Nitrite Attenuates the In Vitro Inflammatory Response of Immune Cells to the SARS-CoV-2 S Protein without Interfering in the Antioxidant Enzyme Activation.

SARS-CoV-2 induces a hyperinflammatory reaction due to the excessive release of cytokines during the immune response. The bacterial endotoxin lipopolysaccharide (LPS) contributes to the low-grade inflammation associated with the metabolic syndrome, enhancing the hyperinflammatory reaction induced by the SARS-CoV-2 infection. The intake of sodium nitrate, a precursor of nitrite and nitric oxide, influences the antioxidant and pro-inflammatory gene expression profile after immune stimulation with LPS in peripheral blood mononuclear cells from metabolic syndrome patients. We aimed to assess the inflammatory and antioxidant responses of immune cells from metabolic syndrome patients to exposure to the SARS-CoV-2 spike protein (S protein) together with LPS and the effect of nitrite in these responses. Whole blood samples obtained from six metabolic syndrome patients were cultured for 16 h at 37 °C with four different media: control medium, control medium plus LPS (100 ng/mL), control medium plus LPS (100 ng/mL) plus S protein (10 ng/mL), and control medium plus LPS (100 ng/mL) plus S protein (10 ng/mL) plus nitrite (5 µM). Immune stimulation with the LPS/S protein enhanced nitrate biosynthesis from nitrite oxidation and probably from additional organic precursors. In vitro incubations with the LPS/S protein enhanced the expression and/or release of pro-inflammatory TNFα, IL-6, IL-1ß, and TLR4, as well as the expression of the anti-inflammatory IL-1ra and IL-10 and antioxidant enzymes. Nitrite attenuated the pro- and anti-inflammatory response induced by the S protein without interfering with the activation of TLR4 and antioxidant enzyme expression, raising the possibility that nitrite could have potential as a coadjutant in the treatment of COVID-19.

A phase 1b randomized, placebo-controlled clinical trial with SNF472 in haemodialysis patients.

AIMS: SNF472 is a calcification inhibitor that is being studied as a novel treatment for calciphylaxis and cardiovascular calcification (CVC). A first study showed acceptable safety and tolerability in a single ascending dose administration in healthy volunteers and a single dose administration in haemodialysis (HD) patients. This study aimed to assess the safety, tolerability, and pharmacokinetics/pharmacodynamics relationship of intravenous SNF472 in HD patients in a multiple ascending dose administration trial with 5 doses tested for 1 week (3 administrations) and 1 dose tested for 4 weeks (12 administrations). METHODS: This double blind, randomized, placebo-controlled Phase 1b study investigated the safety, tolerability, pharmacokinetics and pharmacodynamics of SNF472 after repeated administrations to HD patients for up to 28 days. A pharmacodynamic assessment was performed to evaluate the potential for SNF472 to inhibit hydroxyapatite (HAP) formation. Patients were grouped into 2 cohorts, receiving multiple ascending doses for 1 week (1 to 20 mg/kg, Cohort 1) and 1 dose of 10 mg/kg for 4 weeks (Cohort 2) of intravenous SNF472. RESULTS: Physical status, body weight, cardiorespiratory function, body temperature and laboratory parameters were in the normal range. No clinically relevant effects on heart rate or blood pressure were observed. No abnormal electrocardiogram or QTcB period were reported. The peak plasma concentration (7.6, 16.1, 46.0 and 66.9 µg/mL for 3, 5, 12.5 and 20 mg/kg, respectively) was observed at the end of the 4-hour infusion and thereafter concentrations declined rapidly with half-life between 32 and 65 min. SNF472 at 10 mg/kg inhibited dose dependently HAP crystallization in plasma samples after 28 days of treatment (78% inhibition, P < .001). CONCLUSIONS: SNF472 is safe and well tolerated in HD patients after 2 schemes: multiple ascending doses for 1 week and after repeated dosing of 10 mg/kg for 4 weeks. In both schemes, SNF472 inhibits the induction of HAP crystallization. These results provide support for the use of SNF472 as a novel treatment for CVC in end-stage renal disease.

