Group 2 Innate Lymphoid Cells Protect Mice from Abdominal Aortic Aneurysm Formation via IL5 and Eosinophils.

Development of abdominal aortic aneurysms (AAA) enhances lesion group-2 innate lymphoid cell (ILC2) accumulation and blood IL5. ILC2 deficiency in Rorafl/fl Il7rCre/+ mice or induced ILC2 depletion in Icosfl-DTR-fl/+ Cd4Cre/+ mice expedites AAA growth, increases lesion inflammation, but leads to systemic IL5 and eosinophil (EOS) deficiency. Mechanistic studies show that ILC2 protect mice from AAA formation via IL5 and EOS. IL5 or ILC2 from wild-type (WT) mice, but not ILC2 from Il5-/- mice induces EOS differentiation in bone-marrow cells from Rorafl/fl Il7rCre/+ mice. IL5, IL13, and EOS or ILC2 from WT mice, but not ILC2 from Il5-/- and Il13-/- mice block SMC apoptosis and promote SMC proliferation. EOS but not ILC2 from WT or Il5-/- mice block endothelial cell (EC) adhesion molecule expression, angiogenesis, dendritic cell differentiation, and Ly6Chi monocyte polarization. Reconstitution of WT EOS and ILC2 but not Il5-/- ILC2 slows AAA growth in Rorafl/fl Il7rCre/+ mice by increasing systemic EOS. Besides regulating SMC pathobiology, ILC2 play an indirect role in AAA protection via the IL5 and EOS mechanism.

Dapagliflozin Ameliorates the Formation and Progression of Experimental Abdominal Aortic Aneurysms by Reducing Aortic Inflammation in Mice.

BACKGROUND: Dapagliflozin, a sodium glucose transporter protein-2 (SGLT-2) inhibitor, reduces the risk for cardiovascular diseases. However, the influence of dapagliflozin on nondissecting abdominal aortic aneurysms (AAAs) remains unclear. METHODS: AAAs were created in male C57BL/6 mice via intra-aortic porcine pancreatic elastase (PPE) infusion. Mice were daily treated with dapagliflozin (1 or 5 mg/kg body weight) or an equal volume of vehicle through oral gavage beginning one day prior to PPE infusion for 14 days. To investigate its translational value, dapagliflozin or vehicle was also administered to mice with existing AAAs in another cohort. Aortic diameters were measured prior to (day 0 for baseline) and 14 days after PPE infusion. After sacrifice, mice aortae were collected, and following histological analyses were performed. RESULTS: Dapagliflozin treatment significantly reduced aneurysmal aortic expansion following PPE infusion as compared to vehicle treatment especially at 5 mg/kg body weight (approximately 21% and 33% decreases in 1 and 5 mg/kg treatment groups, respectively). The dose-dependent attenuation of AAAs by dapagliflozin was also confirmed on histological analyses. Dapagliflozin remarkably reduced aortic accumulation of macrophages, CD4+ T cells, and B cells particularly following dapagliflozin treatment at 5 mg/kg. Dapagliflozin treatment also markedly attenuated medial SMC loss. Though the difference was not significant, dapagliflozin treatment tended to attenuate CD8+ T cells and elastin degradation. Dapagliflozin treatment at 5 mg/kg caused a 53% reduction in neovessel density. Furthermore, dapagliflozin treatment mitigated further progress of existing AAAs. CONCLUSION: Dapagliflozin treatment ameliorated PPE-induced AAAs by inhibiting aortic leukocytes infiltration and angiogenesis.

Differences in local immune cell landscape between Q fever and atherosclerotic abdominal aortic aneurysms identified by multiplex immunohistochemistry.

