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Neutrophil chemotaxis is essential in responses to infection and underlies inflammation. In neutrophils, the small GTPase Rac1 has discrete functions at both the leading edge and in the retraction of the trailing structure at the cell’s rear (uropod), but how Rac1 is regulated at the uropod is unknown. Here, we identified a mechanism mediated by the trafficking protein synaptotagmin-like 1 (SYTL1 or JFC1) that controls Rac1-GTP recycling from the uropod and promotes directional migration of neutrophils. JFC1-null neutrophils displayed defective polarization and impaired directional migration to N-formyl-methionine-leucyl-phenylalanine in vitro, but chemoattractant-induced actin remodeling, calcium signaling and Erk activation were normal in these cells. Defective chemotaxis was not explained by impaired azurophilic granule exocytosis associated with JFC1 deficiency. Mechanistically, we show that active Rac1 localizes at dynamic vesicles where endogenous JFC1 colocalizes with Rac1-GTP. Super-resolution microscopy (STORM) analysis shows adjacent distribution of JFC1 and Rac1-GTP, which increases upon activation. JFC1 interacts with Rac1-GTP in a Rab27a-independent manner to regulate Rac1-GTP trafficking. JFC1-null cells exhibited Rac1-GTP accumulation at the uropod and increased tail length, and Rac1-GTP uropod accumulation was recapitulated by inhibition of ROCK or by interference with microtubule remodeling. In vivo, neutrophil dynamic studies in mixed bone marrow chimeric mice show that JFC1−/− neutrophils are unable to move directionally toward the source of the chemoattractant, supporting the notion that JFC1 deficiency results in defective neutrophil migration. Our results suggest that defective Rac1-GTP recycling from the uropod affects directionality and highlight JFC1-mediated Rac1 trafficking as a potential target to regulate chemotaxis in inflammation and immunity.

Significance: Air pollution is a considerable global threat to human health that dramatically increases the risk for cardiovascular pathologies, such as atherosclerosis, myocardial infarction, and stroke. An estimated 4.2 million cases of premature deaths worldwide are attributable to outdoor air pollution. Among multiple other components, airborne particulate matter (PM) has been identified as the major bioactive constituent in polluted air. While PM-related illness was historically thought to be confined to diseases of the respiratory system, overwhelming clinical and experimental data have now established that acute and chronic exposure to PM causes a systemic inflammatory and oxidative stress response that promotes cardiovascular disease. Recent Advances: A large body of evidence has identified an impairment of redox metabolism and the generation of oxidatively modified lipids and proteins in the lung as initial tissue response to PM. In addition, the pathogenicity of PM is mediated by an inflammatory response that involves PM uptake by tissue-resident immune cells, the activation of proinflammatory pathways in various cell types and organs, and the release of proinflammatory cytokines as locally produced tissue response signals that have the ability to affect organ function in a remote manner. Critical Issues: In the present review, we summarize and discuss the functional participation of PM in cardiovascular pathologies and its risk factors with an emphasis on how oxidative stress, inflammation, and immunity interact and synergize as a response to PM. Future Directions: The impact of PM constituents, doses, and novel anti-inflammatory therapies against PM-related illness is also discussed.

There is now overwhelming experimental and clinical evidence that arteriosclerosis is a chronic inflammatory disease. Lessons learned from genome-wide association studies, advanced in vivo imaging techniques, transgenic lineage tracing mice models and clinical interventional studies have shown that both innate and adaptive immune mechanisms can accelerate or curb arteriosclerosis. This article summarizes and discusses the pathogenesis of arteriosclerosis with a focus on the role of the adaptive immune system. Some limitations of animal models are discussed and the need for models that are tailored to better translate to human atherosclerosis and ultimately progress in prevention and treatment are emphasized.

Virtual and augmented reality head-mounted displays (HMDs) are currently heavily relying on spatially tracked input devices (STID) for interaction. These STIDs are all prone to the phenomenon that a discrete input (e.g. button press) will disturb the position of the tracker, resulting in a different selection point during ray-cast interaction (Heisenberg Effect of Spatial Interaction). Besides the knowledge of its existence, there is currently a lack of a deeper understanding of its severity, structure and impact on throughput and angular error during a selection task. In this work, we present a formal evaluation of the Heisenberg effect and the impact of body posture, arm position and STID degrees of freedom on its severity. In a Fitt's Law inspired user study (N=16), we found that the Heisenberg effect is responsible for 30.45% of the overall errors occurring during a pointing task, but can be reduced by 25.4% using a correction function.

The hyperarid parts of the Atacama Desert, N Chile, are among the driest places on Earth, and a number of studies have emphasized the remarkable slowness of Earth surface processes since the late Tertiary. Despite episodic overland flow or flash flood activity, salt-driven shrink-swell processes, dust deposition, and seismic shaking have significantly contributed to the formation of the characteristic landscape. The enigmatic and Atacama-specific zebra stripes are contour-parallel, thin lateral bands of rather angular gravels on hillslopes, characterized by grain sorting, a lateral succession of 1-3 m-wide frontal lobes, and specific wavelengths, occurring generally in areas with lowest rainfall and low to no fog occurrence. While previous investigations suggest that zebra stripes represent fossil evidence of overland flow, a recent study challenged their water-related evolution and emphasized the role of seismicity in their formation, and in shaping the Atacama landscape in general. Similar landforms may also be found on Mars, although related processes may be different to those on Earth.We use UAV-derived orthophotos and digital elevation models, geomorphological surveys and sediment sampling, as well as OSL rock surface dating and cosmogenic nuclide (21Ne, 10Be) analysis of surface clasts to provide new insights into zebra stripe activity and stability in the Atacama Desert. Our investigations show that zebra stripes are found in numerous areas in the hyperarid core of the Atacama, implying a wider distribution of zebra-striped hillslopes than previously suggested. Inter-site comparison illustrates considerable differences between individual zebra stripe sites, and geomorphological characteristics suggest that zebra stripes may be active or inactive forms, depending on their location. Active forms are indicated by well-developed frontal lobes, reduced dust contents, and clearly developed downslope sorting trends with a high percentage of freely floating clasts, whereas inactivity seems to be indicated by poorly visible frontal lobes, high dust contents, poor sorting trends and a high proportion of clasts embedded into the underlying vesicular soil horizon. At the same time, preliminary chronological data based on OSL rock surface dating suggests that active stripes contain clasts with active bleaching fronts at top and bottom sides, pointing to late Pleistocene to Holocene activity and clast overturning. Inactivity, in contrast, is indicated by similar IR50 and pIR225 bleaching curves (equilibrium of bleaching and dosing) in upper clast surfaces and the lack of a bleaching front at the bottom side, pointing to long exposures without clast overturning. Combined with cosmogenic nuclide concentrations, our results allow for inter-site comparisons and a better understanding of (relative) stripe chronologies and activity-stability patterns. Alongside further investigations on the significance of fog, wind, rain and seismicity, future work will aim at constraining time scales on which active/young and inactive/old forms have developed. If these forms are seismicity-related, our study contributes important information on the paleoseismic evolution of the central Atacama.