Hexasodium fytate exposure-response correlations in a randomized, placebo-controlled study of patients on dialysis with cardiovascular calcification.

Background: Patients receiving dialysis have high cardiovascular risk in part due to extensive vascular calcification. In the CaLIPSO study, infusion of hexasodium fytate (SNF472), the hexasodium salt of inositol hexaphosphate, for 52 weeks thrice weekly during hemodialysis significantly reduced progression of coronary artery calcification (CAC). This report examines pharmacokinetic/pharmacodynamic (PK/PD) and exposure-efficacy in CaLIPSO. Methods: We measured hexasodium fytate plasma concentrations (PK) by validated liquid chromatography-mass spectroscopy, and hydroxyapatite crystallization in plasma (PD) by validated spectrophotometry. Analyses included patients evaluable for PK, PD, and CAC change (per-protocol analysis). We developed a simple Emax model for maximum concentration (Cmax) and PD effect, and linear and non-linear Emax models for exposure-efficacy among individual average Cmax and absolute and percent changes in CAC score from baseline to week 52. Results: Among evaluable patients receiving placebo (n = 15), 300 mg (n = 20), or 600 mg (n = 20), average Cmax across visits was not quantifiable (<0.76 µM), 15 µM, and 46 µM, respectively. These results suggest a more-than-proportional increase, without accumulation, with a Cmax ratio of approximately 3 for the doses administered. Average inhibition of hydroxyapatite crystallization was 15%, 61%, and 75%, respectively, and similar across visits. Simple Emax models described 80% maximal effect at exposures >21.9 µM and a plateau in exposure-efficacy above the third quartile of Cmax (≥32 µM). Conclusion: Hexasodium fytate has exposure-dependent effects on hydroxyapatite crystallization and progression of cardiovascular calcification. Simple Emax models show robust relations among exposure, inhibition of hydroxyapatite crystallization, and change in CAC volume. Clinical Trial Registration: https://www.clinicaltrials.gov; identifier NCT02966028.

Antioxidants restore protoporphyrinogen oxidase in variegate porphyria patients.

BACKGROUND: Variegate porphyria (VP) is the result of decreased protoporphyrinogen oxidase (PPOX) activity and results in the accumulation of porphyrins and porphyrin precursors. Our aims were to analyse the basal antioxidant defences and oxidative damage markers and the effects of a diet supplementation with vitamins E and C on the oxidant/antioxidant status and PPOX gene expression in lymphocytes of variegate porphyria (VP) patients. MATERIALS AND METHODS: Twelve women affected by VP and 12 control women participated in a randomized and double-blind crossover study. Each participant took either 50 mg/day vitamin E and 150 mg/day vitamin C or a placebo for 6 months. RESULTS: Lymphocyte PPOX gene expression, together with catalase and glutathione peroxidase activities, was reduced in VP women. No differences were observed in the levels of malondialdehyde and protein carbonyl derivatives. Stimulated lymphocyte H2 O2 production was higher in porphyric women. Supplementation with antioxidant vitamins increased PPOX expression in VP patients. Glutathione reductase (GRd) and superoxide dismutase (SOD) activities were higher in the treatment groups. CONCLUSIONS: Lymphocytes from VP patients show reduced PPOX expression and present a greater susceptibility to producing H2 O2 and impaired H2 O2 detoxifying mechanisms. Supplementation with vitamins E and C restores PPOX expression in VP patients and enhances GRd and SOD activity, suggesting the potential benefits of a diet rich in vitamins E and C in these patients.

Prevention of neutrophil protein oxidation with vitamins C and E diet supplementation without affecting the adaptive response to exercise.

