Background: Dipeptidyl peptidase [DPP]-4 inhibitors are a new class of drug for the treatment of ... more Background: Dipeptidyl peptidase [DPP]-4 inhibitors are a new class of drug for the treatment of hyperglycemia and recent studies revealed anti-inflammatory effects of these gliptins in experimenta...
Vascular oxidative stress, inflammation, and subsequent endothelial dysfunction are consequences ... more 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). Ou...
Oxidative stress plays a key role for the development of cardiovascular, metabolic, and neurodege... more 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 gene...
Reactive oxygen and nitrogen species (RONS) cause oxidative damage, which is associated with endo... more 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 obt...
Background: Reactive oxygen and nitrogen species (RONS such as H2O2, nitric oxide) are generated ... more Background: Reactive oxygen and nitrogen species (RONS such as H2O2, nitric oxide) are generated within the organism. Whereas physiological formation rates confer redox regulation of essential cellular functions and provide the basis for adaptive stress responses, their excessive formation contributes to impaired cellular function or even cell death, organ dysfunction and severe disease phenotypes of the entire organism. Therefore, quantification of RONS formation and knowledge of their tissue/cell/compartment-specific distribution is of great biological and clinical importance. Methods: Here, we used a high-performance/pressure liquid chromatography (HPLC) assay to quantify the superoxide-specific oxidation product of the mitochondria-targeted fluorescence dye triphenylphosphonium-linked hydroethidium (mitoSOX) in biochemical systems and three animal models with established oxidative stress. Type 1 diabetes (single injection of streptozotocin), hypertension (infusion of angiotensin...
The Global Burden of Disease Study identified cardiovascular risk factors as leading causes of gl... more The Global Burden of Disease Study identified cardiovascular risk factors as leading causes of global deaths and life years lost. Endothelial dysfunction represents a pathomechanism that is associated with most of these risk factors and stressors, and represents an early (subclinical) marker/predictor of atherosclerosis. Oxidative stress is a trigger of endothelial dysfunction and it is a hall-mark of cardiovascular diseases and of the risk factors/stressors that are responsible for their initiation. Endothelial function is largely based on endothelial nitric oxide synthase (eNOS) function and activity. Likewise, oxidative stress can lead to the loss of eNOS activity or even “uncoupling” of the enzyme by adverse regulation of well-defined “redox switches” in eNOS itself or up-/down-stream signaling molecules. Of note, not only eNOS function and activity in the endothelium are essential for vascular integrity and homeostasis, but also eNOS in perivascular adipose tissue plays an impo...
Aims CD40 ligand (CD40L) signaling controls vascular oxidative stress and related dysfunction in ... more Aims CD40 ligand (CD40L) signaling controls vascular oxidative stress and related dysfunction in angiotensin-II-induced arterial hypertension by regulating vascular immune cell recruitment and platelet activation. Here we investigated the role of CD40L in experimental hyperlipidemia. Methods and results Male wild type and CD40L−/− mice (C57BL/6 background) were subjected to high fat diet for sixteen weeks. Weight, cholesterol, HDL, and LDL levels, endothelial function (isometric tension recording), oxidative stress (NADPH oxidase expression, dihydroethidium fluorescence) and inflammatory parameters (inducible nitric oxide synthase, interleukin-6 expression) were assessed. CD40L expression, weight, leptin and lipids were increased, and endothelial dysfunction, oxidative stress and inflammation were more pronounced in wild type mice on a high fat diet, all of which was almost normalized by CD40L deficiency. Similar results were obtained in diabetic db/db mice with CD40/TRAF6 inhibitor...
Hyperglycemia associated with inflammation and oxidative stress is a major cause of vascular dysf... more Hyperglycemia associated with inflammation and oxidative stress is a major cause of vascular dysfunction and cardiovascular disease in diabetes. Recent data reports that a selective sodium-glucose co-transporter 2 inhibitor (SGLT2i), empagliflozin (Jardiance ®), ameliorates glucotoxicity via excretion of excess glucose in urine (glucosuria) and significantly improves cardiovascular mortality in type 2 diabetes mellitus (T2DM). The overarching hypothesis is that hyperglycemia and glucotoxicity are upstream of all other complications seen in diabetes. The aim of this study was to investigate effects of empagliflozin on glucotoxicity, β-cell function, inflammation, oxidative stress and endothelial dysfunction in Zucker diabetic fatty (ZDF) rats. Male ZDF rats were used as a model of T2DM (35 diabetic ZDF-Lepr fa/fa and 16 ZDF-Lepr +/+ controls). Empagliflozin (10 and 30 mg/kg/d) was administered via drinking water for 6 weeks. Treatment with empagliflozin restored glycemic control. Empagliflozin improved endothelial function (thoracic aorta) and reduced oxidative stress in the aorta and in blood of diabetic rats. Inflammation and glucotoxicity (AGE/RAGE signaling) were epigenetically prevented by SGLT2i treatment (ChIP). Linear regression analysis revealed a significant inverse correlation of endothelial function with HbA1c, whereas leukocyte-dependent oxidative burst and C-reactive protein (CRP) were positively correlated with HbA1c. Viability of hyperglycemic endothelial cells was pleiotropically improved by SGLT2i. Empagliflozin reduces glucotoxicity and thereby prevents the development of endothelial dysfunction, reduces oxidative stress and exhibits anti-inflammatory effects in ZDF rats, despite persisting hyperlipidemia and hyperinsulinemia. Our preclinical observations provide insights into the mechanisms by which empagliflozin reduces cardiovascular mortality in humans (EMPA-REG trial).
