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Nanoparticles have a promising future in biomedical applications and knowing whether they affect ex vivo vascular reactivity is a necessary step before their use in patients. In this study, we have evaluated the vascular effect of cerium dioxide nanoparticles (CeONPs) on the human saphenous vein in response to relaxing and contractile agonists. In addition, we have measured the protein expression of key enzymes related to vascular homeostasis and oxidative stress. We found that CeONPs increased expression of both SOD isoforms, and the consequent reduction of superoxide anion would enhance the bioavailability of NO explaining the increased vascular sensitivity to sodium nitroprusside in the presence of CeONPs. The NOX4 reduction induced by CeONPs may lead to lower HO synthesis associated with vasodilation through potassium channels explaining the lower vasodilation to bradykinin. In addition, we showed for the first time, that CeONPs increase the expression of ACE2 in human saphenous vein, and it may be the cause of the reduced contraction to angiotensin II. Moreover, we ruled out that CeONPs have effect on the protein expression of eNOS, sGC, BKca channels and angiotensin II receptors or modify the vascular response to noradrenaline, endothelin-1 and TXA analogue. In conclusion, CeONPs show antioxidant properties, and together with their vascular effect, they could be postulated as adjuvants for the treatment of cardiovascular diseases., This work was supported by Carlos III Health Institute (PI22/00424) and the European Regional Development Fund (ERDF ‘‘A way to build Europe’’ grant PI19/00838), by the Ministry of Health of the Valencian Regional Government (PROMETEO/2019/027), and by Universitat de València (UV-INV-AE-1544052).

Food protein or food-derived peptides may regulate blood glucose levels; however, studies have shown inconsistent results. The aim of the present study was to characterize subgroups of individuals with increased risk of type 2 diabetes (T2D) and to investigate the cardiometabolic effects of fish protein in the same subgroups. We first divided participants into high insuliniAUC and low insuliniAUC subjects based on their insulin incremental area under the curve (iAUC) levels after a 2 h oral glucose tolerance test (OGTT), and secondly based on whether they had received 5.2 g salmon fish protein or placebo for 8 weeks, in a previously conducted randomized controlled trial (RCT). We then profiled these groups by analyzing plasma metabolomics and peripheral blood mononuclear cell (PBMC) gene expression. Compared to the low insuliniAUC group, the high insuliniAUC group had higher plasma concentrations of monounsaturated fatty acids (MUFAs) and glycated proteins (GlycA) and lower concentrations of glycine and acetate. After intervention with fish protein compared to placebo, however, only acetate was significantly increased in the low insuliniAUC group. In conclusion, we identified metabolic biomarkers known to be associated with T2D; also, intervention with fish protein did not affect cardiometabolic risk markers in subgroups with increased risk of T2D.

Metformin is an effective drug against type 2 diabetes (T2D), a pathogenesis in which mitochondrial dysfunction is one of the main players. Thus, our first aim was to describe the effect of metform...

Liver fibrosis (LF) occurs as a result of persistent liver injury and can be defined as a pathologic, chronic, wound-healing process in which functional parenchyma is progressively replaced by fibrotic tissue. As a phenomenon involved in the majority of chronic liver diseases, and therefore prevalent, it exerts a significant impact on public health. This impact becomes even more patent given the lack of a specific pharmacological therapy, with LF only being ameliorated or prevented through the use of agents that alleviate the underlying causes. Hepatic stellate cells (HSCs) are fundamental mediators of LF, which, activated in response to pro-fibrotic stimuli, transdifferentiate from a quiescent phenotype into myofibroblasts that deposit large amounts of fibrotic tissue and mediate pro-inflammatory effects. In recent years, much effort has been devoted to understanding the mechanisms through which HSCs are activated or inactivated. Using cell culture and/or different animal models, numerous studies have shown that autophagy is enhanced during the fibrogenic process and have provided specific evidence to pinpoint the fundamental role of autophagy in HSC activation. This effect involves - though may not be limited to - the autophagic degradation of lipid droplets. Several hepatoprotective agents have been shown to reverse the autophagic alteration present in LF, but clinical confirmation of these effects is pending. On the other hand, there is evidence that implicates autophagy in several anti-fibrotic mechanisms in HSCs that stimulate HSC cell cycle arrest and cell death or prevent the generation of pro-fibrotic mediators, including excess collagen accumulation. The objective of this review is to offer a comprehensive analysis of published evidence of the role of autophagy in HSC activation and to provide hints for possible therapeutic targets for the treatment and/or prevention of LF related to autophagy. © 2021 The Authors. The Journal of Pathology published by John Wiley & Sons, Ltd. on behalf of The Pathological Society of Great Britain and Ireland.

