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Stable metal isotopes have received increasing attention as medical biomarkers due to their potential to detect changes in metal metabolism related to diseases. In particular, copper stable isotopes are a powerful tool to identify isotopic variation between tumors and healthy tissue, suggesting application in cancer diagnosis. However, potential mechanisms causing isotope fractionation, such as redox- or bond-forming reactions and interactions of metals during transmembrane import and export, are less well understood. Here, we established an in situ method using laser ablation-multicollector-inductively coupled plasma-mass spectrometry (LA-MC-ICP-MS) to advance our understanding of the underlying processes responsible for isotope fractionation between normal and diseased tissues. Gelatin-based bracketing standards and quality control reference materials, crucial for laser ablation analysis, were developed to allow correction for instrumentally induced isotope fractionation during LA-MC-ICP-MS analysis. Using such matrix-matched standards, the method achieved intermediate precisions for delta values of better than 0.15 ‰ (2 s ) for inorganic reference materials and of better than 0.17 ‰ (2 s ) for biological reference materials. The developed routine was tested on rabbit VX2 liver tumor samples, a model system resembling human hepatocellular carcinoma (HCC) used to study liver cancer. In situ Cu isotope compositions between healthy ( δ NIST 976 65/63 ( Cu ) = -1.5 ‰ to 0.2 ‰) and tumorous ( δ NIST 976 65/63 ( Cu ) = 0.0 ‰ to 1.3 ‰) liver tissue show distinct differences in their isotope ratios. The observed isotopic dichotomy is consistent with previous solution-based MC-ICP-MS work, showing enrichment of heavy 65 Cu in cancer biopsies relative to healthy tissue.

Noise pollution is a known health risk factor and evidence for cardiovascular diseases associated with traffic noise is growing. At least 20% of the European Union's population lives in noise-polluted areas with exposure levels exceeding the recommended limits of the World Health Organization, which is considered unhealthy by the European Environment Agency. This results in the annual loss of 1.6 million healthy life years. Here, we investigated the protective effects of cardiovascular drug interventions against aircraft noise-mediated cardiovascular complications such as elevated oxidative stress or endothelial dysfunction. Using our established mouse exposure model, we applied mean sound pressure levels of 72 dB(A) for 4 d. C57BL/6 mice were treated with the beta-blocker propranolol (15 mg/kg/d s.c. for 5 d) or the alpha-blocker phenoxybenzamine (1.5 mg/kg/d s.c. for 5 d) and noise-exposed for the last 4 d of the drug administration. Short-term noise exposure caused hypertension (measured by tail-cuff blood pressure monitoring) and impaired endothelial function (measured by isometric tension recording in the aorta and video microscopy in cerebral arterioles in response to acetylcholine). Noise also increased markers of oxidative stress and inflammation. Treatment of mice with propranolol and phenoxybenzamine prevented endothelial and microvascular dysfunction, which was supported by a decrease in markers of inflammation and oxidative stress in heart tissue and the brain. Amelioration of noise-induced hypertension (systolic blood pressure) was not observed, whereas pulse pressure was lowered by trend. This study provides a novel perspective mitigating the adverse effects of noise pollution, especially in vulnerable groups with medication, a rationale for further pharmacological human studies.

As the drive towards recycling electronic waste increases, demand for rapid and reliable analytical methodology to analyse the metal content of the waste is increasing, e.g. to assess the value of the waste and to decide the correct recycling routes. Here, we comprehensively assess the suitability of different x-ray fluorescence spectroscopy (XRF)-based techniques as rapid analytical tools for the determination of critical raw materials, such as Al, Ti, Mn, Fe, Co, Ni, Cu, Zn, Nb, Pd and Au, in three electronic waste matrices: printed circuit boards (PCB), light emitting diodes (LED), and lithium (Li)-ion batteries. As validated reference methods and materials to establish metrological traceability are lacking, several laboratories measured test samples of each matrix using XRF as well as other independent complementary techniques (instrumental neutron activation analysis (INAA), inductively coupled plasma mass spectrometry (ICP-MS) and ICP optical emission spectrometry (OES)) as an inter-laboratory comparison (ILC). Results highlighted key aspects of sample preparation, limits of detection, and spectral interferences that affect the reliability of XRF, while additionally highlighting that XRF can provide more reliable data for certain elements compared to digestion-based approaches followed by ICP-MS analysis (e.g. group 4 and 5 metals). A clear distinction was observed in data processing methodologies for wavelength dispersive XRF, highlighting that considering the metals present as elements (rather than oxides) induces overestimations of the mass fractions when compared to other techniques. Eventually, the effect of sample particle size was studied and indicated that smaller particle size (<200 µm) is essential for reliable determinations., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

Background: Epidemiology links noise to increased risk of metabolic diseases like diabetes and obesity. Translational studies in humans and experimental animals showed that noise causes reactive oxygen species (ROS)-mediated cardiovascular damage. The interaction between noise and diabetes, specifically potential additive adverse effects, remains to be determined., Methods and Results: C57BL/6 mice were treated with streptozotocin (i.p. injections, 50 mg/kg/d for 5d) to induce type-1 diabetes, with S961 (subcutaneous osmotic minipumps, 0.57 mg/kg/d for 7d) or fed a high-fat diet (HFD, 20 weeks) to induce type-2 diabetes. Control and diabetic mice were exposed to aircraft noise to an average sound pressure level of 72 dB(A) for 4d. While body weight was unaffected, noise reduced insulin production in all diabetes models. The oral glucose tolerance test showed only an additive aggravation by noise in the HFD model. Noise increased blood pressure and aggravated diabetes-induced aortic, mesenteric, and cerebral arterioles endothelial dysfunction. ROS formation in cerebral arterioles, the aorta, the heart, and isolated mitochondria was consistently increased by noise in all models of diabetes. Mitochondrial respiration was impaired by diabetes and noise, however without additive effects. Noise increased ROS and caused inflammation in adipose tissue in the HFD model. RNA sequencing data and alteration of gene pathway clusters also supported additive damage by noise in the setting of diabetes., Conclusion: In all three models of diabetes, aircraft noise exacerbates oxidative stress, inflammation, and endothelial dysfunction in mice with pre-existing diabetes. Thus, noise may potentiate the already increased cardiovascular risk in diabetic patients., (© The Author(s) 2024. Published by Oxford University Press on behalf of the European Society of Cardiology.)

CD40L-CD40-TRAF signaling plays a role in atherosclerosis progression and affects the pathogenesis of coronary heart disease (CHD). We tested the hypothesis that CD40L-CD40-TRAF signaling is a potential therapeutic target in hyperlipidemia, diabetes, and hypertension. In mouse models of hyperlipidemia plus diabetes (db/db mice) or hypertension (1 mg/kg/d angiotensin-II for 7 days), TRAF6 inhibitor treatment (2.5 mg/kg/d for 7 or 14 days) normalized markers of oxidative stress and inflammation. As diabetes and hypertension are important comorbidities aggravating CHD, we explored whether the CD40L-CD40-TRAF signaling cascade and their associated inflammatory pathways are expressed in CHD patients suffering from comorbidities. Therefore, we analyzed vascular bypass material (aorta or internal mammary artery) and plasma from patients with CHD with diabetes and/or hypertension. Our Olink targeted plasma proteomic analysis using the IMMUNO-ONCOLOGY panel revealed a pattern of step-wise increase for 13/92 markers of low-grade inflammation with significant changes. CD40L or CD40 significantly correlated with 38 or 56 other inflammatory targets. In addition, specific gene clusters that correlate with the comorbidities were identified in isolated aortic mRNA of CHD patients through RNA-sequencing. These signaling clusters comprised CD40L-CD40-TRAF, immune system, hemostasis, muscle contraction, metabolism of lipids, developmental biology, and apoptosis. Finally, immunological analysis revealed key markers correlated with comorbidities in CHD patients, such as CD40L, NOX2, CD68, and 3-nitrotyrosine. These data indicate that comorbidities increase inflammatory pathways in CHD, and targeting these pathways will be beneficial in reducing cardiovascular events in CHD patients with comorbidities., (© 2024. The Author(s).)

Aims: The cytokine interleukin-6 (IL-6) plays a central role in the inflammation cascade as well as cardiovascular disease progression. Since myeloid cells are a primary source of IL-6 formation, we aimed to generate a mouse model to study the role of myeloid cell-derived IL-6 in vascular disease., Methods and Results: Interleukin-6-overexpressing (IL-6 OE ) mice were generated and crossed with LysM-Cre mice, to generate mice (LysM-IL-6 OE mice) overexpressing the cytokine in myeloid cells. Eight- to 12-week-old LysM-IL-6 OE mice spontaneously developed inflammatory colitis and significantly impaired endothelium-dependent aortic relaxation, increased aortic reactive oxygen species (ROS) formation, and vascular dysfunction in resistance vessels. The latter phenotype was associated with decreased survival. Vascular dysfunction was accompanied by a significant accumulation of neutrophils, monocytes, and macrophages in the aorta, increased myeloid cell reactivity (elevated ROS production), and vascular fibrosis associated with phenotypic changes in vascular smooth muscle cells. In addition to elevated Mcp1 and Cxcl1 mRNA levels, aortae from LysM-IL-6 OE mice expressed higher levels of inducible NO synthase and endothelin-1, thus partially accounting for vascular dysfunction, whereas systemic blood pressure alterations were not observed. Bone marrow (BM) transplantation experiments revealed that vascular dysfunction and ROS formation were driven by BM cell-derived IL-6 in a dose-dependent manner., Conclusion: Mice with conditional overexpression of IL-6 in myeloid cells show systemic and vascular inflammation as well as endothelial dysfunction. A decrease in circulating IL-6 levels by replacing IL-6-producing myeloid cells in the BM improved vascular dysfunction in this model, underpinning the relevant role of IL-6 in vascular disease., Competing Interests: Conflict of interest: M.B. received funding and is a consultant for ARCA Biopharma (not related to the presented work). S.K. received funding for consultant lectures from Almiral and Jansson-Cilag (not related to the presented work either)., (© The Author(s) 2024. Published by Oxford University Press on behalf of the European Society of Cardiology.)