Wave transported boulders represent important records of storm and tsunami impact over geological timescales. Their use for hazard assessment requires chronological information that in many cases cannot be achieved by established dating approaches such as radiocarbon and U-Th dating. To fill this gap, we investigated the potential of optically stimulated luminescence rock surface exposure dating for estimating transport ages of coastal boulders. The approach was applied to wave-emplaced calcarenite clasts at the Rabat coast, Morocco. Calibration of the OSL surface exposure model was based on samples with rock surfaces exposed for ~2 years, and OSL exposure ages were evaluated against age control deduced from satellite images. The dating precision is very limited for all boulders due to the local source rock lithology, which contains low amounts of quartz and feldspar and was formed after MIS 5 (OSL signals are not in field saturation). Nevertheless, we propose a robust relative chronology for boulders that are not affected by significant post-depositional erosion and that share surface angle of inclination with the calibration samples. The relative chronology indicates that most boulders were moved by storm waves; these storms lifted boulders with masses of up to ~40 t; indicating that the role of storms for the formation of boulder deposits along the Rabat coast is much more significant than assumed previously. While OSL rock surface exposure dating cannot provide reliable absolute exposure ages for the coastal boulders from Rabat, the approach has great potential for boulder deposits composed of rocks with larger amounts of quartz or feldspar, older formation histories and lower susceptibility for erosion.

Atherosclerosis, the disease underlying stroke, myocardial infarction and ischemic heart failure, is initiated by the subendothelial retention of lipoproteins and cholesterol, which then triggers a non-resolving inflammatory process driving plaque progression in the artery wall. Myeloid cells and in particular macrophages are the primary driver of this process. IL10 is a prototypic anti-inflammatory cytokine made by several immune cells such as macrophages. IL10 is also essential for the protection against atherosclerosis. However, it is not known which cells are responsible for the IL10 production mediating protection from atherosclerosis. Here, we crossed the VertX mouse model, in which an IRES-enhanced green fluorescent protein (eGFP) fusion protein was placed downstream of exon 5 of the IL-10 gene, to the atherosclerosis-prone Apoe−/− background. We found that myeloid cells express high levels of IL10 GFP in VertX-Apoe−/− mice. By scRNAseq we found that macrophages especially inflammatory macrophages in plaques are the main producers of IL10 in atherosclerosis. To address whether IL10 secreted by myeloid cells is essential for the protection, we developed a LyzMCreIL-10flox/flox mouse crossed in the Apoe−/− background and confirmed that macrophages were unable to secrete IL-10. We discovered that the LyzMCreIL-10flox/floxApoe−/− mice developed significantly more atherosclerosis than LyzMCreIL-10+/+Apoe−/−, both on chow and lipid-rich diet. Flow cytometry data and cytokine measurements suggest that the depletion of IL-10 in myeloid cells increases Th17 cells with elevated IL-17a, IL-17f and CCL20 in blood plasma. These data underscore the critical roles of macrophage-derived IL-10 in limiting atherosclerosis. Supported by grants NIH HL 115232, 145241, and HL088093.

Background: CD4 + T cells play an important role in atherosclerosis, but their antigen specificity is poorly understood. Immunization with apolipoprotein B (ApoB, core protein of low density lipoprotein) is known to be atheroprotective in animal models. Here, we report on a human APOB peptide, p18, that is sequence-identical in mouse ApoB and binds to both mouse and human major histocompatibility complex class II molecules. Methods: We constructed p18 tetramers to detect human and mouse APOB-specific T cells and assayed their phenotype by flow cytometry including CD4 lineage transcription factors, intracellular cytokines, and T cell receptor activation. Apolipoprotein E–deficient ( Apoe −/− ) mice were vaccinated with p18 peptide or adjuvants alone, and atherosclerotic burden in the aorta was determined. Results: In human peripheral blood mononuclear cells from donors without cardiovascular disease, p18 specific CD4 + T cells detected by a new human leukocyte antigen-antigen D related-p18 tetramers were mostly Foxp3 + regulatory T cells (Tregs). Donors with subclinical cardiovascular disease as detected by carotid artery ultrasound had Tregs coexpressing retinoic acid-related orphan receptor gamma t or T-bet, which were both almost absent in donors without cardiovascular disease. In Apoe −/− mice, immunization with p18 induced Tregs and reduced atherosclerotic lesions. After peptide restimulation, responding CD4 + T cells identified by Nur77-GFP (green fluorescent protein) were highly enriched in Tregs. A new mouse I-A b -p18 tetramer identified the expansion of p18-specific CD4 + T cells on vaccination, which were enriched for interleukin-10–producing Tregs. Conclusions: These findings show that APOB p18–specific CD4 + T cells are mainly Tregs in healthy donors, but coexpress other CD4 lineage transcription factors in donors with subclinical cardiovascular disease. This study identifies ApoB peptide 18 as the first Treg epitope in human and mouse atherosclerosis.

Vaccination with MHC-II-restricted peptides from Apolipoprotein B (ApoB) with complete and incomplete Freund's adjuvant (CFA/IFA) is known to protect mice from atherosclerosis. This vaccination induces antigen-specific IgG1 and IgG2c antibody responses and a robust CD4 T cell response in lymph nodes. However, CFA/IFA cannot be used in humans. To find a clinically applicable adjuvant, we tested the effect of vaccinating Apoe-deficient mice with ApoB peptide P6 (TGAYSNASSTESASY). In a broad screening experiment, Addavax, a squalene-based oil-in-water adjuvant similar to MF59, was the only adjuvant that showed similar efficacy as CFA/IFA. This was confirmed in a confirmation experiment for both the aortic arch and whole aorta analyzed by en face analysis after atherosclerotic lesion staining. Mechanistically, restimulated peritoneal cells from mice immunized with P6 in Addavax released significant amounts of IL-10. Unlike P6 in CFA/IFA, vaccination with P6 in Addavax did not induce any detectable IgG1 or IgG2c antibodies to P6. These data suggest that squalene-based adjuvants such as MF59 are good candidate adjuvants for developing a clinically effective atherosclerosis vaccine.

Air pollution exposure positively correlates with increased cardiovascular morbidity and mortality rates, mainly due to myocardial infarction (MI). Herein, we aimed to study the metabolic mechanisms underlying this association, focusing on the evaluation of cardiac mitochondrial function and dynamics, together with its impact over MI progression. An initial time course study was performed in BALB/c mice breathing filtered air (FA) or urban air (UA) in whole-body exposure chambers located in Buenos Aires City downtown for up to 16 weeks (n = 8 per group and time point). After 12 weeks, lung inflammatory cell recruitment was evident in UA-exposed mice. Interestingly, impaired redox metabolism, characterized by decreased lung SOD activity and increased GSSG levels and NOX activity, precede local inflammation in this group. At this selected time point, additional mice were exposed to FA or UA (n = 12 per group) and alveolar macrophage PM uptake and nitric oxide (NO) production was observed in UA-exposed mice, together with increased pro-inflammatory cytokine levels (TNF-α and IL-6) in BAL and plasma. Consequently, impaired heart tissue oxygen metabolism and altered mitochondrial ultrastructure and function were observed in UA-exposed mice after 12 weeks, characterized by decreased active state respiration and ATP production rates, and enhanced mitochondrial H

Integrin-based therapeutics have garnered considerable interest in the medical treatment of inflammation. Integrins mediate the fast recruitment of monocytes and neutrophils to the site of inflammation, but are also required for host defense, limiting their therapeutic use. Here, we report a novel monoclonal antibody, anti-M7, that specifically blocks the interaction of the integrin Mac-1 with its pro-inflammatory ligand CD40L, while not interfering with alternative ligands. Anti-M7 selectively reduces leukocyte recruitment in vitro and in vivo. In contrast, conventional anti-Mac-1 therapy is not specific and blocks a broad repertoire of integrin functionality, inhibits phagocytosis, promotes apoptosis, and fuels a cytokine storm in vivo. Whereas conventional anti-integrin therapy potentiates bacterial sepsis, bacteremia, and mortality, a ligand-specific intervention with anti-M7 is protective. These findings deepen our understanding of ligand-specific integrin functions and open a path for a new field of ligand-targeted anti-integrin therapy to prevent inflammatory conditions., Integrin-based therapeutics could block inflammatory processes but they also impair host defence, limiting their usefulness. Here the authors report an anti-Mac1 antibody that blocks its interaction with pro-inflammatory ligand CD40L but not other ligands, and show that it can protect against sepsis in mice.

