M. Lisi

Assessment of vascular function: flow-mediated constriction complements the information of flow-mediated dilatation

Objective: To determine whether vascular function assessed by low-flow-mediated constriction (L-FMC), a novel non-invasive method, complements the information obtained with “traditional” flow-mediated dilatation (FMD). Design and patients: In protocol 1, 12 healthy young volunteers underwent FMD and L-FMC measurements at rest and immediately after isometric exercise of the same hand. In protocol 2, 24 patients with coronary artery disease, 24 with congestive heart failure, 24 hypertensive patients and 64 healthy volunteers were enrolled to undergo L-FMC and FMD measurements. Results: In protocol 1, exercise was associated with mean (SD) increases in radial artery blood flow, diameter and L-FMC (from −5.1 (1.5)% to −7.8 (3.4)%, p<0.05), while FMD was significantly blunted (from 6.0 (2.4)% to 3.0 (3.2)%, p<0.05). In protocol 2, both FMD and L-FMC were blunted in the patient groups. Receiver operating curve analysis showed that, as compared with FMD alone, the combination of L-FMC and FMD significantly improved the sensitivity and specificity in detecting patients diagnosed with cardiovascular disease (p<0.05). Conclusion: In the first protocol, FMD and L-FMC were shown to be reciprocally regulated. A blunted FMD may, in certain cases, be an expression of increased resting vascular activation and not only of impaired endothelial function. In the second protocol, a statistical approach showed that implementation of L-FMC provides a better characterisation than FMD of vascular function in cardiovascular disease. Vascular (endothelial) function is a complex phenomenon which requires a multifaceted approach; it is suggested that a combination of L-FMC and FMD will provide additive and complementary information to “traditional” FMD measurements.

Pentaerythrityl Tetranitrate and Nitroglycerin, but not Isosorbide Mononitrate, Prevent Endothelial Dysfunction Induced by Ischemia and Reperfusion

Background—Short term exposure to nitroglycerin (GTN) has protective properties that are similar to ischemic preconditioning. Whether other organic nitrates such as pentaerithrityl tetranitrate (PETN) and isosorbide mononitrate (ISMN) have similar protective effects has not been explored. Methods and Results—In a randomized, parallel, double blind, controlled trial, 37 healthy young volunteers received no therapy (n=10), transdermal GTN 1.2 mg for 2 hours (n=9), PETN 80 mg (n=9), or ISMN 40 mg (n=9). Twenty-four hours later, endothelium-dependent flow-mediated vasodilation (FMD) was measured before and after local exposure to ischemia and reperfusion (IR). In the no therapy group, IR blunted FMD (FMD after IR: 1.9±0.6%, P<0.05), an effect that was prevented by GTN (FMD after IR: 5.3±1.4%, P<0.05 compared with no therapy). PETN had the same protective effect (FMD after IR: 8.1±1.3%, P<0.05 compared with no therapy), whereas ISMN had no significant pharmacological preconditioning effect (FMD after-IR: 3.6±0.8%, P=ns compared with no therapy). While it blocked the effect of GTN, Vitamin C (n=8) did not modify PETN preconditioning (FMD after IR: 6.3±0.9%, P=ns compared with before IR), showing that this phenomenon is not mediated by oxygen free radical production. In an effort to identify the mechanism of PETN preconditioning, isolated human endothelial cells were incubated with PETN, GTN, or ISMN. Only PETN induced expression of the genes encoding for heme oxygenase and ferritin, which have been involved in ischemic and pharmacological preconditioning. Conclusions—We show important differences among organic nitrates in their capacity to prevent IR-induced endothelial dysfunction. GTN and PETN, but not ISMN, have this preconditioning effect. The potential clinical implications of these data warrant further investigation.

Observations of time-based measures of flow-mediated dilation of forearm conduit arteries: implications for the accurate assessment of endothelial function.