Aim: Protein disulfide isomerases (PDIs) are involved in platelet aggregation and intravascular thrombosis, but their role in regulating endothelial function is unclear. Here, we characterized the involvement of vascular PDIA1 in angiotensin II (Ang II)-induced endothelial dysfunction in mice., Methods: Endothelial dysfunction was induced in C57BL/6JCmd male mice via Ang II subcutaneous infusion, and PDIA1 was inhibited with bepristat. Endothelial function was assessed in vivo with magnetic resonance imaging and ex vivo with a myography, while arterial stiffness was measured as pulse wave velocity. Nitric oxide (NO) bioavailability was measured in the aorta (spin-trapping electron paramagnetic resonance) and plasma (NO 2 - and NO 3 - levels). Oxidative stress, eNOS uncoupling (DHE-based aorta staining), and thrombin activity (thrombin-antithrombin complex; calibrated automated thrombography) were evaluated., Results: The inhibition of PDIA1 by bepristat in Ang II-treated mice prevented the impairment of NO-dependent vasodilation in the aorta as evidenced by the response to acetylcholine in vivo, increased systemic NO bioavailability and the aortic NO production, and decreased vascular stiffness. Bepristat's effect on NO-dependent function was recapitulated ex vivo in Ang II-induced endothelial dysfunction in isolated aorta. Furthermore, bepristat diminished the Ang II-induced eNOS uncoupling and overproduction of ROS without affecting thrombin activity., Conclusion: In Ang II-treated mice, the inhibition of PDIA1 normalized the NO-ROS balance, prevented endothelial eNOS uncoupling, and, thereby, improved vascular function. These results indicate the importance of vascular PDIA1 in regulating endothelial function, but further studies are needed to elucidate the details of the mechanisms involved., (© 2024 The Authors. Acta Physiologica published by John Wiley & Sons Ltd on behalf of Scandinavian Physiological Society.)

Aims: We examined the cardiovascular effects of celiac disease (CeD) in a humanized mouse model, with a focus on vascular inflammation, endothelial dysfunction, and oxidative stress., Methods and Results: NOD.DQ8 mice genetically predisposed to CeD were subjected to a diet regime and oral gavage to induce the disease (gluten group vs. control). We tested vascular function, confirmed disease indicators, and evaluated inflammation and oxidative stress in various tissues. Plasma proteome profiling was also performed. CeD markers were confirmed in the gluten group, indicating increased blood pressure and impaired vascular relaxation. Pro-inflammatory genes were upregulated in this group, with increased CD11b + myeloid cell infiltration and oxidative stress parameters observed in aortic and heart tissue. However, heart function remained unaffected. Plasma proteomics suggested the cytokine interleukin-17A (IL-17A) as a link between gut and vascular inflammation. Cardiovascular complications were reversed by adopting a gluten-free diet., Conclusion: Our study sheds light in the heightened cardiovascular risk associated with active CeD, revealing a gut-to-cardiovascular inflammatory axis potentially mediated by immune cell infiltration and IL-17A. These findings augment our understanding of the link between CeD and cardiovascular disease providing clinically relevant insight into the underlying mechanism. Furthermore, our discovery that cardiovascular complications can be reversed by a gluten-free diet underscores a critical role for dietary interventions in mitigating cardiovascular risks associated with CeD., Competing Interests: Declaration of competing interest All authors declared no conflicts of interest., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

The purpose of this study was to investigate the performances of the tumor response prediction prior to neoadjuvant chemotherapy based on quantitative ultrasound, tumour core-margin, texture derivative analyses, and molecular parameters in a large cohort of patients (n = 208) with locally advanced and earlier-stage breast cancer and combined them to best determine tumour responses with machine learning approach. Two multi-features response prediction algorithms using a k-nearest neighbour and support vector machine were developed with leave-one-out and hold-out cross-validation methods to evaluate the performance of the response prediction models. In a leave-one-out approach, the quantitative ultrasound-texture analysis based model attained good classification performance with 80% of accuracy and AUC of 0.83. Including molecular subtype in the model improved the performance to 83% of accuracy and 0.87 of AUC. Due to limited number of samples in the training process, a model developed with a hold-out approach exhibited a slightly higher bias error in classification performance. The most relevant features selected in predicting the response groups are core-to-margin, texture-derivative, and molecular subtype. These results imply that that baseline tumour-margin, texture derivative analysis methods combined with molecular subtype can potentially be used for the prediction of ultimate treatment response in patients prior to neoadjuvant chemotherapy., (© 2023. The Author(s).)

Large epidemiological studies have shown that traffic noise promotes the development of cardiometabolic diseases. It remains to be established how long these adverse effects of noise may persist in response to a noise-off period. We investigated the effects of acute aircraft noise exposure (mean sound level of 72 dB(A) applied for 4d) on oxidative stress and inflammation mediating vascular dysfunction and increased blood pressure in male C57BL/6 J mice. 1, 2 or 4d of noise cessation after a 4d continuous noise exposure period completely normalized noise-induced endothelial dysfunction of the aorta (measured by acetylcholine-dependent relaxation) already after a 1d noise pause. Vascular oxidative stress and the increased blood pressure were partially corrected, while markers of inflammation (VCAM-1, IL-6 and leukocyte oxidative burst) showed a normalization within 4d of noise cessation. In contrast, endothelial dysfunction, oxidative stress, and inflammation of the cerebral microvessels of noise-exposed mice did not improve at all. These data demonstrate that the recovery from noise-induced damage is more complex than expected demonstrating a complete restoration of large conductance vessel function but persistent endothelial dysfunction of the microcirculation. These findings also imply that longer noise pauses are required to completely reverse noise-induced vascular dysfunction including the resistance vessels., Competing Interests: Declaration of competing interest The authors declare that they have no conflicts of interest with the contents of this article., (Copyright © 2023 The Authors. Published by Elsevier B.V. All rights reserved.)

Aims: Environmental stressors such as traffic noise represent a global threat, accounting for 1.6 million healthy life years lost annually in Western Europe. Therefore, the noise-associated health side effects must be effectively prevented or mitigated. Non-pharmacological interventions such as physical activity or a balanced healthy diet are effective due to the activation of the adenosine monophosphate-activated protein kinase (α1AMPK). Here, we investigated for the first time in a murine model of aircraft noise-induced vascular dysfunction the potential protective role of α1AMPK activated via exercise, intermittent fasting, and pharmacological treatment., Methods and Results: Wild-type (B6.Cg-Tg(Cdh5-cre)7Mlia/J) mice were exposed to aircraft noise [maximum sound pressure level of 85 dB(A), average sound pressure level of 72 dB(A)] for the last 4 days. The α1AMPK was stimulated by different protocols, including 5-aminoimidazole-4-carboxamide riboside application, voluntary exercise, and intermittent fasting. Four days of aircraft noise exposure produced significant endothelial dysfunction in wild-type mice aorta, mesenteric arteries, and retinal arterioles. This was associated with increased vascular oxidative stress and asymmetric dimethylarginine formation. The α1AMPK activation with all three approaches prevented endothelial dysfunction and vascular oxidative stress development, which was supported by RNA sequencing data. Endothelium-specific α1AMPK knockout markedly aggravated noise-induced vascular damage and caused a loss of mitigation effects by exercise or intermittent fasting., Conclusion: Our results demonstrate that endothelial-specific α1AMPK activation by pharmacological stimulation, exercise, and intermittent fasting effectively mitigates noise-induced cardiovascular damage. Future population-based studies need to clinically prove the concept of exercise/fasting-mediated mitigation of transportation noise-associated disease., Competing Interests: Conflict of interest: None declared., (© The Author(s) 2023. Published by Oxford University Press on behalf of the European Society of Cardiology.)

Electronic cigarettes (E-cigarettes) have recently become a popular alternative to traditional tobacco cigarettes. Despite being marketed as a healthier alternative, increasing evidence shows that E-cigarette vapour could cause adverse health effects. It has been postulated that degradation products of E-cigarette liquid, mainly reactive aldehydes, are responsible for those effects. Previously, we have demonstrated that E-cigarette vapour exposure causes oxidative stress, inflammation, apoptosis, endothelial dysfunction and hypertension by activating NADPH oxidase in a mouse model. To better understand oxidative stress mechanisms, we have exposed cultured endothelial cells and macrophages to condensed E-cigarette vapour (E-cigarette condensate) and acrolein. In both endothelial cells (EA.hy 926) and macrophages (RAW 264.7), we have observed that E-cigarette condensate incubation causes cell death. Since recent studies have shown that among toxic aldehydes found in E-cigarette vapour, acrolein plays a prominent role, we have incubated the same cell lines with increasing concentrations of acrolein. Upon incubation with acrolein, a translocation of Rac1 to the plasma membrane has been observed, accompanied by an increase in oxidative stress. Whereas reactive oxygen species (ROS) formation by acrolein in cultured endothelial cells was mainly intracellular, the release of ROS in cultured macrophages was both intra- and extracellular. Our data also demonstrate that acrolein activates the nuclear factor erythroid 2-related factor 2 (Nrf2) antioxidant pathway and, in general, could mediate E-cigarette vapour-induced oxidative stress and cell death. More mechanistic insight is needed to clarify the toxicity associated with E-cigarette consumption and the possible adverse effects on human health., (© 2023. The Author(s).)

