Rodrigo Castillo

A Randomized Controlled Trial to Prevent Post-Operative Atrial Fibrillation by Antioxidant Reinforcement

OBJECTIVES This study was designed to assess whether the reinforcement of the antioxidant system, through n-3 fatty acids plus antioxidant vitamin supplementation, could reduce the incidence of post-operative atrial fibrillation. BACKGROUND Therapy to prevent post-operative atrial fibrillation remains suboptimal. Although oxidative stress plays a key role in the pathogenesis of this arrhythmia, antioxidant reinforcement has produced controversial results. METHODS A total of 203 patients scheduled for on-pump cardiac surgery were randomized to placebo or supplementation with n-3 polyunsaturated fatty acids (2 g/day) (eicosapentaenoic acid:docosahexaenoic acid ratio 1:2), vitamin C (1 g/day), and vitamin E (400 IU/day). The primary outcome was the occurrence of post-operative atrial fibrillation. Secondary outcomes were the biomarkers related to oxidative stress and inflammation. RESULTS Post-operative atrial fibrillation occurred in 10 of 103 patients (9.7%) in the supplemented group versus 32 of 100 patients (32%) in the placebo group (p < 0.001). Early after surgery, placebo patients presented with increased levels of biomarkers of inflammation and oxidative stress, which were markedly attenuated by antioxidant supplementation. The activity of catalase, superoxide dismutase, and glutathione peroxidase in atrial tissue of the supplemented patients was 24.0%, 17.1%, and 19.7% higher than the respective placebo values (p < 0.05). The atrial tissue of patients who developed atrial fibrillation showed NADPH oxidase p47-phox subunit protein and mRNA expression 38.4% and 35.7% higher, respectively, than patients in sinus rhythm (p < 0.05). CONCLUSIONS This safe, well-tolerated, and low-cost regimen, consisting of n-3 polyunsaturated fatty acids plus vitamins C and E supplementation, favorably affected post-operative atrial fibrillation, increased antioxidant potential, and attenuated oxidative stress and inflammation. (Prevention of Post-Operative Atrial Fibrillation: Pathophysiological Characterization of a Pharmacological Intervention Based on a Novel Model of Nonhypoxic Pre-Conditioning; ISRCTN45347268).

Cardioprotection against ischaemia/reperfusion by vitamins C and E plus n-3 fatty acids: molecular mechanisms and potential clinical applications.

The role of oxidative stress in ischaemic heart disease has been thoroughly investigated in humans. Increased levels of ROS (reactive oxygen species) and RNS (reactive nitrogen species) have been demonstrated during ischaemia and post-ischaemic reperfusion in humans. Depending on their concentrations, these reactive species can act either as benevolent molecules that promote cell survival (at low-to-moderate concentrations) or can induce irreversible cellular damage and death (at high concentrations). Although high ROS levels can induce NF-κB (nuclear factor κB) activation, inflammation, apoptosis or necrosis, low-to-moderate levels can enhance the antioxidant response, via Nrf2 (nuclear factor-erythroid 2-related factor 2) activation. However, a clear definition of these concentration thresholds remains to be established. Although a number of experimental studies have demonstrated that oxidative stress plays a major role in heart ischaemia/reperfusion pathophysiology, controlled clinical trials have failed to prove the efficacy of antioxidants in acute or long-term treatments of ischaemic heart disease. Oral doses of vitamin C are not sufficient to promote ROS scavenging and only down-regulate their production via NADPH oxidase, a biological effect shared by vitamin E to abrogate oxidative stress. However, infusion of vitamin C at doses high enough to achieve plasma levels of 10 mmol/l should prevent superoxide production and the pathophysiological cascade of deleterious heart effects. In turn, n-3 PUFA (polyunsaturated fatty acid) exposure leads to enhanced activity of antioxidant enzymes. In the present review, we present evidence to support the molecular basis for a novel pharmacological strategy using these antioxidant vitamins plus n-3 PUFAs for cardioprotection in clinical settings, such as post-operative atrial fibrillation, percutaneous coronary intervention following acute myocardial infarction and other events that are associated with ischaemia/reperfusion.