Type 1 diabetes has been associated with oxidative stress. This study evaluates the rates of oxidative stress, mitochondrial function, leukocyte&ndash, endothelium interactions and adhesion molecules in type 1 diabetic patients. The study population consisted of 52 diabetic patients and 46 body-composition and age-matched controls. We assessed anthropometric and metabolic parameters, oxidative stress and mitochondrial function by evaluating reactive oxygen species (ROS) production, mitochondrial ROS production, mitochondrial membrane potential and superoxide dismutase (SOD) and catalase (CAT) expression in polymorphonuclear leukocytes from type 1 diabetic patients. In addition, we evaluated interactions between leukocytes and human umbilical vein endothelial cells (HUVEC), and serum expression of adhesion molecules (P-selectin, VCAM-1 and ICAM-1), proinflammatory cytokines (IL-6 and TNF&alpha, ) and myeloperoxidase (MPO). HbA1C and glucose levels were higher in diabetic patients than in control subjects, as expected. Mitochondrial function was altered and leukocyte&ndash, endothelium interactions were enhanced in diabetic patients, which was evident in the increase in total and mitochondrial ROS production, higher mitochondrial membrane potential, enhanced leukocyte rolling and adhesion, and decreased rolling velocity. Furthermore, we observed an increase in levels of adhesion molecules P-selectin, VCAM-1, and ICAM-1 in these subjects. In addition, type 1 diabetic patients exhibited an increase in proinflammatory mediators TNF&alpha, and MPO, and a decreased expression of SOD. The enhancement of leukocyte&ndash, endothelium interactions and proinflammatory markers correlated with glucose and HbA1Clevels. Mitochondrial alteration, oxidative stress, and enhanced leukocyte&ndash, endothelium interactions are features of type 1 diabetes and may be related to cardiovascular implications.

Objective In obese patients undergoing caloric restriction, there are several potential mechanisms involved in the improvement of metabolic outcomes. The present study further explores whether caloric restriction can modulate endoplasmic reticulum (ER) stress and mitochondrial function, as both are known to be mechanisms underlying inflammation and insulin resistance (IR) during obesity. Methods A total of 64 obese patients with BMI ≥35 kg/m2 underwent a dietary program consisting of 6 weeks of a very-low-calorie diet followed by 18 weeks of low-calorie diet. We evaluated changes in the metabolic and inflammatory markers -TNFα, hsCRP, complement component 3 (C3c), and retinol binding protein 4 (RBP4)-, in the ER stress markers and modulators -eIF2α-P, sXBP1, ATF6, JNK-P, CHOP, GRP78, and SIRT1-, and in mitochondrial function parameters -mitochondrial reactive oxygen species (mROS), glutathione peroxidase 1 (GPX1), cytosolic Ca2+, and mitochondrial membrane potential. Results The dietary intervention produced an 8.85% weight loss associated with enhanced insulin sensitivity, a less marked atherogenic lipid profile, and a decrease in systemic inflammation (TNFα, hsCRP) and adipokine levels (RBP4 and C3c). Chronic ER stress was significantly reduced (ATF6-CHOP, JNK-P) and expression levels of SIRT1 and GRP78 – a Ca2+-dependent chaperone – were increased and accompanied by the restoration of Ca2+ depots. Furthermore, mROS production and mitochondrial membrane potential improvement were associated with the up-regulation of the antioxidant enzyme GPX1. Conclusions Our data provide evidence that moderate weight loss attenuates systemic inflammation and IR and promotes the amelioration of ER stress and mitochondrial dysfunction, increasing the expression of chaperones, SIRT1 and antioxidant GPX1.

Summary Background & aims It is known that pinitol acts as a mediator of the insulin-signaling pathway, though little is known about its anti-inflammatory effect in human obesity. Therefore, this study aimed to evaluate the effect of pinitol on peripheral blood mononuclear cells (PBMCs) and visceral (VAT) and subcutaneous adipose tissues (SAT), focusing on the involvement of endoplasmic reticulum (ER) stress and sirtuin 1 (SIRT1). Methods In the intervention study, thirteen obese subjects consumed a pinitol-enriched beverage (PEB) for 12 weeks. In the ex vivo study, a biopsy of VAT and SAT was removed from thirty-four obese patients and incubated with D-pinitol for 48 h. Results The consumption of a PEB reduced circulating levels of IL6 and TNFα and increased SIRT1 protein expression in PBMCs. Ex vivo experiments showed a decline in gene expression and protein levels of IL6 and TNFα in SAT and a reduction in ER stress parameters (ATF6 and CHOP), while VAT markers remained unaltered. Differential gene expression profiles revealed an up-regulation of SIRT1 and insulin-signaling pathways in SAT with respect to VAT. Conclusions Our results suggests that pinitol down-regulates the inflammatory pathway which may lead to novel treatment options for obesity and its metabolic disorders.

There is growing focus on mitochondrial impairment and cardiovascular diseases (CVD) in type 2 diabetes (T2D), and the development of novel therapeutic strategies in this context. It is unknown whether mitochondrial-targeting antioxidants such as SS-31 protect sufficiently against oxidative damage in diabetes. We aimed to evaluate if SS-31 modulates SIRT1 levels and ameliorates leukocyte-endothelium interactions, oxidative stress and inflammation in T2D patients. Anthropometric and metabolic parameters were studied in 51 T2D patients and 57 controls. Production of mitochondrial reactive oxygen species (ROS), mitochondrial membrane potential, glutathione content, leukocyte-endothelium interactions, NFκB-p65, TNFα and SIRT1 levels was measured in leukocytes treated or not with SS-31. We observed increased mitochondrial ROS production that was restored by SS-31 treatment. SS-31 also increased mitochondrial membrane potential, glutathione content, SIRT1 levels and leukocyte rolling velocity and reduced rolling flux and adhesion in T2D patients. NFκB-p65 and TNFα, which were enhanced in diabetic patients, were also reduced by SS-31 treatment. Our results reveal that SS-31 exerts beneficial effects on the leukocytes of T2D patients by reducing oxidative stress, leukocyte-endothelium interactions, NFκB and TNFα and by increasing SIRT1 levels. These actions support its use as a potential agent against CVD risk.