Non-alcoholic steatohepatitis (NASH) is characterized by steatosis, panlobular inflammation, live... more Non-alcoholic steatohepatitis (NASH) is characterized by steatosis, panlobular inflammation, liver fibrosis and increased cardiovascular mortality. Dipeptidyl peptidase-4 (DPP-4) inhibitors (gliptins) are indirect glucagon like peptide 1 (GLP-1) agonists with antidiabetic and anti-inflammatory activity, used for the treatment of type 2 diabetes. Their potential and underlying mechanisms to treat metabolic liver inflammation and fibrosis as well as the associated vascular dysfunction remain to be explored. In the methionine/choline deficient (MCD) diet and Mdr2-/- models of NASH and liver fibrosis, treatment with sitagliptin and linagliptin significantly decreased parameters of steatosis and inflammation, which was accompanied by suppression of hepatic transcripts levels reflecting metabolic inflammation and fibrosis like SREBP-1c, FAS, TNFα, iNOS, α-SMA, Col1α1 and MMP-12. Moreover, gliptins reduced the number of liver infiltrating CD11b+Ly6Chi proinflammatory monocytes/macrophages ...
The European Cooperation in Science and Technology (COST) provides an ideal framework to establis... more The European Cooperation in Science and Technology (COST) provides an ideal framework to establish multi-disciplinary research networks. COST Action BM1203 (EU-ROS) represents a consortium of researchers from different disciplines who are dedicated to providing new insights and tools for better understanding redox biology and medicine and, in the long run, to finding new therapeutic strategies to target dysregulated redox processes in various diseases. This report highlights the major achievements of EU-ROS as well as research updates and new perspectives arising from its members. The EU-ROS consortium comprised more than 140 active members who worked together for four years on the topics briefly described below. The formation of reactive oxygen and nitrogen species (RONS) is an established hallmark of our aerobic environment and metabolism but RONS also act as messengers via redox regulation of essential cellular processes. The fact that many diseases have been found to be associat...
Epidemiological studies indicate that traffic noise increases the incidence of coronary artery di... more Epidemiological studies indicate that traffic noise increases the incidence of coronary artery disease, hypertension and stroke. The underlying mechanisms remain largely unknown. Field studies with nighttime noise exposure demonstrate that aircraft noise leads to vascular dysfunction, which is markedly improved by vitamin C, suggesting a key role of oxidative stress in causing this phenomenon. We developed a novel animal model to study the vascular consequences of aircraft noise exposure. Peak sound levels of 85 and mean sound level of 72 dBA applied by loudspeakers for 4 days caused an increase in systolic blood pressure, plasma noradrenaline and angiotensin II levels and induced endothelial dysfunction. Noise increased eNOS expression but reduced vascular NO levels because of eNOS uncoupling. Noise increased circulating levels of nitrotyrosine, interleukine-6 and vascular expression of the NADPH oxidase subunit Nox2, nitrotyrosine-positive proteins and of endothelin-1. FACS analys...
Objective. Oxidative stress and endothelial dysfunction contribute to pulmonary arterial hyperten... more Objective. Oxidative stress and endothelial dysfunction contribute to pulmonary arterial hypertension (PAH). The role of the nitrovasodilator pentaerythritol tetranitrate (PETN) on endothelial function and oxidative stress in PAH has not yet been defined. Methods and Results. PAH was induced by monocrotaline (MCT, i.v.) in Wistar rats. Low (30 mg/kg; MCT30), middle (40 mg/kg; MCT40), or high (60 mg/kg; MCT60) dose of MCT for 14, 28, and 42 d was used. MCT induced endothelial dysfunction, pulmonary vascular wall thickening, and fibrosis, as well as protein tyrosine nitration. Pulmonary arterial pressure and heart/body and lung/body weight ratio were increased in MCT40 rats (28 d) and reduced by oral PETN (10 mg/kg, 24 d) therapy. Oxidative stress in the vascular wall, in the heart, and in whole blood as well as vascular endothelin-1 signaling was increased in MCT40-treated rats and normalized by PETN therapy, likely by upregulation of heme oxygenase-1 (HO-1). PETN therapy improved en...
