Rationale: Ceruloplasmin antioxidant function is mainly related to its ferroxidase I (FeOxI) activity, which influences iron-dependent oxidative and nitrosative radical species generation. Peroxynitrite, whose production is increased in heart failure (HF), can affect ceruloplasmin antioxidant function through amino acid modification. Objective: We investigated the relationship between FeOxI and ceruloplasmin tyrosine and cysteine modification and explored in a cohort of patients with HF the potential clinical relevance of serum FeOxI. Methods and Results: In patients with chronic HF (n=96, 76±9 years; New York Heart Association class, 2.9±0.8) and age-matched controls (n=35), serum FeOxI, FeOxII, ceruloplasmin, nitrotyrosine-bound ceruloplasmin, B-type natriuretic peptide, norepinephrine, and high-sensitivity C-reactive protein were measured, and the patients were followed up for 24 months. Ceruloplasmin, B-type natriuretic peptide, norepinephrine, and high-sensitivity C-reactive protein were increased in HF versus controls. FeOxI was decreased in HF (−20%) and inversely related to nitrotyrosine-bound ceruloplasmin (r, −0.305; P=0.003). In HF, FeOxI lower tertile had a mortality rate doubled compared with middle-higher tertiles. FeOxI emerged as a mortality predictor (hazard ratio, 2.95; 95% confidence intervals [1.29–6.75]; P=0.011) after adjustment for age, sex, hypertension, smoking, sodium level, estimated glomerular filtration rate, and high-sensitivity C-reactive protein. In experimental settings, peroxynitrite incubation of serum samples and isolated purified ceruloplasmin reduced FeOxI activity while increasing ceruloplasmin tyrosine nitration and cysteine thiol oxidation. Reduced glutathione prevented peroxynitrite-induced FeOxI drop, tyrosine nitration, and cysteine oxidation; flavonoid(−)-epicatechin, which prevented ceruloplasmin tyrosine nitration but not cysteine oxidation, partially impeded peroxynitrite-induced FeOxI drop. Conclusions: Reduced activity of serum FeOxI is associated with ceruloplasmin nitration and reduced survival in patients with HF. Both ceruloplasmin tyrosine nitration and cysteine thiol oxidation may be operant in vivo in peroxynitrite-induced FeOxI activity inhibition.

BackgroundAn accurate prognosis prediction represents a key element in chronic heart failure (CHF) management. Seattle Heart Failure Model (SHFM) prognostic power, a validated risk score for predicting mortality in CHF, is improved by adding B-type natriuretic peptide (BNP). We evaluated in a prospective study the incremental value of several biomarkers, linked to different biological domains, on death risk prediction of BNP-added SHFM.MethodsTroponin I (cTnI), norepinephrine, plasma renin activity, aldosterone, high sensitivity-C reactive protein (hs-CRP), tumor necrosis factor-α (TNF-α), interleukin 6 (IL-6), interleukin 2 soluble receptor, leptin, prealbumin, free malondialdehyde, and 15-F2t-isoprostane were measured in plasma from 142 consecutive ambulatory, non-diabetic stable CHF (mean NYHA-class 2.6) patients (mean age 75±8years). Calibration, discrimination, and risk reclassification of BNP-added SHFM were evaluated after individual biomarker addition.ResultsIndividual addition of biomarkers to BNP-added SHFM did not improve death prediction, except for prealbumin (HR 0.49 CI: (0.31–0.76) p=0.002) and cTnI (HR 2.03 CI: (1.20–3.45) p=0.009). In fact, with respect to BNP-added SHFM (Harrell's C-statistic 0.702), prealbumin emerged as a stronger predictor of death showing the highest improvement in model discrimination (+0.021, p=0.033) and only a trend was observed for cTn I (+0.023, p=0.063). These biomarkers showed also the best reclassification statistic (Integrated Discrimination Improvement—IDI) at 1-year (IDI: cTnI, p=0.002; prealbumin, p=0.020), 2-years (IDI: cTnI, p=0.018; prealbumin: p=0.006) and 3-years of follow-up (IDI: cTnI p=0.024; prealbumin: p=0.012).ConclusionsIndividual addition of prealbumin allows a more accurate prediction of mortality of BNP enriched SHFM in ambulatory elderly CHF suggesting its potential use in identifying those at high-risk that need nutritional surveillance.