Nanotechnology has had a significant impact on medicine in recent years, its application being referred to as nanomedicine. Nanoparticles have certain properties with biomedical applications; however, in some situations, they have demonstrated cell toxicity, which has caused concern surrounding their clinical use. In this review, we focus on two aspects: first, we summarize the types of nanoparticles according to their chemical composition and the general characteristics of their use in medicine, and second, we review the applications of nanoparticles in vascular alteration, especially in endothelial dysfunction related to oxidative stress. This condition can lead to a reduction in nitric oxide (NO) bioavailability, consequently affecting vascular tone regulation and endothelial dysfunction, which is the first phase in the development of cardiovascular diseases. Therefore, nanoparticles with antioxidant properties may improve vascular dysfunction associated with hypertension, diabetes mellitus, or atherosclerosis.

Since type 2 diabetes (T2D) is associated with oxidative stress and metformin has been shown to exert a protective role against the said stress, we wondered whether metformin treatment might also modulate endoplasmic reticulum (ER) stress and autophagy in leukocytes of T2D patients. We studied 53 T2D patients (37 of whom had been treated with metformin 1700 mg for at least 1 year) and 30 healthy volunteers. Leukocytes from both groups of T2D patients exhibited increased protein levels of 78-kDa glucose-regulated protein (GRP78) with respect to controls, whereas activating transcription factor 6 (ATF6) was enhanced specifically in nonmetformin-treated T2D, and (s-xbp1) and phosphorylated eukaryotic initiation factor 2α (p-eIF2α) increased only in the metformin-treated group. The autophagy markers beclin1 (becn1), autophagy-related 7 (atg7), and microtubule-associated protein 1A/1B-light chain 3II/I (LC3 II/I) increased in nonmetformin-treated T2D, and metformin treatment reduced mitochondrial superoxide and increased glutathione (GSH) levels. Our observations raise the question of whether metformin treatment could reduce oxidative stress and act as an ER stress modulator in T2D patients by promoting an adaptive unfolded protein response (s-xbp1 and p-eIF2α) in their leukocytes; this was in contrast with nonmetformin-treated patients whose response could be driven by the ATF6-dependent pro-apoptotic pathway. Further, our findings lead to us to form the hypothesis of an autophagy-dependent clearance of misfolded proteins in nonmetformin-treated T2D patients that could be repressed by metformin treatment.-Antioxid. Redox Signal. 28, 1562-1569.

Introduction Low testosterone levels in men are associated with type 2 diabetes and cardiovascular risk. However, the role of testosterone in mitochondrial function and leukocyte-endothelium interactions is unknown. Our aim was to evaluate the relationship between testosterone levels, metabolic parameters, oxidative stress, mitochondrial function, inflammation and leukocyte-endothelium interactions in type 2 diabetic patients. Materials and methods The study was performed in 280 male type 2 diabetic patients and 50 control subjects. Anthropometric and metabolic parameters, testosterone levels, reactive oxygen species (ROS) production, mitochondrial membrane potential, TNFα, adhesion molecules and leukocyte-endothelium cell interactions were evaluated. Results Testosterone levels were lower in diabetic patients. Total and mitochondrial ROS were increased and mitochondrial membrane potential, SOD and GSR expression levels were reduced in diabetic patients. TNFα, ICAM-1 and VCAM-1 levels, leukocyte rolling flux and adhesion were all enhanced in diabetic patients, while rolling velocity was reduced. Testosterone levels correlated negatively with glucose, HOMA-IR, HbA1c, triglycerides, nonHDL-c, ApoB, hs-CRP and AIP, and positively with HDL-c and ApoA1. The multivariable regression model showed that HDL-c, HOMA-IR and age were independently associated with testosterone. Furthermore, testosterone levels correlated positively with membrane potential and rolling velocity and negatively with ROS production, VCAM-1, rolling flux and adhesion. Conclusions Our data highlight that low testosterone levels in diabetic men are related to impaired metabolic profile and mitochondrial function and enhanced inflammation and leukocyte-endothelium cell interaction, which leaves said patients at risk of cardiovascular events.

Polycystic ovary syndrome (PCOS) is associated with insulin resistance, which can lead to metabolic syndrome (MetS). Oxidative stress and leukocyte-endothelium interactions are related to PCOS. Our aim was to evaluate whether the presence of MetS in PCOS patients can influence endoplasmic reticulum (ER) and oxidative stress and leukocyte-endothelium interactions.This was a prospective controlled study conducted in an academic medical center. The study population consisted of 148 PCOS women (116 without/32 with MetS) and 112 control subjects (87 without / 25 with MetS). Metabolic parameters, reactive oxygen species (ROS) production, ER stress markers (GRP78, sXBP1, ATF6), leukocyte-endothelium interactions, adhesion molecules (VCAM-1, ICAM-1, E-Selectin), TNF-α and IL-6 were determined.Total ROS, inflammatory parameters and adhesion molecules were enhanced in the presence of MetS (p0.05), and the PCOS+MetS group showed higher levels of IL-6 and ICAM-1 than controls (p0.05). Increased adhesion and leukocyte rolling flux were observed in PCOS and PCOS+MetS groups vs their respective controls (p0.05). GRP78 protein expression was higher in the PCOS groups (p0.05 vs controls) and sXBP1 was associated with the presence of MetS (p0.05 vs controls without MetS). Furthermore, PCOS+MetS patients exhibited higher GRP78 and ATF6 levels than controls and PCOS patients without MetS (p0.05). In PCOS women, HOMA-IR was positively correlated with ICAM-1 (r=0.501; p0.01), ROS (r=0.604; p0.01), rolling flux (r=0.455;p0.05) and GRP78 (r=0.574; p0.001).Our findings support the hypothesis of an association between altered metabolic status, increased ROS production, ER stress and leukocyte-endothelium interactions in PCOS, all of which are related to vascular complications.

