Altogether, the results suggest a dominant role of IL-1β-induced NETosis in AAA formation.

Objective B cell depletion therapy is widely used for treatment of cancers and autoimmune diseases. B cells are abundant in abdominal aortic aneurysms (AAA), however, it is unknown whether B cell depletion therapy affects AAA growth. Using experimental models of murine AAA, we aim to examine the effect of B cell depletion on AAA formation. Approach and Results Wild-type or Apolipoprotein E knockout mice were treated with mouse monoclonal anti-CD20 or control antibodies and subjected to an elastase perfusion or angiotensin II-infusion model to induce AAA, respectively. Anti-CD20 antibody treatment significantly depleted B1 and B2 cells, and strikingly suppressed AAA growth in both models. B cell depletion resulted in lower circulating IgM levels, but did not affect the levels of IgG or cytokine/chemokine levels. Although the total number of leukocyte remained unchanged in elastase perfused aortas following anti-CD20 antibody treatment, the number of B cell subtypes was significantly lower. Interestingly, plasmacytoid dendritic cells (pDCs) expressing the immunomodulatory enzyme indole 2,3-dioxygenase (IDO) were detected in the aortas of B cell depleted mice. In accordance with an increase in IDO+ pDCs, the number of regulatory T cells was higher while the expression of pro-inflammatory genes was lower in aortas of B cell depleted mice. In a coculture model, presence of B cells significantly lowered the number of IDO+ pDCs without affecting total pDC number. Conclusions The present results demonstrate that B cell depletion protects mice from experimental AAA formation and promotes emergence of an immunosuppressive environment in aorta.

Objectives: Resolvins have been shown to attenuate inflammation, while NETosis, the process of neutrophils releasing neutrophil extracellular traps (NETs), produces increased inflammation. It is hypothesized that treatment of animals with resolvin D1 (RvD1) would reduce abdominal aortic aneurysm (AAA) formation by inhibiting NETosis. Methods: Wild-type (WT) 8–12-week old C57BL/6 male mice (n=47) and apolipoproteinE deficient (ApoE-/) mice (n=20) were used in two models to demonstrate the effects of RvD1 on AAA growth. In the topical elastase AAA model, WT mice were divided into three groups: (1) Deactivated elastase control group: sham surgery was performed using deactivated elastase and mice were intravenously injected with phosphate buffered saline (PBS) once a day until harvest. (2) Elastase group: active elastase was used to induce AAA and mice were injected with PBS daily until harvest. (3) RvD1 treated group: AAA was induced and mice were injected with RvD1 daily until harvest. In the angiotensin II (Ang II) induced AAA model, ApoE−/− mice were fed high fat diet and implanted with osmotic infusion pumps containing Ang II (1000ng/kg/min). The Ang II model was divided into two groups: (1) Ang II control group: Ang II was delivered and mice were injected with PBS daily until harvest. (2) RvD1 treated group: Ang II was delivered and mice were injected with RvD1 daily until harvest. On postoperative days 3, 14, or 28, aortic and blood samples were collected for western blot, histology, cytokine array, enzyme-linked immunosorbent assay, and gelatin zymography after aortic diameter measurement. Results: Day 14 RvD1 treated group demonstrated 42% reduced AAA diameter compared to elastase group (p<0.001). On postoperative day 3, RvD1 treated group showed decreased levels of NETosis markers citrullinated histone H3 (p=0.04) and neutrophil elastase (p=0.002) compared to the Elastase group. Among important cytokines involved in AAA formation, interleukin (IL) 1β was down-regulated (p=0.02), while IL-10, a protective cytokine, was upregulated (p=0.01) in the RvD1 treated group. Active matrix metalloproteinase 2 (MMP2) also decreased in the RvD1 treated group (p= 0.03). The RvD1 treated group in the Ang II AAA model, a second model, demonstrated reduced AAA diameter compared to Ang II control group on day 28 (p<0.046). RvD1 treated group showed decreased levels of citrullinated histone H3 on day 3 (p=0.002). Cytokines INF-γ, IL-1β, CXCL-10, MCP-1, and RANTES were all decreased on day 28 (p<0.05). Conclusion: Resolvin D1-mediated inhibition of NETosis may represent a future medical treatment for the attenuation of AAA growth.

The formation of an abdominal aortic aneurysm (AAA) is characterized by inflammation, macrophage infiltration, and vascular remodeling. In this study, we tested the hypothesis that mesenchymal stromal cell (MSC)-derived extracellular vesicles (EVs) immunomodulate aortic inflammation, to mitigate AAA formation via modulation of microRNA-147. An elastase-treatment model of AAA was used in male C57BL/6 wild-type (WT) mice. Administration of EVs in elastase-treated WT mice caused a significant attenuation of aortic diameter and mitigated proinflammatory cytokines, inflammatory cell infiltration, an increase in smooth muscle cell α-actin expression, and a decrease in elastic fiber disruption, compared with untreated mice. A 10-fold up-regulation of microRNA (miR)-147, a key mediator of macrophage inflammatory responses, was observed in murine aortic tissue in elastase-treated mice compared with controls on d 14. EVs derived from MSCs transfected with miR-147 mimic, but not with miR-147 inhibitor, attenuated aortic diameter, inflammation, and leukocyte infiltration in elastase-treated mice. In vitro studies of human aortic tissue explants and murine-derived CD11b macrophages induced proinflammatory cytokines after elastase treatment, and the expression was attenuated by cocultures with EVs transfected with miR-147 mimic, but not with miR-147 inhibitor. Thus, our findings define a critical role of MSC-derived EVs in attenuation of aortic inflammation and macrophage activation via miR-147 during AAA formation.-Spinosa, M., Lu, G., Su, G., Bontha, S. V., Gehrau, R., Salmon, M. D., Smith, J. R., Weiss, M. L., Mas, V. R., Upchurch, G. R., Sharma, A. K. Human mesenchymal stromal cell-derived extracellular vesicles attenuate aortic aneurysm formation and macrophage activation via microRNA-147.