Background: Chronic Q fever is a zoonosis caused by the bacterium Coxiella burnetii which can manifest as infection of an abdominal aortic aneurysm (AAA). Antibiotic therapy often fails, resulting in severe morbidity and high mortality. Whereas previous studies have focused on inflammatory processes in blood, the aim of this study was to investigate local inflammation in aortic tissue. Methods: Multiplex immunohistochemistry was used to investigate local inflammation in Q fever AAAs compared to atherosclerotic AAAs in aorta tissue specimen. Two six-plex panels were used to study both the innate and adaptive immune systems. Results: Q fever AAAs and atherosclerotic AAAs contained similar numbers of CD68+ macrophages and CD3+ T cells. However, in Q fever AAAs, the number of CD68+CD206+ M2 macrophages was increased, while expression of GM-CSF was decreased compared to atherosclerotic AAAs. Furthermore, Q fever AAAs showed an increase in both the number of CD8+ cytotoxic T cells and CD3+CD8-FoxP3+ regulatory T cells. Finally, Q fever AAAs did not contain any well-defined granulomas. Conclusions: These findings demonstrate that despite the presence of pro-inflammatory effector cells, persistent local infection with C. burnetii is associated with an immune-suppressed microenvironment. Funding: This work was supported by SCAN consortium: European Research Area - CardioVascualar Diseases (ERA-CVD) grant [JTC2017-044] and TTW-NWO open technology grant [STW-14716].

Induction of CD73 prevents death after emergency open aortic surgery for a ruptured abdominal aortic aneurysm: a randomized, double-blind, placebo-controlled study.

Mortality remains high after emergency open surgery for a ruptured abdominal aortic aneurysm (RAAA). The aim of the present study was to assess, if intravenous (IV) Interferon (IFN) beta-1a improve survival after surgery by up-regulating Cluster of differentiation (CD73). This is a multi-center phase II double-blind, 2:1 randomized, parallel group comparison of the efficacy and safety of IV IFN beta-1a vs. placebo for the prevention of death after open surgery for an infra-renal RAAA. All study patients presented a confirmed infra-renal RAAA, survived the primary emergency surgery and were treated with IFN beta-1a (10 µg) or matching placebo for 6 days after surgery. Major exclusion criteria included fatal hemorrhagic shock, chronic renal replacement therapy, diagnosed liver cirrhosis, severe congestive heart failure, advanced malignant disease, primary attempt of endovascular aortic repair (EVAR), and per-operative suprarenal clamping over 30 min. Main outcome measure was all-cause mortality at day 30 (D30) from initial emergency aortic reconstruction. The study was pre-maturely stopped due to a reported drug-drug interaction and was left under-powered. Out of 40 randomized patients 38 were included in the outcome analyses (27 IFN beta-1a and 11 placebo). There was no statistically significant difference between treatment groups at baseline except more open-abdomen and intestinal ischemia was present in the IFN beta-1a arm. D30 all-cause mortality was 22.2% (6/27) in the IFN beta-1a arm and 18.2% (2/11) in the placebo arm (OR 1.30; 95% CI 0.21-8.19). The most common adverse event relating to the IFN beta-1a was pyrexia (20.7% in the IFN beta-1a arm vs. 9.1% in the placebo arm). Patients with high level of serum CD73 associated with survival (P = 0.001) whereas the use of glucocorticoids and the presence of IFN beta-1a neutralizing antibodies associated with a poor CD73 response and survival. The initial aim of the trial, if postoperative INF beta-1a treatment results on better RAAA survival, could not be demonstrated. Nonetheless the anticipated target mechanism up-regulation of CD73 was associated with 100% survival. According to present results the INF beta-1a induced up-regulation of serum CD73 was blocked with both use of glucocorticoids and serum IFN beta-1a neutralizing antibodies. The study was pre-maturely stopped due to interim analysis after a study concerning the use if IV IFN beta-1a in ARDS suggested that the concomitant use of glucocorticoids and IFN beta-1a block the CD73 induction. Trial registration: ClinicalTrials.gov NCT03119701. Registered 19/04/2017 (retrospectively registered).

Hexarelin attenuates abdominal aortic aneurysm formation by inhibiting SMC phenotype switch and inflammasome activation.