Smoking tobacco cigarettes is a significant (cardiovascular) health risk factor. Although the number of tobacco cigarette users declined over the last decades, shisha smoking and e-cigarette vaping partially compensated for this health benefit. E-cigarettes may create highly addicted dual users (vaping and smoking). E-cigarettes seem not to represent a healthier alternative to tobacco smoking, although they may be less harmful. E-cigarette vaping causes oxidative stress, inflammation, endothelial dysfunction, and associated cardiovascular sequelae. This is primarily due to a significant overlap of toxic compounds in the vapor compared to tobacco smoke and, accordingly, a substantial overlap of pathomechanistic features between vaping and smoking. Whereas the main toxins in vapor are reactive aldehydes such as formaldehyde and acrolein, the toxic mixture in smoke is more complex, comprising particulate matter, reactive gases, transition metals, volatile organic compounds, and N-nitrosamines. However, it seems that both lifestyle drugs impair endothelial function to a quite similar extent, which may be due to the role of oxidative stress as the central pathomechanism to mediate endothelial dysfunction and vascular damage. Finally, the main selling argument for e-cigarette use that they help to quit smoking and get rid of nicotine addiction may be false because it seems that e-cigarettes instead trigger the opposite-younger entrance age and more frequent use. With our review, we summarize the adverse health impact of tobacco cigarettes and e-cigarettes, emphasizing the detrimental effects on endothelial function and cardiovascular health., (© 2023. The Author(s).)

Psoriasis is an immune-mediated inflammatory skin disease driven by interleukin-17A (IL-17A) and associated with cardiovascular dysfunction. We used a severe psoriasis mouse model of keratinocyte IL-17A overexpression (K14-IL-17A ind/+ , IL-17A ind/+ control mice) to investigate the activity of neutrophils and a potential cellular interconnection between skin and vasculature. Levels of dermal reactive oxygen species (ROS) and their release by neutrophils were measured by lucigenin-/luminol-based assays, respectively. Quantitative RT-PCR determined neutrophilic activity and inflammation-related markers in skin and aorta. To track skin-derived immune cells, we used PhAM-K14-IL-17A ind/+ mice allowing us to mark all cells in the skin by photoconversion of a fluorescent protein to analyze their migration into spleen, aorta, and lymph nodes by flow cytometry. Compared to controls, K14-IL-17A ind/+ mice exhibited elevated ROS levels in the skin and a higher neutrophilic oxidative burst accompanied by the upregulation of several activation markers. In line with these results psoriatic mice displayed elevated expression of genes involved in neutrophil migration (e.g., Cxcl2 and S100a9) in skin and aorta. However, no direct immune cell migration from the psoriatic skin into the aortic vessel wall was observed. Neutrophils of psoriatic mice showed an activated phenotype, but no direct cellular migration from the skin to the vasculature was observed. This suggests that highly active vasculature-invading neutrophils must originate directly from the bone marrow. Hence, the skin-vasculature crosstalk in psoriasis is most likely based on the systemic effects of the autoimmune skin disease, emphasizing the importance of a systemic therapeutic approach for psoriasis patients., (© 2023 The Authors. BioFactors published by Wiley Periodicals LLC on behalf of International Union of Biochemistry and Molecular Biology.)

Immobilized metal affinity chromatography (IMAC) is a popular and valuable method for the affinity purification of polyhistidine-tagged recombinant proteins. However, it often shows practical limitations, which might require cumbersome optimizations, additional polishing, and enrichment steps. Here, we present functionalized corundum particles for the efficient, economical, and fast purification of recombinant proteins in a column-free format. The corundum surface is first derivatized with the amino silane APTES, then EDTA dianhydride, and subsequently loaded with nickel ions. The Kaiser test, well known in solid-phase peptide synthesis, was used to monitor amino silanization and the reaction with EDTA dianhydride. In addition, ICP-MS was performed to quantify the metal-binding capacity. His-tagged protein A/G (PAG), mixed with bovine serum albumin (BSA), was used as a test system. The PAG binding capacity was around 3 mg protein per gram of corundum or 2.4 mg per 1 mL of corundum suspension. Cytoplasm obtained from different E. coli strains was examined as examples of a complex matrix. The imidazole concentration was varied in the loading and washing buffers. As expected, higher imidazole concentrations during loading are usually beneficial when higher purities are desired. Even when higher sample volumes, such as one liter, were used, recombinant protein down to a concentration of 1 µg/mL could be isolated selectively. Comparing the corundum material with standard Ni-NTA agarose beads indicated higher purities of proteins isolated using corundum. His6-MBP-mSA2, a fusion protein consisting of monomeric streptavidin and maltose-binding protein in the cytoplasm of E. coli , was purified successfully. To show that this method is also suitable for mammalian cell culture supernatants, purification of the SARS-CoV-2-S-RBD-His8 expressed in human Expi293F cells was performed. The material cost of the nickel-loaded corundum material (without regeneration) is estimated to be less than 30 cents for 1 g of functionalized support or 10 cents per milligram of isolated protein. Another advantage of the novel system is the corundum particles' extremely high physical and chemical stability. The new material should be applicable in small laboratories and large-scale industrial applications. In summary, we could show that this new material is an efficient, robust, and cost-effective purification platform for the purification of His-tagged proteins, even in challenging, complex matrices and large sample volumes of low product concentration.

Ischemia-reperfusion (I/R) events are involved in the development of various ocular pathologies, e.g., retinal artery or vein occlusion. We tested the hypothesis that resveratrol is protective against I/R injury in the murine retina. Intraocular pressure (IOP) was elevated in anaesthetized mice to 110 mm Hg for 45 min via a micropipette placed in the anterior chamber to induce ocular ischemia. In the fellow eye, which served as control, IOP was kept at a physiological level. One group received resveratrol (30 mg/kg/day p.o. once daily) starting one day before the I/R event, whereas the other group of mice received vehicle solution only. On day eight after the I/R event, mice were sacrificed and retinal wholemounts were prepared and immuno-stained using a Brn3a antibody to quantify retinal ganglion cells. Reactivity of retinal arterioles was measured in retinal vascular preparations using video microscopy. Reactive oxygen species (ROS) and nitrogen species (RNS) were quantified in ocular cryosections by dihydroethidium and anti-3-nitrotyrosine staining, respectively. Moreover, hypoxic, redox and nitric oxide synthase gene expression was quantified in retinal explants by PCR. I/R significantly diminished retinal ganglion cell number in vehicle-treated mice. Conversely, only a negligible reduction in retinal ganglion cell number was observed in resveratrol-treated mice following I/R. Endothelial function and autoregulation were markedly reduced, which was accompanied by increased ROS and RNS in retinal blood vessels of vehicle-exposed mice following I/R, whereas resveratrol preserved vascular endothelial function and autoregulation and blunted ROS and RNS formation. Moreover, resveratrol reduced I/R-induced mRNA expression for the prooxidant enzyme, nicotinamide adenine dinucleotide phosphate oxidase 2 (NOX2). Our data provide evidence that resveratrol protects from I/R-induced retinal ganglion cell loss and endothelial dysfunction in the murine retina by reducing nitro-oxidative stress possibly via suppression of NOX2 upregulation.

Worldwide, up to 8.8 million excess deaths/year have been attributed to air pollution, mainly due to the exposure to fine particulate matter (PM). Traffic-related noise is an additional contributor to global mortality and morbidity. Both health risk factors substantially contribute to cardiovascular, metabolic and neuropsychiatric sequelae. Studies on the combined exposure are rare and urgently needed because of frequent co-occurrence of both risk factors in urban and industrial settings. To study the synergistic effects of PM and noise, we used an exposure system equipped with aerosol generator and loud-speakers, where C57BL/6 mice were acutely exposed for 3d to either ambient PM (NIST particles) and/or noise (aircraft landing and take-off events). The combination of both stressors caused endothelial dysfunction, increased blood pressure, oxidative stress and inflammation. An additive impairment of endothelial function was observed in isolated aortic rings and even more pronounced in cerebral and retinal arterioles. The increase in oxidative stress and inflammation markers together with RNA sequencing data indicate that noise particularly affects the brain and PM the lungs. The combination of both stressors has additive adverse effects on the cardiovascular system that are based on PM-induced systemic inflammation and noise-triggered stress hormone signaling. We demonstrate an additive upregulation of ACE-2 in the lung, suggesting that there may be an increased vulnerability to COVID-19 infection. The data warrant further mechanistic studies to characterize the propagation of primary target tissue damage (lung, brain) to remote organs such as aorta and heart by combined noise and PM exposure., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022 The Authors. Published by Elsevier B.V. All rights reserved.)

