**3. Remodeling history model. Biological Biodeterminants, BBDs**

Most investigators would agree that the pathogenesis of the abdominal aortic aneurysm (AAA) is multifactorial. There appear to be environmental, genetic, autoimmune, inflammatory, and structural factors.

The term "atherosclerotic AAA" is misleading because it suggests that atherosclerosis is a necessary cause of AAA disease. While some patients with have atherosclerotic occlusive peripheral vascular disease, others have minimal atherosclerotic disease. For this reason, the Joint Committee of the Society for Vascular Surgery recommending that the term "nonspecific AAA" be used since 1991.

The definition of AAA has varied in the literature over the years, but all definitions have in common a specification of the degree of aortic dilatation. So, the definition is a permanent localized dilatation of an artery having at least a 50% increase in diameter compared with the expected normal diameter of the artery or of the diameter of the segment proximal to the dilatation. According to this definition, an infrarenal AAA could then be defined as 3.0 cm if 2.0 cm is the expected maximal diameter of the infrarenalaort in an individual of a specific body scale.

#### *Risk factors*

78 Aneurysm

The majority of studies found in medical literature report this increase in the incidence of aortic aneurismal disease, which is expected in a continuously aging population in developed countries. In spite of significant improvement in surgical procedures and technological advancements in imaging devices in recent years, the associated aneurysm

Currently, the lack of an accurate AAA rupture risk index remains an important problem in the clinical management of the disease. The main clinical criteria in deciding on the treatment of AAA patients are: a) the peak transverse diameter and b) the growth rate. If the peak diameter reaches the upper threshold (5-5.5 cm) or the maximum diameter expansion rate is > 0.5 cm/yr for smaller AAAs the patient may be submitted for surgical intervention, also depending on the state of health and willingness of the patients. The main limitation of this practice is that these criteria, although have a significant empirical basis, can be considered insufficient because they have not a physically sound theoretical basis. This statement should not be surprising; approximately 33% of ruptured AAAs have diameters smaller than 50 mm [5] which is indicative of the complex pathogenesis of the disease

Due to these observations, recently researches have been focused at improving the knowledge and the understanding of the phenomena associated with the formation and evolution of aneurysm pathology in order to define whether other variables could be predictive of rupture. The literature begins to reflect the existence of a consensus that, rather than empirical criteria, the develop of a biomechanical approach based on a multiscale

This chapter examines the basis of the biomechanical approach. The main aim is to support the hypothesis that biomechanical considerations may become into powerful tool for a

This new approach has its foundation in the integration, through appropriate relations, of factors from different natures (biological, structural and geometric) and scales (temporal and dimensional) at the molecular, cellular, tissue and organ levels (from bottom level to top level), which allow to describe, from quantitatively point of view, the aneurysm progression

These defined relations are known as biomechanical factors or biomechanical determinants

The basic premise of the biomechanical approach to estimate the AAA rupture risk, is that this phenomenon follows the principle of material failure, that is, an aneurysm ruptures when the stresses acting on the arterial wall exceeding its failure strength, reflecting the interaction between the arterial wall structural remodelling and the forces generated by

model can be a significant step for the accurate assessment of the rupture risk.

mortality and morbidity rate have also risen concomitantly.

progression that cannot be capture by traditional indicators.

reliable patient-specific prediction of AAA rupture risk.

**2. Biomechanical approach. Method grounds** 

and its rupture potential.

blood flow within the AAA.

(BDs).

The four principal positive risk factors for AAA are smoking, age, male sex, and family history. While smoking clearly seems to be an environmental factor, issues related to addiction and dose-effect responses are doubtless modified by genetic influences. The three principal negative risk factors for AAA are diabetes, female sex, and African-American descent, all of which are genetically determined.

There is a more complicated relationship between plasma lipid levels and the risk of AAA. Blanchard et al [6], failed to show any correlation between cholesterol levels, low-density lipoprotein (LDL) or high-density lipoprotein (HDL) and aneurysm risk (Blanchard). However, it have been showed an increased risk in patients whose plasma cholesterol was high and a protective effect was seen in patients whose serum HDL was high [7]. Low serum HDL gave an increased risk of AAA [8].