Hexarelin, a synthetic growth hormone-releasing peptide, is shown to be protective in cardiovascular diseases such as myocardial infraction and atherosclerosis. However, the functional role of hexarelin in abdominal aortic aneurysm (AAA) remains undefined. The present study determined the effect of hexarelin administration (200 µg/kg twice per day) in a mouse model of elastase-induced abdominal aortic aneurysm. Echocardiography and in situ pictures showed hexarelin decreased infrarenal aorta diameter. Histology staining showed elastin degradation was improved in hexarelin-treated group. Hexarelin rescued smooth muscle cell contractile phenotype with increased α-SMA and decreased MMP2. Furthermore, hexarelin inhibited inflammatory cell infiltration, NLRP3 inflammasome activation and IL-18 production. Particularly, hexarelin suppressed NF-κB signaling pathway which is a key initiator of inflammatory response. These results demonstrated that hexarelin attenuated AAA development by inhibiting SMC phenotype switch and NF-κB signaling mediated inflammatory response.

Temporal and Quantitative Analysis of Aortic Immunopathologies in Elastase-Induced Mouse Abdominal Aortic Aneurysms.

OBJECTIVE: Elastase-induced abdominal aortic aneurysm (AAA) model is widely used for aneurysmal pathogenesis and translational research. However, temporal alternations in aneurysmal histologies remain unknown. This study is aimed at analyzing temporal immunopathologies of aneurysmal aorta following experimental AAA induction. METHODS: Male C57BL/6J mice at the age of 10-14 weeks received intra-aortic infusion of elastase to induce AAAs. Aortic diameters at the baseline and indicated days after AAA induction were measured, and aortae were collected for histopathological analysis. RESULTS: Aorta diameters increased from 0.52 mm at the baseline levels to 0.99 mm, 1.34 mm, and 1.41 mm at days 7, 14, and 28, respectively, corresponding 90%, 158%, and 171% increases over the baseline level. Average aortic diameters did not differ between days 14 and 28. Severe elastin degradation and smooth muscle cell depletion were found at days 14 and 28 as compared to the baseline and day 7. No difference in the scores of medial elastin and SMC destruction was noted between days 14 and 28. Consistent results were found for leukocyte accumulation, neoangiogenesis, and matrix metalloproteinase expression. Twenty-eight days after AAA induction, all aneurysmal pathologies showed an attenuated trend, although most histopathological parameters did no differ between days 14 and 28. CONCLUSION: Our data suggest that almost aneurysmal immunohistopathologies reach maximal 14 days following AAA induction. Analysis of day 14 histologies is sufficient for AAA pathogenesis and translational studies in elastase-induced mouse experimental AAAs.

Tryptophan Catabolism and Inflammation: A Novel Therapeutic Target For Aortic Diseases.

Aortic diseases are the primary public health concern. As asymptomatic diseases, abdominal aortic aneurysm (AAA) and atherosclerosis are associated with high morbidity and mortality. The inflammatory process constitutes an essential part of a pathogenic cascade of aortic diseases, including atherosclerosis and aortic aneurysms. Inflammation on various vascular beds, including endothelium, smooth muscle cell proliferation and migration, and inflammatory cell infiltration (monocytes, macrophages, neutrophils, etc.), play critical roles in the initiation and progression of aortic diseases. The tryptophan (Trp) metabolism or kynurenine pathway (KP) is the primary way of degrading Trp in most mammalian cells, disturbed by cytokines under various stress. KP generates several bioactive catabolites, such as kynurenine (Kyn), kynurenic acid (KA), 3-hydroxykynurenine (3-HK), etc. Depends on the cell types, these metabolites can elicit both hyper- and anti-inflammatory effects. Accumulating evidence obtained from various animal disease models indicates that KP contributes to the inflammatory process during the development of vascular disease, notably atherosclerosis and aneurysm development. This review outlines current insights into how perturbed Trp metabolism instigates aortic inflammation and aortic disease phenotypes. We also briefly highlight how targeting Trp metabolic pathways should be considered for treating aortic diseases.

