortic diameter compared with the normal diameter or a maximum infrarenal aortic diameter ≥30 mm (1). The prevalence of AAA increases with age and ~4% of men >55 years have an AAA (1) and AAAs cause ~1% of all deaths in developed countries (2).

Therapeutic options for AAA are currently limited to surgical or interventional repair to prevent rupture which can be effective for several decades; however, no pharmacologic therapy has been reported to have a significant effect in humans (1,3,4). A delay in the development of AAA treatments is partly due to the absence of an AAA animal model of human AAA (3) and because human AAA tissue can only be obtained during surgical repair, investigation of AAA with human tissue is limited (5). Previous studies have proposed AAA animal models (2,6); however, more evidence of histological features and similarities in shape is needed to establish an effective AAA animal model for the study of human clinical applications. A common animal model for the aneurysmal aortic lesions consists of the transient perfusion of the abdominal aorta with porcine pancreatic elastase (PPE) (2,7-10). Another common model of AAA induction is CaCl₂ perfusion of the aorta (2,4). Other AAA induction methods, including mechanical induction through aortic stretching, patching or injury, have also been reported (2). In certain cases, a combination of the aforementioned methods have been reported (11-13). The present model causes mechanical injury to the aortic intima, making it similar to human AAA. The present study established a novel and reproducible animal model of AAA using mechanical injury. This model is based on the hypothesis that the mechanical injury will initiate the inflammation process and destruction of the elastin and collagen fibers at the abdominal aorta, as observed in human AAA tissue.

This experimental animal study was approved by the Pusan National University Institutional Animal Care and Use Committee (approval no. PNU-2018-1909; Republic of Korea). A total of 22 adult Lewis rats (male; age, 8-12 weeks; Koatech Technology Corporation) weighing ~330 g (range, 256-450 g) were used. Rats were housed and maintained at 25˚C and 50% humidity, with a 12 h light/dark cycle according to the institutional animal protocols. All rats had regular food and water ad libitum with no restrictions on movement. All rats were from a specific pathogen free herd and evaluated as healthy by a veterinarian prior to inclusion. The rats were randomly divided into 2 groups and harvested after 42 (6-week group) and 63 (9-week group) days to investigate the progression of the AAAs. All experimental procedures were performed according to Pusan National University's guidelines for animal care.

The present study aimed to establish evidence for a novel and reproducible animal model of AAA. It was hypothesized that mechanical injury would initiate the inflammation process and destruction of the elastin and collagen fibers at the abdominal aorta, thus mimicking the formation of AAAs in humans. To evaluate this hypothesis, changes in the AAA tissue at 6 and 9 weeks were evaluated by observing macro- and microscopic. The results of the present supported this hypothesis, as the animal models demonstrated similar morphological and microscopic findings as human cases of AAA. This can be seen by how induced AAAs in the present model had a symmetric fusiform shape in the infrarenal area, similar to human AAAs.

The development of new experimental techniques, genetic analysis and robust molecular tools has contributed to the understanding of the pathophysiology of AAAs. While certain cellular mechanisms, such as inflammatory response mechanisms, have been reported to be responsible for AAA formation (14), the application of physical force to put pressure on the heartbeat and blood flow are not commonly used techniques for AAA animal modeling. However, hypertension is positively associated with the incidence of AAAs (15) and physical force, such as that from high blood pressure, could be the cause of mechanical injuries that initiate the inflammation process and the destruction of the elastin and collagen fibers of the abdominal aorta (16). Therefore, a mechanical injury model was used to mimic the physical effects that may lead to AAAs in the present study. Several models were evaluated in pilot studies, including end-to-end anastomosis, out-in-needle injury and in-out-needle injury. Aneurysm was not formed in the end-to-end anastomosis model, and bleeding was not controlled in the out-in-needle injury model. Therefore, the in-out-needle injury model was further evaluated in the present study as this had demonstrated the best results in the pilot studies (data not shown).

There are numerous AAA animal models in use, each of which have drawbacks in the matching of the effects of human AAAs. For example, the porcine pancreatic elastase model is commonly used and AAAs have been previously induced via PPE perfusion in mice (2,8). PPE modeling in rats has been reported to demonstrate similar temporal and pathophysiological findings as those found in humans, such as the correlation of aortic dilation with the state of the inflammatory response, which leads to the destruction of the medial elastic tissue (2,8). In rat PPE models, the rates of successful induction have been high but in larger animals, the success rates have been reported to be lower (9) due to technical difficulties with the model. Specifically, when infusing the PPE, cannulation of the aorta is required and leakage of the PPE solution during perfusion happens frequently, and this process can result in the death of the animal (2).