Objective: To detect the sexual differentiation associated with oxidative stress, hypertension and insulin resistance on an accelerated experimental model of type II diabetes in rat. Methods and Results: Male and female Sprague-Dawley rats (225–250 g) chronically fed 10% of glucose were treated orally either with L-NAME (50 mg/kg/day) alone or with a combination of aspirin (100 mg/kg/day) for 4 weeks. L-NAME treatment in glucose-fed rats progressively increased the systolic blood pressure (with a telemetry device) by 59.3 mmHg (55.9%) in male and by 43.1 mmHg (40.2%) in female compare to control levels, enhanced the aortic and cardiac tissue superoxide production (with the lucigenin-enhanced chemiluminescence method) by 38.2% and 21.9% in male, 26.7% and 10.5% in female, and rose HOMA index by 4.6 times in male and by 2.5 time in female after 4 weeks treatment. The above differences between the male and the females were significant (P < 0.05). Simultaneous oral ASA treatment significantly attenuated the L-NAME plus glucose-induced increases in blood pressure by -34.4 mmHg (-20.8%) in male and by -17.1 mmHg (-11.5%) in female, in the aortic and the cardiac superoxide production (-24.9% and -16.9 % in male, -9.7 % and -8.1 % in female), in HOMA index by -66.0% in male and by -41.9% in female. Conclusions: Male rats are more susceptible to glucose-L-NAME-induced oxidative stress, hypertension, and insulin resistance than females. In addition, the male rats are also more sensitive than female rats to the antioxidant and preventive effects of aspirin against hypertension and diabetes.

Objective. To evaluate the capacity of chronic ASA therapy to prevent cardiac alterations and increased oxidative stress in cardiomyopathic hamsters. Methods and Results. Male Syrian cardiomyopathic and age-matched inbred control hamsters received ASA orally from the age of 60 days. Animals were sacrificed at the age of 150, 250, and 350 days to evaluate the time course of cardiac hypertrophy and cardiovascular tissue superoxide anion (O2-) production. At the age of 150 days, the ventricular weight over body weight ratio, resting heart rate, and cardiovascular O2- production were much higher in cardiomyopathic hamsters than those in control. At the age of 250 days, in addition to the continual deterioration of these parameters with age, the blood pressure started to fall and the signs of heart failure appeared. In these cardiomyopathic hamsters, chronic ASA treatment (a) completely prevented elevated O2- production and the NAD(P)H oxidase activity, (b) significantly slowed down the development of the cardiac hypertrophy and fibrosis. Conclusions. Chronic ASA treatment significantly prevents the deterioration of cardiac function and structure as well as the increased oxidative stress in the cardiomyopathic hamster. Our findings suggest that ASA presents a therapeutic potential to prevent cardiac dysfunction.

The objective of the study was to evaluate the efficacy of an interdisciplinary intervention known as Educoeur in reducing cardiovascular risk and improving health behaviors in people without evidence of cardiovascular disease and to compare the Educoeur program to interventions in a specialized clinic and in usual care family practice. In a parallel, randomized, controlled trial of 185 adults with at least two modifiable cardiovascular risk factors, patients were randomly assigned to either Educoeur, specialized clinic or usual care. Cardiovascular risk, biological and lifestyle measures were assessed at baseline and at 2 years. In Educoeur, measurements were also taken before and after the lifestyle group treatment program. In 12 weeks, patients in Educoeur significantly lowered their cardiovascular risk, weight, body mass index, waist circumference, systolic blood pressure, kilocalories intake and improved their VO2 Max and mental health. Changes remained significant at 2 years. Between group comparisons at 2 years demonstrated that Educoeur was significantly better in reducing cardiovascular risk than interventions in usual care. Together, these results highlight the importance of providing interdisciplinary programs that optimize cardiovascular risk reduction and promote active lifestyles in patients at risk of cardiovascular disease.