Reversal of elastase-induced abdominal aortic aneurysm following the delivery of nanoparticle-based pentagalloyl glucose (PGG) is associated with reduced inflammatory and immune markers.

OBJECTIVE: An Abdominal aortic aneurysm (AAA), a deadly disease in elderly population, is featured by expansion of aortic diameter, degradation and weakening of vasculature. Its common and significant characteristics are disarray and inflammation in vasculature. We tested the hypothesis that the reversal of abdominal aortic aneurysm by pentagalloyl glucose-loaded nanoparticles (PGG-NPs) therapy that targets degraded elastin suppresses inflammatory and immune markers to ameliorate the pathophysiology of the disease in advance stage aneurysm in a porcine pancreatic elastase (PPE)-induced mouse model of AAA. METHODS AND RESULTS: After induction of aneurysm in pathogen-free C57BL/6 male mice by applying PPE peri-adventitially to the abdominal aorta, once a week for two doses of intravenous injections of pentagalloyl glucose-loaded nanoparticles (PGG-NPs) conjugated with elastin targeted antibody were used to reverse the aneurysms. We showed that PGG-NPs therapy could suppress infiltration of macrophages, CD8 and CD4 subsets of T cells, matrix metalloproteinases (MMPs), inflammatory cytokines interferon (IFN-γ) and interleukin (IL)-6 at the local and systemic level. Moreover, such PGG-NPs therapy increases the induction of anti-inflammatory cytokines IL-13, IL-27 and IL-10 at the local and systemic level. The therapy also led to remodeling of elastic lamina at the aneurysm site. CONCLUSION: Nanoparticles-loaded pentagalloyl glucose therapy can be an effective treatment option against advanced stage aneurysms to reverse the disease by ameliorating inflammation and restoring arterial homeostasis.

B Cell-Activating Factor Antagonism Attenuates the Growth of Experimental Abdominal Aortic Aneurysm.

B cell-activating factor (BAFF), part of a tumor necrosis factor family of cytokines, was recently identified as a regulator of atherosclerosis; however, its role in aortic aneurysm has not been determined. Here, the study examined the effect of selective BAFF antagonism using an anti-BAFF antibody (blocks binding of BAFF to receptors BAFF receptor 3, transmembrane activator and CAML interactor, and B-cell maturation antigen) and mBaffR-mFc (blocks binding of BAFF to BAFF receptor 3) on a murine model of abdominal aortic aneurysm (AAA). In a prevention strategy, the antagonists were injected before the induction of AAA, and in an intervention strategy, the antagonists were injected after the induction of AAA. Both strategies attenuated the formation of AAA. In the intervention group, BAFF antagonism depleted most of the mature B-cell subsets in spleen and circulation, leading to enhanced resolution of inflammation in AAA as indicated by decreased infiltration of B cells and proinflammatory macrophages and a reduced number of apoptotic cells. In AAA tissues, B cells and macrophages were found in close contact. In vitro, B cells, irrespective of treatment with BAFF, impaired the efferocytosis activity of macrophages, suggesting a direct innate role of B cells on macrophage function. Altogether, BAFF antagonism affects survival of the mature B cells, promotes resolution of inflammation in the aorta, and attenuates the growth of AAA in mice.

Surgical strategy of IgG4-related inflammatory abdominal aortic aneurysm with preoperative steroid therapy: A case report.

Immunoglobulin G4 (IgG4)-related disease, characterized by high serum IgG4 concentrations and IgG4-positive plasma cell infiltration, often presents as an inflammatory aneurysm. We herein report the case of a 78 year-old man, presenting with elevated inflammatory markers and IgG4 concentrations, who was diagnosed with IgG4-related inflammatory abdominal aortic aneurysm with dense perianeurysmal fibrosis. Before the surgical intervention, steroid therapy was administered to resolve his perianeurysmal inflammatory fibrosis. Half a year after the initiation of steroid therapy, there was an improvement in serum inflammatory markers and IgG4 concentrations, and the perianeurysmal fibrosis had regressed. Thus, we performed a surgical intervention including resection of the aneurysm and interposition with a prosthetic graft. Histopathological examination demonstrated few IgG4-positive plasma cells were distributed in the adventitia, which was suspected to be associated with the preoperative steroid therapy. This case study suggests preoperative steroid therapy is a useful therapeutic strategy for IgG4-related abdominal aortic aneurysm because it allows the use of open surgical procedures with reduced surgical risk.

