Ganglioside GM3 Protects Against Abdominal Aortic Aneurysm by Suppressing Ferroptosis.

BACKGROUND: Abdominal aortic aneurysm (AAA) is a potentially life-threatening vascular condition, but approved medical therapies to prevent AAA progression and rupture are currently lacking. Sphingolipid metabolism disorders are associated with the occurrence and development of AAA. It has been discovered that ganglioside GM3, a sialic acid-containing type of glycosphingolipid, plays a protective role in atherosclerosis, which is an important risk factor for AAA; however, the potential contribution of GM3 to AAA development has not been investigated. METHODS: We performed a metabolomics study to evaluated GM3 level in plasma of human patients with AAA. We profiled GM3 synthase (ST3GAL5) expression in the mouse model of aneurysm and human AAA tissues through Western blotting and immunofluorescence staining. RNA sequencing, affinity purification and mass spectrometry, proteomic analysis, surface plasmon resonance analysis, and functional studies were used to dissect the molecular mechanism of GM3-regulating ferroptosis. We conditionally deleted and overexpressed St3gal5 in smooth muscle cells (SMCs) in vivo to investigate its role in AAA. RESULTS: We found significantly reduced plasma levels of GM3 in human patients with AAA. GM3 content and ST3GAL5 expression were decreased in abdominal aortic vascular SMCs in patients with AAA and an AAA mouse model. RNA sequencing analysis showed that ST3GAL5 silencing in human aortic SMCs induced ferroptosis. We showed that GM3 interacted directly with the extracellular domain of TFR1 (transferrin receptor 1), a cell membrane protein critical for cellular iron uptake, and disrupted its interaction with holo-transferrin. SMC-specific St3gal5 knockout exacerbated iron accumulation at lesion sites and significantly promoted AAA development in mice, whereas GM3 supplementation suppressed lipid peroxidation, reduced iron deposition in aortic vascular SMCs, and markedly decreased AAA incidence. CONCLUSIONS: Together, these results suggest that GM3 dysregulation promotes ferroptosis of vascular SMCs in AAA. Furthermore, GM3 may constitute a new therapeutic target for AAA.

Eugenol restrains abdominal aortic aneurysm progression with down-regulations on NF-κB and COX-2.

Abdominal aortic aneurysm (AAA) is a lethal disease without available medicine for treatment. This study aimed to evaluate the efficiency of eugenol (4-allyl-2-methoxyphenol) against AAA and the underlying mechanism. Eugenol is the major bioactive component of clove. A mouse AAA model was established through porcine pancreatic elastase (PPE) incubation peri-adventitially and 1% 3-aminopropanonitrile (BAPN) diet. Continuous AAA progression from day 0 to day 15 was observed after PPE plus BAPN treatment, according to the AAA diameter and histopathological evaluation. Accompanying with AAA progression, sustained increased expressions of CD68, COX-2 and NF-κB were observed through immunofluorescence assay. After elucidation the efficiency of eugenol against AAA progression by AAA diameter, hematoxylin-eosin staining and orcein staining, the down-regulations of eugenol on COX-2 and NF-κB were further detected by immunohistochemistry and western blot. Eugenol not only blocked AAA expansion and protected the integrity of aortic structure in a dose-dependent manner, but also held high oral bioavailability. Excellent efficiency, high oral bioavailability and down-regulation on COX-2/NF-κB endowed eugenol great potential for future AAA therapy.

Rosmarinic Acid Suppresses Abdominal Aortic Aneurysm Progression in Apolipoprotein E-deficient Mice.

An abdominal aortic aneurysm is a life-threatening cardiovascular disorder caused by dissection and rupture. No effective medicine is currently available for the > 90% of patients whose aneurysms are below the surgical threshold. The present study investigated the impact of rosmarinic acid, salvianolic acid C, or salvianolic acid B on experimental abdominal aortic aneurysms. Abdominal aortic aneurysms were induced in apolipoprotein E-deficient mice via infusion of angiotensin II for 4 wks. Rosmarinic acid, salvianolic acid C, salvianolic acid B, or doxycycline as a positive control was provided daily through intraperitoneal injection. Administration of rosmarinic acid was found to decrease the thickness of the aortic wall, as determined by histopathological assay. Rosmarinic acid also exhibited protection against elastin fragmentation in aortic media and down-regulated cell apoptosis and proliferation in the aortic adventitia. Infiltration of macrophages, T lymphocytes, and neutrophils in aortic aneurysms was found, especially at the aortic adventitia. Rosmarinic acid, salvianolic acid C, or salvianolic acid B inhibited the infiltration on macrophages specifically, but these compounds did not influence T lymphocytes and neutrophils. Expression of matrix metalloproteinase 9 and macrophage migration inhibitory factor significantly increased in aortic aneurysms. Rosmarinic acid and salvianolic acid C decreased the expression of matrix metalloproteinase-9 in media, and rosmarinic acid also tended to reduce migration inhibitory factor expression. Further then, partial least squares-discriminate analysis was used to classify metabolic changes among different treatments. Rosmarinic acid affected most of the metabolites in the biosynthesis of the citrate cycle, fatty acid pathway significantly. Our present study on mice demonstrated that rosmarinic acid inhibited multiple pathological processes, which were the key features important in abdominal aortic aneurysm formation. Further study on rosmarinic acid, the novel candidate for aneurysmal therapy, should be undertaken to determine its potential for clinical use.