Objectives. Theophylline has been shown to delay the onset of myocardial ischemia and to prolong exercise duration. The present study was done to evaluate the mechanisms and actions of intravenous theophylline on the onset of ischemia and exercise duration.Background. The ischemic threshold may be altered by the differential coronary vasodilation induced by endogenous adenosine. Theophylline is a competitive receptor antagonist of adenosine and may have a potential as an anti-ischemic medication.Methods. A double-blind, placebo-controlled crossover trial using an infusion of intravenous theophylline (8.0 ± 2.0 mg/liter) or placebo before exercise in 12 patients was done. Oxygen uptake, heart rate, blood pressure and heart rate-blood pressure product were determined at the onset of ≥0.1-mV ST segment depression and angina pectoris, as well as at peak exercise. The extent of myocardial ischemia was evaluated by electrocardio graphic criteria and quantitation of thallium-201 images at peak exercise.Results. When compared with placebo, theophylline significantly delayed time to the onset of exercise-induced ischemia. Ischemia occurred at a higher heart rate-blood pressure product and oxygen uptake. Exercise duration was prolonged but was not associated with greater ischemia, as determined by oxygen uptake, ST segment depression, angina pectoris and size of thallium-201 defect.Conclusions. It is concluded that theophylline favorably alters myocardial ischemia not only by delaying its onset but also by enabling it to occur at a higher threshold without causing deleterious effects during exercise. The mechanism for the increased ischemic threshold may be through the inhibition of adenosine and the coronary steal phenomenon.

BACKGROUND:Inflammation and oxidative stress have been identified as integral parts in the pathogenesis of hypertension. Cyclo-oxygenase-2 which could elicit inflammation and free radicals generation appears to be a key enzyme in hypertension. Cyclo-oxygenase-2 expression and oxidative stress in cardiovascular tissues are increased in the angiotensin II model.METHODS:Cyclo-oxygenase-1 and cyclo-oxygenase-2 deficient mice and their cultured aortic smooth muscle cells were used to investigate the role of these enzymes in angiotensin II induced superoxide production and hypertension.RESULTS:At resting state, the superoxide production in aortic and cardiac tissues was lower in cyclo-oxygenase-2 deficient than in the wild type or in cyclo-oxygenase-1 deficient mice. Chronic angiotensin II infusion increased the superoxide production in these tissues from both cyclo-oxygenase-deficient and wild-type mice whereas the level in cyclo-oxygenase-2 deficient mice was equivalent to the basal level in wild-type mice. The hypertensive effect of angiotensin II was attenuated in cyclo-oxygenase-2 deficient mice. Aspirin treatment reduced the basal superoxide production and blunted the oxidative and hypertensive effect of angiotensin II in wild type and cyclo-oxygenase-1 deficient mice whereas it lost completely its antioxidative property in angiotensin II-treated aortic smooth muscle cells isolated from cyclo-oxygenase-2 deficient mice.CONCLUSIONS:Cyclo-oxygenase-2 pathway plays a major role in the superoxide generation as well as in the angiotensin II-induced oxidative stress and blood pressure. Cyclo-oxygenase-1 activity didn't show any influence on these parameters. These results suggest that cyclo-oxygenase-2 is involved in the pathogenesis of hypertension.

It has been reported that HMG-CoA reductase inhibitors such as atorvastatin induce vascular smooth muscle cell (SMC) apoptosis in vitro. However, this effect remains to be demonstrated in vivo. The present studies were designed to test the ability of atorvastatin to induce SMC apoptosis in vivo, using the spontaneously hypertensive rat (SHR) as a well-known reference model of SMC apoptosis induction in vivo by cardiovascular drugs including the calcium channel blocker amlodipine. Atorvastatin was administered to SHR for 3 or 6 weeks either alone or together with amlodipine, a drug combination clinically available to patients. Primary endpoints included aortic medial hypertrophy and aortic SMC hyperplasia, internucleosomal DNA fragmentation and expression of the apoptosis regulatory proteins Bax and Bcl-2. The SHR aorta showed no evidence of SMC apoptosis induction by atorvastatin, even at the high dose of 50 mg kg−1 day−1, although the statin significantly reduced oxidative stress after 3 weeks and blood pressure after 6 weeks of administration. Amlodipine-induced regression of aortic hypertophy and aortic SMC hyperplasia were dose- and time-dependent, but there was no interaction between atorvastatin and amlodipine in modulating the primary endpoints. These results do not support the notion that atorvastatin induces SMC apoptosis in the aortic media in vivo.