There is some disagreement in the literature regarding the effect of hypertension on aneurysm risk. The American Veterans study represented the largest of its type and showed hypertension to be an independent risk factor. Taking medication for high blood pressure was a risk factor, whereas hypertension itself was significant in women. Tornwall and Blanchard both showed both systolic and diastolic hypertension to be risks [6]. A study of all men born in Malmo in the year 1914 failed to demonstrate hypertension as a risk factor at all [9]. Experimentally, AAAs artificially induced into hypertensive rats were found to grow larger than those in normotensives [10] and the dilatation correlated well with systolic pressure.

#### *Molecular genetics*

Epidemiologic review indicates an aneurysm gene expression that is typically delayed until at least the sixth decade. There is strong evidence for inherited predisposition, and possibly

an association with generalized arteriomegaly. It have been demonstrated an incidence of 20% aortic aneurysms among first order relatives of aneurysm patients [11]. In [12] was showed genetic linkages, accounting for abdominal aneurysm formation in 50 families, who had clustering of the lesion in two or more first order relatives. Possibly, they possessed a common metabolic disorder affecting the arterial wall.

A retrospective study of hospital patients in Zimbabwe demonstrated a higher incidence of aneurysms among whites than Africans [13]. By using ultrasound screening of first degree relatives demonstrated aortic aneurysms in 20–30% of male siblings over 55 years of age [14]. Case reports of familial aneurysm disease in patients without connective tissue or vascular diseases add validity to the theory of genetic linkage. The occurrence of multiple aneurysms in individuals is consistent with a genetic foundation. Many authors suggest aneurysm disease is a systemic process. Frequently, patients suffer from generalized arteriomegaly; often this is accompanied by multiple aneurysms.

Several cross-linking defects have been associated with aneurysm formation. Tilsonstudied the biochemistry of a collagen component deficiency that predisposes to aneurysms [12]. They evaluated pyridine cross-linkages and found fewer cross-linkages per collagen molecule in human skin samples. This suggests a genetic basis for aneurysm disease. Experiments with sex-linked defects of collagen and elastin demonstrate the blotchy BLO allele. These models exhibit aortic aneurysms and diminished skin tensile strength. The pattern of expression indicates the trait is related to the X chromosome. In [15] it was reviewed the literature and found clear evidence for an independent genetic defect in most AAAs. Their work centered on a genetic analysis of collagen genes. Genetic collagen defects causing architectural defects are established in osteogenesisimperfecta (type I collagen of bone) and chondrodysplasias (type II collagen of cartilage). New evidence implicates mutations in the type III procollagen gene in the pathogenesis of aneurysmal disease. Various mutations have been confirmed in studies of patients with type IV Ehlers–Danlos syndrome (EDS) [16].

Studies of patients with aneurysms clearly demonstrate family linkage, and the data strongly suggest a genetic defect. Statistical analysis supports a recessive inheritance pattern in approximately 10% of men who have aneurysms. Research in this area is active and implicates an autosomal diallelic major locus.

The two genes with the strongest supporting evidence of contribution to the genetic risk for AAA are the CDKN2BAS gene, also known as ANRIL, which encodes an antisense ribonucleic acid that regulates expression of the cyclin-dependent kinase inhibitors CDKN2A and CDKN2B, and DAB2IP, which encodes an inhibitor of cell growth and survival. Functional studies are now needed to establish the mechanisms by which theses genes contribute toward AAA pathogenesis [17].

#### *Structural pathophysiology*

#### *Atherosclerosis*

The traditional view of aneurysm formation is that arterial dilation is a consequence of degenerative atherosclerotic disease, which results in acquired wall weakness. The experienced vascular surgeon is well aware that peripheral arteriosclerosis and aneurysmal disease often coexist. Severe atherosclerotic calcification in the aortoiliac vessels presents a technical challenge in aneurysm surgery. Epidemiologic, radiographic, and histologic data support the association between aneurysm disease and atherosclerosis [18].

AAAs and atherosclerosis share many risk factors and frequently occur simultaneously. The frequency of aortic aneurysms closely parallels the prevalence of atherosclerosis; for example, the low abdominal aneurysm rate in Asia correlates with the decreased incidence of atherosclerosis. Radiographic and histopathologic studies support the link between atherosclerosis and aneurysms. Ultrasound screening of patients with peripheral vascular disease detects a 5.9% rate of AAA, double that of the general population [19]. Studies of patients suffering from coronary and carotid artery occlusive disease detect an aortic aneurysmal rate of 11–13.5% [9]. Histologic evaluations of sections from aortic aneurysms show atherosclerotic changes and thinning of the media.