The authors studied the effects of antioxidant diet supplementation with an almond-based beverage on neutrophil antioxidants, nitrite, and protein oxidative alterations after exercise. Fourteen trained male amateur runners were randomly assigned in a double-blind fashion to receive antioxidant supplementation (152 mg/d vitamin C and 50 mg/d vitamin E) or placebo using an almond-based beverage for 1 mo and participated in a half-marathon race. Blood samples were taken before and after the half-marathon and after 3 hr recovery. Supplementation significantly increased basal neutrophil vitamin C compared with placebo (p < .05). Exercise increased neutrophil vitamin E levels in the supplemented group and decreased vitamin C in both groups after recovery (p < .05). Neutrophil catalase and glutathione peroxidase gene expression and nitrite levels were significantly increased as result of exercise (p < .05). Nitrotyrosine and protein carbonyl derivates increased only in the placebo group after exercise (p < .05), and these values remained high at recovery. No significant differences were evidenced in caspase-3 activity and DNA damage. Antioxidant supplementation with vitamins C and E reduced the exercise-induced oxidation of proteins in neutrophils, without altering the antioxidant adaptive response, as evidenced by the increased catalase and glutathione peroxidase gene expression.

Polyunsaturated and Saturated Oxylipin Plasma Levels Allow Monitoring the Non-Alcoholic Fatty Liver Disease Progression to Severe Stages.

Hepatic fat accumulation is the hallmark of non-alcoholic fatty liver disease (NAFLD). Our aim was to determine the plasma levels of oxylipins, free polyunsaturated fatty acids (PUFA) and markers of lipid peroxidation in patients with NAFLD in progressive stages of the pathology. Ninety 40-60-year-old adults diagnosed with metabolic syndrome were distributed in without, mild, moderate or severe NAFLD stages. The free PUFA and oxylipin plasma levels were determined by the UHPLC-MS/MS system. The plasma levels of oxylipins produced by cyclooxygenases, lipoxygenases and cytochrome P450, such as prostaglandin 2α (PGF2α), lipoxinB4 and maresin-1, were higher in severe NAFLD patients, pointing to the coexistence of both inflammation and resolution processes. The plasma levels of the saturated oxylipins 16-hydroxyl-palmitate and 3-hydroxyl-myristate were also higher in the severe NAFLD patients, suggesting a dysregulation of oxidation of fatty acids. The plasma 12-hydroxyl-estearate (12HEST) levels in severe NAFLD were higher than in the other stages, indicating that the hydroxylation of saturated fatty acid produced by reactive oxygen species is more present in this severe stage of NAFLD. The plasma levels of 12HEST and PGF2α are potential candidate biomarkers for diagnosing NAFLD vs. non-NAFLD. In conclusion, the NAFLD progression can be monitored by measuring the plasma levels of free PUFA and oxylipins characterizing the different NAFLD stages or the absence of this disease in metabolic syndrome patients.

Phytoestrogens enhance antioxidant enzymes after swimming exercise and modulate sex hormone plasma levels in female swimmers.

Our aim was to investigate the effects of diet supplementation with phytoestrogens on sex hormone levels, antioxidant adaptive responses and oxidative damage induced by exercise. Ten female swimmers participated for 26 days in a diet intervention with either a functional beverage rich in vitamins C and E or the same beverage but also supplemented with Lippia citriodora extract (PLX) containing 20 mg/100 ml verbascoside. After the intervention all subjects participated in a swimming session for 30 min maintaining the intensity at about 75-80% of their individual best performance time for a 50-m swim. In lymphocytes, the superoxide dismutase activity increased after exercise, with a higher increase in the PLX group. Swimming increased the erythrocyte activity of glutathione peroxidase and glutathione reductase in the PLX group. Purified glutathione reductase activity increased after an in vitro incubation with PLX. No effects were observed on the lymphocyte levels of malondialdehyde and carbonyls, but exercise increased the percentage of high-damaged lymphocytes 2.8 times in the placebo group and 1.5 times in the PLX group. PLX decreased the levels of 17-ß-estradiol and testosterone and increased the levels of the sex hormone binding globulin. In conclusion, supplementation with phytoestrogens enhances the glutathione-dependent enzyme activities in erythrocytes and the superoxide dismutase activity in lymphocytes in response to exercise. PLX also shows direct antioxidant properties, by increasing glutathione reductase enzyme activity in vitro. Supplementation with phytoestrogens also decreases the plasma steroid hormone levels, pointing towards a possible agonistic effect of verbascoside in the hypothalamic regulation of estradiol synthesis.