Paeonol Ameliorates Abdominal Aortic Aneurysm Progression by the NF-κB Pathway.

OBJECTIVE: Abdominal aortic aneurysm (AAA) is a chronic inflammatory disease characterized by localized progressive dilatation. Currently, paeonol has been shown to possess anti-inflammatory and protective cardiovascular properties. Our study aimed to investigate the potential influences of paeonol on AAA progression. METHODS: Experimental AAAs were created in C57BL/6J mice by intra-aortic infusion of porcine pancreatic elastase, and then intragastrically administered paeonol (20 mg/kg/day) for 14 days. The effects of paeonol on experimental AAA were measured by ultrasound imaging, histopathology, and western blot analyses. RESULTS: Paeonol treatment limited the enlargement of the aneurysmal diameter and alleviated the depletion of elastic fibers and vascular smooth muscle cells (VSMCs). Furthermore, the infiltration of CD68+ macrophages and CD8+ lymphocytes was obviously attenuated after paeonol administration, along with mural neoangiogenesis. Western blot results showed that paeonol inhibited the expression of matrix metalloproteinase (MMP) and the NF-κB pathway activation. CONCLUSIONS: Paeonol might prevent experimental AAA progression by inhibiting the NF-κB pathway, which suggests that it is a potential drug for AAA.

Association of Differential Leukocyte Count With Incident Abdominal Aortic Aneurysm Over 22.5 Years: The ARIC Study.

OBJECTIVE: Leukocytes contribute to the development of abdominal aortic aneurysm (AAA). We evaluated whether associations of differential leukocyte counts with AAA persist after accounting for traditional risk factors of AAA. APPROACH AND RESULTS: Among 11 217 adults from the Atherosclerosis Risk in Communities Study, we evaluated associations of differential leukocyte counts at baseline (1987­1989) with incident AAAs over a median follow-up of 22.5 years, using Cox proportional hazards regression. Each differential leukocyte count was categorized into 5 groups­below normal, tertiles within the normal range, and above normal, with the first tertile serving as the referent. We identified 377 incident AAAs through 2011, using hospital discharge diagnoses, linked Medicare records, or death certificates. At baseline, higher neutrophil, monocyte, and eosinophil counts were associated with higher risk of AAA, independent of smoking, other differential leukocyte counts, and other traditional risk factors. The association with incident AAA was the strongest for above normal neutrophil count, with an adjusted hazard ratio (95% CI) of 2.17 (1.29­3.64). Below normal neutrophil, lymphocyte, eosinophil and basophil counts were associated with higher risk of AAA with adjusted hazard ratio (95% CI) between 1.86 (1.04­3.35) and 1.62 (1.10­2.39). CONCLUSIONS: Higher neutrophil, monocyte, and eosinophil counts in midlife are associated with higher risk of AAA, even after accounting for traditional risk factors such as smoking, obesity, and atherosclerosis. This suggests the need to identify nontraditional risk factors and treatment strategies to mitigate the residual risk of AAA conferred by midlife inflammation. Whether immunosuppression is associated with higher risk of AAA needs further investigation.

NSun2 regulates aneurysm formation by promoting autotaxin expression and T cell recruitment.