The radio-frequency ultrasound backscattered data from tissue is rich in information and can provide important information about tissue state that is not obtained through traditional B-mode imaging. To parameterize the ultrasound backscattered data, the frequency spectrum, i.e., the backscatter coefficient, can be modeled using scattering theory. Models of tissue scattering are often represented by simple discrete geometric shapes, i.e., discrete scattering model. The discrete scattering model provides important insights into how the spatial arrangement of scatterers contributes to the signal spectrum. Another competing model is the continuum scattering model. In this model, the tissue is described as a continuous tissue construct with scatterers that have a continuous impedance change from the background. The continuous model provides a form factor description of the underlying tissue scatterers such as an effective scatterer diameter. In this chapter, we will compare and contrast the two underlying tissue scattering models and how they provide insights into ultrasonic scattering from soft tissues., (© 2023. The Author(s), under exclusive license to Springer Nature Switzerland AG.)

The backscatter coefficient is a fundamental property of tissues, much like the attenuation and sound speed. From the backscatter coefficient, different scatterer properties describing the underlying tissue can be used to characterize tissue state. Furthermore, because the backscatter coefficient is a fundamental property of a tissue, estimation of the backscatter coefficient should be able to be computed with system and operator independence. To accomplish system- and operator-independent estimates of the backscatter coefficient, a calibration spectrum must be obtained at the same system settings as the settings used to scan a tissue. In this chapter, we discuss three approaches to obtaining a calibration spectrum and compare the engineering tradeoffs associated with each approach. In addition, methods for reducing deterministic noise in the backscatter coefficient spectrum are considered and implementation of these techniques is discussed., (© 2023. The Author(s), under exclusive license to Springer Nature Switzerland AG.)

The 18 O/ 16 O ratio of cherts (δ 18 O chert ) increases nearly monotonically by ~15‰ from the Archean to present. Two end-member explanations have emerged: cooling seawater temperature (T SW ) and increasing seawater δ 18 O (δ 18 O sw ). Yet despite decades of work, there is no consensus, leading some to view the δ 18 O chert record as pervasively altered. Here, we demonstrate that cherts are a robust archive of diagenetic temperatures, despite metamorphism and exposure to meteoric fluids, and show that the timing and temperature of quartz precipitation and thus δ 18 O chert are determined by the kinetics of silica diagenesis. A diagenetic model shows that δ 18 O chert is influenced by heat flow through the sediment column. Heat flow has decreased over time as planetary heat is dissipated, and reasonable Archean-modern heat flow changes account for ~5‰ of the increase in δ 18 O chert , obviating the need for extreme T SW or δ 18 O sw reconstructions. The seawater oxygen isotope budget is also influenced by solid Earth cooling, with a recent reconstruction placing Archean δ 18 O SW 5 to 10‰ lower than today. Together, this provides an internally consistent view of the δ 18 O chert record as driven by solid Earth cooling over billion-year timescales that is compatible with Precambrian glaciations and biological constraints and satisfyingly accounts for the monotonic nature of the δ 18 O chert trend.

Tetrahydrobiopterin (BH4) is an essential cofactor of all nitric oxide synthase isoforms, thus determination of BH4 levels can provide important mechanistic insight into diseases. We established a protocol for high-performance liquid chromatography/electrochemical detection (HPLC/ECD)-based determination of BH4 in tissue samples. We first determined the optimal storage and work-up conditions for authentic BH4 and its oxidation product dihydrobiopterin (BH2) under various conditions (pH, temperature, presence of antioxidants, metal chelators, and storage time). We then applied optimized protocols for detection of BH4 in tissues of septic (induced by lipopolysaccharide [LPS]) rats. BH4 standards in HCl are stabilized by addition of 1,4-dithioerythritol (DTE) and diethylenetriaminepentaacetic acid (DTPA), while HCl was sufficient for BH2 standard stabilization. Overnight storage of BH4 standard solutions at room temperature in HCl without antioxidants caused complete loss of BH4 and the formation of BH2. We further optimized the protocol to separate ascorbate and the BH4 tissue sample and found a significant increase in BH4 in the heart and kidney as well as higher BH4 levels by trend in the brain of septic rats compared to control rats. These findings correspond to reports on augmented nitric oxide and BH4 levels in both animals and patients with septic shock.

Ensuring the consistency of spectral-based quantitative ultrasound estimates in vivo necessitates accounting for diffraction, system effects, and propagation losses encountered in the tissue. Accounting for diffraction and system effects is typically achieved through planar reflector or reference phantom methods; however, neither of these is able to account for the tissue losses present in vivo between the ultrasound probe and the region of interest. In previous work, the feasibility of small titanium beads as in situ calibration targets (0.5-2 mm in diameter) was investigated. In this study, the importance of bead size for the calibration signal, the role of multiple echoes coming from the calibration bead, and sampling of the bead signal laterally through beam translation were examined. This work demonstrates that although the titanium beads naturally produce multiple reverberant echoes, time-windowing of the first echo provides the smoothest calibration spectrum for backscatter coefficient calculation. When translating the beam across the bead, the amplitude of the echo decreases rapidly as the beam moves across and past the bead. Therefore, to obtain consistent calibration signals from the bead, lateral interpolation is needed to approximate signals coming from the center of the bead with respect to the beam.

The prevalence and clinical importance of arterial hypertension are still growing. Inorganic nitrite (NO 2 - ) represents an attractive dietary antihypertensive agent, but its metabolism and mode of action, which we aimed to investigate with the present study, are not completely understood. Isolated aortic rings from rats were treated ex vivo with oxidants, and rats were infused in vivo with angiotensin-II. Vascular responses to acetylcholine (ACh) and nitrite were assessed by isometric tension recording. The loss of vasodilatory potency in response to oxidants was much more pronounced for ACh as compared to nitrite ex vivo (but not in vivo with angiotensin-II). This effect may be caused by the redox regulation of conversion to xanthine oxidase (XO). Conventionally raised and germ-free mice were treated with nitrite by gavage, which did not improve ACh-mediated vasodilation, but did increase the plasma levels of S-nitros(yl)ated proteins in the conventionally-raised, but not in the germ-free mice. In conclusion, inorganic nitrite represents a dietary drug option to treat arterial hypertension in addition to already established pharmacological treatment. Short-term oxidative stress did not impair the vasodilatory properties of nitrite, which may be beneficial in cardiovascular disease patients. The gastrointestinal microbiome appears to play a key role in nitrite metabolism and bioactivation.

Transportation noise is recognized as an important cardiovascular risk factor. Key mechanisms are noise-triggered vascular inflammation and oxidative stress with subsequent endothelial dysfunction. Here, we test for adaptation or tolerance mechanisms in mice in response to chronic noise exposure. C57BL/6J mice were exposed to aircraft noise for 0, 4, 7, 14 and 28d at a mean sound pressure level of 72 dB(A) and peak levels of 85 dB(A). Chronic aircraft noise exposure up to 28d caused persistent endothelial dysfunction and elevation of blood pressure. Likewise, reactive oxygen species (ROS) formation as determined by dihydroethidium (DHE) staining and HPLC-based measurement of superoxide formation in the aorta/heart/brain was time-dependently increased by noise. Oxidative burst in the whole blood showed a maximum at 4d or 7d of noise exposure. Increased superoxide formation in the brain was mirrored by a downregulation of neuronal nitric oxide synthase ( Nos3 ) and transcription factor Foxo3 genes, whereas Vcam1 mRNA, a marker for inflammation was upregulated in all noise exposure groups. Induction of a pronounced hearing loss in the mice was excluded by auditory brainstem response audiometry. Endothelial dysfunction and inflammation were present during the entire 28d of aircraft noise exposure. ROS formation gradually increases with ongoing exposure without significant adaptation or tolerance in mice in response to chronic noise stress at moderate levels. These data further illustrate health side effects of long-term noise exposure and further strengthen a consequent implementation of the WHO noise guidelines in order to prevent the development of noise-related future cardiovascular disease., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Frenis, Kalinovic, Ernst, Kvandova, Al Zuabi, Kuntic, Oelze, Stamm, Bayo Jimenez, Kij, Keppeler, Klein, Strohm, Ubbens, Daub, Hahad, Kröller-Schön, Schmeisser, Chlopicki, Eckrich, Strieth, Daiber, Steven and Münzel.)

Voluntary exercise training is an effective way to prevent cardiovascular disease, since it results in increased NO bioavailability and decreased reactive oxygen species (ROS) production. AMP-activated protein kinase (AMPK), especially its α1AMPK subunit, modulates ROS-dependent vascular homeostasis. Since endothelial cells play an important role in exercise-induced changes of vascular signaling, we examined the consequences of endothelial-specific α1AMPK deletion during voluntary exercise training. We generated a mouse strain with specific deletion of α1AMPK in endothelial cells (α1AMPK flox/flox x TekCre + ). While voluntary exercise training improved endothelial function in wild-type mice, it had deleterious effects in mice lacking endothelial α1AMPK indicated by elevated reactive oxygen species production (measured by dihydroethidum fluorescence and 3-nitrotyrosine staining), eNOS uncoupling and endothelial dysfunction. Importantly, the expression of the phagocytic NADPH oxidase isoform (NOX-2) was down-regulated by exercise in control mice, whereas it was up-regulated in exercising α1AMPK flox/flox x TekCre + animals. In addition, nitric oxide bioavailability was decreased and the antioxidant/protective nuclear factor erythroid 2-related factor 2 (Nrf-2) response via heme oxygenase 1 and uncoupling protein-2 (UCP-2) was impaired in exercising α1AMPK flox/flox x TekCre + mice. Our results demonstrate that endothelial α1AMPK is a critical component of the signaling events that enable vascular protection in response to exercise. Moreover, they identify endothelial α1AMPK as a master switch that determines whether the effects of exercise on the vasculature are protective or detrimental.