Effect of lemon verbena supplementation on muscular damage markers, proinflammatory cytokines release and neutrophils' oxidative stress in chronic exercise.

Intense exercise is directly related to muscular damage and oxidative stress due to excessive reactive oxygen species (ROS) in both, plasma and white blood cells. Nevertheless, exercise-derived ROS are essential to regulate cellular adaptation to exercise. Studies on antioxidant supplements have provided controversial results. The purpose of this study was to determine the effect of moderate antioxidant supplementation (lemon verbena extract) in healthy male volunteers that followed a 90-min running eccentric exercise protocol for 21 days. Antioxidant enzymes activities and oxidative stress markers were measured in neutrophils. Besides, inflammatory cytokines and muscular damage were determined in whole blood and serum samples, respectively. Intense running exercise for 21 days induced antioxidant response in neutrophils of trained male through the increase of the antioxidant enzymes catalase, glutathione peroxidase and glutathione reductase. Supplementation with moderate levels of an antioxidant lemon verbena extract did not block this cellular adaptive response and also reduced exercise-induced oxidative damage of proteins and lipids in neutrophils and decreased myeloperoxidase activity. Moreover, lemon verbena supplementation maintained or decreased the level of serum transaminases activity indicating a protection of muscular tissue. Exercise induced a decrease of interleukin-6 and interleukin-1ß levels after 21 days measured in basal conditions, which was not inhibited by antioxidant supplementation. Therefore, moderate antioxidant supplementation with lemon verbena extract protects neutrophils against oxidative damage, decreases the signs of muscular damage in chronic running exercise without blocking the cellular adaptation to exercise.

Dietary Sodium Nitrate Activates Antioxidant and Mitochondrial Dynamics Genes after Moderate Intensity Acute Exercise in Metabolic Syndrome Patients.

Exercise can induce a pro-inflammatory response in aged subjects with metabolic disorders and nitrate supplementation has shown anti-inflammatory effects. We evaluated the influence of dietary nitrate on the response of the antioxidant and mitochondrial dynamics genes to acute exercise in peripheral blood mononuclear cells (PBMCs), as well as the antioxidant and the inflammatory response of PBMCs against immune stimulation. Metabolic syndrome patients participated in a crossover study in which they consumed a beverage containing 16 mM sodium nitrate or a placebo with the same composition without nitrate before performing a submaximal test at 60%-70% of their maximal heart rate for 30 min. The intake of nitrate increased the nitrate plus nitrite plasma levels about 8-fold and induced the upregulation of catalase, superoxide dismutase, glutathione peroxidase, mitofusin 2 and PGC1α in PBMCs after exercise. The gene expression of catalase and TNFα was enhanced by phorbol myristate acetate (PMA) only in the placebo group, while the glutathione peroxidase expression was enhanced by PMA only after nitrate intake. The intake of nitrate by metabolic syndrome patients induces an antioxidant and mitochondrial response to exercise at the same time that it attenuates the pro-inflammatory response to immune stimulation.

Impaired lymphocyte mitochondrial antioxidant defences in variegate porphyria are accompanied by more inducible reactive oxygen species production and DNA damage.

