Gut Microbiota-Derived Trimethylamine N-Oxide Contributes to Abdominal Aortic Aneurysm Through Inflammatory and Apoptotic Mechanisms.

BACKGROUND: Large-scale human and mechanistic mouse studies indicate a strong relationship between the microbiome-dependent metabolite trimethylamine N-oxide (TMAO) and several cardiometabolic diseases. This study aims to investigate the role of TMAO in the pathogenesis of abdominal aortic aneurysm (AAA) and target its parent microbes as a potential pharmacological intervention. METHODS: TMAO and choline metabolites were examined in plasma samples, with associated clinical data, from 2 independent patient cohorts (N=2129 total). Mice were fed a high-choline diet and underwent 2 murine AAA models, angiotensin II infusion in low-density lipoprotein receptor-deficient (Ldlr-/-) mice or topical porcine pancreatic elastase in C57BL/6J mice. Gut microbial production of TMAO was inhibited through broad-spectrum antibiotics, targeted inhibition of the gut microbial choline TMA lyase (CutC/D) with fluoromethylcholine, or the use of mice genetically deficient in flavin monooxygenase 3 (Fmo3-/-). Finally, RNA sequencing of in vitro human vascular smooth muscle cells and in vivo mouse aortas was used to investigate how TMAO affects AAA. RESULTS: Elevated TMAO was associated with increased AAA incidence and growth in both patient cohorts studied. Dietary choline supplementation augmented plasma TMAO and aortic diameter in both mouse models of AAA, which was suppressed with poorly absorbed oral broad-spectrum antibiotics. Treatment with fluoromethylcholine ablated TMAO production, attenuated choline-augmented aneurysm initiation, and halted progression of an established aneurysm model. In addition, Fmo3-/- mice had reduced plasma TMAO and aortic diameters and were protected from AAA rupture compared with wild-type mice. RNA sequencing and functional analyses revealed choline supplementation in mice or TMAO treatment of human vascular smooth muscle cells-augmented gene pathways associated with the endoplasmic reticulum stress response, specifically the endoplasmic reticulum stress kinase PERK. CONCLUSIONS: These results define a role for gut microbiota-generated TMAO in AAA formation through upregulation of endoplasmic reticulum stress-related pathways in the aortic wall. In addition, inhibition of microbiome-derived TMAO may serve as a novel therapeutic approach for AAA treatment where none currently exist.

Calcitriol Supplementation Protects Against Apoptosis and Alleviates the Severity of Abdominal Aortic Aneurysm Induced by Angiotensin II and Anti-TGFß.

The present study aims to assess the effect of vitamin D deficiency (VDD) and its supplementation on the severity of AAA in mice. AAA was induced by AngII and anti-TGF-ß administration. Animals were divided into four groups: Sham, mice with AAA, mice with AAA, and VDD, and mice with AAA supplemented with calcitriol. Blood pressure, echocardiography, abdominal aortic tissues, and plasma samples were monitored for all groups. VDD was associated with enhanced activity of cleaved MMP-9 and elastin degradation and positively correlated with the severity of AAA. Calcitriol supplementation decreased the INFγ/IL-10 ratio and enhanced the Nrf2 pathway. Moreover, Cu/Zn-superoxide dismutase expression and catalase and neutral sphingomyelinase activity were exacerbated in AAA and VDD groups. Furthermore, calcitriol supplementation showed a significantly lower protein expression of caspase-8, caspase-3, Bid, and t-Bid, and prevented the apoptosis of VSMCs treated by AngII and anti-TGF-ß. Calcitriol supplementation may alleviate AAA severity and could be of great interest in the clinical management of AAA. VDD enhances antioxidant enzymes activity and expression, whereas calcitriol supplementation alleviates AAA severity by re-activating Nrf2 and inhibiting apoptotic pathways.

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.

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.

IL (Interleukin)-33 Suppresses Abdominal Aortic Aneurysm by Enhancing Regulatory T-Cell Expansion and Activity.