Pathophysiologic principles also support the concept that atherosclerosis contributes to aneurysm formation. Atherosclerotic plaques may obstruct nutrient diffusion from the lumen to the media. The needs of the media must then be supplied exclusively by vasa vasorum from the adventitia. However, this may be inadequate due to incomplete distribution of vasa vasorum throughout the human arterial system [20]. Aortic vasa vasorum usually arise from the renal arteries, accounting for the relative sparing of the perirenal aorta from aneurysm formation.

Structural changes induced by atherosclerosis may contribute to aneurysm formation. As atherosclerosis progresses in humans, friable type I collagen replaces native type III collagen [21].Thus, the architectural integrity of the vessel is impaired, leading to a predilection to aneurysm formation. An association between aortic aneurysms and atherosclerosis is not surprising since the geometry and hemodynamics of arterial dilation predispose to atherosclerosis formation. Aneurysms have increased in incidence, prevalence, and mortality over the last 30 years, while coronary artery and cerebrovascular diseases have not. The divergence of these diseases in prevalence and mortality indicates that while risk factors are shared, the development of aneurysm disease is not entirely explained by atherosclerosis.

Although the epidemiologic link between the two is strong, it is propose that occlusive atherosclerotic aortic disease and aortic aneurysmal disease are distinct entities [12]. This is based on the different characteristics of these groups including age of onset, male–female ratio, clinical course, and prognosis. Evidence found to correlate with the size and state of aneurysm indicates that aneurysms reflect a heterogeneous disease with multiple forms and etiologic factors.

#### *Autoimmunity*

80 Aneurysm

an association with generalized arteriomegaly. It have been demonstrated an incidence of 20% aortic aneurysms among first order relatives of aneurysm patients [11]. In [12] was showed genetic linkages, accounting for abdominal aneurysm formation in 50 families, who had clustering of the lesion in two or more first order relatives. Possibly, they possessed a

A retrospective study of hospital patients in Zimbabwe demonstrated a higher incidence of aneurysms among whites than Africans [13]. By using ultrasound screening of first degree relatives demonstrated aortic aneurysms in 20–30% of male siblings over 55 years of age [14]. Case reports of familial aneurysm disease in patients without connective tissue or vascular diseases add validity to the theory of genetic linkage. The occurrence of multiple aneurysms in individuals is consistent with a genetic foundation. Many authors suggest aneurysm disease is a systemic process. Frequently, patients suffer from generalized

Several cross-linking defects have been associated with aneurysm formation. Tilsonstudied the biochemistry of a collagen component deficiency that predisposes to aneurysms [12]. They evaluated pyridine cross-linkages and found fewer cross-linkages per collagen molecule in human skin samples. This suggests a genetic basis for aneurysm disease. Experiments with sex-linked defects of collagen and elastin demonstrate the blotchy BLO allele. These models exhibit aortic aneurysms and diminished skin tensile strength. The pattern of expression indicates the trait is related to the X chromosome. In [15] it was reviewed the literature and found clear evidence for an independent genetic defect in most AAAs. Their work centered on a genetic analysis of collagen genes. Genetic collagen defects causing architectural defects are established in osteogenesisimperfecta (type I collagen of bone) and chondrodysplasias (type II collagen of cartilage). New evidence implicates mutations in the type III procollagen gene in the pathogenesis of aneurysmal disease. Various mutations have been confirmed in studies of

Studies of patients with aneurysms clearly demonstrate family linkage, and the data strongly suggest a genetic defect. Statistical analysis supports a recessive inheritance pattern in approximately 10% of men who have aneurysms. Research in this area is active and

The two genes with the strongest supporting evidence of contribution to the genetic risk for AAA are the CDKN2BAS gene, also known as ANRIL, which encodes an antisense ribonucleic acid that regulates expression of the cyclin-dependent kinase inhibitors CDKN2A and CDKN2B, and DAB2IP, which encodes an inhibitor of cell growth and survival. Functional studies are now needed to establish the mechanisms by which theses

The traditional view of aneurysm formation is that arterial dilation is a consequence of degenerative atherosclerotic disease, which results in acquired wall weakness. The

common metabolic disorder affecting the arterial wall.

arteriomegaly; often this is accompanied by multiple aneurysms.

patients with type IV Ehlers–Danlos syndrome (EDS) [16].

implicates an autosomal diallelic major locus.

genes contribute toward AAA pathogenesis [17].