Cellular and molecular mechanisms in the hypoxic tissue: role of HIF-1 and ROS

Reactive oxygen species such as superoxide anion radicals (O2−) and hydrogen peroxide (H2O2) have for long time been recognized as undesirable by‐products of the oxidative mitochondrial generation of adenosine triphosphate (ATP). Recently, these highly reactive species have been associated to important signaling pathways in diverse physiological conditions such as those activated in hypoxic microenvironments. The molecular response to hypoxia requires fast‐acting mechanisms acting within a wide range of partial pressures of oxygen (O2). Intracellular O2 sensing is an evolutionary preserved feature, and the best characterized molecular responses to hypoxia are mediated through transcriptional activation. The transcription factor, hypoxia‐inducible factor 1 (HIF‐1), is a critical mediator of these adaptive responses, and its activation by hypoxia involves O2‐dependent posttranslational modifications and nuclear translocation. Through the induction of the expression of its target genes, HIF‐1 coordinately regulates tissue O2 supply and energetic metabolism. Other transcription factors such as nuclear factor κB are also redox sensitive and are activated in pro‐oxidant and hypoxic conditions. The purpose of this review is to summarize new developments in HIF‐mediated O2 sensing mechanisms and their interactions with reactive oxygen species–generating pathways in normal and abnormal physiology. Copyright

Antioxidant Therapy Reduces Oxidative and Inflammatory Tissue Damage in Patients Subjected to Cardiac Surgery with Extracorporeal Circulation

Ischaemia reperfusion injury is a pathophysiological event that occurs after cardiac surgery with extracorporeal circulation. This clinical event has been associated with the induction of oxidative and inflammatory damage in atrial tissue. Here, we tested whether combined omega 3 polyunsaturated fatty acids (n-3 PUFA)-antioxidant vitamin protocol therapy reduces oxidative and inflammatory cardiac tissue damage. This trial assigned 95 either-sex patients to supplementation with n-3 PUFA (2 g/day), or matching placebo groups, 7 days before on-pump surgery. Antioxidant vitamins C (1 g/day) and E (400 IU/day) or placebo were added from 2 days before surgery until discharge. Blood and atrial tissue samples were obtained during the intervention. Reduced/oxidized glutathione (GSH/GSSG) ratio, malondialdehyde (MDA) and protein carbonylation were determined in atrial tissue. Leucocyte count and high-sensitivity C-reactive protein (hs-CRP) in blood plus nuclear factor (NF)-κappaB activation in atrial tissue served for inflammation assessment. Lipid peroxidation and protein carbonylation were 27.5 and 24% lower in supplemented patients (p < 0.01). GSH/GSSG ratio was 38.1% higher in supplemented patients compared with placebo (p < 0.01). Leucocyte count and serum hs-CRP levels were markedly lower throughout the protocol in supplemented patients (p < 0.01). Atrial tissue NF-κB DNA activation in supplemented patients was 22.5% lower than that in placebo patients (p < 0.05). The combined n-3 PUFA-antioxidant vitamin protocol therapy here proposed reduced the oxidative stress and inflammation biomarkers, in patients undergoing on-pump cardiac surgery.

Use of vitamins C and E as a prophylactic therapy to prevent postoperative atrial fibrillation.

Oxidative stress has been strongly involved in the underlying mechanism of atrial fibrillation, particularly in the arrhythmia occurring in patients undergoing cardiac surgery with extracorporeal circulation (postoperative atrial fibrillation). The ischemia/reperfusion injury thus occurring in the myocardial tissue contributes to the development of tissue remodeling, thought to be responsible for the functional heart impairment. Consequently, structural changes due to the cardiac tissue biomolecules attack by reactive oxygen and/or nitrogen species could account for functional changes in ion channels, transporters, membrane conductance, cytosolic transduction signals, and other events, all associated with the occurrence of arrhythmic consequences. The lack of success and significant side effects of anti-arrhythmic drugs have given rise to attempts aimed to develop alternative novel pharmacologic treatments. On this line, the biological properties of the antioxidant vitamins C and E suggest that they could decrease the vulnerability of the heart to the oxidative damage. Nevertheless, very few studies to assess their anti-arrhythmic effects have been reported in humans. The clinical and experimental evidence supporting the view that the pharmacological use of antioxidant vitamins could contribute to prevent postoperative atrial fibrillation is presented.