Endothelium-dependent flow-mediated dilation (FMD) is measured as the increase in diameter of a conduit artery in response to reactive hyperemia, assessed either at a fixed time point [usually 60-s post-cuff deflation (FMD(60))] or as the maximal dilation during a 5-min continuous, ECG-gated, measurement (FMD(max-cont)). Preliminary evidence suggests that the time between reactive hyperemia and peak dilation (time to FMD(max)) may provide an additional index of endothelial health. We measured FMD(max-cont), FMD(60), and time to FMD(max) in 30 young healthy volunteers, 22 healthy middle-aged adults, 16 smokers, 23 patients with hypertension, 40 patients with coronary artery disease, and 22 patients with heart failure. As previously reported, FMD(max-cont) was similar in healthy cohorts and was significantly blunted in smokers and all patient groups, whereas FMD(60) was significantly blunted only in heart failure patients. There was a wide within-group variability between measures of time to FMD(max) with no significant difference between normal and patient groups. Intra-arterial infusion of the nitric oxide synthase inhibitor N(omega)-monomethyl-l-arginine in eight healthy subjects resulted in a blunting of FMD(max-cont) (P < 0.001) and FMD(60) (P = 0.02) but not time to FMD(max). Both FMD(max-cont) and FMD(60) demonstrated good repeatability in 30 young healthy volunteers studied on two separate occasions (P < 0.01 for both), whereas time to FMD(max) varied widely between visits (P = not significant). In conclusion, although time to FMD(max) does not appear to be a useful adjunctive measure of endothelial health, the use of continuous diameter measurements provides important data in the study of endothelial function in healthy subjects and patients with cardiovascular disease.

Nitroglycerine causes mitochondrial reactive oxygen species production: In vitro mechanistic insights

BACKGROUND Nitroglycerine (GTN) is an organic nitrate that has been used for more than 100 years. Despite its widespread clinical use, several aspects of the pharmacology of GTN remain elusive. In a recent study, the authors of the present study showed that GTN causes opening of the mitochondrial permeability transition pore (mPTP) and mitochondrial production of reactive oxygen species (ROS). OBJECTIVE In the present study, it was tested whether GTN-induced ROS production depends on mitochondrial potassium ATP-dependent channel or mPTP opening, and/or GTN biotransformation. METHODS AND RESULTS Isolated rat heart mitochondria were incubated with succinate (a substrate for complex II) and GTN, causing immediate ROS production, as manifested by chemiluminescence. This ROS production was prevented by concomitant vitamin C incubation. Conversely, inhibitors of potassium ATP-dependent channels, mPTP opening or of GTN biotransformation did not modify ROS production. CONCLUSIONS GTN triggers mitochondrial ROS production independently of the opening of mitochondrial channels and/or of GTN biotransformation. The present data, coupled with previous evidence published by the same authors that GTN causes opening of mPTPs, provide further evidence on the pharmacology of GTN. It is proposed that GTN causes direct uncoupling of the respiratory chain, which determines ROS production and subsequent mPTP opening. The clinical implications of these findings are also discussed.

The mechanism of nitrate-induced preconditioning.

Nitroglycerin (GTN) has been shown, in both human and animal studies, to induce a protective phenotype that limits tissue damage after ischemia and reperfusion. This phenomenon is similar to ischemic preconditioning, and several reports suggest that also the molecular pathways involved in this protective effect of nitrates are the same that determine ischemic preconditioning. Our group conducted a series of studies aimed at investigating, using a human model of endothelial IR injury, the mechanism of nitrate-induced preconditioning and particularly the role of reactive oxygen species formation and of the opening of mitochondrial permeability transition pores. Our data demonstrate that GTN protects the endothelium against postischemic endothelial dysfunction in a mechanism that is mediated by oxygen free radical release and opening of mitochondrial permeability transition pores. In contrast, the protective effect of pentaerithrityl tetranitrate appears to be independent of these mechanisms, and it seems to be mediated by induction of antioxidant genes. Finally, isosorbide mononitrate seems to be devoid of a significant protective effect. These data are summarized and discussed in the present paper.