This study aimed to analyse lymphocyte reactive oxygen species (ROS) production and detoxification mechanisms and the appearance of oxidative damage in variegate porphyria (VP) patients. Twelve women affected by VP and 12 pair-matched healthy control women participated in the study. VP women presented impaired expression of the mitochondrial proteins protoporphyrinogen oxidase, uncoupling protein-3, Bcl-2 and sirtuin 3. Lymphocytes from VP women presented higher H(2)O(2) production than controls after stimulation with phorbol myristate acetate. The inhibition of H(2)O(2) production after in vitro lymphocyte treatment with myxothiazol pointed towards complex III of the mitochondrial respiratory chain as the main contributor of the higher ROS production in porphyric subjects. No differences were observed between VP and control subjects in the levels of DNA damage, assessed by the comet assay method in un-treated lymphocytes. However, DNA damage, expressed both as a percentage of DNA in tail and as the tail moment, was greater in VP women than controls after lymphocyte treatment with H(2)O(2). In conclusion, lymphocytes from VP women showed impaired expression of mitochondrial antioxidant defences but no significant signs of oxidative stress were evidenced in basal, non-stressing conditions; however, lymphocytes of VP women were more susceptible to producing mitochondrial ROS and to suffering oxidative damage when submitted to stressful situations.

The double edge of reactive oxygen species as damaging and signaling molecules in HL60 cell culture.

AIMS: Our aim was to establish the conditions in which reactive oxygen species produce pathological or hormetic effects on HL60 cells. METHODS: HL60 cells were treated with either single bouts (1, 10 and 100 microM) or a sustained production (0.1, 1.0 and 10.0 nM/s) of H(2)O(2). RESULTS: Exposure to 10 and 100 microM H(2)O(2) activated catalase, glutathione peroxidase and glutathione reductase through post-transcriptional mechanisms and induced oxidative modification of proteins. When cells where exposed to sustained H(2)O(2) production, a clear dose-response effect was detected in the activity of the antioxidant enzymes catalase, glutathione peroxidase and Mn-SOD, with higher concentrations of H(2)O(2) inducing greater enzyme activities. Catalase, HO-1, UCP-3, iNOS and PGC-1alpha expressions were activated after sustained exposure to 1 and 10 nM H(2)O(2)/s. Although the antioxidant defenses were activated, oxidative damage appeared in DNA and proteins in cells treated with 1 and 10 nM/s. CONCLUSIONS: HL60 cells respond to exposure to sustained levels of H(2)O(2) in a dose-response manner to H(2)O(2) concentration by activating the expression and activity of the antioxidant machinery, although the activation of the antioxidant defenses is not enough to avoid the appearance of oxidative damage. Of the two designs proposed, continuous exposure seems to be more appropriate to study the antioxidant response of HL60 cells to H(2)O(2).

Variegate porphyria induces plasma and neutrophil oxidative stress: effects of dietary supplementation with vitamins E and C.

Our aim was to analyse the influence of variegate porphyria (VP) on the antioxidant defenses and markers of oxidative damage and inflammation in plasma and neutrophils and the effects of dietary supplementation with vitamins E and C on these parameters in plasma, neutrophils and erythrocytes. Twelve women affected by VP and twelve pair-matched healthy control women participated in a double-blind crossover study. Each participant took 50 mg/d of vitamin E and 150 mg/d of vitamin C, or a placebo, for 6 months, by consuming an almond-based beverage as the vehicle. Women affected by VP presented higher C-reactive protein and malondialdehyde (MDA) circulating levels. Plasma antioxidant defenses were not different between porphyric and control women. Neutrophils from VP women presented decreased catalase (CAT) and glutathione reductase (GR) activities together with increased protein carbonyl levels. Reactive oxygen species (ROS) production from stimulated neutrophils was also higher in porphyric women than their controls. Dietary supplementation was effective in increasing alpha-tocopherol levels in neutrophils and in reducing MDA levels in plasma. Erythrocyte CAT and GR activities were enhanced by the enriched beverage only in the control subjects. In conclusion, women affected by VP present a situation of inflammation, plasma oxidative damage and neutrophils more primed to the oxidative burst, with decreased antioxidant activities and increased ROS production capabilities and protein oxidative damage. Dietary supplementation with vitamin E (50 mg/d) and vitamin C (150 mg/d) for 6 months decreased plasma oxidative damage and enhanced the erythrocyte activities of CAT and GR.