Sodium&ndash, glucose co-transporter 2 inhibitors (iSGLT2) have been linked to a considerable reduction in cardiovascular risk in patients with type 2 diabetes (T2D), but the precise molecular mechanisms are still elusive. We aimed to evaluate the effects of the iSGLT2 empagliflozin on systemic inflammation and its potential antioxidant properties. This is an observational, prospective follow-up study of a cohort of fifteen patients with T2D who received 10 mg/day of empagliflozin according to standard clinical care. Measures at baseline, 12 and 24 weeks were taken. Metabolic and anthropometric parameters were evaluated. Production of mitochondrial superoxide, glutathione content, and glutathione s-reductase and catalase mRNA levels were measured in leukocytes. Serum levels of myeloperoxidase, hs-CRP and IL-10 were determined. In addition to decreased body weight and reduced glucose and HbA1c levels, we observed a reduction in superoxide production in leukocytes of diabetic patients and increased glutathione content, prominently after 24 weeks of empagliflozin treatment. Leukocyte expression of glutathione s-reductase and catalase, and serum levels of IL-10 were enhanced at 24 weeks of empagliflozin treatment. Concomitantly, reduced hs-CRP and myeloperoxidase levels were seen. This study provides evidence of the antioxidant and anti-inflammatory properties of empagliflozin treatment in humans, which may contribute to its beneficial cardiovascular effects.

Mitochondrial dysfunction has been shown to play a central role in the pathophysiology of type 2 diabetes (T2D), and mitochondria-targeted agents such as SS-31 are emerging as a promising strategy for its treatment. We aimed to study the effects of SS-31 on leukocytes from T2D patients by evaluating oxidative stress, endoplasmic reticulum (ER) stress and autophagy. Sixty-one T2D patients and 53 controls were included. Anthropometric and analytical measurements were performed. We also assessed reactive oxygen species (ROS) production, calcium content, the expression of ER stress markers GRP78, CHOP, P-eIF2&alpha, and autophagy-related proteins Beclin1, LC3 II/I, and p62 in leukocytes from T2D and control subjects treated or not with SS-31. Furthermore, we have evaluated the action of SS-31 on leukocyte-endothelium interactions. T2D patients exhibited elevated ROS concentration, calcium levels and presence of ER markers (GRP78 and CHOP gene expression, and GRP78 and P-eIF2&alpha, protein expression), all of which were reduced by SS-31 treatment. SS-31 also led to a drop in BECN1 gene expression, and Beclin1 and LC3 II/I protein expression in T2D patients. In contrast, the T2D group displayed reduced p62 protein levels that were restored by SS-31. SS-20 (with non-antioxidant activity) did not change any analyzed parameter. In addition, SS-31 decreased rolling flux and leukocyte adhesion, and increased rolling velocity in T2D patients. Our findings suggest that SS-31 exerts potentially beneficial effects on leukocytes of T2D patients modulating oxidative stress and autophagy, and ameliorating ER stress.

Obese individuals without metabolic comorbidities are categorized as metabolically healthy obese (MHO). MicroRNAs (miRNAs) may be implicated in MHO. This cross-sectional study explores the link between circulating miRNAs and the main components of metabolic syndrome (MetS) in the context of obesity. We also examine oxidative stress biomarkers in MHO vs. metabolically unhealthy obesity (MUO). We analysed 3536 serum miRNAs in 20 middle-aged obese individuals: 10 MHO and 10 MUO. A total of 159 miRNAs were differentially expressed, of which, 72 miRNAs (45.2%) were higher and 87 miRNAs (54.7%) were lower in the MUO group. In addition, miRNAs related to insulin signalling and lipid metabolism pathways were upregulated in the MUO group. Among these miRNAs, hsa-miR-6796-5p and hsa-miR-4697-3p, which regulate oxidative stress, showed significant correlations with glucose, triglycerides, HbA1c and HDLc. Our results provide evidence of a pattern of differentially expressed miRNAs in obesity according to MetS, and identify those related to insulin resistance and lipid metabolism pathways.

Background & aims: Inositols are thought to be mediators of the insulin signalling pathway. We assessed the effects of inositols on glycaemic control in fasting and postprandial states and evaluated lipoprotein profile and LDL particle size in healthy population. Methods: A 12-week double-blind clinical trial was performed with forty healthy subjects administered either an inositol-enriched beverage (IEB) -containing 2.23 g of inositols in 250 ml- or a sucrose-sweetened beverage (SB) twice a day. Anthropometric measurements, fasting glucose levels, insulin and HOMA-IR index, lipoprotein profile and postprandial glucose concentrations (measured using the continuous glucose monitoring system (CGMS)) were recorded throughout the intervention period. Results: Following the 12-week trial subjects receiving the IEB exhibited a significant decrease in insulin, HOMA-IR and Apo B and an increase in LDL particle size, whereas the SB group showed increases in BMI and fasting glucose concentration. Analysis of postprandial glucose levels at breakfast, lunch and dinner revealed a mean reduction of glucose of approximate to 14% and a significant reduction in the area under the curve at 24 h after consumption of the IEB. Conclusions: Our results show that chronic IEB supplementation induces a significant improvement in carbohydrated metabolism parameters in healthy subjects. (C) 2015 Elsevier Ltd and European Society for Clinical Nutrition and Metabolism. All rights reserved.