Model-based analysis of flow-mediated dilation and intima-media thickness

We present an end-to-end system for the automatic measurement of flow-mediated dilation (FMD) and intima-media thickness (IMT) for the assessment of the arterial function. The video sequences are acquired from a B-mode echographic scanner. A spline model (deformable template) is fitted to the data to detect the artery boundaries and track them all along the video sequence. The a priori knowledge about the image features and its content is exploited. Preprocessing is performed to improve both the visual quality of video frames for visual inspection and the performance of the segmentation algorithm without affecting the accuracy of the measurements. The system allows real-time processing as well as a high level of interactivity with the user. This is obtained by a graphical user interface (GUI) enabling the cardiologist to supervise the whole process and to eventually reset the contour extraction at any point in time. The system was validated and the accuracy, reproducibility, and repeatability of the measurements were assessed with extensive in vivo experiments. Jointly with the user friendliness, low cost, and robustness, this makes the system suitable for both research and daily clinical use.

Acute (but not chronic) smoking paradoxically protects the endothelium from ischemia and reperfusion: insight into the “smoking paradox”

The negative impact of cigarette smoking on the cardiovascular system has been recognized for more than 50 years. Smoking is associated with increased heart rate and sympathetic outflow, inflammation, with accelerated progression of atherosclerosis, enhanced platelet aggregation, decreased fibrinolytic activity, endothelial dysfunction, worsening of renal function [1], and ultimately with increased hospitalizations for myocardial infarction, stroke and heart failure [2, 3]. Importantly, many of these effects are felt to be mediated, at least in part, by increased exposure to reactive oxygen species. This concept is supported by increased plasma markers of oxidative stress and by the observation that administration of antioxidants such as vitamin C reverses the vascular abnormalities associated with smoking [4]. The irrefutable evidence of higher ischemic rates in smokers stands in contrast with several large epidemiological studies demonstrating that, among patients who experience a myocardial infarction, those who smoke appear to have lower mortality and morbidity [5–7]. Some authors have argued that this observation is due to differences between the populations studied [6]; other studies found that in patients with infarction, smoking remains associated with reduced mortality even after controlling for age, infarct size, clinical presentation, blood pressure, cardiovascular risk factors and several other parameters [7, 8]. The complex mechanisms triggered by reactive oxygen species might contribute to explain this smoker’s paradox. While having a crucial role in the pathophysiology and progression of cardiovascular disease, reactive oxygen and nitrogen species are also involved in ischemic preconditioning, a protective phenomenon associated with a reduced susceptibility to ischemia and reperfusion damage [9]. We set out to investigate the impact of smokinginduced oxidative stress on the vascular-endothelial dysfunction induced by ischemia and reperfusion. Radial artery endothelium-dependent flow-mediated dilation (FMD) was measured before and after local ischemia/reperfusion (15 min ischemia by inflating a pneumatic cuff around the upper arm followed by 15 min reperfusion) in 10 healthy subjects (7 males, 25–28 years old) with no cardiac risk factors. These procedures were carried out twice, at 7-day intervals. Twenty-four hours before each visit, subjects underwent, under supervision and in randomized order, two protocols: in the first protocol, they smoked two cigarettes (nicotine content 1.2 mg, tar 16 mg, CO 14 mg) 15 min after receiving 2 g of intravenous vitamin C. In the second protocol, they smoked two cigarettes 15 min after receiving an intravenous placebo (10 ml of normal saline). Subjects were occasional smokers (0–2 cigarettes/month), and none of them had smoked in the previous 7 days. A separate group of 10 smokers (7 males, age 24–28 years, 3–10 cigarettes/day) underwent the same FMD-IR-FMD protocol after a 24-h non-smoking interval. The methods for the assessment of FMD in our laboratory have been previously described [9]. Briefly, the diameter of the radial artery is measured in M. Lisi S. Dragoni M. C. Leone Department of Internal, Cardiovascular and Geriatric Medicine, University of Siena, Siena, Italy

The Tako-Tsubo syndrome: No evidence of peripheral endothelial or microvascular dysfunction