Objective- Inflammation occurs during the progression of abdominal aortic aneurysm (AAA). IL (interleukin)-33 is a pleiotropic cytokine with multiple immunomodulatory effects, yet its role in AAA remains unknown. Approach and Results- Immunoblot, immunohistochemistry, and immunofluorescent staining revealed increased IL-33 expression in adventitia fibroblasts from mouse AAA lesions. Daily intraperitoneal administration of recombinant IL-33 or transgenic IL-33 expression ameliorated periaorta CaPO4 injury- and aortic elastase exposure-induced AAA in mice, as demonstrated by blunted aortic expansion, reduced aortic wall elastica fragmentation, enhanced AAA lesion collagen deposition, attenuated T-cell and macrophage infiltration, reduced inflammatory cytokine production, skewed M2 macrophage polarization, and reduced lesion MMP (matrix metalloproteinase) expression and cell apoptosis. Flow cytometry analysis, immunostaining, and immunoblot analysis showed that exogenous IL-33 increased CD4+Foxp3+ regulatory T cells in spleens, blood, and aortas in periaorta CaPO4-treated mice. Yet, ST2 deficiency muted these IL-33 activities. Regulatory T cells from IL-33-treated mice also showed significantly stronger activities in suppressing smooth muscle cell inflammatory cytokine and chemokine expression, macrophage MMP expression, and in increasing M2 macrophage polarization than those from vehicle-treated mice. In contrast, IL-33 failed to prevent AAA and lost its beneficial activities in CaPO4-treated mice after selective depletion of regulatory T cells. Conclusions- Together, this study established a role of IL-33 in protecting mice from AAA formation by enhancing ST2-dependent aortic and systemic regulatory T-cell expansion and their immunosuppressive activities.

Sesame Extract Attenuates the Degradation of Collagen and Elastin Fibers in the Vascular Walls of Nicotine-administered Mice.

Abdominal aortic aneurysm (AAA) is a vascular disease characterized by the weakening of the vascular walls and the progressive dilation of the abdominal aorta. Nicotine, a primary component of cigarette smoke, is associated with AAA development and rupture. Nicotine induces AAA development by weakening vascular walls. However, little is known about preventive methods using functional food factors for nicotine-induced vascular destruction. Sesamin and sesamolin are functional food factors that are fat-soluble lignans found in Sesamum indicum seeds. Previous reports indicated that sesamin and sesamolin have anti-oxidative and anti-inflammatory effects. In this study, we evaluated the effects of sesamin and sesamolin-rich sesame extract on the weakening of vascular walls in nicotine-administered mice. Sesame extract attenuated the degradation of collagen and elastin fibers caused by nicotine. In addition, sesame extract decreased the area positive for matrix metalloproteinase 12 (MMP-12) and oxidative stress in the vascular walls. These results suggest that sesame extract may decrease the weakening of vascular walls by suppressing the nicotine-induced degradation of collagen and elastin fibers. Sesame extract may be effective in preventing AAA development by decreasing both, MMP-12 expression and oxidative stress in vascular walls.

Ginkgo biloba extracts prevent aortic rupture in angiotensin II-infused hypercholesterolemic mice.