Factor Xa inhibitor rivaroxaban suppresses experimental abdominal aortic aneurysm progression via attenuating aortic inflammation.

OBJECTIVE: Rivaroxaban is a specific factor Xa (FXa) inhibitor for venous thromboembolism treatment. Recently, increasing evidence have reported the beneficial effects of rivaroxaban on treating cardiovascular disorders such as coronary and peripheral artery disease. However, its potential influence on abdominal aortic aneurysm (AAA) remains unclear. This study aims to investigate whether rivaroxaban treatment could attenuate experimental AAA progression and its related mechanisms. APPROACHES AND RESULTS: In human aneurysmal aorta, FXa protein expression was significantly upregulated. Further investigations identified a positive correlation among plasma FXa level, AAA severity (the maximal aortic diameter), and intra-aneurysmal thrombus percentage. In Ang II (angiotensin II)-infused ApoE-/- mice, the administration of high dose rivaroxaban (15 mg/kg/d) for 14 days significantly reduced the maximal aortic diameter, while low dose rivaroxaban (5 mg/kg/d) did not display such a protective role. Although rivaroxaban treatments reduced the incidence of AAA and thrombus formation, these differences did not reach statistical significance. Immunohistochemistry revealed a pronounced aortic remodeling including increased collagen content and enhanced elastin degradation in Ang II-induced AAAs, which was inhibited by high dose rivaroxaban treatment. Further analysis demonstrated that rivaroxaban exerted its protective effects by decreasing leukocyte infiltration, inflammatory cytokines expression, and matrix metalloproteinases (MMPs) expression in the aortic wall. The inhibitory effect of rivaroxaban on aneurysm development was also observed in calcium chloride-induced AAA model. Mechanistically, in human aortic endothelial cells, FXa stimulation increased the expression of inflammatory cytokines (interleukin (IL)-1ß, IL-6, IL-8, monocyte chemoattractant protein-1) and adhesive molecules, which were all reversed by the cotreatment of rivaroxaban. Subsequent monocyte-endothelial cell interaction was enhanced after FXa stimulation and was alleviated by rivaroxaban cotreatment. In addition, FXa induced a significantly heightened expression of MMP2 in human aortic endothelial cells, which was ameliorated by rivaroxaban coadministration. CONCLUSIONS: Rivaroxaban attenuated both angiotensin II- and calcium chloride-induced abdominal aortic aneurysm (AAA) progressions, through inhibiting aortic remodeling and inflammation. Rivaroxaban could be a promising therapeutic agent in attenuating AAA development by counteracting FXa-induced aortic wall inflammation.

Cyclodextrin Prevents Abdominal Aortic Aneurysm via Activation of Vascular Smooth Muscle Cell Transcription Factor EB.

BACKGROUND: Abdominal aortic aneurysm (AAA) is a severe aortic disease with a high mortality rate in the event of rupture. Pharmacological therapy is needed to inhibit AAA expansion and prevent aneurysm rupture. Transcription factor EB (TFEB), a master regulator of autophagy and lysosome biogenesis, is critical to maintain cell homeostasis. In this study, we aim to investigate the role of vascular smooth muscle cell (VSMC) TFEB in the development of AAA and establish TFEB as a novel target to treat AAA. METHODS: The expression of TFEB was measured in human and mouse aortic aneurysm samples. We used loss/gain-of-function approaches to understand the role of TFEB in VSMC survival and explored the underlying mechanisms through transcriptome and functional studies. Using VSMC-selective Tfeb knockout mice and different mouse AAA models, we determined the role of VSMC TFEB and a TFEB activator in AAA in vivo. RESULTS: We found that TFEB is downregulated in both human and mouse aortic aneurysm lesions. TFEB potently inhibits apoptosis in VSMCs, and transcriptome analysis revealed that TFEB regulates apoptotic signaling pathways, especially apoptosis inhibitor B-cell lymphoma 2. B-cell lymphoma 2 is significantly upregulated by TFEB and is required for TFEB to inhibit VSMC apoptosis. We consistently observed that TFEB deficiency increases VSMC apoptosis and promotes AAA formation in different mouse AAA models. Furthermore, we demonstrated that 2-hydroxypropyl-ß-cyclodextrin, a clinical agent used to enhance the solubility of drugs, activates TFEB and inhibits AAA formation and progression in mice. Last, we found that 2-hydroxypropyl-ß-cyclodextrin inhibits AAA in a VSMC TFEB-dependent manner in mouse models. CONCLUSIONS: Our study demonstrated that TFEB protects against VSMC apoptosis and AAA. TFEB activation by 2-hydroxypropyl-ß-cyclodextrin may be a promising therapeutic strategy for the prevention and treatment of AAA.