Background: Extracellular adenosine triphosphate (ATP) binds as a danger signal to purinergic receptor P2X 7 and promotes inflammasome assembly and interleukin-1β expression. We hypothesized a functional role of the signal axis ATP–P2X 7 in inflammasome activation and the chronic inflammation driving atherosclerosis. Methods: P2X 7 -competent and P2X 7 -deficient macrophages were isolated and stimulated with lipopolysaccharide, ATP, or both. To assess whether P2X 7 may have a role in atherosclerosis, P2X 7 expression was analyzed in aortic arches from low density lipoprotein receptor -/- mice consuming a high-cholesterol or chow diet. P2X 7 +/+ and P2X 7 −/− low density lipoprotein receptor −/− mice were fed a high-cholesterol diet to investigate the functional role of P2X 7 knockout in atherosclerosis. Human plaques were derived from carotid endarterectomy and stained against P2X 7 . Results: Lipopolysaccharide or ATP stimulation alone did not activate caspase 1 in isolated macrophages. However, priming with lipopolysaccharide, followed by stimulation with ATP, led to an activation of caspase 1 and interleukin-1β in P2X 7 -competent macrophages. In contrast, P2X 7 -deficient macrophages showed no activation of caspase 1 after sequential stimulation while still expressing a basal amount of interleukin-1β. P2X 7 receptor was higher expressed in murine atherosclerotic lesions, particularly by lesional macrophages. After 16 weeks of a high-cholesterol diet, P2X 7 -deficient mice showed smaller atherosclerotic lesions than P2X 7 -competent mice (0.162 cm 2 ±0.023 [n=9], P2X 7 −/− low density lipoprotein receptor −/− : 0.084 cm 2 ±0.01 [n=11], P =0.004) with a reduced amount of lesional macrophages. In accord with our in vitro findings, lesional caspase 1 activity was abolished in P2X 7 −/− mice. In addition, intravital microscopy revealed reduced leukocyte rolling and adhesion in P2X 7 -deficient mice. Last, we observe increased P2X 7 expression in human atherosclerotic lesions, suggesting that our findings in mice are relevant for human disease. Conclusions: P2X 7 deficiency resolved plaque inflammation by inhibition of lesional inflammasome activation and reduced experimental atherosclerosis. Therefore, P2X 7 represents an interesting potential new target to combat atherosclerosis.

Atopic dermatitis (AD) and psoriasis are driven by alternate type 2 and type 17 immune responses, but some proteins might be critical to both diseases. Here we show that a deficiency of the TNF superfamily molecule TWEAK (TNFSF12) in mice results in defective maintenance of AD-specific T helper type 2 (Th2) and psoriasis-specific Th17 cells in the skin, and impaired expression of disease-characteristic chemokines and cytokines, such as CCL17 and TSLP in AD, and CCL20 and IL-19 in psoriasis. The TWEAK receptor, Fn14, is upregulated in keratinocytes and dermal fibroblasts, and TWEAK induces these cytokines and chemokines alone and in synergy with the signature T helper cytokines of either disease, IL-13 and IL-17. Furthermore, subcutaneous injection of recombinant TWEAK into naive mice induces cutaneous inflammation with histological and molecular signs of both diseases. TWEAK is therefore a critical contributor to skin inflammation and a possible therapeutic target in AD and psoriasis., TWEAK is a TNF family member that binds the NFκB signalling receptor Fn14. Here the authors show that TWEAK is central to skin inflammation in mouse models of atopic dermatitis and psoriasis and causes similar pathology when injected subcutaneously into mice.

Although immunization with major histocompatibility complex (MHC) class II-restricted apolipoprotein B (ApoB) peptides has been shown to be atheroprotective, the mechanism is unclear. Here, we investigated CD4+ T cell populations in immunized atherosclerotic mice. Peptides (16-mers) from mouse ApoB, the core protein of low-density lipoprotein (LDL), were screened for binding to I-Ab by computer prediction and confirmed by radiolabeled peptide competition. Three new peptides, P101 (FGKQGFFPDSVNKALY, 5.5 nM IC50), P102 (TLYALSHAVNSYFDVD, 6.8 nM), and P103 (LYYKEDKTSLSASAAS, 95 nM), were tested in an atherosclerosis model ( Apoe−/− mice on Western diet). Immunization with each of the three peptides (1 time in complete Freund’s adjuvant subcuntaneously and 4 time in incomplete Freund’s adjuvant intraperitoneally) but not with adjuvant alone showed significantly reduced atherosclerotic plaques in the aortic root by serial sections and in the whole aorta by en face staining. There were no differences in body weight, LDL cholesterol, or triglycerides. Peritoneal leukocytes from ApoB peptide-immunized mice, but not control mice, secreted significant amounts of IL-10 (150 pg/ml). Flow cytometry showed that peptide immunization induced IL-10 in 10% of peritoneal CD4+ T cells, some of which also expressed chemokine (C-C motif) receptor 5 (CCR5). Vaccination with ApoB peptides expanded peritoneal FoxP3+ regulatory CD4+ T cells and more than tripled the number of CCR5+FoxP3+ cells. Similar trends were also seen in the draining mediastinal lymph nodes but not in the nondraining inguinal lymph nodes. We conclude that vaccination with MHC class II-restricted autologous ApoB peptides induces regulatory T cells (Tregs) and IL-10, suggesting a plausible mechanism for atheroprotection. NEW & NOTEWORTHY Vaccination against apolipoprotein B (ApoB), the protein of LDL, attracts attention as a novel approach to prevent atherosclerosis. We discovered major histocompatibility complex class II-restricted ApoB peptides, which reduce atherosclerosis and induce IL-10-producing CD4+ T cells and chemokine (C-C motif) receptor 5 expression on regulatory T cells, suggesting that immunization with ApoB peptides inhibits atherosclerosis by inducing anti-inflammatory cytokines.

Aim: The present study analyzes in depth the impact of different calcification patterns on disturbances of the conduction system in transcatheter aortic valve replacement (TAVR) patients. Methods and Results: A total of 169 preprocedural TAVR multislice computed tomography scans from consecutive transfemoral (TF) TAVRs performed between 2014 and 2017 using either Edwards SAPIEN or Medtronic Evolut R valves were retrospectively evaluated. The volume, distribution, and orientation of annular and valvular aortic valve calcification were measured and their impact on postoperative conduction disturbances was determined using linear and logistic regression analyses. The total volume of calcification and distribution at the aortic annulus or valve did not influence the conduction system. Oval calcification of the left aortic cusp was independently associated with an elevated risk for an increase in atrioventricular block degree (+0.6, p = 0.03). Moreover, orthogonal calcifications at the level of the aortic annulus were associated with an increased risk for QRS prolongation (+26 ms, p = 0.004) and an increased risk for permanent pacemaker implantation (OR 4.3, p = 0.03) after TF TAVR. This was more pronounced in patients undergoing TF TAVR using a balloon-expandable Edwards SAPIEN 3 valve (QRS +38.195 ms, p < 0.001; OR permanent pacemaker 15.48, p = 0.013). Conclusion: Orthogonal annular calcification confers an increased risk for conduction disturbances after TAVR. This is even more pronounced after implantation of balloon-expandable valves.