Abdominal aortic aneurysms (AAAs) are a chronic vascular disease characterized by pathological luminal dilation. Aortic rupture is the fatal consequence of AAAs. Ginkgo biloba extracts (GBEs), a natural herb extract widely used as food supplements, drugs, and cosmetics, has been reported to suppress development of calcium chloride-induced AAAs in mice. Calcium chloride-induced AAAs do not rupture, while angiotensin II (AngII)-induced AAAs in mice have high rate of aortic rupture, implicating potentially different mechanisms from calcium chloride-induced AAAs. This study aimed to determine whether GBE would improve aortic dilation and rupture rate of AngII-induced AAAs. Male apolipoprotein E (apoE) -/- mice were infused with AngII and administered either GBE or its major active ingredients, flavonoids and ginkgolides, individually or in combination. To determine the effects of GBE in mice with established AAAs, male apoE-/- mice were firstly infused with AngII for 28 days to develop AAAs, and then administered either GBE or vehicle in mice with established AAAs, which were continuously infused with AngII for another 56 days. GBE, but not the two major active components separately or synergistically, prevented aortic rupture, but not aortic dilation. The protection of GBE from aortic rupture was independent of systolic blood pressure, lipid, and inflammation. GBE also did not attenuate either aortic rupture or progressive aortic dilation in mice with established AAAs. GBE did not reduce the atherosclerotic lesion areas, either. In conclusion, GBE prevents aortic rupture in AngII-infused hypercholesterolemic mice, but only in the early phase of the disease development.

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.

Tamsulosin attenuates abdominal aortic aneurysm growth.

BACKGROUND: Tamsulosin, an α1A-adrenergic receptor inhibitor, is prescribed to treat benign prostatic hyperplasia in men >60 years of age, the same demographic most susceptible to abdominal aortic aneurysm. The goal of this study was to investigate the effect of tamsulosin on abdominal aortic aneurysm pathogenesis. METHODS: Abdominal aortic aneurysms were induced in WT C57BL/6 male mice (n = 9-18/group), using an established topical elastase abdominal aortic aneurysm model. Osmotic pumps were implanted in mice 5 days before operation to create the model, administering either low dose (0.125 µg/day tamsulosin), high dose (0.250µg/day tamsulosin), or vehicle treatments with and without topical application of elastase. Blood pressures were measured preoperatively and on postoperative days 0, 3, 7, and 14. On postoperative day 14, aortic diameter was measured before harvest. Sample aortas were prepared for histology and cytokine analysis. RESULTS: Measurements of systolic blood pressure did not differ between groups. Mice treated with the low dose of tamsulosin and with the high dose of tamsulosin showed decreased aortic diameter compared with vehicle-treated control (93% ± 24 versus 94% ± 30 versus 132% ± 24, respectively; P = .0003, P = .0003). Cytokine analysis demonstrated downregulation of pro-inflammatory cytokines in both treatment groups compared with the control (P < .05). Histology exhibited preservation of elastin in both low- and high-dose tamsulosin-treated groups (P = .0041 and P = .0018, respectively). CONCLUSION: Tamsulosin attenuates abdominal aortic aneurysm formation with increased preservation of elastin and decreased production of pro-inflammatory cytokines. Further studies are necessary to elucidate the mechanism by which tamsulosin attenuates abdominal aortic aneurysm pathogenesis.

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.

Role of myeloperoxidase in abdominal aortic aneurysm formation: mitigation by taurine.

Oxidative stress plays a fundamental role in abdominal aortic aneurysm (AAA) formation. Activated polymorphonuclear leukocytes (or neutrophils) are associated with AAA and express myeloperoxidase (MPO), which promotes inflammation, matrix degradation, and other pathological features of AAA, including enhanced oxidative stress through generation of reactive oxygen species. Both plasma and aortic MPO levels are elevated in patients with AAA, but the role of MPO in AAA pathogenesis has, heretofore, never been investigated. Here, we show that MPO gene deletion attenuates AAA formation in two animal models: ANG II infusion in apolipoprotein E-deficient mice and elastase perfusion in C57BL/6 mice. Oral administration of taurine [1% or 4% (wt/vol) in drinking water], an amino acid known to react rapidly with MPO-generated oxidants like hypochlorous acid, also prevented AAA formation in the ANG II and elastase models as well as the CaCl2 application model of AAA formation while reducing aortic peroxidase activity and aortic protein-bound dityrosine levels, an oxidative cross link formed by MPO. Both MPO gene deletion and taurine supplementation blunted aortic macrophage accumulation, elastin fragmentation, and matrix metalloproteinase activation, key features of AAA pathogenesis. Moreover, MPO gene deletion and taurine administration significantly attenuated the induction of serum amyloid A, which promotes ANG II-induced AAAs. These data implicate MPO in AAA pathogenesis and suggest that studies exploring whether taurine can serve as a potential therapeutic for the prevention or treatment of AAA in patients merit consideration.NEW & NOTEWORTHY Neutrophils are abundant in abdominal aortic aneurysm (AAA), and myeloperoxidase (MPO), prominently expressed in neutrophils, is associated with AAA in humans. This study demonstrates that MPO gene deletion or supplementation with the natural product taurine, which can scavenge MPO-generated oxidants, can prevent AAA formation, suggesting an attractive potential therapeutic strategy for AAA.