Oxidative stress as a novel target in pediatric sepsis management

Sepsis with secondary multisystem organ dysfunction syndrome is the leading cause of death in the pediatric intensive care unit. Increased reactive oxygen species may influence circulating and endothelial cells, contributing to inflammatory tissue injury and explaining the tissue hypoxia paradigm based on microvascular dysfunction. An impaired mitochondrial cellular oxygen utilization, rather than inadequate oxygen delivery, was claimed to play a more important role in the development of multisystem organ dysfunction syndrome. Anyway, it seems plausible that reactive oxygen species can mediate the pathophysiologic processes occurring in sepsis. However, the consensus guidelines for the management of patients with these conditions do not include the enhancement of antioxidant potential. Therefore, further investigation is needed to support interventions aimed to attenuate the severity of the systemic compromise by abrogating the mechanism of oxidative damage. Antioxidant supplementation currently in use lacks a mechanistic support. Specific pharmacologic targets, such as mitochondria or Nicotinamide Adenine Dinucleotide Phosphate-Oxidase (NADPH) oxidase system, need to be explored. Furthermore, the early recognition of oxidative damage in these seriously ill patients and the usefulness of oxidative stress biomarkers to define a cut point for more successful therapeutic antioxidant interventions to be instituted would offer a new strategy to improve the outcome of critically ill children.

Prevention of atrial fibrillation following cardiac surgery: basis for a novel therapeutic strategy based on non-hypoxic myocardial preconditioning.

Atrial fibrillation is the most common complication of cardiac surgical procedures performed with cardiopulmonary bypass. It contributes to increased hospital length of stay and treatment costs. At present, preventive strategies offer only suboptimal benefits, despite improvements in anesthesia, surgical technique, and medical therapy. The pathogenesis of postoperative atrial fibrillation is considered to be multifactorial. However oxidative stress is a major contributory factor representing the unavoidable consequences of ischemia/reperfusion cycle occurring in this setting. Considerable evidence suggests the involvement of reactive oxygen species (ROS) in the pathogenic mechanism of this arrhythmia. Interestingly, the deleterious consequences of high ROS exposure, such as inflammation, cell death (apoptosis/necrosis) or fibrosis, may be abrogated by a myocardial preconditioning process caused by previous exposure to moderate ROS concentration known to trigger survival response mechanisms. The latter condition may be created by n-3 PUFA supplementation that could give rise to an adaptive response characterized by increased expression of myocardial antioxidant enzymes and/or anti-apoptotic pathways. In addition, a further reinforcement of myocardial antioxidant defenses could be obtained through vitamins C and E supplementation, an intervention also known to diminish enzymatic ROS production. Based on this paradigm, this review presents clinical and experimental evidence supporting the pathophysiological and molecular basis for a novel therapeutic approach aimed to diminish the incidence of postoperative atrial fibrillation through a non-hypoxic preconditioning plus a reinforcement of the antioxidant defense system in the myocardial tissue.

Effect of Carvedilol and Nebivolol on Oxidative Stress‐related Parameters and Endothelial Function in Patients with Essential Hypertension

Oxidative stress and endothelial dysfunction have been associated with essential hypertension (EH) mechanisms. The purpose of this study was to evaluate the effect of carvedilol and nebivolol on the oxidative stress‐related parameters and endothelial function in patients with EH. The studied population included 57 patients, either sex, between 30 and 75 years of age, with mild‐to‐moderate EH complications. Participants were randomized to receive either carvedilol (12.5 mg) (n = 23) or nebivolol (5 mg) (n = 21) for 12 weeks. Measurements included; 24‐hr ambulatory blood pressure (BP), flow‐mediated dilatation, levels of nitric oxide estimated as nitrite – a nitric oxide metabolite ( NO 2 ) – in plasma, and oxidative stress‐related parameters in plasma and erythrocyte. EH patients who were treated with nebivolol or carvedilol showed systolic BP reductions of 17.4 and 19.9 mmHg, respectively, compared with baseline values (p < 0.01). Diastolic BP was reduced by 13.7 and 12.8 mmHg after the treatment with ebivolol and carvedilol, respectively (p < 0.01) (fig. 2B). Nebivolol and carvedilol showed 7.3% and 8.1% higher endothelium‐dependent dilatation in relation to baseline values (p < 0.05). Ferric‐reducing ability of plasma (FRAP) and reduced glutathione/oxidized glutathione (GSSH) ratio showed 31.5% and 29.6% higher levels in the carvedilol group compared with basal values; however, nebivolol‐treated patients did not show significant differences after treatment. On the other hand, the NO 2 plasma concentration was not modified by the administration of carvedilol. However, nebivolol enhanced these levels in 62.1% after the treatment. In conclusion, this study demonstrated the antihypertensive effect of both beta‐blockers. However, carvedilol could mediate these effects by an increase in antioxidant capacity and nebivolol through the raise in NO 2 concentration. Further studies are needed to determine the molecular mechanism of these effects.