Glutathione (γ-L-glutamyl-L-cysteinyl-glycine, GSH) is a tripeptide that is part of the antioxidant defense system and contributes to numerous redox-regulatory processes. In vivo , reduced GSH and oxidized glutathione disulfide (GSSG) are present in redox equilibrium and their ratio provides important information on the cellular redox state. Here, we compared three different methods for in vivo quantification of glutathione in tissues of hypertensive rats, an accepted animal model of oxidative stress. In the present study, we used hypertensive rats (infusion of 1 mg/kg/d angiotensin-II for 7 days) to determine the levels of reduced GSH and/or GSH/GSSG ratios in different tissue samples. We used an HPLC-based method with direct electrochemical detection (HPLC/ECD) and compared it with Ellman's reagent (DTNB) dependent derivatization of reduced GSH to the GS-NTB adduct and free NTB (UV/Vis HPLC) as well as with a commercial GSH/GSSG assay (Oxiselect). Whereas all three methods indicated overall a decreased redox state in hypertensive rats, the assays based on HPLC/ECD and DTNB derivatization provided the most significant differences. We applied a direct, fast and sensitive method for electrochemical GSH detection in tissues from hypertensive animals, and confirmed its reliability for in vivo measurements by head-to-head comparison with two other established assays. The HPLC/ECD but not DTNB and Oxiselect assays yielded quantitative GSH data but all three assays reflected nicely the qualitative redox changes and functional impairment in hypertensive rats. However, especially our GSH/GSSG values are lower than reported by others pointing to problems in the work-up protocol.

Environmental exposures represent a significant health hazard, which cumulatively may be responsible for up to 2/3 of all chronic non-communicable disease and associated mortality (Global Burden of Disease Study and The Lancet Commission on Pollution and Health), which has given rise to a new concept of the exposome: the sum of environmental factors in every individual's experience. Noise is part of the exposome and is increasingly being investigated as a health risk factor impacting neurological, cardiometabolic, endocrine, and immune health. Beyond the well-characterized effects of high-intensity noise on cochlear damage, noise is relatively well-studied in the cardiovascular field, where evidence is emerging from both human and translational experiments that noise from traffic-related sources could represent a risk factor for hypertension, ischemic heart disease, diabetes, and atherosclerosis. In the present review, we comprehensively discuss the current state of knowledge in the field of noise research. We give a brief survey of the literature documenting experiments in noise exposure in both humans and animals with a focus on cardiovascular disease. We also discuss the mechanisms that have been uncovered in recent years that describe how exposure to noise affects physiological homeostasis, leading to aberrant redox signaling resulting in metabolic and immune consequences, both of which have considerable impact on cardiovascular health. Additionally, we discuss the molecular pathways of redox involvement in the stress responses to noise and how they manifest in disruptions of the circadian rhythm, inflammatory signaling, gut microbiome composition, epigenetic landscape and vessel function., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Frenis, Kuntic, Hahad, Bayo Jimenez, Oelze, Daub, Steven, Münzel and Daiber.)

Background: Diabetic vasculopathy plays an important role in the pathophysiology of coronary artery disease (CAD) with oxidative stress as a strong mediator. This study aims to elucidate the underlying pathomechanisms of diabetic cardiac vasculopathy leading to coronary disease with an emphasis on the role of oxidative stress. Therefore, novel insights into antioxidant pathways might contribute to new strategies in the treatment and prevention of diabetic CAD., Methods: In 20 patients with insulin-dependent or non-insulin dependent diabetes mellitus (IDDM/NIDDM) and 39 non-diabetic (CTR) patients, myocardial markers of oxidative stress, vasoactive proteins, endothelial nitric oxide synthase (eNOS), activated phosphorylated eNOS ( p -eNOS), and antioxidant enzymes, e.g., tetrahydrobiopterin generating dihydrofolate reductase (DHFR), heme oxygenase (HO-1), as well as serum markers of inflammation, e.g., E-selectin, interleukin-6 (IL-6), and lipid metabolism, e.g., high- and low-density lipoptrotein (HDL- and LDL-cholesterol) were determined in specimens of right atrial tissue and in blood samples from type 2 diabetic and non-diabetic patients undergoing coronary artery bypass graft (CABG) surgery., Results: IDDM/NIDDM increased markers of inflammation (e.g., E-selectin, p = 0.005 and IL-6, p = 0.051), decreased the phosphorylated myocardial p -eNOS ( p = 0.032), upregulated the myocardial stress response protein HO-1 ( p = 0.018), and enhanced the serum LDL-/HDL-cholesterol ratio ( p = 0.019). However, the oxidative stress markers in the myocardium and the expression of vasoactive proteins (eNOS, DHFR) showed only marginal adverse changes in patients with IDDM/NIDDM., Conclusion: Dyslipidemia and myocardial inflammation seem to be the major determinants of diabetic CAD complications. Dysregulation in pro-oxidative enzymes might be attributable to the severity of CAD and oxidative stress levels in all included patients undergoing CABG.

This article contains supporting information on data collection for the research article entitled "Aircraft noise exposure drives the activation of white blood cells and induces microvascular dysfunction in mice" by Eckrich et al. We found that noise-induced stress triggered microvascular dysfunction via involvement of innate immune-derived reactive oxygen species. In this article, we present the instrumentation of mice with dorsal skinfold chambers for in vivo microscopic imaging of blood flow, interaction of leukocytes with the vascular wall (also by fluorescent labelling of blood cells) and vessel diameter. In addition, we explain the preparation of cerebral arterioles for measurement of vascular reactivity in vitro .•visualization of noise-dependent effects in dorsal skinfold chamber.• in vivo microscopy of noise-dependent activation of white blood cells.•analysis of noise-dependent microvascular dysfunction in dorsal skinfold chamber and cannulated cerebral arterioles., Competing Interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© 2021 The Authors. Published by Elsevier B.V.)

Epidemiological studies showed that traffic noise has a dose-dependent association with increased cardiovascular morbidity and mortality. Whether microvascular dysfunction contributes significantly to the cardiovascular health effects by noise exposure remains to be established. The connection of inflammation and immune cell interaction with microvascular damage and functional impairment is also not well characterized. Male C57BL/6J mice or gp91phox -/y mice with genetic deletion of the phagocytic NADPH oxidase catalytic subunit (gp91phox or NOX-2) were used at the age of 8 weeks, randomly instrumented with dorsal skinfold chambers and exposed or not exposed to aircraft noise for 4 days. Proteomic analysis (using mass spectrometry) revealed a pro-inflammatory phenotype induced by noise exposure that was less pronounced in noise-exposed gp91phox -/y mice. Using in vivo fluorescence microscopy, we found a higher number of adhesive leukocytes in noise-exposed wild type mice. Dorsal microvascular diameter (by trend), red blood cell velocity, and segmental blood flow were also decreased by noise exposure indicating microvascular constriction. All adverse effects on functional parameters were normalized or improved at least by trend in noise-exposed gp91phox -/y mice. Noise exposure also induced endothelial dysfunction in cerebral microvessels, which was associated with higher oxidative stress burden and inflammation, as measured using video microscopy. We here establish a link between a pro-inflammatory phenotype of plasma, activation of circulating leukocytes and microvascular dysfunction in mice exposed to aircraft noise. The phagocytic NADPH oxidase was identified as a central player in the underlying pathophysiological mechanisms., (Copyright © 2021 The Author(s). Published by Elsevier B.V. All rights reserved.)

Ischemia/reperfusion (I/R) events are involved in the pathophysiology of numerous ocular diseases. The purpose of this study was to test the hypothesis that betulinic acid protects from I/R injury in the mouse retina. Ocular ischemia was induced in mice by increasing intraocular pressure (IOP) to 110 mm Hg for 45 min, while the fellow eye served as a control. One group of mice received betulinic acid (50 mg/kg/day p.o. once daily) and the other group received the vehicle solution only. Eight days after the I/R event, the animals were killed and the retinal wholemounts and optic nerve cross-sections were prepared and stained with cresyl blue or toluidine blue, respectively, to count cells in the ganglion cell layer (GCL) of the retina and axons in the optic nerve. Retinal arteriole responses were measured in isolated retinas by video microscopy. The levels of reactive oxygen species (ROS) were assessed in retinal cryosections and redox gene expression was determined in isolated retinas by quantitative PCR. I/R markedly reduced cell number in the GCL and axon number in the optic nerve of the vehicle-treated mice. In contrast, only a negligible reduction in cell and axon number was observed following I/R in the betulinic acid-treated mice. Endothelial function was markedly reduced and ROS levels were increased in retinal arterioles of vehicle-exposed eyes following I/R, whereas betulinic acid partially prevented vascular endothelial dysfunction and ROS formation. Moreover, betulinic acid boosted mRNA expression for the antioxidant enzymes SOD3 and HO-1 following I/R. Our data provide evidence that betulinic acid protects from I/R injury in the mouse retina. Improvement of vascular endothelial function and the reduction in ROS levels appear to contribute to the neuroprotective effect.