Abdominal aortic aneurysm (AAA) is characterized by inflammatory cell infiltration and aggravated by hyperhomocysteinemia (HHcy). It is unknown whether the homocysteine (Hcy)-activated RNA methyltransferase NOP2/Sun domain family member 2 (NSun2) is associated with AAA. Here, we found that NSun2 deficiency significantly attenuated elastase-induced and HHcy-aggravated murine AAA with decreased T cell infiltration in the vessel walls. T cell labeling and adoptive transfer experiments confirmed that NSun2 deficiency inhibited the chemotaxis of vessels to T cells. RNA sequencing of endothelial cells showed that Hcy induced the accumulation of various metabolic enzymes of the phospholipid PC-LPC-LPA metabolic pathway, especially autotaxin (ATX). In the elastase-induced mouse model of AAA, ATX was specifically expressed in the endothelium and the plasma ATX concentration was upregulated and even higher in the HHcy group, which were decreased dramatically by NSun2 knockdown. In vitro Transwell experiments showed that ATX dose-dependently promoted T cell migration. HHcy may upregulate endothelial ATX expression and secretion and in turn recruit T cells into the vessel walls to induce vascular inflammation and consequently accelerate the pathogenesis of AAA. Mechanistically, secreted ATX interacted with T cells by binding to integrin α4, which subsequently activated downstream FAK/Src-RhoA signaling pathways and then induced T cell chemokinesis and adhesion. ATX overexpression in the vessel walls reversed the inhibited development of AAA in the NSun2-deficient mice. Therefore, NSun2 mediates the development of HHcy-aggravated AAA primarily by increasing endothelial ATX expression, secretion and T cell migration, which is a novel mechanism for HHcy-aggravated vascular inflammation and pathogenesis of AAA.

Memory T cells of patients with abdominal aortic aneurysm differentially expressed micro RNAs 21, 92a, 146a, 155, 326 and 663 in response to Helicobacter pylori and Lactobacillus acidophilus.

Regarding the role of micro RNAs (miRNA) in the proliferation and differentiation of T cells as well as the controversy around the role of bacteria in the pathogenesis of abdominal aortic aneurysm (AAA), the effects of Helicobacter pylori (Hp) and Lactobacillus acidophilus (La) were investigated in the induction of miRNAs and apoptosis in CD4+ memory T (Tem) cells of AAA patients and controls. Signature atherosclerosis miRNAs 21, 92a, 146a, 155, 326 and 663 were measured in the sera and tissues of AAA patients and control. PBMCs separately and in co-culture with HUVEC were treated with Hp-water-extract (HpWE) and La-conditioned-medium (LaCM). Apoptosis and miRNA levels were assessed in the isolated Tem by flowcytometry and real-time-PCR. In single-culture, HpWE increased apoptosis and miR-155 and LaCM decreased apoptosis and increased miR-21. In co-culture, apoptosis decreased in both groups in response to CagA+HpWE. Also, all miRNAs increased in patients Tem but in controls, only miR- 146a and 21 showed changes. Although, apoptosis was similar in Tem of patients and controls, the effects of Hp and La were different on the induction of apoptosis and miRNAs and also these bacteria showed different impacts in single and co-culture conditions. Beyond the direct effects of these bacteria on the pathogenesis of diseases, their effects on miRNAs expression may shed light on their roles in the development and the prevention of AAA.

Prevention of CaCl2-induced aortic inflammation and subsequent aneurysm formation by the CCL3-CCR5 axis.

Inflammatory mediators such as cytokines and chemokines are crucially involved in the development of abdominal aortic aneurysm (AAA). Here we report that CaCl2 application into abdominal aorta induces AAA with intra-aortic infiltration of macrophages as well as enhanced expression of chemokine (C-C motif) ligand 3 (CCL3) and MMP-9. Moreover, infiltrating macrophages express C-C chemokine receptor 5 (CCR5, a specific receptor for CCL3) and MMP-9. Both Ccl3-/- mice and Ccr5-/- but not Ccr1-/- mice exhibit exaggerated CaCl2-inducced AAA with augmented macrophage infiltration and MMP-9 expression. Similar observations are also obtained on an angiotensin II-induced AAA model. Immunoneutralization of CCL3 mimics the phenotypes observed in CaCl2-treated Ccl3-/- mice. On the contrary, CCL3 treatment attenuates CaCl2-induced AAA in both wild-type and Ccl3-/- mice. Consistently, we find that the CCL3-CCR5 axis suppresses PMA-induced enhancement of MMP-9 expression in macrophages. Thus, CCL3 can be effective to prevent the development of CaCl2-induced AAA by suppressing MMP-9 expression.