Niacin protects against abdominal aortic aneurysm formation via GPR109A independent mechanisms: role of NAD+/nicotinamide.

AIMS: Chronic adventitial and medial infiltration of immune cells play an important role in the pathogenesis of abdominal aortic aneurysms (AAAs). Nicotinic acid (niacin) was shown to inhibit atherosclerosis by activating the anti-inflammatory G protein-coupled receptor GPR109A [also known as hydroxycarboxylic acid receptor 2 (HCA2)] expressed on immune cells, blunting immune activation and adventitial inflammatory cell infiltration. Here, we investigated the role of niacin and GPR109A in regulating AAA formation. METHODS AND RESULTS: Mice were supplemented with niacin or nicotinamide, and AAA was induced by angiotensin II (AngII) infusion or calcium chloride (CaCl2) application. Niacin markedly reduced AAA formation in both AngII and CaCl2 models, diminishing adventitial immune cell infiltration, concomitant inflammatory responses, and matrix degradation. Unexpectedly, GPR109A gene deletion did not abrogate the protective effects of niacin against AAA formation, suggesting GPR109A-independent mechanisms. Interestingly, nicotinamide, which does not activate GPR109A, also inhibited AAA formation and phenocopied the effects of niacin. Mechanistically, both niacin and nicotinamide supplementation increased nicotinamide adenine dinucleotide (NAD+) levels and NAD+-dependent Sirt1 activity, which were reduced in AAA tissues. Furthermore, pharmacological inhibition of Sirt1 abrogated the protective effect of nicotinamide against AAA formation. CONCLUSION: Niacin protects against AAA formation independent of GPR109A, most likely by serving as an NAD+ precursor. Supplementation of NAD+ using nicotinamide-related biomolecules may represent an effective and well-tolerated approach to preventing or treating AAA.

Involvement of macrophage-derived exosomes in abdominal aortic aneurysms development.

BACKGROUND AND AIMS: Abdominal aortic aneurysm (AAA) is characterized by infiltration of inflammatory cells, extracellular matrix (ECM) degradation, and dysfunction of vascular smooth muscle cells (VSMCs). Recent studies reported that exosomes mediate intercellular communication and are involved in different diseases. Whether exosomes play a role in AAA is poorly understood. Hence, this study evaluated the function of exosomes in AAA development. METHODS: The presence of exosomes in human and calcium phosphate (CaPO4)-induced AAA tissues was determined by immunofluorescence staining of CD63 and Alix. GW4869, an inhibitor of exosome biogenesis, was intraperitoneally injected into CaPO4-induced AAA tissues to evaluate the effects of exosomal inhibition on AAA development. To explore the underlying mechanisms, the human monocytic cell line THP-1 was differentiated into macrophages, and exosomes were collected from macrophages. VSMCs were treated with macrophage-derived exosomes, and the expression of matrix metalloproteinase-2 (MMP-2) was evaluated. The activation of mitogen-activated protein kinases (MAPKs) pathways was also investigated in vitro and in vivo. RESULTS: Exosomes were detected in the adventitia of aneurysmal tissues obtained from humans and mice. They were mainly expressed in clusters of macrophages. Intraperitoneal injection of GW4869 for two weeks significantly attenuated the progression of CaPO4-induced AAA, preserved elastin integrity and decreased MMP-2 expression. Similarly, administration of GW4869 suppressed the systemic and aneurysmal exosome generation. In vitro, treatment with macrophage-derived exosomes elevated MMP-2 expression in human VSMCs, while pre-treatment with GW4869 abolished these effects. It was also found that JNK and p38 pathways mediated the production of MMP-2 in VSMCs following treatment with macrophage-derived exosomes. CONCLUSIONS: This study suggests that exosomes derived from macrophages are involved in the pathogenesis of AAA. Macrophage-derived exosomes trigger MMP-2 expression in VSMC via JNK and p38 pathways. GW4869 supplementation attenuates CaPO4-induced AAA in mice.