Arterial hypertension is one of the major health risk factors leading to coronary artery disease, stroke or peripheral artery disease. Dietary uptake of inorganic nitrite (NO 2 - ) and nitrate (NO 3 - ) via vegetables leads to enhanced vascular NO bioavailability and provides antihypertensive effects. The present study aims to understand the underlying vasoprotective effects of nutritional NO 2 - and NO 3 - co-therapy in mice with angiotensin-II (AT-II)-induced arterial hypertension. High-dose AT-II (1 mg/kg/d, 1w, s. c.) was used to induce arterial hypertension in male C57BL/6 mice. Additional inorganic nitrite (7.5 mg/kg/d, p. o.) or nitrate (150 mg/kg/d, p. o.) were administered via the drinking water. Blood pressure (tail-cuff method) and endothelial function (isometric tension) were determined. Oxidative stress and inflammation markers were quantified in aorta, heart, kidney and blood. Co-treatment with inorganic nitrite, but not with nitrate, normalized vascular function, oxidative stress markers and inflammatory pathways in AT-II treated mice. Of note, the highly beneficial effects of nitrite on all parameters and the less pronounced protection by nitrate, as seen by improvement of some parameters, were observed despite no significant increase in plasma nitrite levels by both therapies. Methemoglobin levels tended to be higher upon nitrite/nitrate treatment. Nutritional nitric oxide precursors represent a non-pharmacological treatment option for hypertension that could be applied to the general population (e.g. by eating certain vegetables). The more beneficial effects of inorganic nitrite may rely on superior NO bioactivation and stronger blood pressure lowering effects. Future large-scale clinical studies should investigate whether hypertension and cardiovascular outcome in general can be influenced by dietary inorganic nitrite therapy., (Copyright © 2021 The Author(s). Published by Elsevier Inc. All rights reserved.)

Aims: Oxidative stress and inflammation contribute to atherogenesis. Rac1 GTPase regulates pro-oxidant NADPH oxidase activity, reactive oxygen species (ROS) formation, actin cytoskeleton organization and monocyte adhesion. We investigated the vascular effects of pharmacological inhibition of Rac1 GTPase in mice. Methods and Results: We treated wild-type and apolipoprotein E-deficient (ApoE -/- ) mice with Clostridium sordellii lethal toxin (LT), a Rac1 inhibitor, and assessed vascular oxidative stress, expression and activity of involved proteins, endothelial function, macrophage infiltration, and atherosclerosis development. LT-treated wild-type mice displayed decreased vascular NADPH oxidase activity and ROS production. Therapeutic LT doses had no impact on behavior, food intake, body weight, heart rate, blood pressure, vascular and myocardial function, differential blood count, and vascular permeability. ApoE -/- mice were fed a cholesterol-rich diet and were treated with LT or vehicle. LT treatment led to decreased aortic Rac1 GTPase activity, NADPH oxidase activity and ROS production, but had no impact on expression and membrane translocation of NADPH oxidase subunits and RhoA GTPase activity. LT-treated mice showed improved aortic endothelium-dependent vasodilation, attenuated atherosclerotic lesion formation and reduced macrophage infiltration of atherosclerotic plaques. Concomitant treatment of cholesterol-fed ApoE -/- mice with LT, the specific synthetic Rac1 inhibitor NSC 23766 or simvastatin comparably reduced aortic Rac1 activity, NADPH oxidase activity, oxidative stress, endothelial dysfunction, atherosclerosis development, and macrophage infiltration. Conclusions: These findings identify an important role of the small GTPase Rac1 in atherogenesis and provide a potential target for anti-atherosclerotic therapy., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Zimmer, Goody, Oelze, Ghanem, Mueller, Laufs, Daiber, Jansen, Nickenig and Wassmann.)

Sepsis causes high mortality in the setting of septic shock. LEADER and other trials revealed cardioprotective and anti-inflammatory properties of glucagon-like peptide-1 (GLP-1) analogs like liraglutide (Lira). We previously demonstrated improved survival in lipopolysaccharide (LPS)-induced endotoxemia by inhibition of GLP-1 degradation. Here we investigate the effects of Lira in the polymicrobial sepsis model of cecal ligation and puncture (CLP). C57BL/6J mice were intraperitoneally injected with Lira (200 µg/kg/d; 3 days) and sepsis induced by CLP after one day of GLP-1 analog treatment. Survival and body temperature were monitored. Aortic vascular function (isometric tension recording), protein expression (immunohistochemistry and dot blot) and gene expression (qRT-PCR) were determined. Endothelium-dependent relaxation in the aorta was impaired by CLP and correlated with markers of inflammation (e.g., interleukin 6 and inducible nitric oxide synthase) and oxidative stress (e.g., 3-nitrotyrosine) was higher in septic mice, all of which was almost completely normalized by Lira therapy. We demonstrate that the GLP-1 analog Lira ameliorates sepsis-induced endothelial dysfunction by the reduction of vascular inflammation and oxidative stress. Accordingly, the findings suggest that the antioxidant and anti-inflammatory effects of GLP-1 analogs may be a valuable tool to protect the cardiovascular system from dysbalanced inflammation in polymicrobial sepsis.

Superoxide formation is a hallmark of cardiovascular disease with the involvement of different tissues and cell types. Identification of the cellular sources and subcellular localization of superoxide formation is important to understand the underlying disease pathomechanisms. In the present study, we used HPLC quantification of the superoxide-specific oxidation products of hydroethidine (HE or DHE) and its derivative hydropropidine (HPr + ) for measurement of intra- and extracellular superoxide formation in isolated leukocytes and tissues of hypertensive rats. Superoxide generation by isolated leukocytes from human subjects as well as tissue samples of hypertensive rats (infusion of angiotensin-II for 7 days) was investigated using HPr + and HE fluorescent probes with HPLC or plate reader detection. Both fluorescent dyes were used to test for intra- and extracellular superoxide formation using the supernatant or cell/tissue pellet for analysis. We demonstrate the correlation of impaired functional parameters (blood pressure, vascular function, and oxidative burst) and increased superoxide formation in different organ systems of hypertensive rats using the HPr + /HPLC method. In the cell model, the differences between HE and HPr + and especially the advantage of the extracellular specificity of HPr + , due to its cell impermeability, became evident. Plate reader-based assays showed much higher background signal and were inferior to HPLC based methods. In conclusion, the HPr + /HPLC assay for superoxide determination is highly reliable in isolated immune cells and an animal model of arterial hypertension. In particular, the cell impermeability of HPr + made it possible to differentiate between intra- and extracellular superoxide formation., (Copyright © 2021 Elsevier Inc. All rights reserved.)

Aircraft noise induces vascular and cerebral inflammation and oxidative stress causing hypertension and cardiovascular/cerebral dysfunction. With the present studies, we sought to determine the role of myeloid cells in the vascular vs. cerebral consequences of exposure to aircraft noise. Toxin-mediated ablation of lysozyme M + (LysM + ) myeloid cells was performed in LysMCre iDTR mice carrying a cre-inducible diphtheria toxin receptor. In the last 4d of toxin treatment, the animals were exposed to noise at maximum and mean sound pressure levels of 85 and 72 dB(A), respectively. Flow cytometry analysis revealed accumulation of CD45 + , CD11b + , F4/80 + , and Ly6G - Ly6C + cells in the aortas of noise-exposed mice, which was prevented by LysM + cell ablation in the periphery, whereas brain infiltrates were even exacerbated upon ablation. Aircraft noise-induced increases in blood pressure and endothelial dysfunction of the aorta and retinal/mesenteric arterioles were almost completely normalized by ablation. Correspondingly, reactive oxygen species in the aorta, heart, and retinal/mesenteric vessels were attenuated in ablated noise-exposed mice, while microglial activation and abundance in the brain was greatly increased. Expression of phagocytic NADPH oxidase (NOX-2) and vascular cell adhesion molecule-1 (VCAM-1) mRNA in the aorta was reduced, while NFκB signaling appeared to be activated in the brain upon ablation. In sum, we show dissociation of cerebral and peripheral inflammatory reactions in response to aircraft noise after LysM + cell ablation, wherein peripheral myeloid inflammatory cells represent a dominant part of the pathomechanism for noise stress-induced cardiovascular effects and their central nervous counterparts, microglia, as key mediators in stress responses.

Vascular oxidative stress, inflammation, and subsequent endothelial dysfunction are consequences of traditional cardiovascular risk factors, all of which contribute to cardiovascular disease. Environmental stressors, such as traffic noise and air pollution, may also facilitate the development and progression of cardiovascular and metabolic diseases. In our previous studies, we investigated the influence of aircraft noise exposure on molecular mechanisms, identifying oxidative stress and inflammation as central players in mediating vascular function. The present study investigates the role of heme oxygenase-1 (HO-1) as an antioxidant response preventing vascular consequences following exposure to aircraft noise. C57BL/6J mice were treated with the HO-1 inducer hemin (25 mg/kg i.p.) or the NRF2 activator dimethyl fumarate (DMF, 20 mg/kg p.o.). During therapy, the animals were exposed to noise at a maximum sound pressure level of 85 dB(A) and a mean sound pressure level of 72 dB(A). Our data showed a marked protective effect of both treatments on animals exposed to noise for 4 days by normalization of arterial hypertension and vascular dysfunction in the noise-exposed groups. We observed a partial normalization of noise-triggered oxidative stress and inflammation by hemin and DMF therapy, which was associated with HO-1 induction. The present study identifies possible new targets for the mitigation of the adverse health effects caused by environmental noise exposure. Since natural dietary constituents can achieve HO-1 and NRF2 induction, these pathways represent promising targets for preventive measures.