Type 2 diabetes can increase the risk of skeletal muscle dysfunction and, consequently, that of cardiovascular diseases, including coronary artery disease and stroke. It is also related to a reduced capacity for exercise, but the underlying mechanism is only partially understood. There are several factors that contribute to the development of skeletal muscle dysfunction, of which oxidative stress and mitochondrial dysfunction are among the most important. This review discusses the role of oxidative stress in the development and progression of skeletal and cardiac dysfunction associated with diabetes. It also provides an overview of the potential actions of antioxidants in general and mitochondria-targeted antioxidants in particular in the treatment of muscle dysfunction in type 2 diabetes.

The rising prevalence of obesity and type 2 diabetes (T2D) is a growing concern worldwide. New discoveries in the field of metagenomics and clinical research have revealed that the gut microbiota plays a key role in these metabolic disorders. The mechanisms regulating microbiota composition are multifactorial and include resistance to stress, presence of pathogens, diet, cultural habits and general health conditions. Recent evidence has shed light on the influence of microbiota quality and diversity on mitochondrial functions. Of note, the gut microbiota has been shown to regulate crucial transcription factors, coactivators, as well as enzymes implicated in mitochondrial biogenesis and metabolism. Moreover, microbiota metabolites seem to interfere with mitochondrial oxidative/nitrosative stress and autophagosome formation, thus regulating the activation of the inflammasome and the production of inflammatory cytokines, key players in chronic metabolic disorders. This review focuses on the association between intestinal microbiota and mitochondrial function and examines the mechanisms that may be the key to their use as potential therapeutic strategies in obesity and T2D management.

Aim: The primary objective of this pilot study was to evaluate the effect of non-surgical periodontal treatment. The secondary aim was to evaluate the effect of dietary therapy on both parameters of oxidative stress in leukocytes and leukocyte-endothelial cell interactions in an obese population. Methods: This was a pilot study with a before-and-after design. Forty-nine obese subjects with periodontitis were randomized by means of the minimization method and assigned to one of two groups, one of which underwent dietary therapy while the other did not. All the subjects underwent non-surgical periodontal treatment. We determined periodontal, inflammatory and oxidative stress parameters&mdash, total reactive oxygen species (ROS), superoxide production, intracellular Ca2+, mitochondrial membrane potential and superoxide dismutase (SOD) activity. We also evaluated interactions between leukocytes and endothelium cells&mdash, velocity, rolling flux and adhesion&mdash, at baseline and 12 weeks after intervention. Results: Periodontal treatment improved the periodontal health of all the patients, with a reduction in serum retinol-binding protein 4 (RBP4), total superoxide production and cytosolic Ca2+ in leukocytes. In the patients undergoing dietary therapy, there were less leukocyte adhesion to the endothelium, an effect that was accompanied by a decrease in TNF&alpha, P-selectin and total ROS and an increase in SOD activity. Conclusions: Whereas non-surgical periodontal treatment induces an improvement in leukocyte homeostasis, dietary therapy as an adjuvant reduces systemic inflammation and increases antioxidant status which, in turn, modulates leukocyte-endothelium dynamics.

Background:To evaluate the relationship between leukocyte‐endothelial cellinteractions and oxidative stress parameters in non‐diabetic patients with differentgrades of obesity.Material and methods:For this cross‐sectional study, 225 subjects wererecruited from January 1, 2014 to December 31, 2016 and divided into groupsaccording to BMI (40 kg/m²). We determined clin-ical parameters, systemic inflammatory markers, soluble cellular adhesion mole-cules, leukocyte‐endothelium cell interactions—rolling flux, velocity and adhesion—, oxidative stress parameters—total ROS, total superoxide, glutathione—andmitochondrial membrane potential in leukocytes.Results:We verified that HOMA‐IR and hsCRP increased progressively as obe-sity developed, whereas A1c, IL6 and TNFαwere augmented in the BMI > 40kg/m² group. The cellular adhesion molecule sP‐selectin was increased in patientswith obesity, while sICAM, total ROS, total superoxide and mitochondrial mem-brane potential were selectively higher in the BMI > 40 kg/m² group. Obesityinduced a progressive decrease in rolling velocity and an enhancement of rollingflux and leukocyte adhesion.Conclusion:Our findings reveal that endothelial dysfunction markers are alteredin human obesity and are associated with proinflammatory cytokines andincreased oxidative stress parameters.