Supplemental Digital Content is available in the text., Background: Throughout the inflammatory response that accompanies atherosclerosis, autoreactive CD4+ T-helper cells accumulate in the atherosclerotic plaque. Apolipoprotein B100 (apoB), the core protein of low-density lipoprotein, is an autoantigen that drives the generation of pathogenic T-helper type 1 (TH1) cells with proinflammatory cytokine secretion. Clinical data suggest the existence of apoB-specific CD4+ T cells with an atheroprotective, regulatory T cell (Treg) phenotype in healthy individuals. Yet, the function of apoB-reactive Tregs and their relationship with pathogenic TH1 cells remain unknown. Methods: To interrogate the function of autoreactive CD4+ T cells in atherosclerosis, we used a novel tetramer of major histocompatibility complex II to track T cells reactive to the mouse self-peptide apo B978-993 (apoB+) at the single-cell level. Results: We found that apoB+ T cells build an oligoclonal population in lymph nodes of healthy mice that exhibit a Treg-like transcriptome, although only 21% of all apoB+ T cells expressed the Treg transcription factor FoxP3 (Forkhead Box P3) protein as detected by flow cytometry. In single-cell RNA sequencing, apoB+ T cells formed several clusters with mixed TH signatures that suggested overlapping multilineage phenotypes with pro- and anti-inflammatory transcripts of TH1, T helper cell type 2 (TH2), and T helper cell type 17 (TH17), and of follicular-helper T cells. ApoB+ T cells were increased in mice and humans with atherosclerosis and progressively converted into pathogenic TH1/TH17-like cells with proinflammatory properties and only a residual Treg transcriptome. Plaque T cells that expanded during progression of atherosclerosis consistently showed a mixed TH1/TH17 phenotype in single-cell RNA sequencing. In addition, we observed a loss of FoxP3 in a fraction of apoB+ Tregs in lineage tracing of hyperlipidemic Apoe–/– mice. In adoptive transfer experiments, converting apoB+ Tregs failed to protect from atherosclerosis. Conclusions: Our results demonstrate an unexpected mixed phenotype of apoB-reactive autoimmune T cells in atherosclerosis and suggest an initially protective autoimmune response against apoB with a progressive derangement in clinical disease. These findings identify apoB autoreactive Tregs as a novel cellular target in atherosclerosis.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Diabetes worsens atherosclerosis progression and leads to a defect in repair of arteries after cholesterol reduction, a process termed regression. Empagliflozin reduces blood glucose levels via inhibition of the sodium glucose cotransporter 2 (SGLT-2) in the kidney and has been shown to lead to a marked reduction in cardiovascular events in humans. To determine whether glucose lowering by empagliflozin accelerates atherosclerosis regression in a mouse model, male C57BL/6J mice were treated intraperitoneally with LDLR- and SRB1- antisense oligonucleotides and fed a high cholesterol diet for 16 weeks to induce severe hypercholesterolemia and atherosclerosis progression. At week 14 all mice were rendered diabetic by streptozotocin (STZ) injections. At week 16 a baseline group was sacrificed and displayed substantial atherosclerosis of the aortic root. In the remaining mice, plasma cholesterol was lowered by switching to chow diet and treatment with LDLR sense oligonucleotides to induce atherosclerosis regression. These mice then received either empagliflozin or vehicle for three weeks. Atherosclerotic plaques in the empagliflozin treated mice were significantly smaller, showed decreased lipid and CD68+ macrophage content, as well as greater collagen content. Proliferation of plaque resident macrophages and leukocyte adhesion to the vascular wall were significantly decreased in empagliflozin-treated mice. In summary, plasma glucose lowering by empagliflozin improves plaque regression in diabetic mice.

Introduction Many clinical and experimental observations have established an association between visceral obesity and chronic adipose tissue inflammation. Potent pro-inflammatory mediators such as TNFα, CD40 and IL-1β are regulated by Tumor necrosis factor (TNF) receptor-associated factors (TRAFs). TRAF5 deficiency accelerates atherogenesis in mice by increasing inflammatory leukocyte recruitment. Since inflammatory cell invasion is also a prerequisite of adipose tissue inflammation, we tested the hypothesis that deficient TRAF5 signaling aggravates adipose tissue inflammation and its metabolic complications in a murine diet-induced obesity (DIO) model. Purpose We aimed to clarify the role of TRAF5 in adipose tissue inflammation and metabolic syndrome. Methods TRAF5−/− mice and gender- and age-matched wild-type (WT) mice consumed a high fat diet (HFD, 45%kcal from fat) or a matched low-fat diet (LFD, 10%kcal from fat) for 18 weeks to induce DIO and adipose tissue inflammation. All mice were then subjected to subsequent analysis, including glucose and insulin tolerance testing, body composition assessment by MRI imaging, flow cytometry, gene expression of different tissues, plasma analysis and histology. Finally, we studied if TRAF5 expression was associated with metabolic syndrome in humans by analyzing plasma and adipocytes samples from 62 patients of the Tumor-Necrosis-Factor Receptor Associated in Cardiovascular Risk Study (TRAFICS). Results TRAF5 expression was significantly attenuated in isolated WT-adipocytes and WT-macrophages after 18 weeks of HFD compared to LFD-fed controls. TRAF5−/− mice on HFD gained significantly more weight compared to TRAF5-competent mice and showed an aggravated metabolic phenotype, including impaired insulin tolerance, hyperinsulinemia and increased fasting glucose plasma levels. The weight gain in TRAF5−/− mice was attributable to a significant increase in adipose tissue and liver weight. Further analysis of the visceral adipose tissue revealed enhanced macrophage accumulation and increased pro-inflammatory CD11c+ subset polarization in HFD-fed TRAF5−/− mice. In line with an increased migratory capacity of inflammatory cells, we observed enhanced peritoneal invasion of leukocytes and subsets in TRAF5−/− mice. Accordingly, TRAF5 deficiency increased inflammatory cytokine expression and ameliorated parameters of insulin sensitivity in adipose tissue. Finally, patients with metabolic syndrome displayed decreased TRAF5 expression in blood and adipocytes compared to humans without metabolic syndrome. Conclusion We show that genetic deficiency of TRAF5 aggravates metabolic syndrome in murine diet-induced obesity. Enhanced accumulation of leukocytes subsets in adipose tissue serves as the likely mechanism. We conclude that TRAF5 signaling properties may favorably affect metabolic disease. Acknowledgement/Funding Forschungskommission Medizinische Fakultät Universität Freiburg, MOTI-VATE Promotionskolleg der Medizinischen Fakultät Freiburg (EKFS)

Background Guidelines recommend cholesterol lowering for primary and secondary prevention of cardiovascular disease. While lipid lowering has been reported to induce plaque regression, the underlying mechanisms have remained speculative. Purpose We hypothesize that lipid uptake triggers local macrophage proliferation in the plaque, and conversely, statin treatment inhibits local macrophage proliferation leading to plaque regression. Methods Mixed bone marrow chimeras were generated in LDLR−/− mice reconstituted with wild type and scavenger receptor deficient or cholesterol exporter deficient bone marrow cells to study cell autonomous effects on macrophage proliferation. APOE*3-Leiden.huCETP mice with established atherosclerosis were randomized to three groups: Continued cholesterol diet, cholesterol diet supplemented with 0.01% atorvastatin, and cholesterol free diet for 4 weeks to study mechanisms of plaque regression. Results Proliferation of scavenger receptor A and CD36 deficient macrophages with impaired lipid uptake was reduced by 30–50% in the plaque, while ABCA1/ABCG1 exporter deficiency resulted in cholesterol overloading and apoptosis. Oral atorvastatin treatment decreased total plasma cholesterol levels by 50% to the same extend as cholesterol free diet feeding in APOE*3-Leiden.huCETP. Cholesterol lowering resulted in a 50% reduction in local macrophage proliferation and plaque regression with reduced macrophage and lipid contents and increased collagen. GFP bone marrow reconstitution of APOE*3-Leiden.huCETP mice in which the aortas were shielded from irradiation showed infiltrating monocytes to contribute only 11% to the plaque macrophage pool during plaque progression, thereby underscoring the relevance of targeting macrophage proliferation for plaque regression. Finally, rates of macrophage proliferation in human carotid artery plaques correlated with serum LDL-cholesterol levels, in line with our experimental studies. Conclusion Foam cell formation in atherosclerotic plaques triggers their proliferation. Targeting macrophage proliferation leads to plaque regression.