L-citrulline-malate influence over branched chain amino acid utilization during exercise.

Exhaustive exercise induces disturbances in metabolic homeostasis which can result in amino acid catabolism and limited L-arginine availability. Oral L-citrulline supplementation raises plasma L-arginine concentration and augments NO-dependent signalling. Our aim was to evaluate the effects of diet supplementation with L-citrulline-malate prior to intense exercise on the metabolic handle of plasma amino acids and on the products of metabolism of arginine as creatinine, urea and nitrite and the possible effects on the hormonal levels. Seventeen voluntary male pre-professional cyclists were randomly assigned to one of two groups: control or supplemented (6 g L-citrulline-malate 2 h prior exercise) and participated in a 137-km cycling stage. Blood samples were taken in basal conditions, 15 min after the race and 3 h post race (recovery). Most essential amino acids significantly decreased their plasma concentration as a result of exercise; however, most non-essential amino acids tended to significantly increase their concentration. Citrulline-malate ingestion significantly increased the plasma concentration of citrulline, arginine, ornithine, urea, creatinine and nitrite (p < 0.05) and significantly decreased the isoleucine concentration from basal measures to after exercise (p < 0.05). Insulin levels significantly increased after exercise in both groups (p < 0.05) returning to basal values at recovery. Growth hormone increased after exercise in both groups, although the increase was higher in the citrulline-malate supplemented group (p < 0.05). L-citrulline-malate supplementation can enhance the use of amino acids, especially the branched chain amino acids during exercise and also enhance the production of arginine-derived metabolites such as nitrite, creatinine, ornithine and urea.

A Novel Ex Vivo Model of Aortic Valve Calcification. A Preliminary Report.

Background: No pharmacological treatment exists to prevent or stop the calcification process of aortic valves causing aortic stenosis. The aim of this study was to develop a robust model of induced calcification in whole aortic valve leaflets which could be suitable for studies of the basic mechanisms and for testing potentially inhibitory drugs. Methods: Pig hearts were obtained from a commercial abattoir. The aortic valve leaflets were dissected free and randomized between experimental groups. Whole leaflets were cultured in individual wells. Two growth media were used for cultivation: standard growth medium and an antimyofibroblastic growth medium. The latter was employed to inhibit contraction of the leaflet into a ball-like structure. Calcification was induced in the growth medium by supplementation with an osteogenic medium. Leaflets were cultivated for four weeks and medium was changed every third day. To block calcification, the inhibitor SNF472 (a formulation of the hexasodium salt of myo-inositol hexaphosphate hexasodium salt) was used at concentrations between 1 and 100 µM. After cultivation for four weeks the leaflets were snap frozen in liquid nitrogen and kept at -80 °C until blind assessment of the calcium amount in leaflets by inductively coupled plasma optical emission spectroscopy. For statistical analysis, a Kruskal-Wallis test with Dunn's post-test was applied. Results: Osteodifferentiation with calcium accumulation was in principle absent when standard medium was used. However, when the antimyofibroblastic medium was used, a strong calcium accumulation was induced (p = 0.006 compared to controls), and this was blocked in a dose-dependent manner by the calcification inhibitor SNF472 (p = 0.008), with an EC50 of 3.3 µM. Conclusion: A model of experimentally induced calcification in cultured whole leaflets from porcine aortic valves was developed. This model can be useful for studying the basic mechanisms of valve calcification and to test pharmacological approaches to inhibit calcification.

Oral Administration of Sodium Nitrate to Metabolic Syndrome Patients Attenuates Mild Inflammatory and Oxidative Responses to Acute Exercise.