Reactive oxygen and nitrogen species (RONS such as H 2 O 2 , nitric oxide) confer redox regulatio... more Reactive oxygen and nitrogen species (RONS such as H 2 O 2 , nitric oxide) confer redox regulation of essential cellular functions (e.g. differentiation, proliferation, migration, apoptosis), initiate and catalyze adaptive stress responses. In contrast, excessive formation of RONS caused by impaired breakdown by cellular antioxidant systems and/or insufficient repair of the resulting oxidative damage of biomolecules may lead to appreciable impairment of cellular function and in the worst case to cell death, organ dysfunction and severe disease phenotypes of the entire organism. Therefore, the knowledge of the severity of oxidative stress and tissue specific localization is of great biological and clinical importance. However, at this level of investigation quantitative information may be enough. For the development of specific drugs, the cellular and subcellular localization of the sources of RONS or even the nature of the reactive species may be of great importance, and accordingly, more qualitative information is required. These two different philosophies currently compete with each other and their different needs (also reflected by different detection assays) often lead to controversial discussions within the redox research community. With the present review we want to shed some light on these different philosophies and needs (based on our personal views), but also to defend some of the traditional assays for the detection of RONS that work very well in our hands and to provide some guidelines how to use and interpret the results of these assays. We will also provide an overview on the "new assays" with a brief discussion on their strengths but also weaknesses and limitations.
Excessive inflammation in sepsis causes microvascular thrombosis and thrombocytopenia associated ... more Excessive inflammation in sepsis causes microvascular thrombosis and thrombocytopenia associated with organ dysfunction and high mortality. The present studies aimed to investigate whether dipeptidyl peptidase-4 (DPP-4) inhibition and supplementation with glucagon-like peptide-1 (GLP-1) analogues improves endotoxaemia-associated microvascular thrombosis via immunomodulatory effects. Endotoxaemia was induced in C57BL/6 J mice by single injection of lipopolysaccharide (LPS) (17.5 for survival and 10 mg/kg for all other studies). For survival studies therapy was started 6 h after LPS injection. For all other studies drugs were administrated 48 h prior to LPS treatment. Endotoxaemic control mice showed severe thrombocytopenia, microvascular thrombosis in the pulmonary circulation (fluorescence imaging), increased lactate dehydrogenase (LDH)-activity, endothelial dysfunction and increased markers of inflammation in the aorta and whole blood (e.g. leukocyte-dependent oxidative burst, nitr...
Nitroglycerin (GTN) and other organic nitrates are widely used vasodilators. Their side effects a... more Nitroglycerin (GTN) and other organic nitrates are widely used vasodilators. Their side effects are development of nitrate tolerance and endothelial dysfunction. Given the potential of GTN to induce nitro-oxidative stress, we investigated the interaction between nitro-oxidative DNA damage and vascular dysfunction in experimental nitrate tolerance. Cultured endothelial hybridoma cells (EA.hy 926) and Wistar rats were treated with GTN (ex vivo: 10-1000 µM; in vivo: 10, 20 and 50 mg/kg/day for 3 days, s.c.). The level of DNA strand breaks, 8-oxoguanine and O (6)-methylguanine DNA adducts was determined by Comet assay, dot blot and immunohistochemistry. Vascular function was determined by isometric tension recording. DNA adducts and strand breaks were induced by GTN in cells in vitro in a concentration-dependent manner. GTN in vivo administration leads to endothelial dysfunction, nitrate tolerance, aortic and cardiac oxidative stress, formation of DNA adducts, stabilization of p53 and a...
Cardiovascular diseases are major contributors to global deaths and disability-adjusted life year... more Cardiovascular diseases are major contributors to global deaths and disability-adjusted life years, with hypertension as a significant risk factor for all cause death. The endothelium that lines the inner wall of the vasculature regulates essential haemostatic functions such as regulation of the vascular tone, circulation of blood cells, inflammation and platelet activity. Endothelial dysfunction is an early predictor of atherosclerosis and future cardiovascular events. We review the prognostic value of measurements of endothelial function, the clinical techniques for its determination, mechanisms leading to endothelial dysfunction and the therapeutic treatment of endothelial dysfunction. Since vascular oxidative stress and inflammation are major determinants of endothelial function, we also address current antioxidant and anti-inflammatory therapies. In light of recent data that speak against the prognostic value of endothelial function in healthy human cohorts, we also discuss alt...
