The contribution of reactive oxygen species and p38 mitogen-activated protein kinase to myofilament oxidation and progression of heart failure in rabbits

Background and purpose:

The formation of reactive oxygen species (ROS) is increased in heart failure (HF). However, the causal and mechanistic relationship of ROS formation with contractile dysfunction is not clear in detail. Therefore, ROS formation, myofibrillar protein oxidation and p38 MAP kinase activation were related to contractile function in failing rabbit hearts.

Experimental approach and key results:

Three weeks of rapid left ventricular (LV) pacing reduced LV shortening fraction (SF, echocardiography) from 32 ± 1% to 13 ± 1%. ROS formation, as assessed by dihydroethidine staining, increased by 36 ± 8% and was associated with increased tropomyosin oxidation, as reflected by dimer formation (dimer to monomer ratio increased 2.28 ± 0.66-fold in HF vs. sham, P < 0.05). Apoptosis (TdT-mediated dUTP nick end labelling staining) increased more than 12-fold after 3 weeks of pacing when a significant increase in the phosphorylation of p38 MAP kinase and HSP27 was detected (Western blotting). Vitamins C and E abolished the increases in ROS formation and tropomyosin oxidation along with an improvement of LVSF (19 ± 1%, P < 0.05 vs. untreated HF) and prevention of apoptosis, but without modifying p38 MAP kinase activation. Inhibition of p38 MAP kinase by SB281832 counteracted ROS formation, tropomyosin oxidation and contractile failure, without affecting apoptosis.

Conclusions and implications:

Thus, p38 MAP kinase activation appears to be upstream rather than downstream of ROS, which impacts on LV function through myofibrillar oxidation. p38 MAP kinase inhibition is a potential target to prevent or treat HF.     

"One-stop-shop" ultrasound diagnosis of functional, structural and physicomechanical properties of the brachial artery

BACKGROUND: The pathogenesis of atherosclerosis comprises endothelial dysfunction, thickening as well as impaired compliance of the arterial vessel wall. Early assessment of these alterations of the vessel wall at the same site of the vascular tree has yet been hampered by the lack of highly sensitive diagnostic approaches suitable for clinical routine. We therefore aimed to develop and validate a single non-invasive examination of the brachial artery for simultaneous and highly accurate measurement of functional, structural and physicomechanical parameters of the brachial artery. PATIENTS AND METHODS: 20 healthy individuals were investigated using high resolution ultrasound. Flow-mediated dilation (FMD), fractional diameter changes (FDC) and intima-media-thickness (IMT) were measured in the same segment of the brachial artery. Coefficients of variation, day-to-day-variability, between- and within-observer-variability were investigated in 5 individuals. All measurements were performed manually and by an automated PC-based analyzing system. RESULTS: Mean values for all measured parameters were 7.65 +/- 0.8% for FMD, 0.02 +/- 0.002 for FDC, 0.351 +/- 0.007 mm for IMT and followed an even distribution throughout the study population. Automated analysis of coefficient of variation, day-to-day-, between- and within-observer variabilities were: 0. 78%, 1.3%, 0.8%, 0.8% (FMD); 4.7%, 2.8%, 4.2%, 2.7% (FDC); 1.8%, 1.1%, 1.9%, 1.1% (IMT). Coefficient of variation, day-to-day-, between- and within-observer variabilities for the manual readings were significantly higher. CONCLUSIONS: Functional, structural and physicomechanical parameters of the brachial artery can be quantified consecutively, time-saving and highly reproducibly as an "one-stop-shop" in a single session using high resolution ultrasound with digitized post-processing. This highlights the future possibility of early, sensitive and non-invasive diagnostic testing of vascular function in patients prone to vascular disease.     

The angiotensin–calcineurin–NFAT pathway mediates stretch-induced up-regulation of matrix metalloproteinases-2/-9 in atrial myocytes