Objective We aim to assess the effect of metformin treatment on metabolic parameters, endothelial function and inflammatory markers in polycystic ovary syndrome (PCOS) subjects. Methods The study population consisted of 40 reproductive-age women with PCOS, who underwent treatment with metformin during a 12-week period, and their corresponding matched controls (n = 44). We evaluated endocrinological parameters, adhesion molecules (vascular cell adhesion molecule 1 (VCAM-1), intercellular cell adhesion molecule 1 (ICAM-1) and E-selectin) and proinflammatory cytokines (interleukin 6 (IL-6) and tumor necrosis factor alpha (TNFα)) in serum. In addition, interactions between human umbilical vein endothelial cells and polymorphonuclear (PMN) cells were assessed by flow chamber microscopy. In addition, a group of type 2 diabetes patients who underwent treatment with metformin during a 12-week period was incorporated into the study. Results Metformin produced beneficial effects on PCOS patients by decreasing polymorphonuclear (PMN) rolling flux and adhesion. It also decreased levels of ICAM-1, E-selectin, IL-6 and ΤΝFα. In addition, metformin induced an improvement of endocrine and anthropometric parameters in PCOS subjects by reducing glucose, follicle-stimulating hormone (FSH) and androstendione, and by increasing dehydroepiandrosterone-sulfate (DHEA-S). Metformin also had beneficial effects in type 2 diabetic subjects by reducing body weight, waist circumference and PMN adhesion, and by increasing PMN rolling velocity. Conclusion Our results highlight the modulating effect of metformin on leukocyte/endothelium interactions. These findings may explain the potential beneficial effect of metformin in reducing the risk of vascular events in PCOS patients and in insulin resistance conditions.

The aim of this study was to evaluate the efficacy of a bakery product enriched with dietary fibre and L-carnitine in countering glucose homoeostasis and insulin sensitivity in patients with and without MetS. This was a 12-week, double-blind, randomized placebo-controlled trial employing fifty-four subjects (28 with MetS and 26 without MetS). After one month (run-in period), subjects were divided into two intervention groups: one received dietary fibre (5.59 g of soluble fibre and 9.49 g of insoluble fibre) plus 2325 mg of L-carnitine enriched bread (n = 26) and the other received placebo bread (n = 28). Anthropometric measurements, biochemical parameters inflammatory markers and LDL subfractions were analysed before and after intervention. Patients with MetS who received the enriched product exhibited a significant decrease in insulin (12.4%), C-peptide (8.9%), HOMA-IR index (14.8%) and in percentage of small and dense LDL. In addition, a significant correlation was found between percentage of change in glucose and baseline glucose levels (r = -0.471; p = 0.017). No changes were detected in either lipid profile or inflammatory markers in patients with or without MetS at any point of the experimental period. In conclusion, dietary fibre plus L-carnitine supplementation improves hydrocarbonated metabolism parameters and insulin resistance, leading to a healthier atherogenic profile in patients with MetS. (C) 2015 Elsevier Ltd. All rights reserved.

Mitochondrial function and specifically its implication in cellular redox/oxidative balance is fundamental in controlling the life and death of cells, and has been implicated in a wide range of human pathologies. In this context, mitochondrial therapeutics, particularly those involving mitochondria-targeted antioxidants, have attracted increasing interest as potentially effective therapies for several human diseases. For the past 10 years, great progress has been made in the development and functional testing of molecules that specifically target mitochondria, and there has been special focus on compounds with antioxidant properties. In this review, we will discuss several such strategies, including molecules conjugated with lipophilic cations (e.g.,triphenylphosphonium) or rhodamine, conjugates of plant alkaloids, amino-acid- and peptide-based compounds, and liposomes. This area has several major challenges that need to be confronted. Apart from antioxidants and other redox active molecules, current research aims at developing compounds that arecapable of modulating other mitochondria-controlled processes, such as apoptosis and autophagy. Multiple chemically different molecular strategies have been developed as delivery tools that offer broad opportunities for mitochondrial manipulation. Additional studies, and particularly in vivo approaches under physiologically relevant conditions, are necessary to confirm the clinical usefulness of these molecules. This study was financed by grants PI13/1025, PI11/00327, CIBER CB06/04/0071, PROMETEOII/2014/035, and GV/2014/118 and by the European Regional Development Fund (ERDF). V.M.V. is recipient of a contract from the Ministry of Health of the Valencian Regional Government (CES10/030).

Background and Purpose The non-nucleoside analogue reverse transcriptase inhibitor efavirenz is associated with hepatic toxicity and metabolic disturbances. Although the mechanisms involved are not clear, recent evidence has pinpointed a specific mitochondrial action of efavirenz accompanied by the induction of an endoplasmic reticulum (ER) stress/unfolded protein response in human hepatic cells. The aim of this study was to further investigate the involvement of this organelle by evaluating efavirenz's effects in cells lacking functional mitochondria (rho°) and comparing them with those of the typical mitotoxic agent rotenone, a standard complex I inhibitor, and the ER stress inducer thapsigargin.

Type 2 diabetes (T2D) is a metabolic disorder characterized by hyperglycemia and insulin resistance in which oxidative stress is thought to be a primary cause. Considering that mitochondria are the main source of ROS, we have set out to provide a general overview on how oxidative stress is generated and related to T2D. Enhanced generation of reactive oxygen species (ROS) and oxidative stress occurs in mitochondria as a consequence of an overload of glucose and oxidative phosphorylation. Endoplasmic reticulum (ER) stress plays an important role in oxidative stress, as it is also a source of ROS. The tight interconnection between both organelles through mitochondrial-associated membranes (MAMs) means that the ROS generated in mitochondria promote ER stress. Therefore, a state of stress and mitochondrial dysfunction are consequences of this vicious cycle. The implication of mitochondria in insulin release and the exposure of pancreatic β-cells to hyperglycemia make them especially susceptible to oxidative stress and mitochondrial dysfunction. In fact, crosstalk between both mechanisms is related with alterations in glucose homeostasis and can lead to the diabetes-associated insulin-resistance status. In the present review, we discuss the current knowledge of the relationship between oxidative stress, mitochondria, ER stress, inflammation, and lipotoxicity in T2D.