There has been some dispute regarding reaction products formed at physiological peroxynitrite flu... more There has been some dispute regarding reaction products formed at physiological peroxynitrite fluxes in the nanomolar range with phenolic molecules, when used to predict the behavior of protein-bound aromatic amino acids like tyrosine. Previous data showed that at nanomolar fluxes of peroxynitrite, nitration of these phenolic compounds was outcompeted by dimerization (e.g. biphenols or dityrosine). Using 3-morpholino sydnonimine (Sin-1), we created low fluxes of peroxynitrite in our reaction setup to demonstrate that salicylaldehyde displays unique features in the detection of physiological fluxes of peroxynitrite, yielding detectable nitration but only minor dimerization products. By means of HPLC analysis and detection at 380 nm we could identify the expected nitration products 3-and 5-nitrosalicylaldehyde, but also novel nitrated products. Using mass spectrometry, we also identified 2-nitrophenol and a not fully characterized nitrated dimerization product. The formation of 2-nitrophenol could proceed either by primary generation of a phenoxy radical, followed by addition of the NO 2-radical to the various resonance structures, or by addition of the peroxynitrite anion to the polarized carbonyl group with subsequent fragmentation of the adduct (as seen with carbon dioxide). Interestingly, we observed almost no 3-and 5-nitrosalicylic acid products and only minor dimerization reaction. Our results disagree with the previous general assumption that nitration of low molecular weight phenolic compounds is always outcompeted by dimerization at nanomolar peroxynitrite fluxes and highlight unique features of salicylaldehyde as a probe for physiological concentrations of peroxynitrite.
Cardiovascular diseases are associated with and/or caused by oxidative stress. This concept has b... more Cardiovascular diseases are associated with and/or caused by oxidative stress. This concept has been proven by using the approach of genetic deletion of reactive species producing (pro-oxidant) enzymes as well as by the overexpression of reactive species detoxifying (antioxidant) enzymes leading to a marked reduction of reactive oxygen and nitrogen species (RONS) and in parallel to an amelioration of the severity of diseases. Likewise, the development and progression of cardiovascular diseases is aggravated by overexpression of RONS producing enzymes as well as deletion of antioxidant RONS detoxifying enzymes. Thus, the consequences of the interaction (redox crosstalk) of superoxide/hydrogen peroxide produced by mitochondria with other ROS producing enzymes such as NADPH oxidases (Nox) are of outstanding importance and will be discussed including the consequences for endothelial nitric oxide synthase (eNOS) uncoupling as well as the redox regulation of the vascular function/tone in general (soluble guanylyl cyclase, endothelin-1, prostanoid synthesis). Pathways and potential mechanisms leading to this crosstalk will be analysed in detail and highlighted by selected examples from the current literature including hypoxia, angiotensin 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" will be discussed providing evidence that mitochondria represent key players and amplifiers of the burden of oxidative stress.
Introduction: Sepsis is an inflammatory state. Proinflammatory enzymes like inducible-NO Synthase... more Introduction: Sepsis is an inflammatory state. Proinflammatory enzymes like inducible-NO Synthase (iNOS) and NADPH-Oxidase (NOX) are activated and influence endothelial function via oxidative and nitrosative stress. Inhibition of dipeptidyl-peptidase 4 (DPP4i) is a new approach to treat diabetic patients. Matubara et al. (Circulation, 2012) showed a positive influence of DPP4i on endothelial function in an animal model Hypothesis: DPP4 inhibition influences endothelial function, hemostasis and survival in a rodent model of septic shock Methods: C57/BL6j mice were pretreated with different DPP4i (linagliptin, sitagliptin) and GLP-1-analogue (GLP1a) liraglutide for 3 days. Aorta and heart tissue was used for western-blotting and rtPCR. Blood was examined for coagulation parameters (aPTT, PT), cell count and oxidative stress Results: Treatment with DPP4i and GLP1-a improved survival and blood pressure of septic animals significantly. LPS-induced oxidative stress and nitrosylated hemoglobin (Hb-NO) levels wer...
International journal of molecular sciences, Jan 5, 2015
Cardiovascular disease is a leading cause of death and reduced quality of life worldwide. Arteria... more Cardiovascular disease is a leading cause of death and reduced quality of life worldwide. Arterial vessels are a primary target for endothelial dysfunction and atherosclerosis, which is accompanied or even driven by increased oxidative stress. Recent research in this field identified different sources of reactive oxygen and nitrogen species contributing to the pathogenesis of endothelial dysfunction. According to lessons from the past, improvement of endothelial function and prevention of cardiovascular disease by systemic, unspecific, oral antioxidant therapy are obviously too simplistic an approach. Source- and cell organelle-specific antioxidants as well as activators of intrinsic antioxidant defense systems might be more promising. Since basic research demonstrated the contribution of different inflammatory cells to vascular oxidative stress and clinical trials identified chronic inflammatory disorders as risk factors for cardiovascular events, atherosclerosis and cardiovascular...