Omega-3 fatty acids decrease oxidative stress and inflammation in macrophages from patients with small abdominal aortic aneurysm.

Abdominal aortic aneurysm (AAA) is associated with inflammation and oxidative stress, the latter of which contributes to activation of macrophages, a prominent cell type in AAA. Omega-3 polyunsaturated fatty acids (n-3 PUFAs) have been reported to limit oxidative stress in animal models of AAA. The aim of this study was to evaluate the effect of the n-3 PUFA docosahexaenoic acid (DHA) on antioxidant defence in macrophages from patients with AAA. Cells were obtained from men with small AAA (diameter 3.0-4.5 cm, 75 ± 6 yr, n = 19) and age- matched male controls (72 ± 5 yr, n = 41) and incubated with DHA for 1 h before exposure to 0.1 µg/mL lipopolysaccharide (LPS) for 24 h. DHA supplementation decreased the concentration of tumour necrosis factor-α (TNF-α; control, 42.1 ± 13.6 to 5.1 ± 2.1 pg/ml, p < 0.01; AAA, 25.2 ± 9.8 to 1.9 ± 0.9 pg/ml, p < 0.01) and interleukin-6 (IL-6; control, 44.9 ± 7.7 to 5.9 ± 2.0 pg/ml, p < 0.001; AAA, 24.3 ± 5.2 to 0.5 ± 0.3 pg/ml, p < 0.001) in macrophage supernatants. DHA increased glutathione peroxidase activity (control, 3.2 ± 0.3 to 4.1 ± 0.2 nmol/min/ml/µg protein, p = 0.004; AAA, 2.3 ± 0.5 to 3.4 ± 0.5 nmol/min/ml/µg protein, p = 0.008) and heme oxygenase-1 mRNA expression (control, 1.5-fold increase, p < 0.001). The improvements in macrophage oxidative stress status serve as a stimulus for further investigation of DHA in patients with AAA.

n-3 PUFAs improve erythrocyte fatty acid profile in patients with small AAA: a randomized controlled trial.

Abdominal aortic aneurysm (AAA) is an important cause of death in older adults, which has no current drug therapy. Inflammation and abnormal redox status are believed to be key pathogenic mechanisms for AAA. In light of evidence correlating inflammation with aberrant fatty acid profiles, this study compared erythrocyte fatty acid content in 43 AAA patients (diameter 3.0-4.5 cm) and 52 healthy controls. In addition, the effect of omega-3 PUFA (n-3 PUFA) supplementation on erythrocyte fatty acid content was examined in a cohort of 30 AAA patients as part of a 12 week randomized placebo-controlled clinical trial. Blood analyses identified associations between AAA and decreased linoleic acid (LA), and AAA and increased Δ6-desaturase activity and biosynthesis of arachidonic acid (AA) from LA. Omega-3 PUFA supplementation (1.5 g DHA + 0.3 g EPA/day) decreased red blood cell distribution width (14.8 ± 0.4% to 13.8 ± 0.2%; P = 0.003) and levels of pro-inflammatory n-6 PUFAs (AA, 12.46 ± 0.23% to 10.14 ± 0.3%, P < 0.001; adrenic acid, 2.12 ± 0.13% to 1.23 ± 0.09%; P < 0.001). In addition, Δ-4 desaturase activity increased (DHA/docosapentaenoic acid ratio, 1.85 ± 0.14 to 3.93 ± 0.17; P < 0.001) and elongase 2/5 activity decreased (adrenic acid/AA ratio, 0.17 ± 0.01 to 0.12 ± 0.01; P < 0.01) following supplementation. The findings suggest that n-3 PUFAs improve fatty acid profiles and ameliorate factors associated with inflammation in AAA patients.

Inhibitory effects of cycloastragenol on abdominal aortic aneurysm and its related mechanisms.

BACKGROUND AND PURPOSE: Abdominal aortic aneurysm (AAA) is a degenerative disease affecting human health, but there are no safe and effective medications for AAA therapy. Cycloastragenol (CAG), derived from Astragali Radix, has various pharmacological effects. However, whether CAG can protect against AAA remains elusive. In this study, we investigated whether CAG has an inhibitory effect on AAA and its related mechanism. EXPERIMENTAL APPROACH: The AAA mouse model was induced by incubating the abdominal aorta with elastase. CAG was administered by gavage at different doses beginning on the same day or 14 days after inducing AAA to explore its preventive or therapeutic effects respectively. The preventive effects of CAG on AAA were verified in another AAA mouse model induced by angiotensin II in ApoE-/- mouse. In vitro experiments were implemented on rat vascular smooth muscle cells (VSMCs) stimulated by TNF-α. KEY RESULTS: Compared to the control AAA model group, CAG (125 mg·kg-1 body weight day-1 ) reduced the incidence of AAA, the dilatation of aorta and elastin degradation in media in both mouse models of AAA. CAG suppressed the inflammation, oxidation, phenotype switch and apoptosis in TNF-α-stimulated VSMCs, ameliorated the expression and activity of MMPs and decreased the activation of the ERK/JNK signalling pathway. CAG also inhibited the degradation of elastin in TNF-α-stimulated VSMCs. CONCLUSION AND IMPLICATIONS: CAG presents protective effects against AAA through down-regulation of the MAPK signalling pathways and thus attenuates inflammation, oxidation, VSMC phenotype switch and apoptosis and the expression of MMPs as well as increasing elastin biosynthesis.