Macrophage pyroptosis is mediated by immunoproteasome subunit ß5i (LMP7) in abdominal aortic aneurysm.

Macrophages contribute to abdominal aortic aneurysm (AAA), but the effect of macrophage on AAA formation is not totally understood. Recent research proved that macrophage pyroptosis plays an important role in many cardiovascular disease. However, whether macrophage pyroptosis is involved in AAA and its mechanism remains unknown. In this study, we found that the pyroptosis significantly increased in AAA tissues. ß5i inhibitor PR-957 treatment or ß5i deficiency markedly ameliorated AAA formation and decreased the pyroptosis. Pyroptosis were also significantly attenuated in bone marrow derived macrophages (BMDM) from ß5i-/- mice compared with the control group when they were subjected to OXLDL. Mechanistically, ß5i may promote activation of NFκB which augment NLRP3 expression. In conclusion, this study suggested macrophages pyroptosis are involved in AAA and inhibition or knockout of ß5i decreased macrophage pyroptosis via IκB/NFκB pathway.

Prevention of Progression of Aortic Aneurysm by Peptide Vaccine Against Ang II (Angiotensin II) in a Rat Model.

There is no proven medical therapy to inhibit the progression of abdominal aortic aneurysm (AAA) in the clinical setting. To develop a novel therapeutic approach for the treatment of AAA, we focused on vaccination targeting Ang II (angiotensin II) and assessed the effect of an Ang II peptide vaccine on the progression of AAA using a rat model. Ang II peptide was conjugated with KLH (keyhole limpet hemocyanin) carrier protein to induce a sufficient immune response. Male rats were subcutaneously immunized with Ang II-KLH with an adjuvant on days 0, 14, and 28. Aortic dilatation was induced by intraluminal incubation with elastase on day 35. Treatment with Ang II vaccine successfully induced the production of a high titer of anti-Ang II antibodies. Immunization with Ang II vaccine resulted in a significant reduction in expansion of the aortic diameter compared with control rats, without a blood pressure-lowering effect. Four weeks after operation, the increase in Ang II in the aneurysm wall was significantly inhibited by treatment with Ang II vaccine. Inhibition of Ang II action led to suppression of the inflammatory response in the AAA wall through attenuation of the NFκB (nuclear factor kappa B) and c-jun N-terminal kinase signaling cascades. Treatment with Ang II vaccine inhibited accumulation of macrophages in the AAA wall. In addition, expression of TNF-α (tumor necrosis factor alpha) and activation of MMP (matrix metalloproteinase)-2 and MMP-9 were also inhibited by treatment with Ang II vaccine, resulting in protection against the destruction of elastic fibers. This vaccine therapy could become a potent therapeutic option to treat patients with AAA.

Simultaneous molecular MRI of extracellular matrix collagen and inflammatory activity to predict abdominal aortic aneurysm rupture.

Abdominal aortic aneurysm (AAA) is a life-threatening vascular disease with an up to 80% mortality in case of rupture. Current biomarkers fail to account for size-independent risk of rupture. By combining the information of different molecular probes, multi-target molecular MRI holds the potential to enable individual characterization of AAA. In this experimental study, we aimed to examine the feasibility of simultaneous imaging of extracellular collagen and inflammation for size-independent prediction of risk of rupture in murine AAA. The study design consisted of: (1) A outcome-based longitudinal study with imaging performed once after one week with follow-up and death as the end-point for assessment of rupture risk. (2) A week-by-week study for the characterization of AAA development with imaging after 1, 2, 3 and 4 weeks. For both studies, the animals were administered a type 1 collagen-targeted gadolinium-based probe (surrogate marker for extracellular matrix (ECM) remodeling) and an iron oxide-based probe (surrogate marker for inflammatory activity), in one imaging session. In vivo measurements of collagen and iron oxide probes showed a significant correlation with ex vivo histology (p < 0.001) and also corresponded well to inductively-coupled plasma-mass spectrometry and laser-ablation inductively-coupled plasma mass spectrometry. Combined evaluation of collagen-related ECM remodeling and inflammatory activity was the most accurate predictor for AAA rupture (sensitivity 80%, specificity 100%, area under the curve 0.85), being superior to information from the individual probes alone. Our study supports the feasibility of a simultaneous assessment of collagen-related extracellular matrix remodeling and inflammatory activity in a murine model of AAA.