Background: Dipeptidyl peptidase [DPP]-4 inhibitors are a new class of drug for the treatment of ... more Background: Dipeptidyl peptidase [DPP]-4 inhibitors are a new class of drug for the treatment of hyperglycemia and recent studies revealed anti-inflammatory effects of these gliptins in experimenta...
Vascular oxidative stress, inflammation, and subsequent endothelial dysfunction are consequences ... more 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). Ou...
Oxidative stress plays a key role for the development of cardiovascular, metabolic, and neurodege... more 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 gene...
Reactive oxygen and nitrogen species (RONS) cause oxidative damage, which is associated with endo... more 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 obt...
Background: Reactive oxygen and nitrogen species (RONS such as H2O2, nitric oxide) are generated ... more Background: Reactive oxygen and nitrogen species (RONS such as H2O2, nitric oxide) are generated within the organism. Whereas physiological formation rates confer redox regulation of essential cellular functions and provide the basis for adaptive stress responses, their excessive formation contributes to impaired cellular function or even cell death, organ dysfunction and severe disease phenotypes of the entire organism. Therefore, quantification of RONS formation and knowledge of their tissue/cell/compartment-specific distribution is of great biological and clinical importance. Methods: Here, we used a high-performance/pressure liquid chromatography (HPLC) assay to quantify the superoxide-specific oxidation product of the mitochondria-targeted fluorescence dye triphenylphosphonium-linked hydroethidium (mitoSOX) in biochemical systems and three animal models with established oxidative stress. Type 1 diabetes (single injection of streptozotocin), hypertension (infusion of angiotensin...
The Global Burden of Disease Study identified cardiovascular risk factors as leading causes of gl... more The Global Burden of Disease Study identified cardiovascular risk factors as leading causes of global deaths and life years lost. Endothelial dysfunction represents a pathomechanism that is associated with most of these risk factors and stressors, and represents an early (subclinical) marker/predictor of atherosclerosis. Oxidative stress is a trigger of endothelial dysfunction and it is a hall-mark of cardiovascular diseases and of the risk factors/stressors that are responsible for their initiation. Endothelial function is largely based on endothelial nitric oxide synthase (eNOS) function and activity. Likewise, oxidative stress can lead to the loss of eNOS activity or even “uncoupling” of the enzyme by adverse regulation of well-defined “redox switches” in eNOS itself or up-/down-stream signaling molecules. Of note, not only eNOS function and activity in the endothelium are essential for vascular integrity and homeostasis, but also eNOS in perivascular adipose tissue plays an impo...
Aims CD40 ligand (CD40L) signaling controls vascular oxidative stress and related dysfunction in ... more Aims CD40 ligand (CD40L) signaling controls vascular oxidative stress and related dysfunction in angiotensin-II-induced arterial hypertension by regulating vascular immune cell recruitment and platelet activation. Here we investigated the role of CD40L in experimental hyperlipidemia. Methods and results Male wild type and CD40L−/− mice (C57BL/6 background) were subjected to high fat diet for sixteen weeks. Weight, cholesterol, HDL, and LDL levels, endothelial function (isometric tension recording), oxidative stress (NADPH oxidase expression, dihydroethidium fluorescence) and inflammatory parameters (inducible nitric oxide synthase, interleukin-6 expression) were assessed. CD40L expression, weight, leptin and lipids were increased, and endothelial dysfunction, oxidative stress and inflammation were more pronounced in wild type mice on a high fat diet, all of which was almost normalized by CD40L deficiency. Similar results were obtained in diabetic db/db mice with CD40/TRAF6 inhibitor...
Hyperglycemia associated with inflammation and oxidative stress is a major cause of vascular dysf... more Hyperglycemia associated with inflammation and oxidative stress is a major cause of vascular dysfunction and cardiovascular disease in diabetes. Recent data reports that a selective sodium-glucose co-transporter 2 inhibitor (SGLT2i), empagliflozin (Jardiance ®), ameliorates glucotoxicity via excretion of excess glucose in urine (glucosuria) and significantly improves cardiovascular mortality in type 2 diabetes mellitus (T2DM). The overarching hypothesis is that hyperglycemia and glucotoxicity are upstream of all other complications seen in diabetes. The aim of this study was to investigate effects of empagliflozin on glucotoxicity, β-cell function, inflammation, oxidative stress and endothelial dysfunction in Zucker diabetic fatty (ZDF) rats. Male ZDF rats were used as a model of T2DM (35 diabetic ZDF-Lepr fa/fa and 16 ZDF-Lepr +/+ controls). Empagliflozin (10 and 30 mg/kg/d) was administered via drinking water for 6 weeks. Treatment with empagliflozin restored glycemic control. Empagliflozin improved endothelial function (thoracic aorta) and reduced oxidative stress in the aorta and in blood of diabetic rats. Inflammation and glucotoxicity (AGE/RAGE signaling) were epigenetically prevented by SGLT2i treatment (ChIP). Linear regression analysis revealed a significant inverse correlation of endothelial function with HbA1c, whereas leukocyte-dependent oxidative burst and C-reactive protein (CRP) were positively correlated with HbA1c. Viability of hyperglycemic endothelial cells was pleiotropically improved by SGLT2i. Empagliflozin reduces glucotoxicity and thereby prevents the development of endothelial dysfunction, reduces oxidative stress and exhibits anti-inflammatory effects in ZDF rats, despite persisting hyperlipidemia and hyperinsulinemia. Our preclinical observations provide insights into the mechanisms by which empagliflozin reduces cardiovascular mortality in humans (EMPA-REG trial).