Since the introduction of augmented reality (AR) technology, in-situ instructions for manual tasks have been a central use case for a large body of previous work. However, most implementations provide identical sets of instructions to each user disregarding the user's current mental load. This is a major issue since previous work has shown the importance and potential of an adapted instruction fidelity for manual tasks such as playing an instrument. To implement a low-cost mental load adaptation for AR instructions, we evaluated a mobile off-the-shelf electroencephalographic (EEG) device for its suitability and feasibility to measure mental load while wearing a video see-through AR head-mounted display (HMD). In a first user experiment (n=12), data of EEG power band values and proprietary performance metrics of the manufacturer were collected and analysed regarding their validity to estimate the user's mental load. Our results indicate that our setup successfully induced different levels of mental effort. The proprietary performance metrics, however, only partially reflected the participants' current mental effort and require further analysis.

Background Inflammation drives atherosclerosis and its complications. Anti-inflammatory therapy with interleukin 1 beta (IL-1β) antibody reduces cardiovascular events in patients with elevated high-sensitive C-reactive protein (hsCRP). This study aims to identify the share of patients with coronary heart disease (CHD) and residual inflammation who may benefit from anti-inflammatory therapy. Methods hsCRP and low-density lipoprotein (LDL) levels were determined in 2741 all-comers admitted to the cardiological ward of our tertiary referral hospital between June 2016 and June 2018. Patients without CHD, with acute coronary syndrome, chronic or recurrent systemic infection, use of immunosuppressant or anti-inflammatory agents, chronic inflammatory diseases, chemotherapy, terminal organ failure, traumatic injury and pregnancy were excluded. Results 856 patients with stable CHD were included. 42.7% of those had elevated hsCRP ≥ 2 mg/l. Within the group of patients with LDL-cholesterol

Background: Platelets store large amounts of serotonin that they release during thrombus formation or acute inflammation. This facilitates hemostasis and modulates the inflammatory response. Methods: Infarct size, heart function, and inflammatory cell composition were analyzed in mouse models of myocardial reperfusion injury with genetic and pharmacological depletion of platelet serotonin. These studies were complemented by in vitro serotonin stimulation assays of platelets and leukocytes in mice and men, and by measuring plasma serotonin levels and leukocyte activation in patients with acute coronary syndrome. Results: Platelet-derived serotonin induced neutrophil degranulation with release of myeloperoxidase and hydrogen peroxide (H2O2) and increased expression of membrane-bound leukocyte adhesion molecule CD11b, leading to enhanced inflammation in the infarct area and reduced myocardial salvage. In patients hospitalized with acute coronary syndrome, plasmatic serotonin levels correlated with CD11b expression on neutrophils and myeloperoxidase plasma levels. Long-term serotonin reuptake inhibition - reported to protect patients with depression from cardiovascular events - resulted in the depletion of platelet serotonin stores in mice. These mice displayed a reduction in neutrophil degranulation and preserved cardiac function. In line, patients with depression using serotonin reuptake inhibition, presented with suppressed levels of CD11b surface expression on neutrophils and lower myeloperoxidase levels in blood. Conclusions: Taken together, we identify serotonin as a potent therapeutic target in neutrophil-dependent thromboinflammation during myocardial reperfusion injury. Fil: Mauler, Maximilian. No especifíca; Fil: Herr, Nadine. No especifíca; Fil: Schoenichen, Claudia. No especifíca; Fil: Witsch, Thilo. No especifíca; Fil: Marchini, Timoteo Oscar. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Bioquímica y Medicina Molecular. Universidad de Buenos Aires. Facultad Medicina. Instituto de Bioquímica y Medicina Molecular; Argentina Fil: Härdtner, Carmen. No especifíca; Fil: Koentges, Christoph. No especifíca; Fil: Kienle, Korbinian. Max Planck Institute Of Immunobiology And Epigenetics; Alemania Fil: Ollivier, Véronique. Inserm; Francia Fil: Schell, Maximilian. No especifíca; Fil: Dorner, Ludwig. No especifíca; Fil: Wippel, Christopher. No especifíca; Fil: Stallmann, Daniela. No especifíca; Fil: Normann, Claus. No especifíca; Fil: Bugger, Heiko. No especifíca; Fil: Walther, Paul. Universitat Ulm; Alemania Fil: Wolf, Dennis. La Jolla Institute for Allergy and Immunology; Estados Unidos Fil: Ahrens, Ingo. No especifíca; Fil: Lämmermann, Tim. Max Planck Institute Of Immunobiology And Epigenetics; Alemania Fil: Ho-Tin-Noé, Benoît. Inserm; Francia Fil: Ley, Klaus. La Jolla Institute for Allergy and Immunology; Estados Unidos Fil: Bode, Christoph. No especifíca; Fil: Hilgendorf, Ingo. No especifíca; Fil: Duerschmied, Daniel. No especifíca

Using 10× Genomics-based single-cell RNA sequencing on digested atherosclerotic mouse aortas, we discovered an unexpected immune cell population whose transcriptome was similar to thymic CD4+CD8+ cells (DPs). Flow cytometry showed DP presence in the aortic arch (AA) and mediastinal adipose tissue (MAT), but not in lymph nodes, spleen, blood or other tissues. DPs in MAT or AA did not change with atherosclerosis in Apoe−/− mice. Fluorescence microscopy of cleared whole mounts placed DPs in the AA adventitia and near microvessels in MAT. MAT and AA DPs were absent in Rag2−/− mice and in day3 and 21 thymectomized mice. Deep transcriptional profiling from sorted thymic, MAT, and AA DPs showed hundreds of differentially expressed genes. While thymic DPs were enriched for TCR signaling pathways, extra-thymic DPs were enriched for stress responses and cell cycle pathways. Pulse-chase experiments demonstrated a five-day turnover of MAT and AA DPs. Functionally, sorted MAT DPs developed successfully into conventional T cells (with a bias to CD4) in hanging drop cultures. Pharmacologic inhibition of cell migration by S1P1 blockage with FTY720 did not affect the presence of these cells, suggesting that migration of extra-thymic DPs does not require passage into blood. Transplantation of congenically marked thymi under the kidney capsule of thymectomized mice reconstituted T cells in the inguinal lymph node, but failed to reconstitute extra-thymic DPs. The discovery of two new, transcriptionally distinct subsets of extrathymic DPs reveals a novel, unexpected local escape mechanism from the thymus. Their association with blood vessels suggests possible angiotrophic functions.

Sterile inflammation of visceral fat, provoked by dying adipocytes, links the metabolic syndrome to cardiovascular disease. Danger-associated molecular patterns, such as adenosine triphosphate (ATP), are released by activated or dying cells and orchestrate leukocyte infiltration and inflammation via the purinergic receptor P2Y2. The gene expression of ATP receptor P2Y2 did not change in several tissues in the course of obesity, but was increased within epididymal fat. Adipose tissue from P2Y 2 −/− mice consuming high-fat diet (HFD) contained less crown-like structures with a reduced frequency of adipose tissue macrophages (ATMs). This was likely due to decreased leukocyte migration because of missing VCAM-1 exposition on P2Y2 deficient hypertrophic adipose tissue endothelial cells. Accordingly, P2Y 2 −/− mice showed blunted traits of the metabolic syndrome: they gained less weight compared to P2Y 2 +/+ controls, while intake of food and movement behaviour remained unchanged. Liver and adipose tissue were smaller in P2Y 2 −/− animals. Insulin tolerance testing (ITT) performed in obese P2Y 2 −/− mice revealed a better insulin sensitivity as well as lower plasma C-peptide and cholesterol levels. We demonstrate that interfering with somatic P2Y2 signalling prevents excessive immune cell deposition in diet-induced obesity (DIO), both attenuating adipose tissue inflammation and ameliorating the metabolic phenotype. Thus, blocking the P2Y2 cascade may be a promising strategy to limit metabolic disease and its sequelae.