*Structural pathophysiology* 

*Atherosclerosis* 

Autoimmunity may precipitate the inflammatory cascade. Aneurysm aortic extract was studied and noted to contain large quantities of IgG. Further studies revealed that the IgG from AAA patients was present and reactive against various proteins present in the

aneurysmal aorta [22]. One of the initial putative autoantigen extracts was an 80-kDa dimer, designated aortic aneurysm associated protein-40 (AAAP-40). AAAP-40 was reactive with 79% (11 of 14) of AAA IgG preparations, and 11% (1 of 9) of controls *(p =* 0.002) (Gregory). Other autoantigens have subsequently been found, and are currently under investigation in our laboratory. Evidence continues to accumulate to support the notion that autoimmunity may play an important role in aneurysmal degeneration of the aorta. Some of these autoantigens are absent in the external iliac artery, perhaps explaining why this artery rarely becomes aneurismal.

Triggering of autoimmunity can be brought about by autoantigens or molecular mimics. For example, molecular mimicry may occur with cytomegalovirus and clone 1. Also, rabbit antibody against *Treponemapallidum*and herpes simplex have been shown to bind to the adventitial elastin-associated microfibrils. The putative autoantigen AAAP-40 has homologies with *Treponemapallidum*and herpes. The hypothesis is that there are epitopes in the microbial proteins that are similar to the AAAP-40, thereby triggering an autoimmune response. Tanaka et al [23] detected herpes simplex viral DNA in 12 of 44 AAA specimens, compared with 1 of 10 normal subjects.

#### *Inflammation*

The normal aorta has few inflammatory cells within in its wall. An influx of CD3+ cells and lymphocytes is seen in AAA tissues Although 66% of all lymphocytes in AAAs are in the adventitia, polyclonal B-lymphocytes are abundant in the media. IgG is elevated in AAA specimens. In [24] it was showed an inflammatory infiltrate in the adventitia in 68% of 156 AAA resection specimens examined retrospectively. Macrophages are found throughout the wall of AAA specimens. The macrophage Fc receptors regulate the secretion of proteinases by receptor specific mechanisms. Phagocytes produce proteinases such as elastase and collagenase. On the other hand, it have been implicated the collagenase, stromelysin, and gelatinase-B (MMP-1,3,9) in the destruction of the aorta matrix [25]. Cytokines are released by inflammatory cells and smooth muscle cells in the aorta. They are predominantly: interleukin 1 (IL-1), IL-6, IL-8, monocyte chemoattractant protein (MCP-1), tumor necrosis factor (TNF), and interferon (IFN). These cytokines, to varying degrees, cause MMP expression, TIMP reduction, induction of prostaglandin synthesis, lymphocyte proliferation, and chemotaxis. An autoimmune or inflammatory cascade, as proposed in some etiologies of AAAs, is perpetuated via the use of cytokines [26].

#### *Enzymatic degradation*

The elastin: collagen ratio has consistently been shown to be reduced in AAAs when ompared with normal aortas, leading to loss of elasticity and weakening of the aneurysmal wall. This may not be simply due to increased elastin degradation, as Minion et al. have shown that the total elastin content of the aneurismal wall may actually increase, but that the corresponding increase in collagen is much greater (Minion). Despite this evidence, there is little doubt that proteolysis plays an important role in aneurysm development. Aneurysmal disease differs from stenotic disease by the intensity of proteolytic activity within the extracellular matrix. The established association with chronic lung disease supports the argument that elastolysis is a major contributory factor, and indeed this is an area in which there has been much research. For some time, the cause of elastin degradation remained unknown, but even as early as 1980 when it was described increased collagenase activity [27]. In 1991, it was found a spectrum of collagenase activity in the aortic wall of both atherosclerotic and aneurysmal vessels ranging from 55–92 kDa [29].