Little is known regarding how the kidney shifts from a sodium and water reclaiming state (antinatriuresis) to a state where sodium and water are eliminated (natriuresis). In human renal proximal tubule cells (RPTCs), sodium reabsorption is decreased by the dopamine D1-like receptors (D1R/D5R) and the angiotensin type 2 receptor (AT2R), while the angiotensin type 1 receptor increases sodium reabsorption. Aberrant control of these opposing systems is thought to lead to sodium retention and subsequently hypertension. We show that D1R/D5R stimulation increased plasma membrane AT2R 4-fold via a D1R-mediated, cAMP-coupled, and PP2A-dependent specific signaling pathway. D1R/D5R stimulation also reduced the ability of angiotensin II to stimulate phospho-ERK, an effect that was partially reversed by an AT2R antagonist. Fenoldopam did not increase AT2R recruitment in RPTCs with D1Rs uncoupled from adenylyl cyclase, suggesting a role of cAMP in mediating these events. D1Rs and AT2Rs heterodimerized and cooperatively increased cAMP and cGMP production, PP2A activation, sodium-potassium-ATPase internalization and sodium transport inhibition. These studies shed new light on the regulation of renal sodium transport by the dopaminergic and angiotensin systems and potential new therapeutic targets for selectively treating hypertension.

Pannexin-1 (Panx1) channels have been shown to regulate leukocyte trafficking and tissue inflammation but the mechanism of Panx1 in chronic vascular diseases like abdominal aortic aneurysms (AAA) is unknown. Here we demonstrate that Panx1 on endothelial cells, but not smooth muscle cells, orchestrate a cascade of signaling events to mediate vascular inflammation and remodeling. Mechanistically, Panx1 on endothelial cells acts as a conduit for ATP release that stimulates macrophage activation via P2X7 receptors and mitochondrial DNA release to increase IL-1β and HMGB1 secretion. Secondly, Panx1 signaling regulates smooth muscle cell-dependent intracellular Ca2+ release and vascular remodeling via P2Y2 receptors. Panx1 blockade using probenecid markedly inhibits leukocyte transmigration, aortic inflammation and remodeling to mitigate AAA formation. Panx1 expression is upregulated in human AAAs and retrospective clinical data demonstrated reduced mortality in aortic aneurysm patients treated with Panx1 inhibitors. Collectively, these data identify Panx1 signaling as a contributory mechanism of AAA formation.

A reproducible model of aortic aneurysm rupture was developed with a high incidence of abdominal and thoracic aortic aneurysm. This model should enable further studies investigating the pathogenesis of aortic rupture, as well as allow for targeted strategies to prevent human aortic aneurysm rupture.

Objective: Few large animal models exist for the study of aortic aneurysms. βaminopropionitrile (BAPN) is a compound known to cause aortic aneurysms by inhibiting lysyl oxidase, a collagen cross-linking enzyme. It is hypothesized that BAPN plus aneurysm induction surgery would result in significant aneurysm formation in swine with biologic properties similar to human disease. Methods: Initial experiments were performed in uncastrated male swine not treated with BAPN (surgery alone). Subsequently, uncastrated male swine were fed BAPN (0.15 grams/kilogram) for 7 days before undergoing surgery: the infrarenal aorta was circumferentially dissected and measured, balloon dilated, perfused with elastase (500 units) and Type 1 collagenase (8000 units), with extraluminal elastase application. In the BAPN groups, daily BAPN feedings continued until swine harvest at postoperative days 7, 14, and 28. Results: Swine undergoing surgery alone (n=12) had significantly less dilation at 28 days compared with BAPN + surgery swine (51.9±29.2% (0-100%) vs. 113.5±30.2% (52.9-146.2%); p<0.0003). Mean aortic dilation in animals undergoing treatment with surgery and BAPN was 86.9%±47.4% (range, 55.6-157.1%), 105.4%±58.1% (50-133.3%), and 113.5%±30.2% (52.9-146.2%) at 7, 14, and 28 days, respectively. In the BAPN + surgery group, significant elastolysis was present at all time points, while aortic wall collagen content was not significantly different. Smooth muscle cells were significantly depleted at 14 and 28 days, and M1 macrophages were increased at 14 and 28 days (p<0.05, all). Matrix metalloproteinase 2 was elevated at 7 days