X-ray fluoroscopy is the gold standard for coronary diagnostics and intervention. Magnetic resonance imaging is a radiation-free alternative to x-ray with excellent soft tissue contrast in arbitrary slice orientation. Here, we assessed real-time MRI-guided coronary interventions from femoral access using newly designed MRI technologies. Six Goettingen minipigs were used to investigate coronary intervention using real-time MRI. Catheters were custom-designed and equipped with an active receive tip-coil to improve visibility and navigation capabilities. Using modified standard clinical 5 F catheters, intubation of the left coronary ostium was successful in all animals. For the purpose of MR-guided coronary interventions, a custom-designed 8 F catheter was used. In spite of the large catheter size, and therefore limited steerability, intubation of the left coronary ostium was successful in 3 of 6 animals within seconds. Thereafter, real-time guided implantation of a non-metallic vascular scaffold into coronary arteries was possible. This study demonstrates that real-time MRI-guided coronary catheterization and intervention via femoral access is possible without the use of any contrast agents or radiation, including placement of non-metallic vascular scaffolds into coronary arteries. Further development, especially in catheter and guidewire technology, will be required to drive forward routine MR-guided coronary interventions as an alternative to x-ray fluoroscopy.

The scarcity of established input methods for augmented reality (AR) head-mounted displays (HMD) motivates us to investigate the performance envelopes of two easily realisable solutions: indirect cursor control via a smartwatch and direct control by in-air touch. Indirect cursor control via a smartwatch has not been previously investigated for AR HMDs. We evaluate these two techniques for carrying out three fundamental user interface actions: target acquisition, goal crossing, and circular steering. We find that in-air is faster than smartwatch (p < 0.001) for target acquisition and circular steering. We observe, however, that in-air selection can lead to discomfort after extended use and suggest that smartwatch control offers a complementary alternative.

Purpose of review The immune system plays a critical role in the development and modulation of atherosclerosis. New high-parameter technologies, including mass cytometry (CyTOF) and single-cell RNA sequencing (scRNAseq), allow for an encompassing analysis of immune cells. Unexplored marker combinations and transcriptomes can define new immune cell subsets and suggest their functions. Here, we review recent advances describing the immune cells in the artery wall of mice with and without atherosclerosis. We compare technologies and discuss limitations and advantages. Recent findings Both CyTOF and scRNAseq on leukocytes from digested aortae show 10-30 immune cell subsets. Myeloid, T, B and natural killer cells were confirmed. Although cellular functions can be inferred from RNA-Seq data, some subsets cannot be identified based on current knowledge, suggesting they may be new cell types. CyTOF and scRNAseq each identified four B-cell subsets and three macrophage subsets in the atherosclerotic aorta. Limitations include cell death caused by enzymatic digestion and the limited depth of the scRNAseq transcriptomes. Summary High-parameter methods are powerful tools for uncovering leukocyte diversity. CyTOF is currently more powerful at discerning leukocyte subsets in the atherosclerotic aorta, whereas scRNAseq provides more insight into their likely functions.

This paper is about AMIDAR, a processor concept for an adaptive processor that uses a Coarse Grained Reconfigurable Array as a configurable accelerator to dynamically speed up hot spots of arbitrary applications. We describe the concepts and implementation details and we summarize some central findings of more than 15 years of research around this topic. Eventually, we demonstrate a fully working hardware implementation and show the potential effect of a configurable hardware accelerator.

Background: Integrins, such as Mac-1 (alphaMbeta2, CD11b/CD18), drive myeloid cell recruitment and inflammation in cardiovascular disease. Although integrins have generated interest as therapeutic targets in acute and chronic inflammation, they also contribute to host defense, regeneration, and haemostasis. To overcome these limitations, we have designed a novel antibody that targets a distinct region within the Mac-1 major ligand-binding (I-) domain required for binding of its ligand CD40L. Here, we describe the consequences of a ligand-specific inhibition and its efficacy in mice. Methods and Results: To generate an antibody specifically targeting the interaction of Mac-1 with CD40L, mice were immunized with a peptide containing the CD40L-binding motif EQLKKSKTL. Antibody clones were screened and tested for specificity. One clone, termed anti-M7, showed specific peptide binding and blocked the binding of the activated, open conformation of Mac-1 to CD40L, but not to alternative Mac-1 ligands suggesting ligand specificity. Anti-M7 highly effectively limited leukocyte recruitment in vitro and to the mesentery and the omentum in intravital confocal microscopy. While conventional anti-Mac-1 antibodies induced outside-in activation of the integrin, anti-M7 did not provoke MAP-kinase dependent activation and expression of inflammatory cytokines in macrophages. Following cecal-ligation and puncture (CLP) sepsis in mice, a conventional anti-Mac-1 antibody inhibited leukocyte accumulation, but potentiated bacteremia and limited overall survival. In contrast, anti-M7 limited peritoneal leukocyte accumulation, reduced bacterial titers, and improved survival during CLP. Conclusion: We present proof-of-concept demonstrating selective inhibition of the interaction between the leukocyte integrin Mac-1 and CD40L by a novel monoclonal antibody. This antibody shows several advantages over conventional anti-integrin therapy, and prevented inflammation and sepsis-related mortality in mice. This novel approach gives new insight into ligand-specific integrin pathways and merits further evaluation as a selective anti-inflammatory therapy in cardiovascular disease.

Background: The signaling pathways linking inflammatory and thrombotic circuits in platelets are only poorly understood. Here, we tested the role of the inflammatory signaling adapter TRAF-1, which bundles TNF-, TLR, and IL1-signaling, in platelets. Methods and Results: To establish a role for platelet expressed TRAF-1, we verified its expression in in vitro generated mouse thrombi in immunohistochemistry and western blot. Blood clots generated from blood of Traf1 -deficient mice were smaller, suggestive of a defective plasmatic coagulation. In accord, tail bleeding time was increased by ~4-fold in Traf1 -/- mice. Genetically chimeric mice generated by bone marrow transplantations with a selective deficiency of Traf1 in bone-marrow-derived leukocytes showed no changes in bleeding time, suggesting that the prolonged bleeding time in Traf1 -/- mice was regulated by vascular/stromal cells. In a gene expression array of Traf1 -/- endothelial cells, several factors that regulate coagulation, including fibrinogen, tumor-homing peptide (F3), and Von Willebrand factor (vWF) were reduced. In addition to this vascular phenotype, expression of P-selectin and the activation epitope Jon/A induced by in vitro ADP and thrombin stimulation was reduced in TRAF-1-deficient platelets. As a consequence, in vivo thrombus-generation in the mesenterium was delayed with an enhanced rate of emboli in Traf1 -/- mice – an effect that was confirmed in mice transferred with TRAF1 -/- platelets, and in mice with a selective deficiency of Traf1 in bone-marrow-derived cells. Finally, we demonstrate TRAF-1 protein expression in human coronary thrombi and the presence of TRAF1 -transcripts in RNA-sequencing of human platelets. TRAF1 -mRNA expression was down-regulated in collagen and TRAP stimulated human platelets and correlated with the gene expression of several upstream platelet-receptors, including EDA2R , RELT , and CD137. Conclusion: We present the novel finding that the pro-inflammatory signaling adapter TRAF-1 is expressed in mouse and human thrombi and participates in coagulation and thrombosis by vascular and platelet-mediated pathways. These findings emphasize the connection of inflammatory signaling and haemostasis.