The beneficial effects of exercise for the treatment and prevention of metabolic syndrome pathologies have been related to its anti-inflammatory and antioxidant effects. Dietary nitrate supplementation is an emerging treatment strategy to alleviate the symptoms of metabolic syndrome affections and to improve vascular function. In this double-blind crossover trial, metabolic syndrome patients performed two exercise tests for 30 min at 60-70% maximal heart rate after the intake of a placebo or a nitrate-enriched beverage. Acute exercise increased the plasma concentration of TNFα, intercellular adhesion molecule ICAM1, PGE1, PGE2 and the newly detected 16-hydroxypalmitic acid (16-HPAL) in metabolic syndrome patients. The cytokine and oxylipin production by peripheral blood mononuclear cells (PBMCs) and neutrophils could be responsible for the plasma concentrations of TNFα and IL6, but not for the plasma concentration of oxylipins nor its post-exercise increase. The intake of sodium nitrate 30 min before exercise increased the concentration of nitrate and nitrite in the oral cavity and plasma and reduced the oxygen cost of exercise. Additionally, nitrate intake prevented the enhancing effects of acute exercise on the plasma concentration of TNFα, ICAM1, PGE1, PGE2 and 16-HPAL, while reducing the capabilities of PBMCs and neutrophils to produce oxylipins.

A novel assay to measure calcification propensity: from laboratory to humans.

Cardiovascular calcification (CVC) contributes to morbidity and mortality in patients undergoing dialysis. We examined the pharmacodynamic effects of SNF472, a calcification inhibitor, on plasma calcium phosphate crystallization using spectrometric measurements, and its correlations with effects on CVC in rats or humans. Rats (N = 38) injected with vitamin D (days 1-3) to induce CVC were infused with saline or SNF472 (days 1-12). Inhibition of CVC was 50-65% with SNF472 3 mg/kg and ~ 80% with SNF472 10 or 30 mg/kg. SNF472 dose-dependently inhibited calcium phosphate crystallization, which correlated with inhibition of CVC (r = 0.628, P = 0.005). In patients with calciphylaxis (N = 14), infusion of SNF472 (~ 7 mg/kg) during hemodialysis for 12 weeks inhibited calcium phosphate crystallization by nearly 70%. In patients with CVC (N = 274), infusion of SNF472 during hemodialysis for 52 weeks inhibited calcium phosphate crystallization (placebo: 15%; 300 mg: 61%; 600 mg: 75%), which correlated with inhibition of CVC (r = 0.401, P = 0.003). These findings show a direct correlation between inhibition of calcium phosphate crystallization in plasma and inhibition of CVC both in a rat model and in humans, supporting the use of the pharmacodynamic assay in clinical trials as a potentially predictive tool to evaluate the activity of calcification inhibitors.

Calorie Restriction Improves Physical Performance and Modulates the Antioxidant and Inflammatory Responses to Acute Exercise.

Our aim was to characterize the effects of calorie restriction on the anthropometric characteristics and physical performance of sportsmen and to evaluate the effects of calorie restriction and acute exercise on mitochondria energetics, oxidative stress, and inflammation. Twenty volunteer taekwondo practitioners undertook a calorie restriction of 30-40% on three alternate days a week for one month. Eleven volunteer sportsmen participated as controls. Both groups performed an energy efficiency test to evaluate physical performance, and samples were taken before and after exercise. The total weight of participants significantly decreased (5.9%) after calorie restriction, while the efficiency of work and the contributions of fat to obtain energy were enhanced by calorie restriction. No significant differences induced by acute exercise were observed in individual non-esterified fatty acid percentage or oxidative stress markers. Calorie restriction downregulated the basal gene expression of nitric oxide synthase, antioxidant enzymes, mitochondrial uncoupling proteins, and repairing stress proteins, but it enhanced the expression of sirtuins in peripheral blood mononuclear cells. In conclusion, one month of calorie restriction decreases body weight and increases physical performance, enhancing energy efficiency, moderating the antioxidant and inflammatory basal gene expression, and influencing its response to acute exercise.