82 Aneurysm

becomes aneurismal.

*Inflammation* 

*Enzymatic degradation* 

compared with 1 of 10 normal subjects.

of AAAs, is perpetuated via the use of cytokines [26].

aneurysmal aorta [22]. One of the initial putative autoantigen extracts was an 80-kDa dimer, designated aortic aneurysm associated protein-40 (AAAP-40). AAAP-40 was reactive with 79% (11 of 14) of AAA IgG preparations, and 11% (1 of 9) of controls *(p =* 0.002) (Gregory). Other autoantigens have subsequently been found, and are currently under investigation in our laboratory. Evidence continues to accumulate to support the notion that autoimmunity may play an important role in aneurysmal degeneration of the aorta. Some of these autoantigens are absent in the external iliac artery, perhaps explaining why this artery rarely

Triggering of autoimmunity can be brought about by autoantigens or molecular mimics. For example, molecular mimicry may occur with cytomegalovirus and clone 1. Also, rabbit antibody against *Treponemapallidum*and herpes simplex have been shown to bind to the adventitial elastin-associated microfibrils. The putative autoantigen AAAP-40 has homologies with *Treponemapallidum*and herpes. The hypothesis is that there are epitopes in the microbial proteins that are similar to the AAAP-40, thereby triggering an autoimmune response. Tanaka et al [23] detected herpes simplex viral DNA in 12 of 44 AAA specimens,

The normal aorta has few inflammatory cells within in its wall. An influx of CD3+ cells and lymphocytes is seen in AAA tissues Although 66% of all lymphocytes in AAAs are in the adventitia, polyclonal B-lymphocytes are abundant in the media. IgG is elevated in AAA specimens. In [24] it was showed an inflammatory infiltrate in the adventitia in 68% of 156 AAA resection specimens examined retrospectively. Macrophages are found throughout the wall of AAA specimens. The macrophage Fc receptors regulate the secretion of proteinases by receptor specific mechanisms. Phagocytes produce proteinases such as elastase and collagenase. On the other hand, it have been implicated the collagenase, stromelysin, and gelatinase-B (MMP-1,3,9) in the destruction of the aorta matrix [25]. Cytokines are released by inflammatory cells and smooth muscle cells in the aorta. They are predominantly: interleukin 1 (IL-1), IL-6, IL-8, monocyte chemoattractant protein (MCP-1), tumor necrosis factor (TNF), and interferon (IFN). These cytokines, to varying degrees, cause MMP expression, TIMP reduction, induction of prostaglandin synthesis, lymphocyte proliferation, and chemotaxis. An autoimmune or inflammatory cascade, as proposed in some etiologies

The elastin: collagen ratio has consistently been shown to be reduced in AAAs when ompared with normal aortas, leading to loss of elasticity and weakening of the aneurysmal wall. This may not be simply due to increased elastin degradation, as Minion et al. have shown that the total elastin content of the aneurismal wall may actually increase, but that the corresponding increase in collagen is much greater (Minion). Despite this evidence, there is little doubt that proteolysis plays an important role in aneurysm development. Aneurysmal disease differs from stenotic disease by the intensity of proteolytic activity within the extracellular matrix. The established association with chronic lung disease Importantly, although the collagenase activity was limited, it increased dramatically when tissue inhibitors of metalloproteinases (TIMPs) were destroyed. In [30], it wasalso described the increased expression of a 92 kDagelatinase in AAAs when compared with both normal aortas and aorto-occlusive disease, and localized this to the area around infiltrating macrophages. This gelatinase is part of a family of zinc-dependent proteolytic enzymes, the matrix metalloproteinases (MMPs), now known as MMP9. In the same year, Freestone et al [31],further elucidated the relative amounts of both MMP9 and MMP2 by a combination of gelatinzymography and immunoblotting. This study demonstrated that the principal gelatinase in smaller aneurysms was MMP2, but that in larger aneurysms MMP9 predominated. McMillan et al [21],investigated mRNA levels for MMPs in AAAs and found that MMP9 was maximally expressed in moderate diameter (5–6.9 cm) rather than large (>7 cm) or small (<4 cm) aneurysms. These findings suggested that whilst MMP9 was responsible for the rapid growth that was seen in this size of aneurysm, other enzymes were responsible for initiation and rupture. Pyoet al.'s paper elegantly proves a link between MMP9 and aneurysm pathogenesis by looking at the effect of inhibiting it both pharmacologically and by targeted gene disruption [32]. Mice that were deficient in the MMP9 gene failed to develop aneurysms as their wild-type counterparts did when subjected to elastase perfusion of the aorta. Bone marrow transplants from each group to the other reversed the response to elastase infusion, demonstrating that the expression of MMP9 by inflammatory cells is crucial to aneurysm development. Other MMPs have also been implicated in the development of AAAs, particularly MMP1 and MMP3. Vine and Powell also found immunoreactive MMP1 in extracts from AAAs (Vine). And more recently the expression of MMP3, as measured by reverse transcriptase polymerase chain reaction (rt-PCR), was found to be elevated in AAAs when compared to aorto-occlusive disease.