Dietary DNA Attenuates the Degradation of Elastin Fibers in the Aortic Wall in Nicotine-Administrated Mice.

Abdominal aortic aneurysm (AAA) is a vascular disease characterized by chronic inflammation in the infrarenal aorta. Epidemiologic data have clearly linked tobacco smoking to aneurysm formation and a faster rate of expansion. It suggested that nicotine, one of the main ingredients of tobacco, has been suggested to be associated with AAA development and rupture. In the condition where no established drugs are available; therefore, an effective approach to prevent the vascular damage from nicotine consumption may be the use of dietary functional food factors. However, little is known about the relationship between dietary components and AAA. In this study, we estimated the effect of dietary deoxyribonucleic acid (DNA) on the vascular wall. After habituation for 5 d, the mice were divided into four groups: control diet and distilled water group (C), DNA-Na diet and distilled water group (DNA), control diet and 0.5 mg/mL nicotine solution group (C-Nic), DNA-Na diet, and 0.5 mg/mL nicotine solution group (DNA-Nic). The dietary DNA attenuated the degradation of elastin fibers induced by nicotine administration. The areas stained positive for MMP-2 in the DNA-Nic group were significantly suppressed compared to C-Nic mice. These data suggest that the dietary DNA may prevent the weakening of the aortic wall via inhibition of the MMP-2-dependent pathway. In conclusion, we have revealed the protective effect of dietary DNA on the vascular pathology of nicotine-administrated mice. A nucleic acid-rich diet might be useful for people who consume nicotine via smoking, chewing tobacco, or nicotine patches.

A diet enriched with tree nuts reduces severity of atherosclerosis but not abdominal aneurysm in angiotensin II-infused apolipoprotein E deficient mice.

BACKGROUND AND AIMS: Diets enriched with tree nuts have been demonstrated to reduce the risk of atherosclerosis-related cardiovascular events. Abdominal aortic aneurysm (AAA) shares common risk factors with atherosclerosis and AAA patients commonly have atherosclerosis related cardiovascular events. AAA has some distinct pathological and clinical characteristics to those of atherosclerosis. No previous study has examined the effect of a diet enriched with tree nuts on experimental or clinical AAA. This study investigated the effect of a diet enriched with tree nuts on the development and severity of AAA within an experimental rodent model. METHODS: Male apolipoprotein E deficient mice were allocated to a diet enriched with tree nuts or control diet for 56 days (n = 17 per group). After 28 days, all mice were infused with angiotensin II whilst being maintained on their respective diets. The primary outcome was AAA severity assessed by the supra-renal aortic diameter, measured by ultrasound and ex vivo morphometric analysis. The severity of atherosclerosis was assessed by computer-aided analysis of Sudan IV stained aortic arches and sections of brachiocephalic arteries prepared with Van Gieson's stain. RESULTS: The diet enriched with tree nuts did not influence aortic diameter or aortic rupture incidence. Mice receiving the diet enriched with tree nuts had significantly less atherosclerosis within the brachiocephalic artery (p = 0.033) but not in the aortic arch. CONCLUSIONS: This experimental study suggests that a diet enriched with tree nuts does not reduce the severity of AAA, but does reduce the severity of atherosclerosis within the brachiocephalic artery. The study was not powered to identify a moderate effect of the diet on the primary outcome and therefore this cannot be excluded.

Effects of osteoprotegerin/TNFRSF11B in two models of abdominal aortic aneurysms.