The Tako-Tsubo syndrome or left ventricular apical ballooning syndrome is a recently described condition characterized by precordial pain, ST-segment elevation and T wave abnormalities in the left precordial leads of the ECG as well as akinesia of all apical segments of the left ventricle in the absence of coronary artery disease at angiography [1]. While its pathogenesis remains incompletely understood, the Tako-Tsubo syndrome has been proposed to be determined by catecholamine-induced endothelial dysfunction and subsequent impaired microvascular reactivity at the level of the mid and apical segments of the left ventricle [2]. We report a case of a 73-years-old woman with history of hypertension who was admitted to the Emergency Department for oppressive chest pain radiated to the right arm and the back. Upon admission, the patient showed high blood pressure levels (160/110 mmHg); her ECG showed sinus rhythm and Twave inversion in leads II, III, aVF, V5 and V6. Troponin T was 0.39 ng/ml (reference range, b0.06 ng/ml) and increased to 0.61 ng/ml 6 h later, when a minor elevation of CPK and CPK-MB was also observed. Therapy with lowmolecular weight heparin and double antiplatelet agents (aspirin 100 mg+clopidogrel 50 mg) was started. The day after, the patient was asymptomatic but ECG showed a 1– 2 mm STsegment elevation in leads V2, V3 and deep Twave inversion in leads I, II, III, AVF and V4–V6. Echocardiogram

Mitral regurgitation severity correlates with symptoms and extent of left atrial dysfunction : effect of mitral valve repair

Recent trends in diagnostic work-up among unselected patients newly diagnosed with heart failure : a Swedish population-based studyMitral regurgitation severity correlates with symptoms and extent of left atrial dysfunction : effect of mitral valve repairHeart failure can occur in any age, no depend on sex, but in men and women the mechanism, even if is the same, the fact is that the compromise on pumping function is diferent. AIM We realized a follow-up with 100 female patients during hospitalization with heart failure as a mean diagnostic. These are patient between 60 and 75 years old, with different pathologies: diabetes mellitus, arterial hypertension, obesity, atrial fibrillation, and hypothyroidism. We observed their treatment comparing with a control group (100 men in heart failure) by administering vasodilator and diuretic drugs. Performed echocadiography doppler control, daily renal function, NT pro BNP levels control, oxide nitric response. Results: We observed that ventricular dilation, hypertrophy as tachycardia is more typical in men. Our group demonstrated very fast response to beta blockers and diuretics. The ejection fraction increased in 10-15% faster than in control group. Oxide nitric had not the result we expected. But in men the effect is very high. NT pro BNP levels no were increased as a control group. Recovering renal function in women during heart failure depends on risk factors as diabetes mellitus, obesity, more characteristics for women. Conclusions: In women heart failure has the same mechanism that in men, but more of the cardiac compensatory mechanisms during heart failure as Frank-Starling mechanism, ventricular dilation or hypertrophy and tachycardia present more complications in men; women recover sinus rhythm faster than men, hypertrophy is not characteristic and dilation recovers EF as pumping function is near normal. We do not observed increased sympathetic adrenergic activity in our patients and increased vagal activity to heart. Renin-angiotensin-aldosterone and antidiuretic hormone systems in women is compensated by vasoconstriction improving ventricular stroke volume by reducing afterload on the ventricle. Table 3. NT-pro BN characteristics NT-proBNP cutoff value of 125 pg/mL had the best sensitivity-to-specificity ratio and NPV to rule out asymptomatic LV moderate to severe diastolic or systolic dysfunction in patients at risk for heart failure: 1. Men younger than 60 years (sensitivity, 87.5%; specificity, 92.7%; NPV, 99.5%; positive predictive value [PPV], 33.3%) 2. Women younger than 60 years (sensitivity, 100%; specificity, 84.1%; NPV, 100%; PPV, 33.3%) 3. Men at least age 60 years (sensitivity, 100%; specificity, 77.1%; NPV, 100%; PPV, 32.5%) 4. Women at least age 60 years (sensitivity, 100%; specificity, 69.9%; NPV, 100%; PPV, 21%)