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.

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.

Antioxidant Vitamin C attenuates experimental abdominal aortic aneurysm development in an elastase-induced rat model.

BACKGROUND: We investigated the hypothesis that an antioxidant, Vitamin C, could attenuate abdominal aortic aneurysm (AAA) development in a rat model. METHODS: An AAA model induced by intraluminal infusion was created in 36 male Sprague Dawley rats, which were randomly distributed into three groups: Sham (saline infused, placebo treated), Control (elastase infused, placebo treated), and Vitamin C (elastase infused, vitamin C treated). Vitamin C and placebo were intraperitoneally injected, initiating 1 wk before the infusion and continuing throughout the study. The aortic dilatation ratio was measured, and aortic tissues were further examined using biochemical and histologic techniques. RESULTS: Vitamin C attenuated the development of AAA, decreasing maximal aortic diameter by 25.8% (P < 0.05) and preserving elastin lamellae (P < 0.05). Vitamin C also decreased 8-hydroxyguanine (a marker of oxidative damage to DNA) and 8-isoprostane content (a marker of oxidative stress) in aortic tissues (P < 0.05, respectively). The proteins of matrix metalloproteinase (MMP)-2, MMP-9, and interleukin 6 were markedly downregulated (P < 0.05, respectively), accompanied with notably reduced messenger RNA expression of tumor necrosis factor-α, MMP-2/9, and interleukin 1ß (P < 0.05, respectively). However, messenger RNA of tissue inhibitors of metalloproteinase-1 and tissue inhibitors of metalloproteinase-2 were both significantly upregulated in Vitamin C group. Vitamin C treatment had no significant effect on systolic blood pressure (P > 0.05). CONCLUSIONS: Vitamin C attenuated AAA development in an elastase-induced rat model via crucial protective effect, which was mediated by an increased level of antioxidant in cooperation with preserving elastin lamellae, inhibiting matrix-degrading proteinases and suppressing inflammatory responses.

Orally administered dipeptidyl peptidase-4 inhibitor (alogliptin) prevents abdominal aortic aneurysm formation through an antioxidant effect in rats.