Non-alcoholic steatohepatitis (NASH) is characterized by steatosis, panlobular inflammation, live... more Non-alcoholic steatohepatitis (NASH) is characterized by steatosis, panlobular inflammation, liver fibrosis and increased cardiovascular mortality. Dipeptidyl peptidase-4 (DPP-4) inhibitors (gliptins) are indirect glucagon like peptide 1 (GLP-1) agonists with antidiabetic and anti-inflammatory activity, used for the treatment of type 2 diabetes. Their potential and underlying mechanisms to treat metabolic liver inflammation and fibrosis as well as the associated vascular dysfunction remain to be explored. In the methionine/choline deficient (MCD) diet and Mdr2-/- models of NASH and liver fibrosis, treatment with sitagliptin and linagliptin significantly decreased parameters of steatosis and inflammation, which was accompanied by suppression of hepatic transcripts levels reflecting metabolic inflammation and fibrosis like SREBP-1c, FAS, TNFα, iNOS, α-SMA, Col1α1 and MMP-12. Moreover, gliptins reduced the number of liver infiltrating CD11b+Ly6Chi proinflammatory monocytes/macrophages ...
The European Cooperation in Science and Technology (COST) provides an ideal framework to establis... more The European Cooperation in Science and Technology (COST) provides an ideal framework to establish multi-disciplinary research networks. COST Action BM1203 (EU-ROS) represents a consortium of researchers from different disciplines who are dedicated to providing new insights and tools for better understanding redox biology and medicine and, in the long run, to finding new therapeutic strategies to target dysregulated redox processes in various diseases. This report highlights the major achievements of EU-ROS as well as research updates and new perspectives arising from its members. The EU-ROS consortium comprised more than 140 active members who worked together for four years on the topics briefly described below. The formation of reactive oxygen and nitrogen species (RONS) is an established hallmark of our aerobic environment and metabolism but RONS also act as messengers via redox regulation of essential cellular processes. The fact that many diseases have been found to be associat...
Epidemiological studies indicate that traffic noise increases the incidence of coronary artery di... more Epidemiological studies indicate that traffic noise increases the incidence of coronary artery disease, hypertension and stroke. The underlying mechanisms remain largely unknown. Field studies with nighttime noise exposure demonstrate that aircraft noise leads to vascular dysfunction, which is markedly improved by vitamin C, suggesting a key role of oxidative stress in causing this phenomenon. We developed a novel animal model to study the vascular consequences of aircraft noise exposure. Peak sound levels of 85 and mean sound level of 72 dBA applied by loudspeakers for 4 days caused an increase in systolic blood pressure, plasma noradrenaline and angiotensin II levels and induced endothelial dysfunction. Noise increased eNOS expression but reduced vascular NO levels because of eNOS uncoupling. Noise increased circulating levels of nitrotyrosine, interleukine-6 and vascular expression of the NADPH oxidase subunit Nox2, nitrotyrosine-positive proteins and of endothelin-1. FACS analys...
Objective. Oxidative stress and endothelial dysfunction contribute to pulmonary arterial hyperten... more Objective. Oxidative stress and endothelial dysfunction contribute to pulmonary arterial hypertension (PAH). The role of the nitrovasodilator pentaerythritol tetranitrate (PETN) on endothelial function and oxidative stress in PAH has not yet been defined. Methods and Results. PAH was induced by monocrotaline (MCT, i.v.) in Wistar rats. Low (30 mg/kg; MCT30), middle (40 mg/kg; MCT40), or high (60 mg/kg; MCT60) dose of MCT for 14, 28, and 42 d was used. MCT induced endothelial dysfunction, pulmonary vascular wall thickening, and fibrosis, as well as protein tyrosine nitration. Pulmonary arterial pressure and heart/body and lung/body weight ratio were increased in MCT40 rats (28 d) and reduced by oral PETN (10 mg/kg, 24 d) therapy. Oxidative stress in the vascular wall, in the heart, and in whole blood as well as vascular endothelin-1 signaling was increased in MCT40-treated rats and normalized by PETN therapy, likely by upregulation of heme oxygenase-1 (HO-1). PETN therapy improved en...