Osteoprotegerin (OPG), additionally termed tumor necrosis factor receptor superfamily member 11B, is produced by vascular smooth muscle cells (VSMCs) and endothelial cells in the vasculature, and its release may be modulated by pro­inflammatory cytokines, including interleukin­1ß and tumor necrosis factor­α. The present study investigated the effects of treatment with low­dose human recombinant OPG on abdominal aortic aneurysm (AAA) development in mice. Mice were treated with 1 µg human recombinant OPG four times (or vehicle) for 2 weeks prior to inducing AAA. A total of two different models for inducing AAA were used to investigate the hypothesis as to whether OPG is involved in key events of AAA development, using osmotic mini­pumps with angiotensin II in apolipoprotein­E (ApoE­/­) mice for 28 days or using periaortic application of CaCl2 on the aorta in C57Bl/6J mice for 14 days. OPG was continuously administered during the experimental period. Histological staining using Masson's trichrome, Verhoeff's van­Gieson and picro­sirius red, in addition to reverse transcription­quantitative polymerase chain reaction analysis of various markers, were used to analyze phenotypic alterations. Treatment with OPG had no inhibitory effect on AAA development in the angiotensin II model in ApoE­/­ mice, which developed suprarenal aneurysms, although it increased vessel wall thickness of the aorta and total collagen in C57Bl/6J mice using the CaCl2 model that induced infrarenal dilation of the aorta. Treatment with OPG did not inhibit aneurysm development and key events, including inflammation, extracellular matrix or VSMC remodeling, in aortas from OPG­treated mice with periaortic treatment with CaCl2. The results indicated that mice treated with low levels of human recombinant OPG may have a more stable aneurysmal phenotype due to compensatory production of collagen and increased vessel wall thickness of the aorta, potentially protecting the aneurysm from rupture. Further studies investigating rupture models of AAA in addition to using higher levels of OPG are require to verify this speculation. Furthermore, treatment with low levels of OPG in patients with AAA may represent a novel therapeutic strategy for the treatment of AAA as well as attenuate the adverse effects associated with the administration of normal and high dosages of OPG.

Translational Relevance and Recent Advances of Animal Models of Abdominal Aortic Aneurysm.

Human abdominal aortic aneurysm (AAA) pathophysiology is not yet completely understood. In conductance arteries, the insoluble extracellular matrix, synthesized by vascular smooth muscle cells, assumes the function of withstanding the intraluminal arterial blood pressure. Progressive loss of this function through extracellular matrix proteolysis is a main feature of AAAs. As most patients are now treated via endovascular approaches, surgical AAA specimens have become rare. Animal models provide valuable complementary insights into AAA pathophysiology. Current experimental AAA models involve induction of intraluminal dilation (nondissecting AAAs) or a contained intramural rupture (dissecting models). Although the ideal model should reproduce the histological characteristics and natural history of the human disease, none of the currently available animal models perfectly do so. Experimental models try to represent the main pathophysiological determinants of AAAs: genetic or acquired defects in extracellular matrix, loss of vascular smooth muscle cells, and innate or adaptive immune response. Nevertheless, most models are characterized by aneurysmal stabilization and healing after a few weeks because of cessation of the initial stimulus. Recent studies have focused on ways to optimize existing models to allow continuous aneurysmal growth. This review aims to discuss the relevance and recent advances of current animal AAA models. VISUAL OVERVIEW: An online visual overview is available for this article.

Prevention of abdominal aortic aneurysm progression by oral administration of green tea polyphenol in a rat model.

OBJECTIVE: Inflammation-mediated elastin destruction in the aortic medial layer is related to progression of abdominal aortic aneurysm (AAA). Epigallocatechin-3-gallate (EGCG), a major component of green tea polyphenols, reportedly increases elastin synthesis in vitro and may possess anti-inflammatory effects. We used a rat model to investigate whether EGCG could prevent AAA progression. METHODS: AAA was induced with administration of intraluminal elastase and extraluminal CaCl2 in male rats. Rats were randomly divided into a control group (n = 30) and an EGCG group (n = 30). In the EGCG group, an EGCG solution (20 mg/d) was administered orally to each rat from 2 weeks before AAA induction and continued 4 weeks beyond induction. RESULTS: The abdominal aortic diameter was significantly smaller in the EGCG group than in the control group on day 28 (2.9 ± 0.2 vs 2.3 ± 0.1 mm; P < .0001). The medial layer wall thickness and elastin content were significantly greater in the EGCG group than in the control group on day 28 (68.4 ± 13.6 vs 46.7 ± 13.4 µm [P < .001] and 20.3 ± 4.6 vs 9.5 ± 3.6% [P < .0001], respectively). Gene expression levels of tropoelastin and lysyl oxidase were significantly higher in the EGCG group immediately before AAA induction, indicating promoted elastoregeneration by EGCG administration (tropoelastin: 0.59 ± 0.36 control vs 1.24 ± 0.36 EGCG [P < .05], lysyl oxidase: 0.77 ± 0.45 control vs 1.34 ± 0.4 EGCG [P < .05]) (fold increase). Gene expression levels of inflammatory cytokines, including tumor necrosis factor-α and interleukin-1ß, were significantly downregulated in the EGCG group (1.82 ± 0.71 vs 0.97 ± 0.59 [P < .05] and 3.91 ± 3.24 vs 0.89 ± 0.59 [P < .05], respectively). On day 7, gene expression levels and gelatinolytic activity of matrix metalloproteinase 9 were significantly lower in the EGCG group (1.41 ± 0.86 vs 0.51 ± 0.42 [P < .05] and 1.00 ± 0.17 vs 0.29 ± 0.12 [P < .0001], respectively), whereas gene expression levels of tissue inhibitors of metalloproteinase-1 were significantly higher in the EGCG group (0.96 ± 0.11 vs 1.14 ± 0.09; P < .05). CONCLUSIONS: EGCG attenuated AAA progression in a rat model by preserving the aortic thickness and elastin content of the medial layer through regeneration of elastin, as mediated by anti-inflammatory effects, and subsequent reduction of matrix metalloproteinase activity.