Our aim was to investigate the role played by endothelial nitric oxide (NO) during acute vascular response to hypoxia, as a modulator of both vascular tone (through guanylate cyclase (sGC) activation) and mitochondrial O2 consumption (through competitive inhibition of cytochrome-c-oxydase (CcO)). Organ bath experiments were performed and O2 consumption (Clark electrode) was determined in isolated aorta, mesenteric and pulmonary arteries of rats and eNOS-knockout mice. All pre-contracted vessels exhibited a triphasic hypoxic response consisting of an initial transient contraction (not observed in vessels from eNOS-knockout mice) followed by relaxation and subsequent sustained contraction. Removal of the endothelium, inhibition of eNOS (by L-NNA) and inhibition of sGC (by ODQ) abolished the initial contraction without altering the other two phases. The initial hypoxic contraction was observed in the presence of L-NNA + NO-donors. L-NNA and ODQ increases O2 consumption in hypoxic vessels and increases the arterial tone in normoxia but not in hypoxia. When L-NNA + mitochondrial inhibitors (cyanide, rotenone or myxothiazol) were added, the increase in tone was similar in normoxic and hypoxic vessels, which suggests that inhibition of the binding of NO to reduced CcO restored the action of NO on sGC. Conclusion A complex equilibrium is established between NO, sGC and CcO in vessels in function of the concentration of O2: as O2 falls, NO inhibition of mitochondrial O2 consumption increases and activation of sGC decreases, thus promoting a rapid increase in tone in both pulmonary and systemic vessels, which is followed by the triggering of NO-independent vasodilator/vasoconstrictor mechanisms.

Introduction Testosterone undecanoate (T) treatment is common in female‐to‐male transsexuals (FtMs) but can induce impairment of mitochondrial function and oxidative stress. Aim The effect of T treatment on the mitochondrial function and redox state of leukocytes of FtMs subjects was evaluated. Methods This was an observational study conducted in a university hospital. Fifty‐seven FtMs were treated with T (1,000 mg) for 12 weeks, after which anthropometric and metabolic parameters and mitochondrial function were evaluated. Main Outcome Measures Anthropometric and metabolic parameters were evaluated. Mitochondrial function was studied by assessing mitochondrial oxygen (O 2 ) consumption, membrane potential, reactive oxygen species (ROS) production, glutathione levels (GSH), and the reduced glutathione/oxidized glutathione (GSH)/(GSSG) ratio in polymorphonuclear cells. Results T treatment led to mitochondrial impairment in FtMs as a result of a decrease in mitochondria O 2 consumption, the membrane potential, GSH levels, and the (GSH)/(GSSG) ratio and an increase in ROS production. Mitochondrial O 2 consumption and membrane potential negatively correlated with T levels, which was further confirmed that the T treatment had induced mitochondrial dysfunction. T also produced a significant increase in total testosterone, free androgenic index, and atherogenic index of plasma, and a decrease in sex hormone‐binding globulin and high‐density lipoprotein cholesterol. Conclusions Treatment of FtMs with T can induce impairment of mitochondrial function and a state of oxidative stress. This effect should be taken into account in order to modulate possible comorbidities in these patients. Victor VM, Rocha M, Banuls C, Rovira‐Llopis S, Gomez M, and Hernandez‐Mijares A. Mitochondrial impairment and oxidative stress in leukocytes after testosterone administration to female‐to‐male transsexuals. J Sex Med 2014;11:454–461.

Background & Aims ER stress is associated with a growing number of liver diseases, including drug-induced hepatotoxicity. The non-nucleoside analogue reverse transcriptase inhibitor Efavirenz, a cornerstone of the multidrug strategy employed to treat HIV1 infection, has been related to the development of various adverse events, including metabolic disturbances and hepatic toxicity, the mechanisms of which remain elusive. Recent evidence has pinpointed a specific mitochondrial effect of Efavirenz in human hepatic cells. This study assesses the induction of ER stress by Efavirenz in the same model and the implication of mitochondria in this process. Methods Primary human hepatocytes and Hep3B were treated with clinically relevant concentrations of Efavirenz and parameters of ER stress were studied using standard cell biology techniques. Results ER stress markers, including CHOP and GRP78 expression (both protein and mRNA), phosphorylation of eIF2α, and presence of the spliced form of XBP1 were upregulated. Efavirenz also enhanced cytosolic Ca 2+ content and induced morphological changes in the ER suggestive of ER stress. This response was greatly attenuated in cells with altered mitochondrial function (Rho°). The effects of Efavirenz on the ER, and particularly in regard to the mitochondrial involvement, differed from those elicited by a standard pharmacological ER stressor. Conclusions This newly discovered mechanism of cellular insult involving ER stress and UPR response may help comprehend the hepatic toxicity that has been associated with the widespread and life-long use of Efavirenz. In addition, the specificity of the actions of Efavirenz observed expands our knowledge of the mechanisms that trigger ER stress and shed some light on the mitochondria/ER interplay in drug-induced hepatic challenge.