Reactive oxygen and nitrogen species (RONS such as H 2 O 2 , nitric oxide) confer redox regulatio... more Reactive oxygen and nitrogen species (RONS such as H 2 O 2 , nitric oxide) confer redox regulation of essential cellular functions (e.g. differentiation, proliferation, migration, apoptosis), initiate and catalyze adaptive stress responses. In contrast, excessive formation of RONS caused by impaired breakdown by cellular antioxidant systems and/or insufficient repair of the resulting oxidative damage of biomolecules may lead to appreciable impairment of cellular function and in the worst case to cell death, organ dysfunction and severe disease phenotypes of the entire organism. Therefore, the knowledge of the severity of oxidative stress and tissue specific localization is of great biological and clinical importance. However, at this level of investigation quantitative information may be enough. For the development of specific drugs, the cellular and subcellular localization of the sources of RONS or even the nature of the reactive species may be of great importance, and accordingly, more qualitative information is required. These two different philosophies currently compete with each other and their different needs (also reflected by different detection assays) often lead to controversial discussions within the redox research community. With the present review we want to shed some light on these different philosophies and needs (based on our personal views), but also to defend some of the traditional assays for the detection of RONS that work very well in our hands and to provide some guidelines how to use and interpret the results of these assays. We will also provide an overview on the "new assays" with a brief discussion on their strengths but also weaknesses and limitations.
Excessive inflammation in sepsis causes microvascular thrombosis and thrombocytopenia associated ... more Excessive inflammation in sepsis causes microvascular thrombosis and thrombocytopenia associated with organ dysfunction and high mortality. The present studies aimed to investigate whether dipeptidyl peptidase-4 (DPP-4) inhibition and supplementation with glucagon-like peptide-1 (GLP-1) analogues improves endotoxaemia-associated microvascular thrombosis via immunomodulatory effects. Endotoxaemia was induced in C57BL/6 J mice by single injection of lipopolysaccharide (LPS) (17.5 for survival and 10 mg/kg for all other studies). For survival studies therapy was started 6 h after LPS injection. For all other studies drugs were administrated 48 h prior to LPS treatment. Endotoxaemic control mice showed severe thrombocytopenia, microvascular thrombosis in the pulmonary circulation (fluorescence imaging), increased lactate dehydrogenase (LDH)-activity, endothelial dysfunction and increased markers of inflammation in the aorta and whole blood (e.g. leukocyte-dependent oxidative burst, nitr...
Nitroglycerin (GTN) and other organic nitrates are widely used vasodilators. Their side effects a... more Nitroglycerin (GTN) and other organic nitrates are widely used vasodilators. Their side effects are development of nitrate tolerance and endothelial dysfunction. Given the potential of GTN to induce nitro-oxidative stress, we investigated the interaction between nitro-oxidative DNA damage and vascular dysfunction in experimental nitrate tolerance. Cultured endothelial hybridoma cells (EA.hy 926) and Wistar rats were treated with GTN (ex vivo: 10-1000 µM; in vivo: 10, 20 and 50 mg/kg/day for 3 days, s.c.). The level of DNA strand breaks, 8-oxoguanine and O (6)-methylguanine DNA adducts was determined by Comet assay, dot blot and immunohistochemistry. Vascular function was determined by isometric tension recording. DNA adducts and strand breaks were induced by GTN in cells in vitro in a concentration-dependent manner. GTN in vivo administration leads to endothelial dysfunction, nitrate tolerance, aortic and cardiac oxidative stress, formation of DNA adducts, stabilization of p53 and a...
Cardiovascular diseases are major contributors to global deaths and disability-adjusted life year... more Cardiovascular diseases are major contributors to global deaths and disability-adjusted life years, with hypertension as a significant risk factor for all cause death. The endothelium that lines the inner wall of the vasculature regulates essential haemostatic functions such as regulation of the vascular tone, circulation of blood cells, inflammation and platelet activity. Endothelial dysfunction is an early predictor of atherosclerosis and future cardiovascular events. We review the prognostic value of measurements of endothelial function, the clinical techniques for its determination, mechanisms leading to endothelial dysfunction and the therapeutic treatment of endothelial dysfunction. Since vascular oxidative stress and inflammation are major determinants of endothelial function, we also address current antioxidant and anti-inflammatory therapies. In light of recent data that speak against the prognostic value of endothelial function in healthy human cohorts, we also discuss alt...