Cyclooxygenase-2 Inhibitors as a Therapeutic Target in Inflammatory Diseases.

Inflammation plays a crucial role in the development of many complex diseases and disorders including autoimmune diseases, metabolic syndrome, neurodegenerative diseases, and cardiovascular pathologies. Prostaglandins play a regulatory role in inflammation. Cyclooxygenases are the main mediators of inflammation by catalyzing the initial step of arachidonic acid metabolism and prostaglandin synthesis. The differential expression of the constitutive isoform COX-1 and the inducible isoform COX-2, and the finding that COX-1 is the major form expressed in the gastrointestinal tract, lead to the search for COX-2-selective inhibitors as anti-inflammatory agents that might diminish the gastrointestinal side effects of traditional non-steroidal anti-inflammatory drugs (NSAIDs). COX-2 isoform is expressed predominantly in inflammatory cells and decidedly upregulated in chronic and acute inflammations, becoming a critical target for many pharmacological inhibitors. COX-2 selective inhibitors happen to show equivalent efficacy with that of conventional NSAIDs, but they have reduced gastrointestinal side effects. This review would elucidate the most recent findings on selective COX-2 inhibition and their relevance to human pathology, concretely in inflammatory pathologies characterized by a prolonged pro-inflammatory status, including autoimmune diseases, metabolic syndrome, obesity, atherosclerosis, neurodegenerative diseases, chronic obstructive pulmonary disease, arthritis, chronic inflammatory bowel disease and cardiovascular pathologies.

Therapeutic Effects of Hyperbaric Oxygen in the Process of Wound Healing.

Chronic and non-healing wounds, especially diabetic foot ulcers and radiation injuries, imply remarkable morbidity with a significant effect on the quality of life and a high sanitary cost. The management of these wounds requires complex actions such as surgical debris, antibiotic treatment, dressings and even revascularization. These wounds are characterized by poor oxygen supply resulting in inadequate oxygenation of the affected tissue. The adjuvant treatment with hyperbaric oxygen therapy (HBOT) may increase tissue oxygenation favoring the healing of wounds which do not respond to the usual clinical care. The increase in the partial pressure of oxygen contributes to cover the energy demands necessary for the healing process and reduces the incidence of infections. Moreover, the increase in oxygen leads to the production of reactive species with hormetic activity, acting on signaling pathways that modulate the synthesis of inflammation mediators, antioxidants and growth factors which can contribute to the healing process. Studies performed with cell cultures and in animal models seem to demonstrate the beneficial effects of HBOT. However, clinical trials do not show such conclusive results; thus, additional randomized placebo-controlled studies are necessary to determine the real efficacy of HBOT and the mechanism of action for various types of wounds.

Peripheral Blood Mononuclear Cells Antioxidant Adaptations to Regular Physical Activity in Elderly People.

Regular physical activity prescription is a key point for healthy aging and chronic disease management and prevention. Our aim was to evaluate the antioxidant defense system and the mitochondrial status in peripheral blood mononuclear cells (PBMCs) and the level of oxidative damage in plasma in active, intermediate and inactive elderly. In total, 127 healthy men and women >55 years old participated in the study and were classified according on their level of declared physical activity. A more active lifestyle was accompanied by lower weight, fat mass and body mass index when compared to a more sedentary life-style. Active participants exhibited lower circulating PBMCs than inactive peers. Participants who reported higher levels of exercise had increased antioxidant protein levels when compared to more sedentary partakers. Carbonylated protein levels exhibited similar behavior, accompanied by a significant raise in expression of cytochrome c oxidase subunit IV in PBMCs. No significant changes were found in the activities of antioxidant enzymes and in the expression of structural (MitND5) and mitochondrial dynamic-related (PGC1α and Mitofusins1/2.) proteins. Active lifestyle and daily activities exert beneficial effects on body composition and it enhances the antioxidant defenses and oxidative metabolism capabilities in PBMCs from healthy elderly.