Aim  During atrial fibrillation, arterial hypertension and systolic or diastolic heart failure, atrial myocytes are exposed to increased baseline stretch. Atrial stretch has been shown to induce cellular hypertrophy and extracellular matrix remodeling (ECM) via angiotensin-II dependent pathways and the matrix metalloproteinases system (MMPs). We hypothesized that atrial myocytes exposed to static stretch may increase their ECM remodeling activity via up-regulation of MMP-2/-9. We then tested the hypothesis that the membrane bound angiotensin-II type 1 (AT1) receptor and the intracellular calcineurin (Cn)-NFAT signaling pathway are potential mediators of stretch-induced MMP alterations, since Cn-NFAT is one important contributor to myocyte hypertrophy. Methods and results  Neonatal rat atrial myocytes (NRAM) were cultured under conditions of static stretch by 21%. The differential effects of selective AT1 receptor blockade by losartan, Cn blockade by Cyclosporine-A (CsA) or NFAT inhibition by 11R-VIVIT (VIV), were analyzed. Stretch resulted in a significant up-regulation of active-MMP-2/-9 protein amount (active-MMP-2 ng/μg: control 8.95 ± 0.64 vs. stretch 13.11 ± 0.74 / active-MMP-9 ng/μg: control 1.45 ± 0.18 vs. stretch 1.94 ± 0.21, all n = 5) and enzyme activity (MMP-2 in %: control 1 ± 0.0 vs. stretch 1.87 ± 0.25, n = 7) associated with a significant increase of the membrane-type-1-MMP (MT1-MMP) protein expression (MT1-MMP in %: control 1 ± 0.0 vs. stretch 2.17 ± 0.21, n = 8). These observations were accompanied by an activation of the Cn-NFAT pathway (Cn-activity in nmol PO₄ release/20 μg protein/30 min: control 0.37 ± 0.08 vs. stretch 0.65 ± 0.09, n = 3 / NFATc1-DNA binding activity in %: control 1 ± 0.0 vs. stretch 1.53 ± 0.17, n = 3). Losartan, CsA or VIV abolished stretch-induced alterations in MMP-2/-9 and MT1-MMP expression and enzyme activity by normalizing the Cn-activity and the DNA binding activity of NFATc1. Conclusion  Our results present new insights in molecular mechanisms of ECM remodeling activity of atrial myocytes exposed to static stretch. The AT1-Cn-NFAT pathway is a potential mediator of MMP activation. Returned for 1. Revision: 15 August 2008 1. Revision received: 17 November 2008 E. Saygili and O. R. Rana contributed equally to this work.     

Epicardial adipose tissue is a robust measure of increased risk of myocardial infarction – a meta-analysis on over 6600 patients and rationale for the EPIC-ACS study

Background:Epicardial adipose tissue (EAT) surrounds the heart and the coronary vessels. EAT produces pro- and anti-inflammatory cytokines. Several studies have already documented the association of EAT and cardiovascular risk factors as well as coronary artery disease manifestations. Currently computed tomography (CT) is considered the gold standard for measurement of 3-dimensional volume of EAT. In addition, echocardiography might be an easy accessible alternative in particular in an emergency setting.Methods:We performed a metaanalysis of existing studies describing the differences of EAT in patients with and without myocardial infarction. We used established databases and were searching for “epicardial adipose tissue” or “pericardial adipose tissue” and “myocardial infarction”, “coronary events”, or “acute coronary syndrome”. We included over 6600 patients from 7 studies. Random effect models were calculated and all analyses were performed by using the Review Manager 5.3.Results:Patients with myocardial infarction had 37% (confidence interval [0.21-0.54], P value <.001)] higher measures of EAT compared to patients without myocardial infarction. Comparing studies using echocardiography vs CT for assessment of EAT thickness, similar relative differences in EAT with wide overlap of confidence intervals were observed (for echocardiography: 0.4 [0.04-0.76], for CT: 0.36 [0.16-0.57], P value <.001 for both).Conclusions:Patients with myocardial infarction have more EAT as compared to patients without myocardial infarction independently of the used imaging modality. Further prospective studies are needed to evaluate, how quantification of EAT in clinical routine can improve patients management.     

Circulating NO pool in humans

Nitric oxide (NO) generated from L-arginine by NO synthases in the endothelium and in other cells plays a central role in several aspects of vascular biology and has been linked to many regulatory functions in mammalian cells. Whereas for a long time the signaling actions of NO in the vasculature have been thought to be short-lived as a result of the rapid reaction of NO with hemoglobin, recent studies changed the biochemical thinking of NO. NO is not anymore the paracrine agent with only local effects, but, like a hormone, it disseminates throughout the body. Thus, a circulating pool of NO exists, opening new considerable pharmacological and therapeutical avenues in the diagnosis and therapy of cardiovascular diseases. In this review we briefly discuss the major routes of NO metabolism and transport in the mammalian circulation, considering plasma, red blood cell and tissue compartments separately, with a special focus on the implication of the circulating NO pool in clinical research.     

Cellular targets and mechanisms of nitros(yl)ation: an insight into their nature and kinetics in vivo