Global epidemiological studies reported a shift from maternal/infectious communicable diseases to chronic non-communicable diseases and a major part is attributable to atherosclerosis and metabolic disorders. Accordingly, ischemic heart disease was identified as a leading risk factor for global mortality and morbidity with a prevalence of 128 million people. Almost 9 million premature deaths can be attributed to ischemic heart disease and subsequent acute myocardial infarction and heart failure, also representing a substantial socioeconomic burden. As evidenced by typical oxidative stress markers such as lipid peroxidation products or oxidized DNA/RNA bases, the formation of reactive oxygen species by various sources (NADPH oxidases, xanthine oxidase and mitochondrial resperatory chain) plays a central role for the severity of ischemia/reperfusion damage. The underlying mechanisms comprise direct oxidative damage but also adverse redox-regulation of kinase and calcium signaling, inflammation and cardiac remodeling among others. These processes and the role of reactive oxygen species are discussed in the present review. We also present and discuss potential targets for redox-based therapies that are either already established in the clinics (e.g. guanylyl cyclase activators and stimulators) or at least successfully tested in preclinical models of myocardial infarction and heart failure (mitochondria-targeted antioxidants). However, reactive oxygen species have not only detrimental effects but are also involved in essential cellular signaling and may even act protective as seen by ischemic pre- and post-conditioning or eustress - which makes redox therapy quite challenging., (Copyright © 2020 Elsevier Inc. All rights reserved.)

The COVID-19 pandemic disrupted the world in 2020 by spreading at unprecedented rates and causing tens of thousands of fatalities within a few months. The number of deaths dramatically increased in regions where the number of patients in need of hospital care exceeded the availability of care. Many COVID-19 patients experience Acute Respiratory Distress Syndrome (ARDS), a condition that can be treated with mechanical ventilation. In response to the need for mechanical ventilators, designed and tested an emergency ventilator (EV) that can control a patient's peak inspiratory pressure (PIP) and breathing rate, while keeping a positive end expiratory pressure (PEEP). This article describes the rapid design, prototyping, and testing of the EV. The development process was enabled by rapid design iterations using additive manufacturing (AM). In the initial design phase, iterations between design, AM, and testing enabled a working prototype within one week. The designs of the 16 different components of the ventilator were locked by additively manufacturing and testing a total of 283 parts having parametrically varied dimensions. In the second stage, AM was used to produce 75 functional prototypes to support engineering evaluation and animal testing. The devices were tested over more than two million cycles. We also developed an electronic monitoring system and with automatic alarm to provide for safe operation, along with training materials and user guides. The final designs are available online under a free license. The designs have been transferred to more than 70 organizations in 15 countries. This project demonstrates the potential for ultra-fast product design, engineering, and testing of medical devices needed for COVID-19 emergency response., Competing Interests: The study was funded by University of Illinois Urbana-Champaign and by Carle Foundation Hospital. GD is principal at Tekmill Inc., which worked on the project at the direction of and under contract to the University of Illinois Urbana-Champaign. SE, AM, and BR are employees of Creative Thermal Solutions Inc., which worked on the project at the direction of and under contract to the University of Illinois Urbana-Champaign. DB, MN, JO, SR, and CW are employees at Fast Radius Inc., which worked on the project at the direction of and under contract to the University of Illinois Urbana-Champaign. WPK is Professor at University of Illinois Urbana-Champaign and Chief Scientist at Fast Radius Inc., which manufactured prototypes used in this study. WPK performed the work in as an employee at the University of Illinois Urbana-Champaign and received no compensation from Fast Radius. This project was conducted in accordance with conflict of management policies at both organizations. This does not alter our adherence to PLOS ONE policies on sharing data and materials. There are no other patents, products in development or marketed products associated with this research to declare.

Background: The superoxide-generating enzyme nicotinamide adenine dinucleotide phosphate (NADPH) oxidase (NOX2 or gp91phox, the phagocytic isoform) was reported as a major source of oxidative stress in various human diseases. Genetic deletion is widely used to study the impact of NOX2-derived reactive oxygen species (ROS) on disease development and progression in various animal models. Here, we investigate why NOX2 knockout mice show no NOX2 activity but express NOX2 mRNA and protein., Methods and Results: Oxidative burst (NOX2-dependent formation of ROS) was measured by L-012-based chemiluminescence and was largely absent in whole blood of NOX2 knockout mice. Protein expression was still detectable in different tissues of the NOX2 knockout mice, at the expected and a slightly lower molecular weight (determined by Western blot). The NOX2 gene was even largely enhanced at its expressional level in NOX2 knockout mice. RNA sequencing revealed a modified NOX2 mRNA in the knockout mice that is obviously translated to a truncated inactive mutant enzyme., Conclusion: Although the commercial NOX2 knockout mice display no considerable enzymatic NOX2 activity, expression of the NOX2 gene (when using standard primers) and protein (when using antibodies binding to the carboxy-terminal end) can still be detected, which may lead to confusion among investigators.

We have demonstrated that exposing human breast tumour xenografts to ultrasound-stimulated microbubbles enhances tumour cell death and vascular disruption resulting from hyperthermia treatment. The aim of this study was to investigate the effect of varying the hyperthermia and ultrasound-stimulated microbubbles treatment parameters in order to optimize treatment bioeffects. Human breast cancer (MDA-MB-231) tumour xenografts in severe combined immunodeficiency (SCID) mice were exposed to varying microbubble concentrations (0%, 0.1%, 1% or 3% v/v) and ultrasound sonication durations (0, 1, 3 or 5 min) at 570 kPa peak negative pressure and central frequency of 500 kHz. Five hours later, tumours were immersed in a 43°C water bath for varying hyperthermia treatment durations (0, 10, 20, 30, 40, 50 or 60 minutes). Results indicated a significant increase in tumour cell death reaching 64 ± 5% with combined treatment compared to 11 ± 3% and 26 ± 5% for untreated and USMB-only treated tumours, respectively. A similar but opposite trend was observed in the vascular density of the tumours receiving the combined treatment. Optimal treatment parameters were found to consist of 40 minutes of heat with low power ultrasound treatment microbubble parameters of 1 minute of sonification and a 1% microbubble concentration., Competing Interests: The authors have declared that no competing interests exist.

Aims: Electronic (e)-cigarettes have been marketed as a 'healthy' alternative to traditional combustible cigarettes and as an effective method of smoking cessation. There are, however, a paucity of data to support these claims. In fact, e-cigarettes are implicated in endothelial dysfunction and oxidative stress in the vasculature and the lungs. The mechanisms underlying these side effects remain unclear. Here, we investigated the effects of e-cigarette vapour on vascular function in smokers and experimental animals to determine the underlying mechanisms., Methods and Results: Acute e-cigarette smoking produced a marked impairment of endothelial function in chronic smokers determined by flow-mediated dilation. In mice, e-cigarette vapour without nicotine had more detrimental effects on endothelial function, markers of oxidative stress, inflammation, and lipid peroxidation than vapour containing nicotine. These effects of e-cigarette vapour were largely absent in mice lacking phagocytic NADPH oxidase (NOX-2) or upon treatment with the endothelin receptor blocker macitentan or the FOXO3 activator bepridil. We also established that the e-cigarette product acrolein, a reactive aldehyde, recapitulated many of the NOX-2-dependent effects of e-cigarette vapour using in vitro blood vessel incubation., Conclusions: E-cigarette vapour exposure increases vascular, cerebral, and pulmonary oxidative stress via a NOX-2-dependent mechanism. Our study identifies the toxic aldehyde acrolein as a key mediator of the observed adverse vascular consequences. Thus, e-cigarettes have the potential to induce marked adverse cardiovascular, pulmonary, and cerebrovascular consequences. Since e-cigarette use is increasing, particularly amongst youth, our data suggest that aggressive steps are warranted to limit their health risks., (© The Author(s) 2019. Published by Oxford University Press on behalf of the European Society of Cardiology.)

Arterial hypertension is the most important risk factor for the development of cardiovascular disease. Recently, aircraft noise has been shown to be associated with elevated blood pressure, endothelial dysfunction, and oxidative stress. Here, we investigated the potential exacerbated cardiovascular effects of aircraft noise in combination with experimental arterial hypertension. C57BL/6J mice were infused with 0.5 mg/kg/d of angiotensin II for 7 days, exposed to aircraft noise for 7 days at a maximum sound pressure level of 85 dB(A) and a mean sound pressure level of 72 dB(A), or subjected to both stressors. Noise and angiotensin II increased blood pressure, endothelial dysfunction, oxidative stress and inflammation in aortic, cardiac and/or cerebral tissues in single exposure models. In mice subjected to both stressors, most of these risk factors showed potentiated adverse changes. We also found that mice exposed to both noise and ATII had increased phagocytic NADPH oxidase (NOX-2)-mediated superoxide formation, immune cell infiltration (monocytes, neutrophils and T cells) in the aortic wall, astrocyte activation in the brain, enhanced cytokine signaling, and subsequent vascular and cerebral oxidative stress. Exaggerated renal stress response was also observed. In summary, our results show an enhanced adverse cardiovascular effect between environmental noise exposure and arterial hypertension, which is mainly triggered by vascular inflammation and oxidative stress. Mechanistically, noise potentiates neuroinflammation and cerebral oxidative stress, which may be a potential link between both risk factors. The results indicate that a combination of classical (arterial hypertension) and novel (noise exposure) risk factors may be deleterious for cardiovascular health., Competing Interests: Declaration of competing interest Nothing to declare., (Copyright © 2020 The Authors. Published by Elsevier B.V. All rights reserved.)