Atherosclerosis, an inflammatory disease of large arteries, is - through its clinical manifestations stroke and myocardial infarction - globally the leading cause of morbidity and mortality. The interplay of pro- and anti-inflammatory leukocytes in the aorta modulates and drives atherosclerosis. Although cells of the innate and adaptive immune system are found in atherosclerotic plaques, their phenotypic and functional diversity is poorly understood. Here, we applied single cell RNA-sequencing (scRNAseq) and mass cytometry (CyTOF) to assess leukocyte diversity in depth, thus defining an immune cell atlas in atherosclerosis. Single cell transcriptional profiling of aortic leukocytes from 20-week old chow (CD) and western diet (WD) fed Apoe -/- and Ldlr -/- mice revealed 11 phenotypically different leukocyte clusters. Atherosclerotic aortas exhibited enhanced leukocyte diversity, whilst WD further changed the abundance of leukocyte subpopulations. Gene set enrichment analysis of single cells established that multiple pathways, e.g. for lipid metabolism, proliferation, and cytokine secretion, pertained to particular leukocyte clusters. Applying a novel 35-marker CyTOF panel with metal-labelled antibodies confirmed the phenotypic diversity of aortic leukocytes. Among lymphocytes, we detected three principal B-cell subsets defined by scRNAseq, CyTOF, and flow cytometry. These B cell subsets harbor distinct surface marker expression, functional gene pathways, and ex vivo cytokine production. Finally, we used leukocyte cluster gene signatures to assess leukocyte frequencies in 121 human plaques by a transcriptomic deconvolution strategy. This approach revealed a similar immune cell complexity in human carotid plaques with a higher percentage of monocytes and macrophages. In addition, the frequency of genetically defined leukocyte populations in carotid plaques predicted cardiovascular events in patients. The definition of leukocyte diversity by high-dimensional analyses enables a fine-grained analysis of aortic leukocyte subsets, reveals new immunological mechanisms and cell-type specific pathways, and may result in novel diagnostic risk stratification tools.

Rationale: Atherosclerosis is a chronic inflammatory disease that is driven by the interplay of pro- and anti-inflammatory leukocytes in the aorta. Yet, the phenotypic and transcriptional diversity of aortic leukocytes is poorly understood. Objective: We characterized leukocytes from healthy and atherosclerotic mouse aortas in-depth by single-cell RNA-sequencing and mass cytometry (cytometry by time of flight) to define an atlas of the immune cell landscape in atherosclerosis. Methods and Results: Using single-cell RNA-sequencing of aortic leukocytes from chow diet– and Western diet–fed Apoe −/− and Ldlr −/− mice, we detected 11 principal leukocyte clusters with distinct phenotypic and spatial characteristics while the cellular repertoire in healthy aortas was less diverse. Gene set enrichment analysis on the single-cell level established that multiple pathways, such as for lipid metabolism, proliferation, and cytokine secretion, were confined to particular leukocyte clusters. Leukocyte populations were differentially regulated in atherosclerotic Apoe −/− and Ldlr −/− mice. We confirmed the phenotypic diversity of these clusters with a novel mass cytometry 35-marker panel with metal-labeled antibodies and conventional flow cytometry. Cell populations retrieved by these protein-based approaches were highly correlated to transcriptionally defined clusters. In an integrated screening strategy of single-cell RNA-sequencing, mass cytometry, and fluorescence-activated cell sorting, we detected 3 principal B-cell subsets with alterations in surface markers, functional pathways, and in vitro cytokine secretion. Leukocyte cluster gene signatures revealed leukocyte frequencies in 126 human plaques by a genetic deconvolution strategy. This approach revealed that human carotid plaques and microdissected mouse plaques were mostly populated by macrophages, T-cells, and monocytes. In addition, the frequency of genetically defined leukocyte populations in carotid plaques predicted cardiovascular events in patients. Conclusions: The definition of leukocyte diversity by high-dimensional analyses enables a fine-grained analysis of aortic leukocyte subsets, reveals new immunologic mechanisms and cell-type–specific pathways, and establishes a functional relevance for lesional leukocytes in human atherosclerosis.

Rationale: The coincidence of inflammation and metabolic derangements in obese adipose tissue has sparked the concept of met-inflammation. Previous observations, however, suggest that inflammatory pathways may not ultimately cause dysmetabolism. Objective: We have revisited the relationship between inflammation and metabolism by testing the role of TRAF (tumor necrosis receptor-associated factor)-1, an inhibitory adapter of inflammatory signaling of TNF (tumor necrosis factor)-a, IL (interleukin)-1ß, and TLRs (toll-like receptors). Methods and Results: Mice defcient for TRAF-1, which is expressed in obese adipocytes and adipose tissue lymphocytes, caused an expected hyperinflammatory phenotype in adipose tissue with enhanced adipokine and chemokine expression, increased leukocyte accumulation, and potentiated proinflammatory signaling in macrophages and adipocytes in a mouse model of diet-induced obesity. Unexpectedly, TRAF-1-/-mice were protected from metabolic derangements and adipocyte growth, failed to gain weight, and showed improved insulin resistance-an effect caused by increased lipid breakdown in adipocytes and UCP (uncoupling protein)-1-enabled thermogenesis. TRAF-1-dependent catabolic and proinflammatory cues were synergistically driven by ß3-adrenergic and inflammatory signaling and required the presence of both TRAF-1-defcient adipocytes and macrophages. In human obesity, TRAF-1-dependent genes were upregulated. Conclusions: Enhancing TRAF-1-dependent inflammatory pathways in a gain-of-function approach protected from metabolic derangements in diet-induced obesity. These fndings identify TRAF-1 as a regulator of dysmetabolism in mice and humans and question the pathogenic role of chronic inflammation in metabolism. Fil: Michel, Nathaly Anto. University of Freiburg. University Medical Center; Alemania Fil: Colberg, Christian. University of Freiburg. University Medical Center; Alemania Fil: Buscher, Konrad. La Jolla Institute for Allergy and Immunology; Estados Unidos Fil: Sommer, Björn. Universitat Erlangen-Nuremberg; Alemania Fil: Pramod, Akula Bala. La Jolla Institute for Allergy and Immunology; Estados Unidos Fil: Ehinger, Erik. La Jolla Institute for Allergy and Immunology; Estados Unidos Fil: Dufner, Bianca. University of Freiburg. University Medical Center; Alemania Fil: Hoppe, Natalie. University of Freiburg. University Medical Center; Alemania Fil: Pfeiffer, Katharina. University of Freiburg. University Medical Center; Alemania Fil: Marchini, Timoteo Oscar. University of Freiburg. University Medical Center; Alemania. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Bioquímica y Medicina Molecular. Universidad de Buenos Aires. Facultad Medicina. Instituto de Bioquímica y Medicina Molecular; Argentina Fil: Willecke, Florian. University of Freiburg. University Medical Center; Alemania Fil: Stachon, Peter. University of Freiburg. University Medical Center; Alemania Fil: Hilgendorf, Ingo. University of Freiburg. University Medical Center; Alemania Fil: Heidt, Timo. University of Freiburg. University Medical Center; Alemania Fil: Von Zur Muhlen, Constantin. University of Freiburg. University Medical Center; Alemania Fil: Von Elverfeldt, Dominik. University of Freiburg. University Medical Center; Alemania Fil: Pfeifer, Dietmar. University of Freiburg; Alemania Fil: Schüle, Roland. University of Freiburg; Alemania Fil: Kintscher, Ulrich. University of Freiburg; Alemania Fil: Brachs, Sebastian. Center For Cardiovascular Research; Alemania Fil: Ley, Klaus. La Jolla Institute for Allergy and Immunology; Estados Unidos Fil: Bode, Christoph. University of Freiburg. University Medical Center; Alemania Fil: Zirlik, Andreas. University of Freiburg. University Medical Center; Alemania Fil: Wolf, Dennis. La Jolla Institute for Allergy and Immunology; Estados Unidos