Objective: Cyclo-oxygenase (COX)-2 overreaction is implicated in numerous human diseases and could elicit inflammation and free radicals generation. Angiotensin II (AngII) stimulates COX-2 expression and induces oxidative stress in cardiovascular tissues. Aspirin reduces superoxide (O2-) production and prevents AngII-induced oxidative stress and hypertension. The present study was designed to evaluate the role of the COX-2 pathway in the antioxidative effect of aspirin. Methods: The COX-1 (COX-1-/-) or COX-2 (COX-2-/-) knock-out mice and the cultured aortic smooth muscle cells (SMCs) from these animals were treated either with aspirin or AngII for 14 days (in vivo) or for 48 hours (cell culture). The O2- production was measured with the lucigenin-enhanced chemiluminescence method. Results: In vivo study. As observed on wild type mice, aspirin treatment reduced and AngII treatment increased the cardiovascular tissue O2- production in COX-1-/- mice. AngII treatment also significantly increased the blood pressure in those mice. At variance from wild type or COX-1-/- mice, AngII treatment increased only moderately the O2- production and blood pressure in COX-2-/- mice. Histological analysis showed the presence of hypertrophy on renal arterioles in untreated COX-2-/- mice and in AngII-treated wild type or COX-1-/- mice. Study on cultured SMC. Aspirin treatment reduced the AngII-enhanced O2- production on cultured aortic SMCs from wild type mice by 23% and COX-1-/- mice by 30%. The basal O2- level produced by SMCs from COX-2-/- mice was significantly lower than that from wild type or COX-1-/- mice. AngII infusion brought the O2¡¥B production rate back to the basal level of WT mice. Aspirin treatment neither reduced the basal O2- level nor inhibited the AngII-enhanced O2- production on cultured SMCs from COX-2-/- mice. Conclusions: a) The COX-2 pathway plays an important role in the redox balance in cardiovascular tissue. b) The COX-2 pathway appears to be responsible in great part for the antioxidative and cardiovascular protective effect of aspirin against AngII.

To study the role of the serotoninergic 5-HT2B receptor in the development of cardiac hypertrophy and its link with left ventricular superoxide anion generation in a mouse model of angiotensin II-induced hypertension. Wild-type and 5-HT2B receptor knock-out (KO) mice were perfused with angiotensin II (0.2 mg.kg-1.d-1) for 14 days with or without SB215505 (1 mg.kg-1.d-1), an antagonist of the 5-HT2B receptor. Heart rate and blood pressure were measured by tail-cuff plethysmography. Cardiac hypertrophy was evaluated by echocardiography and direct measurement of heart weight. Superoxide anion production and maximal NAD(P)H oxidase activity were measured by a chemiluminescence method using lucigenin. Superoxide anion production was also measured in primary left ventricular fibroblasts cell cultures. Angiotensin II increased superoxide anion production (+32 %), the maximal activity of NAD(P)H oxidase (+84 %) in left ventricle of wild-type mice concomitantly with the arterial blood pressure (+37mmHg) and the heart/body weight ratio (+17 %). A pharmacological blockade (SB215505) or a genetic suppression of the 5-HT2B receptor prevented the increased superoxide anion production and cardiac hypertrophy but had no effect on cardiac hemodynamics or blood pressure. Angiotensin II also increased NAD(P)H oxidase activity in cultured cardiac fibroblasts and this increase was prevented by SB215505. The 5-HT2B receptor is a new potential target for the prevention of cardiac hypertrophy and its associated superoxide anion production. Cells of the extracellular matrix could possibly be involved in this mechanism.