There is mounting evidence that the established paradigm of nitric oxide (NO) biochemistry, from formation through NO synthases, over interaction with soluble guanylyl cyclase, to eventual disposal as nitrite/nitrate, represents only part of a richer chemistry through which NO elicits biological signaling. Additional pathways have been suggested that include interaction of NO-derived metabolites with thiols and metals to form S-nitrosothiols (RSNOs) and metal nitrosyls. Despite the overwhelming attention paid in this regard to RSNOs, little is known about the stability of these species, their significance outside the circulation, and whether other nitros(yl)ation products are of equal importance. We here show that N-nitrosation and heme-nitrosylation are indeed as ubiquitous as S-nitrosation in vivo and that the products of these reactions are constitutively present throughout the organ system. Our study further reveals that all NO-derived products are highly dynamic, have fairly short lifetimes, and are linked to tissue oxygenation and redox state. Experimental evidence further suggests that nitroso formation occurs substantially by means of oxidative nitrosylation rather than NO autoxidation, explaining why S-nitrosation can compete effectively with nitrosylation. Moreover, tissue nitrite can serve as a significant extravascular pool of NO during brief periods of hypoxia, and tissue nitrate/nitrite ratios can serve as indicators of the balance between local oxidative and nitrosative stress. These findings vastly expand our understanding of the fate of NO in vivo and provide a framework for further exploration of the significance of nitrosative events in redox sensing and signaling. The findings also raise the intriguing possibility that N-nitrosation is directly involved in the modulation of protein function.     

L-arginine preferentially dilates stenotic segments of coronary arteries thereby increasing coronary flow

Background and objectives. Oxidized LDL cholesterol and cytokines increase arginase and decrease nitric oxide (NO) synthase expression in human endothelial cells, leading to a decrease in NO production. In arteriosclerotic plaques, characterized by increased oxidized LDL and cytokine levels, a sustained local NO reduction might enhance sensitivity of the downstream guanylyl cyclase system towards an acute NO increase. We tested whether application of the NO synthase substrate l ‐arginine (l ‐arg, 150 μmol min^?1) or the NO donor isosorbide dinitrate (ISDN; 0.3 mg) preferentially dilates stenotic coronary artery segments (CS) subsequently increasing poststenotic coronary blood flow (CBF) in patients with coronary artery disease (CAD). Design, setting and subjects. Changes in coronary diameter and circumferential surface area were assessed by quantitative coronary angiography (QCA) in a nonstenotic upstream segment, the CS, downstream the CS and in a reference vessel (n = 24). CBF was estimated in a subset of 13 patients by QCA and intracoronary Doppler. Results. CS ranged from 62% to 89% (77 ± 5%). l ‐arg increased minimal luminal diameter of the stenotic segment from 0.98 ± 0.06 to 1.14 ± 0.07 mm (P < 0.05) without affecting other coronary segments. Poststenotic CBF increased by 24 ± 3%. ISDN dilated all segments again with a predominance of CS (25 ± 4%) and increased poststenotic CBF by 38 ± 9%. In a multifactorial anova , a medication with an angiotensin‐converting enzyme inhibitor (decreasing inflammation and radical formation) and a ratio of LDL/HDL <3.5 were predictive for an l ‐arg‐induced dilation. Conclusion. The increase in poststenotic CBF without affecting nondiseased arteries highlights the therapeutic potential of l ‐arg in patients with CAD.     

Myoglobin’s novel role in nitrite-induced hypoxic vasodilation

Hypoxic vasodilation represents a key physiological response of the cardiovascular system to low tissue oxygen tension, adjusting local blood flow to meet the metabolic requirements in tissue. Vasodilation occurs by nitric oxide (NO) activation of the cyclic guanosine monophosphate (cGMP) signaling pathway in vascular smooth muscle cells. Under normoxia, NO is formed by the well-known endothelial NO synthase (eNOS) system while under hypoxia NO is generated from nitrite. We have unraveled the heme-protein myoglobin in vascular smooth muscle cells as a major source of NO generation by reduction of endogenous nitrite under hypoxia. This mediates hypoxic vasodilation under physiological conditions without direct involvement of eNOS and independently of effects on cardiac function.     

Kommentar zu den Leitlinien der Europäischen Gesellschaft für Kardiologie (ESC) zur Therapie des akuten Herzinfarkts bei Patienten mit ST-Streckenhebung (STEMI)

In August 2012 amended guidelines of the European Society of Cardiology on the acute therapy patients with persisting ST segment elevation myocardial infarction (STEMI) were published to replace the guidelines from the year 2008. The major changes affect patients with atypical electrocardiogram (ECG) alterations with signs and symptoms of persistent ischemia who need rapid treatment. The treatment of patients with cardiac arrest has for the first time been given a chapter on its own with emphasis on the necessity for immediate invasive diagnostics. The chapter on logistics emphasizes the necessity for regional myocardial infarction networks in cooperation with general physicians, emergency services and hospitals with and without interventional possibilities for rapid reperfusion therapy. Primary percutaneous coronary intervention (PCI) remains the preferred reperfusion therapy and should be carried out within the first 90 min following first medical contact and by short symptom duration even within 60 min. For antithrombotic therapy the new platelet aggregation inhibitors prasugrel and ticagrelor as well as bivalirudin for acute anticoagulation are emphasized. Detailed information is given on secondary prevention with an emphasis on physical activity and lifestyle modifications.