Environmental pollution and non-chemical stressors such as mental stress or traffic noise exposure are increasingly accepted as health risk factors with substantial contribution to chronic noncommunicable diseases (e.g. cardiovascular, metabolic and mental). Whereas the mechanisms of air pollution-mediated adverse health effects are well characterized, the mechanisms of traffic noise exposure are not completely understood, despite convincing clinical and epidemiological evidence for a significant contribution of environmental noise to overall mortality and disability. The initial mechanism of noise-induced cardiovascular, metabolic and mental disease is well defined by the "noise reaction model" and consists of neuronal activation involving the hypothalamic-pituitary-adrenal (HPA) axis as well as the sympathetic nervous system, followed by a classical stress response via cortisol and catecholamines. Stress pathways are initiated by noise-induced annoyance and sleep deprivation/fragmentation. This review highlights the down-stream pathophysiology of noise-induced mental stress, which is based on an induction of inflammation and oxidative stress. We highlight the sources of reactive oxygen species (ROS) involved and the known targets for noise-induced oxidative damage. Part of the review emphasizes noise-triggered uncoupling/dysregulation of endothelial and neuronal nitric oxide synthase (eNOS and nNOS) and its central role for vascular dysfunction., Competing Interests: Declaration of competing interest The authors declare that they have no conflicts of interest with the contents of this article., (Copyright © 2020 The Authors. Published by Elsevier B.V. All rights reserved.)

Oxidative stress plays a key role for the development of cardiovascular, metabolic, and neurodegenerative disease. This concept has been proven by using the approach of genetic deletion of reactive oxygen and nitrogen species (RONS) producing, pro-oxidant enzymes as well as by the overexpression of RONS detoxifying, antioxidant enzymes leading to an amelioration of the severity of diseases. Vice versa, the development and progression of cardiovascular diseases is aggravated by overexpression of RONS producing enzymes as well as deletion of RONS detoxifying enzymes. We have previously identified cross talk mechanisms between different sources of RONS, which can amplify the oxidative stress-mediated damage. Here, the pathways and potential mechanisms leading to this cross talk are analyzed in detail and highlighted by selected examples from the current literature and own data including hypoxia, angiotensin II (AT-II)-induced hypertension, nitrate tolerance, aging, and others. The general concept of redox-based activation of RONS sources via "kindling radicals" and enzyme-specific "redox switches" as well as the interaction with redox-sensitive inflammatory pathways are discussed. Here, we present evidence for the existence of such cross talk mechanisms in the setting of diabetes and critically assess their contribution to the severity of diabetic complications.

Reactive oxygen and nitrogen species (RONS) cause oxidative damage, which is associated with endothelial dysfunction and cardiovascular disease, but may also contribute to redox signaling. Therefore, their precise detection is important for the evaluation of disease mechanisms. Here, we compared three different methods for the detection of 3-nitrotyrosine (3-NT), a marker of nitro-oxidative stress, in biological samples. Nitrated proteins were generated by incubation with peroxynitrite or 3-morpholino sydnonimine (Sin-1) and subjected to total hydrolysis using pronase, a mixture of different proteases. The 3-NT was then separated by high performance liquid chromatography (HPLC) and quantified by electrochemical detection (ECD, CoulArray) and compared to classical methods, namely enzyme-linked immunosorbent assay (ELISA) and dot blot analysis using specific 3-NT antibodies. Calibration curves for authentic 3-NT (detection limit 10 nM) and a concentration-response pattern for 3-NT obtained from digested nitrated bovine serum albumin (BSA) were highly linear over a wide 3-NT concentration range. Also, ex vivo nitration of protein from heart, isolated mitochondria, and serum/plasma could be quantified using the HPLC/ECD method and was confirmed by LC-MS/MS. Of note, nitro-oxidative damage of mitochondria results in increased superoxide (O 2 •- ) formation rates (measured by dihydroethidium-based HPLC assay), pointing to a self-amplification mechanism of oxidative stress. Based on our ex vivo data, the CoulArray quantification method for 3-NT seems to have some advantages regarding sensitivity and selectivity. Establishing a reliable automated HPLC assay for the routine quantification of 3-NT in biological samples of cell culture, of animal and human origin seems to be more sophisticated than expected., Competing Interests: Abbreviations: 3-NT, 3-nitrotyrosine; Sin-1, 3-morpholino sydnonimine; PN, peroxynitrite; ROS, reactive oxygen species; Reactive oxygen and nitrogen species (RONS); MnSOD, manganese superoxide dismutase (mitochondrial isoform); HPLC/ECD, high performance liquid chromatography with electrochemical detection; enzyme-linked immunosorbent assay (ELISA) O2•–, superoxide; BSA, bovine serum albumin; mitoSOX, triphenylphosphonium-linked dihydroethidium; NOS, nitric oxide synthase; dTH, dithionite; CoulArray, coulometric array system; retention time (RT).

Exposure to traffic noise is associated with stress and sleep disturbances. The World Health Organization (WHO) recently concluded that road traffic noise increases the risk for ischemic heart disease and potentially other cardiometabolic diseases, including stroke, obesity, and diabetes. The WHO report focused on whole-day noise exposure, but new epidemiological and translational field noise studies indicate that nighttime noise, in particular,is an important risk factor for cardiovascular disease (CVD) through increased levels of stress hormones and vascular oxidative stress, leading to endothelial dysfunction and subsequent development of various CVDs. Novel experimental studies found noise to be associated with oxidative stress-induced vascular and brain damage, mediated by activation of the NADPH oxidase, uncoupling of endothelial and neuronal nitric oxide synthase, and vascular/brain infiltration with inflammatory cells. Noise-induced pathophysiology was more pronounced in response to nighttime as compared with daytime noise. This review focuses on the consequences of nighttime noise.

Background: Large epidemiological studies point towards a link between the incidence of arterial hypertension, ischaemic heart disease, metabolic disease and exposure to traffic noise, supporting the role of noise exposure as an independent cardiovascular risk factor. We characterised the underlying molecular mechanisms leading to noise-dependent adverse effects on the vasculature and myocardium in an animal model of aircraft noise exposure and identified oxidative stress and inflammation as central players in mediating vascular and cardiac dysfunction. Here, we studied the impact of noise-induced oxidative DNA damage on vascular function in DNA-repair deficient 8-oxoguanine glycosylase knockout ( Ogg1 -/- ) mice. Methods and results: Noise exposure (peak sound levels of 85 and mean sound level of 72 dB(A) applied for 4d) caused oxidative DNA damage (8-oxoguanine) and enhanced NOX-2 expression in C57BL/6 mice with synergistic increases in Ogg1 -/- mice (shown by immunohistochemistry). A similar pattern was found for oxidative burst of blood leukocytes and other markers of oxidative stress (4-hydroxynonenal, 3-nitrotyrosine) and inflammation (cyclooxygenase-2). We observed additive impairment of noise exposure and genetic Ogg1 deficiency on endothelium-independent relaxation (nitroglycerine), which may be due to exacerbated oxidative DNA damage leading to leukocyte activation and oxidative aldehyde dehydrogenase inhibition. Conclusions: The finding that chronic noise exposure causes oxidative DNA damage in mice is worrisome since these potential mutagenic lesions could contribute to cancer progression. Human field studies have to demonstrate whether oxidative DNA damage is also found in urban populations with high levels of noise exposure as recently shown for workers with high occupational noise exposure.

Objective: Cardiovascular outcome trials demonstrated that GLP-1 (glucagon-like peptide-1) analogs including liraglutide reduce the risk of cardiovascular events in type 2 diabetes mellitus. Whether GLP-1 analogs reduce the risk for atherosclerosis independent of glycemic control is challenging to elucidate as the GLP-1R (GLP-1 receptor) is expressed on different cell types, including endothelial and immune cells. Approach and Results: Here, we reveal the cardio- and vasoprotective mechanism of the GLP-1 analog liraglutide at the cellular level in a murine, nondiabetic model of arterial hypertension. Wild-type (C57BL/6J), global ( Glp1r -/- ), as well as endothelial ( Glp1r f lox/floxxCdh5 cre ) and myeloid cell-specific knockout mice ( Glp1r flox/flox xLysM cre ) of the GLP-1R were studied, and arterial hypertension was induced by angiotensin II. Liraglutide treatment normalized blood pressure, cardiac hypertrophy, vascular fibrosis, endothelial dysfunction, oxidative stress, and vascular inflammation in a GLP-1R-dependent manner. Mechanistically, liraglutide reduced leukocyte rolling on the endothelium and infiltration of myeloid Ly6G - Ly6C + and Ly6G + Ly6C + cells into the vascular wall. As a consequence, liraglutide prevented vascular oxidative stress, reduced S-glutathionylation as a marker of eNOS (endothelial NO synthase) uncoupling, and increased NO bioavailability. Importantly, all of these beneficial cardiovascular effects of liraglutide persisted in myeloid cell GLP-1R-deficient ( Glp1r flox/flox xLysM cre ) mice but were abolished in global ( Glp1r -/- ) and endothelial cell-specific ( Glp1r flox/flox xCdh5 cre ) GLP-1R knockout mice., Conclusions: GLP-1R activation attenuates cardiovascular complications of arterial hypertension by reduction of vascular inflammation through selective actions requiring the endothelial but not the myeloid cell GLP-1R.