The preventive effect of fish oil on abdominal aortic aneurysm development.

Abdominal aortic aneurysm (AAA) is a vascular disease involving gradual dilation of the abdominal aorta and high rupture-related mortality rates. AAA is histologically characterized by oxidative stress, chronic inflammation, and extracellular matrix degradation in the vascular wall. We previously demonstrated that aortic hypoperfusion could cause the vascular inflammation and AAA formation. However, the preventive method for hypoperfusion-induced AAA remains unknown. In this study, we evaluated the effect of fish oil on AAA development using a hypoperfusion-induced AAA animal model. Dilation of the abdominal aorta in the fish oil administration group was smaller than in the control group. Collagen destruction and oxidative stress were suppressed in the fish oil administration group than in the control group. These results suggested that fish oil could prevent the development of AAA induced by hypoperfusion.

Zinc Prevents Abdominal Aortic Aneurysm Formation by Induction of A20-Mediated Suppression of NF-κB Pathway.

Chronic inflammation and degradation of elastin are the main processes in the development of abdominal aortic aneurysm (AAA). Recent studies show that zinc has an anti-inflammatory effect. Based on these, zinc may render effective therapy for the treatment of the AAA. Currently, we want to investigate the effects of zinc on AAA progression and its related molecular mechanism. Rat AAA models were induced by periaortic application of CaCl2. AAA rats were treated by daily intraperitoneal injection of ZnSO4 or vehicle alone. The aorta segments were collected at 4 weeks after surgery. The primary rat aortic vascular smooth muscle cells (VSMCs) were stimulated with TNF-α alone or with ZnSO4 for 3 weeks. The results showed that zinc supplementation significantly suppressed the CaCl2-induced expansion of the abdominal aortic diameter, as well as a preservation of medial elastin fibers in the aortas. Zinc supplementation also obviously attenuated infiltration of the macrophages and lymphocytes in the aortas. In addition, zinc reduced MMP-2 and MMP-9 production in the aortas. Most importantly, zinc treatment significantly induced A20 expression, along with inhibition of the NF-κB canonical signaling pathway in vitro in VSMCs and in vivo in rat AAA. This study demonstrated, for the first time, that zinc supplementation could prevent the development of rat experimental AAA by induction of A20-mediated inhibition of the NF-κB canonical signaling pathway.

Inhibiting the Th17/IL-17A-related inflammatory responses with digoxin confers protection against experimental abdominal aortic aneurysm.

OBJECTIVE: T helper 17 cells and interleukin-17A have been implicated in the progression of abdominal aortic aneurysm (AAA). Retinoic acid-related orphan receptor gamma thymus, the master transcription factor of T helper 17 cell differentiation, is selectively antagonized by digoxin. However, the effect of antagonizing retinoic acid-related orphan receptor gamma thymus on AAA has not been investigated. APPROACH AND RESULTS: We used human aortic sample analysis and 2 different experimental AAA models: (a) Angiotensin II (Ang II)-induced ApoE(-/-) male mice (Ang II/APOE model) and (b) porcine pancreatic elastase perfusion C57BL/6 mice (porcine pancreatic elastase/C57 model). In the Ang II/APOE model, all mice (n=80) were divided into 4 groups: sham group (saline+0.5% dimethyl sulfoxide treatment), control group (Ang II+0.5% dimethyl sulfoxide treatment), low-dose group (Ang II+low-dose digoxin, 20 µg/d per mouse), and high-dose group (Ang II+high-dose digoxin, 40 µg/d per mouse). All treatments began on day 0 after surgery. Efficacy was determined via aortic diameter and systolic blood pressure measurements, histopathology and protein expression, and flow cytometry analysis when euthenized. Human aortic tissue analysis showed that both interleukin-17A and retinoic acid-related orphan receptor gamma thymus increased in AAA tissues. The low-dose and high-dose groups had AAA incidences of 60% and 35%, respectively, compared with 70% in the control group. The T helper 17- and interleukin-17A-related inflammatory responses were dose-dependently attenuated by digoxin treatment. Digoxin was also highly effective in the porcine pancreatic elastase/C57 model. CONCLUSIONS: Digoxin attenuates experimental AAA progression in a model-independent manner. Antagonizing retinoic acid-related orphan receptor gamma thymus activity by digoxin may become a novel strategy for nonsurgical AAA treatment.