Circulatory CD4-Positive T-Lymphocyte Imbalance Mediated by Homocysteine-Induced AIM2 and NLRP1 Inflammasome Upregulation and Activation Is Associated with Human Abdominal Aortic Aneurysm.

INTRODUCTION: CD4-positive T lymphocytes (CD4 cells) play a significant role in human abdominal aortic aneurysm (AAA). However, we know little about the role of the different CD4 subtypes. OBJECTIVE: We aimed to discover the circulatory CD4 phenotypic marker profile and the roles of the newly found T helper cell 9 (Th9) and follicular helper T cells (Tfh) and that of inflammasomes in CD4 cells from AAA patients. METHODS: We extracted CD4 cells from 30 AAA patients and 21 age-matched controls. Phenotype-specific transcription factors (TFs) and inflammasomes were analyzed with qRT-PCR. RESULTS: Th17-, Th1-, Th9-, and Tfh-specific TFs and inflammasome components NLRP1 (NLR family pyrin domain-containing 1), NLRP3, NLRC4, AIM2 (absent in melanoma 2), PYCARD (apoptosis speck-like protein containing a CARD), and CASP1 (caspase 1) were upregulated, and T regulatory cell- and Th2-specific TFs were downregulated in the patients' peripheral blood CD4 cells. Homocysteine was involved in Tfh and Th17 imbalance by AIM2 and NLRP1 inflammasome upregulation. Blood total cholesterol level correlated positively with NLRP1 expression, and blood low-density lipoprotein level correlated negatively with FOXP3 expression. CONCLUSIONS: Inflammasome-induced CD4 cell imbalance was involved in AAA. We thought that AAA might be a consequence of synergism between systemic immune imbalance and local autoimmunity.

Ex vivo magnetic particle imaging of vascular inflammation in abdominal aortic aneurysm in a murine model.

Abdominal aortic aneurysms (AAAs) are currently one of the leading causes of death in developed countries. Inflammation is crucial in the disease progression, having a substantial impact on various determinants in AAAs development. Magnetic particle imaging (MPI) is an innovative imaging modality, enabling the highly sensitive detection of magnetic nanoparticles (MNPs), suitable as surrogate marker for molecular targeting of vascular inflammation. For this study, Apolipoprotein E-deficient-mice underwent surgical implantation of osmotic minipumps with constant Angiotensin II infusion. After 3 and 4 weeks respectively, in-vivo-magnetic resonance imaging (MRI), ex-vivo-MPI and ex-vivo-magnetic particle spectroscopy (MPS) were performed. The results were validated by histological analysis, immunohistology and laser ablation-inductively coupled plasma-mass spectrometry. MR-angiography enabled the visualization of aneurysmal development and dilatation in the experimental group. A close correlation (R = 0.87) with histological area assessment was measured. Ex-vivo-MPS revealed abundant iron deposits in AAA samples and ex-vivo histopathology measurements were in good agreement (R = 0.76). Ex-vivo-MPI and MPS results correlated greatly (R = 0.99). CD68-immunohistology stain and Perls'-Prussian-Blue-stain confirmed the colocalization of macrophages and MNPs. This study demonstrates the feasibility of ex-vivo-MPI for detecting inflammation in AAA. The quantitative ability for mapping MNPs establishes MPI as a promising tool for monitoring inflammatory progression in AAA in an experimental setting.