Background— The aim of this study is to examine the relationship between time in the therapeutic range (TTR) and clinical outcomes in heart failure patients in sinus rhythm treated with warfarin. Methods and Results— We used data from the Warfarin versus Aspirin in Reduced Cardiac Ejection Fraction (WARCEF) trial to assess the relationship of TTR with the WARCEF primary outcome (ischemic stroke, intracerebral hemorrhage, or death), with death alone, ischemic stroke alone, major hemorrhage alone, and net clinical benefit (primary outcome and major hemorrhage combined). Multivariable Cox models were used to examine how the event risk changed with TTR and to compare the high TTR, low TTR, and aspirin-treated patients, with TTR being treated as a time-dependent covariate. A total of 2217 patients were included in the analyses; among whom 1067 were randomized to warfarin and 1150 were randomized to aspirin. The median (interquartile range) follow-up duration was 3.6 (2.0–5.0) years. Mean (±SD) age was 61±11.3 years, with 80% being men. The mean (±SD) TTR was 57% (±28.5%). Increasing TTR was significantly associated with reduction in primary outcome (adjusted P P =0.001), and improved net clinical benefit (adjusted P P =0.082 for ischemic stroke and adjusted P =0.109 for major hemorrhage). Conclusions— In patients with heart failure in sinus rhythm, increasing TTR is associated with better outcome and improved net clinical benefit. Patients in whom good quality anticoagulation can be achieved may benefit from the use of anticoagulants. Clinical Trial Registration— URL: http://www.clinicaltrials.gov . Unique identifier: NCT00041938.

Sterile inflammation of visceral fat, provoked by dying adipocytes, links the metabolic syndrome to cardiovascular disease. Danger-associated molecular patterns, such as adenosine triphosphate (ATP), are released by activated or dying cells and orchestrate leukocyte infiltration and inflammation via the purinergic receptor P2Y

A substantial proportion of cardiovascular mortality is caused by atherosclerosis of large and midsized arteries that supply the brain. Stroke is the main complication of cerebral atherosclerosis and ranks second as cause of death worldwide.1 Recruitment of some myeloid cell subpopulations to atherosclerotic lesions can drive plaque inflammation and predispose to stroke. After a stroke, myeloid cells may also be harmful by directly aggravating the reperfusion injury to the brain. The latter possibility is of significant clinical relevance because epidemiological data have suggested that increased numbers of circulating leukocytes, particularly monocytes and neutrophils, after stroke correlate with poor outcomes.2 However, the functional mechanistic links between monocytosis and neutrophilia and stroke were unknown. Article, see p 407 In the current issue of Circulation Research , Courties, Nahrendorf et al addressed this intriguing question and elucidate how stroke speeds up the turnover of hematopoietic stem cells (HSCs) to specifically generate myeloid cell populations and release them into the blood circulation.3 In their study, the authors have chosen an established mouse stroke model, in which the middle cerebral artery was transiently occluded. After this procedure, the numbers of monocytes and neutrophils resident in the bone marrow increased within 3 days, an effect caused by enhanced proliferation of myeloid progenitors, such as granulocyte-macrophage progenitors and monocyte/dendritic cell progenitors. Interestingly, this effect seems to be specific for myeloid cells because neither noninflammatory Ly6Clow monocytes, nor B cells or lymphoid progenitors showed increased turnover. In fact, the numbers of nonmyeloid cells, as well as expression of their lineage-characterizing factors, such as IL-7 receptor, decreased, whereas the myeloid transcription factor PU-1 increased, suggesting a specific effect on the myeloid lineage. In seeking the most upstream cause of this myeloid bias—a state defined by an increased myeloid-to-lymphoid cell ratio in the bone marrow—the authors …

Introduction: Inflammation drives atherosclerosis and its complications. Thus, CANTOS as the first anti-inflammatory outcome trial in this population produced positive results. However, it is unclear how many patients qualify for an anti-inflammatory therapy in everyday practice. Hypothesis: This study analyzes how many patients with coronary heart disease (CHD) on guideline conform therapy show an increased residual inflammatory as opposed to an increased residual lipid risk in order to define the need for an anti-inflammatory treatment in a real world setting. Methods: High sensitive C-reactive protein (hsCRP) and low density lipoprotein (LDL) levels were determined in 700 all comer patients between June 2016 and June 2017 in our center. Patients lacking CHD, such with chronic-inflammatory diseases, acute inflammation, and on immunosuppressive medication were excluded. Patients were divided in the following groups: elevated hsCRP (≥2mg/dl), normal hsCRP ( Results: From 700 patients 221 fulfilled the inclusion and exclusion criteria. HsCRP was increased in 45% of these patients. Patients with on target LDL Levels showed lower hsCRP concentrations than those with off target values of LDL confirming a positive association between both (1,92mg/dl vs. 3,15mg/dl, p=0.005). However, despite guideline-conform LDL-control 34% of patients with a LDL-cholesterol Conclusions: A substantial part of patients with CHD shares a residual inflammatory risk defining a need for an anti-inflammatory therapy. Residual inflammation is particularly prevalent in patients with heart failure and diabetes.

Platelets can be found on the surface of inflamed and ruptured atherosclerotic plaques. Thus, targeting of activated platelets may allow for molecular imaging of vulnerable atherosclerotic lesions. We here investigated microbubbles (MB) functionalized with the selectin ligand sialyl Lewisa individually (MBsLea) or dually with sLea and an antibody targeting ligand-induced binding sites of the activated GPIIb/IIIa receptor (MBDual). Assessed by in vitro flow chamber, targeted MB exhibited increased adhesion to platelets as compared to MBControl. While MBsLea rolled slowly on the platelets’ surface, MBDual enhanced the percentage of firm adhesion. In vivo, MB were investigated by ultrasound in a model of ferric chloride induced non-occlusive carotid artery thrombosis. MBsLea and MBDual revealed a higher ultrasound mean acoustic intensity than MBControl (p Dual demonstrated no additional increase in mean signal intensity as compared to MBsLea. The degree of carotid artery stenosis on histology correlated well with the ultrasound acoustic intensity of targeted MB (p in vivo models.

Calcineurin inhibitor toxicity (CNT) is a frequent occurrence in transplanted renal grafts and autochthone kidneys from patients undergoing long-term treatment with calcineurin inhibitors, notably cyclosporin A (CsA) and tacrolimus. Here, we show an indispensable role of the tumor necrosis factor superfamily (TNFS) molecule TNF-related weak inducer of apoptosis (TWEAK) (TNFSF12) in the pathogenesis of acute CNT lesions in mice. A deficiency in TWEAK resulted in limited tubulotoxicity after CsA exposure, which correlated with diminished expression of inflammatory cytokines and reduced intraparenchymal infiltration with immune cells. We further identified tubular epithelial cells of the kidney as major targets of CsA activity and found that Fn14 (tumor necrosis factor receptor superfamily 12A), the receptor for TWEAK, is a highly CsA-inducible gene in these cells. Correlating with this, CsA pretreatment sensitized tubular epithelial cells specifically to the pro-inflammatory activities of recombinant TWEAK in vitro. Moreover, injection of rTWEAK alone into mice induced moderate disease similar to CsA, and rTWEAK combined with CsA resulted in synergistic nephrotoxicity. These findings support the importance of tubular epithelial cells as cellular targets of CsA toxicity and introduce TWEAK as a critical contributor to CNT pathogenesis.