Suppression of experimental abdominal aortic aneurysms in the mice by treatment with Ginkgo biloba extract (EGb 761).

ETHNOPHARMACOLOGICAL RELEVANCE: Ginkgo biloba extract (EGb 761) is widely used to treat cerebral disorders. Clinical trials have demonstrated therapeutic benefits of EGb 761 in various vascular diseases. Because the potential pathophysiological mechanisms appear similar to those involved in aneurysmal degeneration, we postulated that EGb 761 might affect the development and progression of experimental abdominal aortic aneurysm (AAA). This study was aimed to investigate whether EGb 761 influences the development of experimental AAAs, and to explore the underlying mechanisms. MATERIAL AND METHODS: C57/BL6 mice underwent abluminal application of CaCl2 to the abdominal aorta followed by gavages with either 200mg/kg EGb 761 per day or vehicle. Six weeks after AAA induction, aortic tissue was excised for further examinations. RESULTS: EGb 761 treatment reduced the aneurysm size compared with vehicle-treated controls. EGb 761 had no effect on hemodynamics or macrophage infiltration in the aortic wall. However, nuclear factor κB protein levels were decreased in the aortas of EGb 761 treated animals. The increased ROS production, SOD and CAT activities, and mRNA expression of p47phox nicotinamide adenine dinucleotide phosphate oxidase were attenuated by EGb 761 treatment. Moreover, administration of EGb 761 preserved the destruction of the wavy morphology of the elastin during AAA formation. Zymographic activity of matrix metalloproteinase (MMP)-9 and MMP-2 was lowered in EGb 761 treated mice. CONCLUSIONS: These results suggest that treatment with EGb 761 in mice prevented the development of CaCl2-induced AAA. The possible mechanisms include decreased oxidative damage and inflammation, preservation of aortic wall architecture, and altered MMPs activities.

[Effects of buyang huanwu decoction on the sarcoplasmic reticulum calcium uptake in abdominal aortic constriction induced myocardial hypertrophic rats].

UNLABELLED: objective: To investigate effects of buyang huanwu decoction (BYHWD) on the rats' myocardial hypertrophic model induced by abdominal aortic constriction, and to clarify the molecular regulatory mechanisms for sarcoplasmic reticulum calcium uptake. METHODS: Hypertrophic myocardium rat model was induced by abdominal aorta constriction (AAC). Four weeks after modeling, rats were randomly divided into the sham-operation group (Group S), the AAC model group (Group M), the Enalapril group (Group E), and the BYHWD treatment group (Group BYHWD), respectively. The left ventricular systolic pressure (LVSP), left ventricular end-diastolic pressure (LVEDP), + dp/dtmax,-dp/dtmax, cardiac output (CO), heart mass index (HMI), and left ventricular mass index (LVMI) were observed in each group after 12-week medication. The serum contents of the atrial natriuretic peptide (ANP) and brain natriuretic peptide (BNP) were detected using ELISA. The SERCA2a activity, the ex- pressions of SERCA2a, phospholamban (PLN), and PLN phosphorylation were observed finally. RESULTS: Compared with Group S, LVSP and LVEDP significantly increased,-dp/dtmax and CO obviously decreased, the myocardial tissue was obviously thickened, the serum contents of ANP and BNP increased, the activity and expression of SERCA2a decreased, the SERCA2a/PLN ratio and PLN phosphorylation degree decreased in Group M (all P <0.05). Compared with Group M, LVEDP obviously decreased, -dp/dtmax and CO obviously increased, the hypertrophy myocardial tissue was obviously lessened, the serum contents of ANP and BNP decreased, the activity of SERCA2a increased, the relative expression contents of SERCA2a, Ser16, and Thrl7 were elevated in Group BYHWD (all P <0.05). BYHWD had significant roles in elevating the SERCA2a/PLN ratio and PLN phosphorylation degree (P <0.05). CONCLUSION: BYHWD could significantly improve hemodynamics of heart failure rats, elevate CO, lessen cardiac hypertrophy, and improve the capabilities for sarcoplasmic reticulum calcium uptake.