To determine the contribution of nitric oxide (NO) in cardiovascular remodeling associated to hypertension and insulin resistance, male Sprague-Dawley rats received tap water supplemented or not (control), with 10% D-glucose (G) and/or 50 mg x kg(-1) x d(-1) L-NAME to inhibit NO synthase (G-LN or LN) for 4 weeks. Systolic blood pressure increased by 12%, 26%, and 39% with G, LN, and G-LN treatments, respectively. Hyperinsulinemia and insulin resistance (homeostasis model assessment index) occurred in G-treated rats (P < 0.05) and were further increased in G-LN (P < 0.01). Plasma adrenaline concentrations were markedly increased in all treated groups, especially in G-LN (P < 0.01), whereas noradrenaline was increased in G-treated rats only. Whereas no cardiac hypertrophy or fibrosis was detected, aortic hypertrophy occurred in LN and G-LN rats (P < 0.001) without smooth muscle hyperplasia. Superoxide anion formation was increased in the aorta of all treated groups (P < 0.01) and in the heart of LN (P < 0.05), but reduced nicotinamide adenine dinucleotide phosphate (NAD(P)H) oxidase activity was not affected. In conclusion, the loss of the wide-range protective effects of NO, the increased vascular oxidative stress, and the sympathoadrenal hyperactivity are among the contributing factors leading to the exacerbation of hypertension and insulin resistance in G-LN. These factors were sufficient to cause vascular but not cardiac hypertrophy.

We established previously that 5-HT 2B receptors are involved in cardiac hypertrophy through the regulation of hypertrophic cytokines in cardiac fibroblasts. Moreover, the generation of reactive oxygen species and tumor necrosis factor-α through the activation of reduced nicotinamide-adenine dinucleotide phosphate [NAD(P)H] oxidase has been implicated in cardiac hypertrophy. In this study, we investigated whether 5-HT 2B receptors could be involved in the development of cardiac hypertrophy associated with superoxide anion production. Therefore, we measured the effects of serotonergic 5-HT 2B receptor blockade on left-ventricular superoxide anion generation in 2 established pharmacological models of cardiac hypertrophy, ie, angiotensin II and isoproterenol infusions in mice. Angiotensin II infusion for 14 days increased superoxide anion concentration (+32%), NAD(P)H oxidase maximal activity (+84%), and p47 phox NAD(P)H oxidase subunit expression in the left ventricle together with hypertension (+37 mm Hg) and cardiac hypertrophy (+17% for heart weight:body weight). The 5-HT 2B receptor blockade by a selective antagonist (SB215505) prevented the increase in cardiac superoxide generation and hypertrophy. Similarly, infusion for 5 days of isoproterenol increased left-ventricular NAD(P)H oxidase activity (+48%) and cardiac hypertrophy (+31%) that were prevented by the 5-HT 2B receptor blockade. Finally, in the primary culture of left-ventricular cardiac fibroblasts, angiotensin II and isoproterenol stimulated NAD(P)H oxidase activity. This activation was prevented by SB215505. These findings suggest that the 5-HT 2B receptor may represent a new target to reduce cardiac hypertrophy and oxidative stress. Its blockade affects both angiotensin II and β-adrenergic trophic responses without significant hemodynamic alteration.

Diabetes is known to be associated with cardiovascular complications and polyneuropathic pain. This study reports on the expression and function of kinin B1 receptor (B1R) in rats treated with 10% glucose for 12 weeks, a model of insulin resistance. Beneficial effects of the non-peptide B1R antagonist SSR240612 (10 mg/kg/day) given by gavage during the last week were determined on the following parameters: tactile and cold allodynia (hindpaw plantar stimulation), systolic blood pressure (tail cuff plethysmography). At the end of the protocol, expression of B1R (mRNA and binding sites) and MnSOD (mRNA) was measured by real time PCR and/or autoradiography. Plasma glucose and insulin were measured in overnight fasted rats as well as aortic superoxide anion (chemiluminescence). Results: Glucose-fed rats exhibited tactile and cold allodynia, hyperglycemia, hyperinsulinemia, hypertension, increased aortic superoxide anion and insulin resistance (Homa index). B1R (mRNA and binding sites) was significantly increased in the spinal cord and renal cortex. All these abnormalities, including B1R overexpression, were normalized or reduced by SSR240612. Furthermore, the mRNA of the antioxidant enzyme MnSOD was significantly increased in spinal cord and renal cortex of rats treated with SSR240612 when compared to glucose-fed rat treated with vehicle. Conclusion: This study provides pharmacological and molecular evidence that B1R is involved in cardiovascular, metabolic and sensory abnormalities that occurred in a model of insulin resistance. Part of the beneficial effects of B1R antagonist appears to be due to decreased oxidative stress and increased of anti-oxidant defence. Thus, the novel generation of orally active B1R antagonists offers promising therapeutic value in the treatment of diabetic complications.