There has been some dispute regarding reaction products formed at physiological peroxynitrite flu... more There has been some dispute regarding reaction products formed at physiological peroxynitrite fluxes in the nanomolar range with phenolic molecules, when used to predict the behavior of protein-bound aromatic amino acids like tyrosine. Previous data showed that at nanomolar fluxes of peroxynitrite, nitration of these phenolic compounds was outcompeted by dimerization (e.g. biphenols or dityrosine). Using 3-morpholino sydnonimine (Sin-1), we created low fluxes of peroxynitrite in our reaction setup to demonstrate that salicylaldehyde displays unique features in the detection of physiological fluxes of peroxynitrite, yielding detectable nitration but only minor dimerization products. By means of HPLC analysis and detection at 380 nm we could identify the expected nitration products 3-and 5-nitrosalicylaldehyde, but also novel nitrated products. Using mass spectrometry, we also identified 2-nitrophenol and a not fully characterized nitrated dimerization product. The formation of 2-nitrophenol could proceed either by primary generation of a phenoxy radical, followed by addition of the NO 2-radical to the various resonance structures, or by addition of the peroxynitrite anion to the polarized carbonyl group with subsequent fragmentation of the adduct (as seen with carbon dioxide). Interestingly, we observed almost no 3-and 5-nitrosalicylic acid products and only minor dimerization reaction. Our results disagree with the previous general assumption that nitration of low molecular weight phenolic compounds is always outcompeted by dimerization at nanomolar peroxynitrite fluxes and highlight unique features of salicylaldehyde as a probe for physiological concentrations of peroxynitrite.
Cardiovascular diseases are associated with and/or caused by oxidative stress. This concept has b... more Cardiovascular diseases are associated with and/or caused by oxidative stress. This concept has been proven by using the approach of genetic deletion of reactive species producing (pro-oxidant) enzymes as well as by the overexpression of reactive species detoxifying (antioxidant) enzymes leading to a marked reduction of reactive oxygen and nitrogen species (RONS) and in parallel to an amelioration of the severity of diseases. Likewise, the development and progression of cardiovascular diseases is aggravated by overexpression of RONS producing enzymes as well as deletion of antioxidant RONS detoxifying enzymes. Thus, the consequences of the interaction (redox crosstalk) of superoxide/hydrogen peroxide produced by mitochondria with other ROS producing enzymes such as NADPH oxidases (Nox) are of outstanding importance and will be discussed including the consequences for endothelial nitric oxide synthase (eNOS) uncoupling as well as the redox regulation of the vascular function/tone in general (soluble guanylyl cyclase, endothelin-1, prostanoid synthesis). Pathways and potential mechanisms leading to this crosstalk will be analysed in detail and highlighted by selected examples from the current literature including hypoxia, angiotensin 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" will be discussed providing evidence that mitochondria represent key players and amplifiers of the burden of oxidative stress.
Introduction: Sepsis is an inflammatory state. Proinflammatory enzymes like inducible-NO Synthase... more Introduction: Sepsis is an inflammatory state. Proinflammatory enzymes like inducible-NO Synthase (iNOS) and NADPH-Oxidase (NOX) are activated and influence endothelial function via oxidative and nitrosative stress. Inhibition of dipeptidyl-peptidase 4 (DPP4i) is a new approach to treat diabetic patients. Matubara et al. (Circulation, 2012) showed a positive influence of DPP4i on endothelial function in an animal model Hypothesis: DPP4 inhibition influences endothelial function, hemostasis and survival in a rodent model of septic shock Methods: C57/BL6j mice were pretreated with different DPP4i (linagliptin, sitagliptin) and GLP-1-analogue (GLP1a) liraglutide for 3 days. Aorta and heart tissue was used for western-blotting and rtPCR. Blood was examined for coagulation parameters (aPTT, PT), cell count and oxidative stress Results: Treatment with DPP4i and GLP1-a improved survival and blood pressure of septic animals significantly. LPS-induced oxidative stress and nitrosylated hemoglobin (Hb-NO) levels wer...
International journal of molecular sciences, Jan 5, 2015
Cardiovascular disease is a leading cause of death and reduced quality of life worldwide. Arteria... more Cardiovascular disease is a leading cause of death and reduced quality of life worldwide. Arterial vessels are a primary target for endothelial dysfunction and atherosclerosis, which is accompanied or even driven by increased oxidative stress. Recent research in this field identified different sources of reactive oxygen and nitrogen species contributing to the pathogenesis of endothelial dysfunction. According to lessons from the past, improvement of endothelial function and prevention of cardiovascular disease by systemic, unspecific, oral antioxidant therapy are obviously too simplistic an approach. Source- and cell organelle-specific antioxidants as well as activators of intrinsic antioxidant defense systems might be more promising. Since basic research demonstrated the contribution of different inflammatory cells to vascular oxidative stress and clinical trials identified chronic inflammatory disorders as risk factors for cardiovascular events, atherosclerosis and cardiovascular...
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Papers by Andreas Daiber