The CaCl₂ model is another frequently used model (2). Exposing the aorta to CaCl₂ leads to the destruction of the elastin fibers of the wall of the aorta, the infiltration of inflammatory cells and AAA induction. However, periaortic CaCl₂ application does not reliably induce AAA formation, with success varying from 80-100% of the models developing aneurysms (2,10,17).

An angiotensin II model in mice is technically less challenging than others mentioned here and there is no need for major surgical manipulation. It has been previously reported that AAA is encouraged in hyperlipidemic apolipoprotein E-deficient mice with continuous subcutaneous angiotensin II infusion (5,18). However, the induced AAAs in this model have certain differences when compared with human AAAs such as a significantly higher incidence of aortic dissection and rupture, and more frequently observed suprarenal AAAs, while humans present a much higher incidence of infrarenal AAAs (2).

The xenograft model uses implanted xenograft arterial tissues to generate AAAs using the rejection of the implanted tissue by the immune response (7,19). Models which use knockout animals, which focus on the disruption of one or more gene alleles resulting in the creation of strain-deficient animals have also been previously reported (2).

The mechanical injury model used in the present study has three distinct advantages. First, the model had high reproducibility. At 9 weeks, the size of the aorta increased by up to 133% in 88% of the rats (8/11). Reproducibility is one of the most important conditions to meet in animal models so that additional studies can be performed reliably. Second, the gross and microscopic findings of the model were very similar to findings previously reported in human AAAs. The induced AAAs in this model had a symmetric fusiform shape in the infrarenal area, similar to human AAAs. However, in other animal models, saccular shaped AAAs (20) or suprarenal AAAs (5) were reported to have been induced. Because the different shapes could indicate different blood flow and progression, gross similarities are important to consider. In the present model, important microscopic findings were found such as the destruction of the tunica media and the elastic fibers, neovascularization, infiltration of lymphocytes and intraluminal mural hematomas (19). In particular, the histological characteristic of broken elastin in the formation of an aneurysm is similar to that of human aneurysm. Finally, the model described in the present study can be reproduced at a very low cost as it did not require special drugs or equipment. Only typical vascular surgical instruments were needed to perform the surgical intervention and because the rats were allowed to live without subsequent manipulation, there was no additional cost. Furthermore, no paraplegia occurred in the short operation time of 30-50 min.

The limitation of the model comes from the surgical intervention which required microsurgical skills, which can be difficult for inexperience staff to follow. Improper surgical technique may result in animal deaths or unsuccessful induction of AAAs, which could cause reproducibility to decrease and costs to increase. Furthermore, the present study only performed a 9-week investigation. Because the AAA diameters increased from 6- to 9-weeks, it could be hypothesized that the AAAs would grow larger over time, however this and whether they eventually rupture like human AAAs was not confirmed. Before starting to use this model, it is recommended that researchers should be trained on animal experiments and vascular management. Furthermore, the present study did not yield clear results for the inflammatory response and confirmation of the inflammatory response requires further microscopic examination in future studies.

In the 6-week group, it was demonstrated that the aortic diameter did not increase in 3/10 animals. They typically had an initial weight of <300 g, which was 18% less than the mean weight of 365 g. This suggested the need to investigate the relationship between weight and hemodynamic parameters such as blood pressure and cardiac output. Further studies are also required to elucidate the timeframe for the occurrence of AAAs and what the effect of proteins expressed by different genes on the formation of aneurysms in mice with and without aneurysms are. Whether the use of certain drugs can prevent the formation of aneurysms, delay the continued increase of an aneurysm until rupture or even make the aneurysm shrink also requires evaluation. Future studies, are also required to assess the long-term results of this model while measuring the AAAs with ultrasound. This model should also be trialed in larger animals although there have been many reports of failures when applied to larger animals (2,21). However, a trial in larger animals is needed to confirm that the AAAs in the present model are similar to human AAAs with infrarenal symmetric fusiform shape.

In conclusion, the mechanical injury-induced AAA animal model reported in the present study demonstrated very similar morphological and microscopic findings to those of human AAAs. The model showed high reproducibility, with 60% induction in the 6-week group and 88% induction in the 9-week group. Though the present study was performed in rodents, this method could provide evidence for AAA treatment if it can be applied to middle and large sized animals.