Molecular mechanisms of gastrointestinal protection by quercetin against indomethacin-induced damage: role of NF-κB and Nrf2.

The aim of this study was to determine the gastrointestinal protection by quercetin against indomethacin-induced oxidative stress and inflammation, with specific interest in studying the underlying molecular mechanisms. We hypothesized that the quercetin-protective effect relies on its antioxidant and antiinflammatory properties. Rats were pretreated with quercetin (50- or 100-mg/kg, ig single dose), 30 min before INDO administration (40-mg/kg ig single dose). Caco-2 cells were treated with INDO (250 and 500 μM) in the absence or presence of quercetin (10 μg/ml). Quercetin prevented the decrease in nuclear translocation of Nrf2, a key regulator of the antioxidant response, and the increase in reactive oxygen species levels induced by INDO by inhibiting the enhancement of NADPH oxidase and xanthine oxidase activities as well as the reduction in superoxide dismutase and glutathione peroxidase activities in gastric and ileal tissues. Quercetin also prevented INDO-induced ICAM-1 and P-selectin expressions and the increase of myeloperoxidase activity in gastric and ileal tissues and NF-κB activation and IL-8 production in Caco-2 cells. Quercetin did not affect the inhibition of TNFα-mediated production of prostaglandin E2 induced by INDO in Caco-2 cells. The protective effects of quercetin observed in the gastric and ileal mucosa of rats as well as in Caco-2 cells relied on the ability of this flavonol to prevent NF-κB activation and increase Nrf2 translocation. This study supports the concept that quercetin may be useful in the prevention and/or treatment of nonsteroidal antiinflammatory drug-associated side effects, without interfering with their therapeutic efficacy.

3,4-dihydroxyphenylacetic acid, a microbiota-derived metabolite of quercetin, protects against pancreatic β-cells dysfunction induced by high cholesterol

Cholesterol plays an important role in inducing pancreatic β-cell dysfunction, characterized by an impaired insulin secretory response to glucose, representing a hallmark of the transition from pre-diabetes to diabetes. 3,4 dihydroxyphenylacetic acid (ES) is a scarcely studied microbiota-derived metabolite of quercetin with antioxidant properties. The aim of this study was to determine the protective effect of ES against apoptosis, mitochondrial dysfunction and oxidative stress induced by cholesterol in Min6 pancreatic β-cells. Cholesterol decreased viability, induced apoptosis and mitochondrial dysfunction by reducing complex I activity, mitochondrial membrane potential, ATP levels and oxygen consumption. Cholesterol promoted oxidative stress by increasing cellular and mitochondrial reactive oxygen species and lipid peroxidation and decreasing antioxidant enzyme activities; in addition, it slightly increased Nrf2 translocation to the nucleus. These events resulted in the impairment of the glucose-induced insulin secretion. ES increased Nrf2 translocation to the nucleus and protected pancreatic β-cells against impaired insulin secretion induced by cholesterol by preventing oxidative stress, apoptosis and mitochondrial dysfunction. Nrf2 activation seems to be involved in the mechanisms underlying the antioxidant protection exerted by ES in addition to preventing the disruption of antioxidant enzymatic defenses. Although additional in vivo experiments are required, this metabolite is suggested as a promising drug target for the prevention of the pathological development from a pre-diabetic to a diabetic state.