Matrix metalloproteinase 13 is a recently described enzyme also known as collagenase-3 and its expression is tightly regulated. Whilst MMP13 was not expressed at all in normal tissue, it was found in atherosclerotic disease and in significantly higher concentrations in AAAs. Expression was localized to medial smooth muscle cells in the aortic tissue, and could also be detected in human vascular smooth muscle cells in culture. Membrane type MMP1 (MT MMP1) is an activator of MMP2 and was found to be increased in aneurismal aorta when compared to normal or atherosclerotic aorta. Membrane type MMP1 was localized to aortic smooth muscle cells and macrophages in aneurysmal tissue by immunohistochemical analysis. The ability to activate MMP2 was confirmed by the addition of radiolabelled pro-MMP2, and determination of the subsequent amount of radiolabelled active MMP2. In vivo, the activity of MMPs is tightly controlled by their natural inhibitors, the TIMPs. In 2000, it was demonstrated that TIMP-1 bound to both the monomeric and dimeric forms of MMP9, whereas TIMP-2 bound only to the active form. Whilst it has been shown that the TIMPs are present in large quantities in AAAs, it has been suggested that itis an imbalance between MMPs and TIMPs that leads to the net increase in proteolysis seen. Tamarinaet al also showed that the TIMP: MMP ratio was actually decreased in AAAs, despite an absolute increase in TIMP levels [33].

Whilst there has been considerable work published in the area of collagenases and other metalloproteinases in AAAs, less is known about the role of serine proteases. Elastases of approximately 20–30 kDa have been demonstrated in the inner aspect of the media in AAAs. This elastase works best in the alkaline range, and is inhibited by α-1 anti-trypsin. The fact that it is also inhibited by phenylmethylsulphonyl fluoride (PMSF) confirms that it is indeed a serine protease. Five distinct serine proteases have been separated by gel electrophoresis from aortic aneurysm tissue, suggesting there is a spectrum of enzymes at work. In addition to MMPs and serine proteases, there is also the cysteine protease group. These differ from serine proteases by the substitution of an Asn residue for an Asp in the catalytic triad. Cathepsins S and K are examples of this type of elastase and have been shown to be produced in abundance by smooth muscle cells in atheroma. They are inhibited by cystatin C, the expression of which is governed by a polymorphism of its signal peptide. As discussed previously, patients in whom the cathepsins were not inhibited displayed faster growing aneurysms.

AAA is a multifactorial disease with genetic risk factors and an immunologic component. Immune cells, including macrophages, neutrophils, mast cells, B- and T- lymphocytes, along with vascular smooth muscle cells and adventitial fibroblasts, produce cytokines and enzymes, promoting an inflammatory reaction, extracellular matrix degradation, and neovascularization. Among the different enzymes secreted by immune and stromal cells, matrix metalloproteinase (MMP)-2, MMP-9, MMP-12, cathepsins, and neutrophil elastase cause medial degeneration. Chymase causes smooth muscle cell apoptosis, and MMP-3, MMP-8, and MMP-13 cause adventitial collagen degradation, promoting abdominal aortic aneurysm rupture [34].