This study assessed the effects of an inositol-enriched beverage (IEB) on blood glucose levels and inflammation status in subjects with an impaired fasting glucose (IFG) state according to body mass index (BMI). This was a 12 week, double-blind, randomized, controlled trial employing forty-four IFG subjects (fasting glucose levels 100-125 mg dl-1) that were divided into two intervention groups: one receiving a IEB (n = 24) containing mainly pinitol (2.0 g twice a day), and the other a sweetened beverage based on sucrose (SB; n = 20). Anthropometric and biochemical measurements, postprandial and fasting nocturnal glycaemia (continuous glucose monitoring system), and inflammatory parameters (IL-6 and TNF-α) were analyzed at baseline and after intervention according to BMI (non-obese: BMI < 30 kg m-2 or obese: BMI ≥ 30 kg m-2). Non-obese subjects who consumed IEB exhibited a significant decrease in insulin (-14.4%), HOMA-IR index (-15.1%) and percentage of glucose change after postprandial and fasting nocturnal periods (-10.0% and -10.3%, respectively) compared with the SB group (-2.35% and 10.2%, respectively) although they did not show any change in inflammatory cytokine levels. By contrast, obese subjects who consumed IEB showed a smaller variation in glucose levels after nocturnal fasting (-4.34%) and a marked decrease in IL-6 and TNF-α (p < 0.05). These findings support that consumption of IEB in prediabetic subjects produces a response that is dependent on BMI, with a clear improvement of insulin resistance and postprandial and nocturnal glycemia in non-obese subjects and a marked anti-inflammatory response in obese subjects.

Ceria-supported gold nanoparticles are prepared exhibiting peroxidase activity and acting as radical traps. Au/CeO2 shows a remarkable biocompatibility as demonstrated by measuring cellular viability, proliferation, and lack of apoptosis for two human cell lines (Hep3B and HeLa). The antioxidant activity of Au/CeO2 against reactive oxygen species (ROS) is demonstrated by studying the cellular behavior of Hep3B and HeLa in a model of cellular oxidative stress. It is determined that Au/CeO2 exhibits higher antioxidant activity than glutathione, the main cytosolic antioxidant compound, and its CeO2 carrier. Overall the result presented here shows the potential of implementing well-established nanoparticulated gold catalysts with remarkable biocompatibility in cellular biology., Financial support by the Spanish ministry of Science is (CTQ2006-06785 and CTQ2007-67805-AR07, PI10/1195, AP192/11) is gratefully acknowledged. RM also thanks Spanish Ministry of Education for a postgraduate scholarship. VMV is recipient of Regional Ministry of Health of the Valencian Community and Carlos III Health Institute contract (ACOMP/2012/045 and CES10/030). NA is a recipient of VALi+D contract (APOSTD/2011/049).

Background The measurement of small dense low-density lipoprotein (sdLDL) particles is relevant when assessing cardiovascular risk. However, there is as yet no referenced method for the determination of LDL subfractions or a standardized comparison of the methods currently available. Therefore, the aim of this study was to compare the pattern of LDL particles measured by polyacrylamide tube gel electrophoresis (PTGE) and polyacrylamide gradient gel electrophoresis (PGGE) and to correlate the results with triglyceride concentration. Materials and methods Serum samples were collected from 177 patients. Lipid profile and LDL particle size were assessed using PTGE and PGGE. Results Pearson correlation and kappa index revealed a very good agreement between the methods. There was 81.3% concordance for classification of sdLDL particles and 97.2% concordance for classification of large LDL when PTGE and PGGE were compared. LDL size correlated with triglyceride in subjects with triglyceride levels > 116 mg/dl, pointing to a high CAD risk, as reflected by their higher prevalence of pattern B. Conclusions PTGE correlates favourably and is in very good agreement with PGGE. The determination of LDL particle size may be an appropriate analytical procedure to estimate CAD risk in patients with high triglyceride levels.

Summary Background & aims Phytosterols (PS) lower LDLc, but their effect on metabolic syndrome (MetS) remains unknown. We evaluated whether low-fat milk enriched with PS improves cardiovascular risk factors in these patients. Methods A randomised parallel trial employing 24 moderate-hypercholesterolaemic MetS patients and consisting of two 3-month intervention phases. After a 3-month healthy diet, patients were divided into two intervention groups: diet (n = 10) and diet + PS (n = 14) (2 g/day). A control group of 24 moderate-hypercholesterolaemic patients without MetS (matched in age and BMI) underwent the same procedure. Results Neither dietary intervention nor enrichment of PS induced any improvement in the serum lipoprotein profile of MetS patients. By contrast, in the non-MetS population, a healthy diet effectively reduced TC, LDLc, non-HDLc and Apo B-100, with further decreases in TC (6.9%), LDLc (10.5%), non-HDLc (10.3%), Apo B-100 (6.2%) and Apo B-100/ApoA-I ratio (11.6%) being observed when PS were administered. No differences in LDL diameter, hsCRP or homocysteine were detected in any of the groups after consuming PS. This supplementation produced a significant increase in PS levels only in the non-MetS population. Conclusions PS therapy appears to be of little value to MetS patients, likely due to its reduced intestinal cholesterol absorption. The efficacy of PS as hypocholesterolaemic agents is thus limited.