OBJECTIVE: Dipeptidyl peptidase-4 (DPP-4) inhibitor, a novel antidiabetic drug, has a cardioprotective effect on ischemia-reperfusion injury through an antioxidant effect. However, the effect of DPP-4 inhibitor on aneurysm formation has not been investigated. We aimed to test the hypothesis that the DPP-4 inhibitor, alogliptin, attenuates vascular oxidative stress and thus inhibits abdominal aortic aneurysm (AAA) formation. METHODS: AAAs were created with intraluminal elastase and extraluminal calcium chloride in 36 male rats. Rats were divided into three groups: a low dose of alogliptin group (group LD; 1 mg/kg/d), a high-dose group (group HD; 3 mg/kg/d), and a control group (group C, water). Alogliptin was administered by gastric gavage once daily beginning 3 days before surgery. On day 7 after aneurysm preparation, reactive oxygen species (ROS) expression was semiquantified by dihydroethidium staining, and the oxidation product of DNA produced by ROS, 8-hydroxydeoxyguanosine (8-OHdG), was measured by immunohistochemical staining. Blood glucose concentrations were measured. Hematoxylin and eosin and elastica Van Gieson stainings were performed on day 28, and the AAA dilatation ratio was calculated. RESULTS: On day 7 (six in each group), dihydroethidium staining of the aneurysm wall showed a reduced level of ROS expression (4.6 ± 0.6 in group C, 2.7 ± 0.3 in group LD, and 1.7 ± 0.5 in group HD; P < .0001) and showed fewer 8-OHdG-positive cells in alogliptin-treated samples (138.1 ± 7.4 cells in group C, 102.5 ± 4.5 cells in group LD, and 66.1 ± 4.5 cells in group HD; P < .0001) The treatment significantly reduced messenger RNA expression of matrix metalloproteinases (MMPs) in aneurysm walls (relative expression: MMP-2: 2.1 ± 0.4 in group C, 1.3 ± 0.3 in group LD, and 0.9 ± 0.2 in group HD; P < .001; MMP-9: 2.0 ± 0.5 in group C, 0.3 ± 0.3 in group LD, and 0.3 ± 0.2 in group HD; P < .001). On day 28 (six in each group), the aortic wall in groups LD and HD was less dilated (dilatation ratio: 199.2% ± 11.8% in group C, 159.6% ± 2.8% in group LD, and 147.1% ± 1.9% in group HD; P < .02 group C vs HD) and had higher elastin content than in group C. The difference in blood glucose levels among the three groups was not significant. CONCLUSIONS: The DPP-4 inhibitor, alogliptin, attenuates aneurysm formation and expansion dose-dependently in a rat AAA model via an antioxidative action.

Hyperhomocysteinemia exaggerates adventitial inflammation and angiotensin II-induced abdominal aortic aneurysm in mice.

RATIONALE: A number of epidemiological studies have suggested an association of hyperhomocysteinemia (HHcy) and abdominal aortic aneurysm (AAA), but discrepancies exist. In addition, we lack direct evidence supporting a causal role. OBJECTIVE: We determined the association and contribution of HHcy to AAA formation. METHODS AND RESULTS: We first performed a meta-analysis of studies involving 1489 subjects and found a strong association of HHcy and AAA (odds ratio, 7.39). Next, we used angiotensin II-infused male apolipoprotein E-deficient mice and tested whether HHcy contributes to AAA pathogenesis. Homocysteine (Hcy) supplement (1.8 g/L) in drinking water resulted in mild HHcy. Intriguingly, HHcy greatly increased the incidence of angiotensin II-induced AAA and aortic dissection in apolipoprotein E-deficient mice (vehicle versus Hcy: 50% versus 100%; P<0.05). Histology indicated HHcy markedly exaggerated aortic adventitial inflammation. Increased levels of proinflammatory interleukin-6 and monocyte chemoattractant protein-1 were preferentially colocalized within adventitial fibroblasts in HHcy plus angiotensin II mice, which suggested the importance of adventitial fibroblasts activation in Hcy-aggravated AAA. Hcy sequentially stimulated adventitial fibroblasts transformation into myofibroblasts, secretion of interleukin-6 and monocyte chemoattractant protein-1, and consequent recruitment of monocytes/macrophages to adventitial fibroblasts, which was abolished by the NADPH oxidase inhibitor diphenyliodonium. NADPH oxidase 4, but not other homologs of NADPH oxidase, was significantly upregulated by Hcy in adventitial fibroblasts, whereas NADPH oxidase 4 small interfering RNA silencing diminished Hcy-induced adventitial fibroblasts activation. Finally, folic acid supplement (0.071 µg/g per day) markedly reduced HHcy-aggravated angiotensin II-induced AAA formation in apolipoprotein E-deficient mice. CONCLUSIONS: HHcy may aggravate AAA formation at least partially via activating adventitial fibroblast NADPH oxidase 4.