#### *Oxidative stress*

The action of reactive oxygen species has been implicated in the etiology of many disease processes. In particular, the effect of oxidative stress on many aspects of vascular biology has come under intense scrutiny over the past few years. The addition of antioxidants significantly reduced the activity of MMP9, whereas the addition of inhibitors of protein kinase C had no effect. These results suggest that the increased proteolytic activity seen in the extracellular matrix in patients with diabetes mellitus is due, at least in part, to the effects of oxidation, and may help to explain a link between aneurysm formation and oxidative stress. A further series of aortic banding experiments have demonstrated that in areas of high pressure there is an up-regulation of endothelial nitric oxide synthase (eNOS) when compared with tissues downstream of the artificial coarctation [35].

Measuring nitrotyrosine in the same tissues gave some indication of the degree of nitric oxide breakdown and sequestration by reactive oxygen species. In the areas above the banding (heart, brain and thoracic aorta) the levels of nitrotyrosine were much higher than in areas not exposed to high pressures (distal aorta). The inactivation of nitric oxide due to oxidative damage in areas of high pressure is another indication of vascular endothelial dysfunction, which may contribute to the pathogenesis of aneurysms. Combining the in vitro elastase perfusion rat model of Anidjaret al [10],with modern cDNAmicroassay analysis,looked at the expression of 8799 genes in rats with induced aortic aneurysms, and compared them with genes expressed in rats that had undergone sham operations [36]. Using this technique they were able to identify over 200 genes whose expression had more than doubled in the aneurysm group. Significantly, this included many genes reflecting an increase in oxidative stress, notably hemeoxygenase, inducible nitric oxide synthase (iNOS), 12-lipoxygenase and heart cytochrome C oxidase, subunit VIa. Conversely, antioxidant genes such as superoxide dismutase, reduced NAD-cytochrome b-5 reductase and glutathione S reductase werefound to be down-regulated. These two complementary findings both point to oxidative stress playing a major role in AAA development.

#### *Infection*

84 Aneurysm

increase in TIMP levels [33].

faster growing aneurysms.

aneurysm rupture [34].

*Oxidative stress* 

present in large quantities in AAAs, it has been suggested that itis an imbalance between MMPs and TIMPs that leads to the net increase in proteolysis seen. Tamarinaet al also showed that the TIMP: MMP ratio was actually decreased in AAAs, despite an absolute

Whilst there has been considerable work published in the area of collagenases and other metalloproteinases in AAAs, less is known about the role of serine proteases. Elastases of approximately 20–30 kDa have been demonstrated in the inner aspect of the media in AAAs. This elastase works best in the alkaline range, and is inhibited by α-1 anti-trypsin. The fact that it is also inhibited by phenylmethylsulphonyl fluoride (PMSF) confirms that it is indeed a serine protease. Five distinct serine proteases have been separated by gel electrophoresis from aortic aneurysm tissue, suggesting there is a spectrum of enzymes at work. In addition to MMPs and serine proteases, there is also the cysteine protease group. These differ from serine proteases by the substitution of an Asn residue for an Asp in the catalytic triad. Cathepsins S and K are examples of this type of elastase and have been shown to be produced in abundance by smooth muscle cells in atheroma. They are inhibited by cystatin C, the expression of which is governed by a polymorphism of its signal peptide. As discussed previously, patients in whom the cathepsins were not inhibited displayed

AAA is a multifactorial disease with genetic risk factors and an immunologic component. Immune cells, including macrophages, neutrophils, mast cells, B- and T- lymphocytes, along with vascular smooth muscle cells and adventitial fibroblasts, produce cytokines and enzymes, promoting an inflammatory reaction, extracellular matrix degradation, and neovascularization. Among the different enzymes secreted by immune and stromal cells, matrix metalloproteinase (MMP)-2, MMP-9, MMP-12, cathepsins, and neutrophil elastase cause medial degeneration. Chymase causes smooth muscle cell apoptosis, and MMP-3, MMP-8, and MMP-13 cause adventitial collagen degradation, promoting abdominal aortic

The action of reactive oxygen species has been implicated in the etiology of many disease processes. In particular, the effect of oxidative stress on many aspects of vascular biology has come under intense scrutiny over the past few years. The addition of antioxidants significantly reduced the activity of MMP9, whereas the addition of inhibitors of protein kinase C had no effect. These results suggest that the increased proteolytic activity seen in the extracellular matrix in patients with diabetes mellitus is due, at least in part, to the effects of oxidation, and may help to explain a link between aneurysm formation and oxidative stress. A further series of aortic banding experiments have demonstrated that in areas of high pressure there is an up-regulation of endothelial nitric oxide synthase (eNOS)

Measuring nitrotyrosine in the same tissues gave some indication of the degree of nitric oxide breakdown and sequestration by reactive oxygen species. In the areas above the

when compared with tissues downstream of the artificial coarctation [35].

Infected aortic aneurysms are uncommon, and infrequently have their pathological features been described. Panneton and Edwards evaluated clinical and histopathologic features in patients undergoing surgical repair of infected aneurysms of the descending thoracic or abdominal aorta over a 24-year period [37]. The results showed that among cases with an identifiable causative organism, staphylococcus accounted for 30%, streptococcus for 20%, salmonella for 20%, Escherichia coli for 15%, and other organisms for 15%.

During recent years, attention has been paid to the role of atypical bacterial infections, including Chlamydia and Helicobacter pylori, in the process of atherogenesis and arterial disease development. The reported rates of detection within atherosclerotic lesions by PCR vary widely. Regarding Chlamydia, several studies hypothesized this organism as a possible source of vascular disease, including carotid, coronary, and aortic pathology. Its role in the pathogenesis of aortic aneurysms, however, has been controversial. Sodecket al [38],investigated the presence of C. pneumoniae in 148 tissue samples excised from control and diseased aortas. DNA of C. pneumoniae, C. trachomatis and C. psittaci were assessed by highly sensitive and specific real time polymerase chain reaction (PCR). C. trachomatis-DNA was detected in 1/65 diseased patients and in none of 83 controls (P=0.43). In a similar study, surgical specimens derived from aneurysm or aorta fragments were investigated for C. pneumoniae utilizing PCR. In asymptomatic aneurysms, DNA was found in 9 cases (29%), and in ruptured aneurysms in 14 cases (49%). In the control group, C. pneumoniae DNA was not detected in the aortic wall. Conflicting data has failed to show a clear relationship between chlamydia infection and aortic pathology.

Cytomegalovirus (CMV)-induced arterial disease has also been linked to aortic pathology. To further elucidate the mechanism by which CMV may promote atherosclerosis, Westphalet al.(Westhpal), studied the expression pattern of cellular inflammatory and proliferative signals in the aortic wall of CMV (+) and CMV (−) patients undergoing coronary artery bypass grafting (CABG). CMV-DNA in smooth muscle cells was thought to

induce local growth factor expression as well as endothelial activation, both of which can promote the progression of atherosclerosis. Since traditional atherogenic risk factors increase the likelihood of aortic CMV manifestation, CMV may play a crucial role in mediating the progression of atherosclerosis. The persistent expression of CMV-gene in the vessel wall plays a role in the vascular cellular response, including progression of atherosclerosis or vasculitis in vivo. Kilicet al [39], performed PCR analysis to demonstrate the relationship between CMV and atheromathosis at the aortic wall. CMV DNA was found in 37.9% atherosclerotic and 32.7% non-atherosclerotic vascular wall specimens.

#### *Vitamin E deficiency*

Studies have pointed to an inverse relationship between vitamin E (a-tocopherol) levels and the incidence of arterial disease. Vitamin E is an important lipid-soluble antioxidant that localizes to the hydrophobic area of biologic membranes [40]. In terms of AAA, it is hypothesized that activated polymorphonuclear cells (PMNs) release proteinases which degrade the aortic wall matrix. These same PMNs would also release oxidative enzymes, generating toxic oxygen species such as hydrogen peroxide which would lead to lipid peroxidation. Vitamin E is considered a specific, though indirect, index of in vivo peroxidation. They also showed that a small group of AAA patients had decreased vitamin E levels but not decreased vitamin E/total lipid ratios compared with controls (coronary artery disease and normal patients). Accordingly, the AAA patients may be under increased oxidative stress (e.g., increased inflammation or PMN activation) but do not have decreased concentrations of plasma vitamin E carriers.

This analysis reveals how the biological information associated with AAA pathogenesis constitute the foundation on which can be defined the destructive remodeling of the aortic wall and its influence in AAA rupture.
