The preventive and therapeutic consumption of meat enriched with carob fruit extract, rich in phenolic compounds, improves colonic antioxidant status in late-stage T2DM rats.

Oxidative stress is prevalent in Type 2 Diabetes Mellitus (T2DM) and has been associated with high meat consumption. Carob Fruit Extract (CFE) contains phenolic compounds, making it a suitable functional ingredient. Current study aims to evaluate the effect of CFE-enriched meat (CFE-meat) consumption on the antioxidant status of proximal and distal colon, and its relationship with fecal phenolic compounds in late-stage T2DM rats. Three groups of eight rats were studied: 1) D, fed control-meat; 2) ED, fed CFE-meat since the beginning of the study; 3) DE, fed CFE-meat after confirming T2DM. CFE-meat consumption reduces colonic oxidative stress mainly in the proximal section and helps to ameliorate glutathione metabolism and antioxidant score. Difference between ED and DE groups were associated with colon homeostasis and T2DM progression suggesting greater fermentation but lower absorption in the DE group. CFE appears as a promising tool to improve the antioxidant status observed in late-stage T2DM.

Analysis of immunohistomorphological changes in the colonic mucosa in a high-saturated fat and high-cholesterol fed streptozotocin/nicotinamide diabetic rat model.

The mucosal surface of gastrointestinal tract is lined with epithelial cells that establish an effective barrier between the lumen and internal environment through intercellular junctions, preventing the passage of potentially harmful substances. The "intestinal barrier function" consist of a defensive system that prevent the passage of antigens, toxins, and microbial products, while maintains the correct development of the epithelial barrier, the immune system and the acquisition of tolerance toward dietary antigens and intestinal microbiota. Intestinal morphology changes subsequent to nutritional variations, stress, aging or diseases, which can also affect the composition of the microbiota, altering the homeostasis of the intestine. A growing body of evidence suggests that alterations in intestinal barrier function favor the development of exaggerated immune responses, leading to metabolic endotoxemia, which seems to be the origin of many chronic metabolic diseases such as type 2 diabetes mellitus (T2DM). Although the mechanisms are still unknown, the interaction between dietary patterns, gut microbiota, intestinal mucosa, and metabolic inflammation seems to be a key factor for the development of T2DM, among other diseases. This chapter details the different techniques that allow evaluating the morphological and molecular alterations that lead of the intestinal barrier dysfunction in a T2DM experimental model. To induce both diabetic metabolic disturbances and gut barrier disruption, Wistar rats were fed a high-saturated fat and high-cholesterol diet and received a single dose of streptozotocin/nicotinamide. This animal model may contribute to clarify the understanding of the role of intestinal barrier dysfunction on the late-stage T2DM etiology.

Dietary Supplementation with a Cocoa-Carob Blend Modulates Gut Microbiota and Prevents Intestinal Oxidative Stress and Barrier Dysfunction in Zucker Diabetic Rats.

We have recently developed a cocoa-carob blend (CCB) rich in polyphenols with antidiabetic properties. In this study, we investigated whether its benefits could be related to gut health and gut microbiota (GM) composition and the likely phenolic metabolites involved. Zucker diabetic fatty rats were fed on a standard or a CCB-rich diet for 12 weeks. Intestinal barrier structure and oxidative and inflammatory biomarkers were analyzed in colonic samples. GM composition and phenolic metabolites were evaluated from feces. The results show that CCB improved mucin and tight-junction proteins and counteracted gut oxidative stress and inflammation by regulating sirtuin-1 and nuclear factor erythroid 2-related factor 2 (Nrf2) levels. CCB also modulated the composition of the GM, showing increases in Akkermansia and Bacteroides and decreases in Ruminococcus genera. Correlation analysis strengthened the associations between these genera and improved pathological variables in diabetic animals. Moreover, 12 phenolic metabolites were identified in CCB feces, being2,3-dihydroxybenzoic and 3,4,5-trihydroxybenzoic acids significantly associated with increased levels of Akkermansia and Oscillospira genera. Our findings support the potential use of CCB to prevent intestinal damage and dysbiosis in T2D, which would help to delay the progression of this pathology.

The bitter taste receptor (TAS2R) agonist denatonium promotes a strong relaxation of rat corpus cavernosum.

Bitter taste receptors (TAS2R) are found in numerous extra-oral tissues, including smooth muscle (SM) cells in both vascular and visceral tissues. Upon activation, TAS2R stimulate the relaxation of the SM. Nitric oxide (NO)/cyclic guanosine monophosphate (cGMP) signaling pathway is involved in penile erection, and type 5 phosphodiesterase (PDE5) inhibitors, a cGMP-specific hydrolase are used as first-line treatments for erectile dysfunction (ED). Nevertheless, PDE5 inhibitors are ineffective in a considerable number of patients, prompting research into alternative pharmacological targets for ED. Since TAS2R agonists regulate SM contractility, this study investigates the role of TAS2Rs in rat corpus cavernosum (CC). We performed immunohistochemistry to detect TAS2R10, isometric force recordings for TAS2R agonists denatonium and chloroquine, the slow-release H2S donor GYY 4137, the NO donor SNAP, the ß-adrenoceptor agonist isoproterenol and electrical field stimulation (EFS), as well as measurement of endogenous hydrogen sulfide (H2S) production. The immunofluorescence staining indicated that TAS2R10 was broadly expressed in the CC SM and to some extent in the nerve fibers. Denatonium, chloroquine, SNAP, and isoproterenol cause potent dose-dependent SM relaxations. H2S production was decreased by NO and H2S synthase inhibitors, while it was enhanced by denatonium. In addition, denatonium increased the relaxations induced by GYY 4137 and SNAP but failed to modify EFS- and isoproterenol-induced responses. These results suggest neuronal and SM TAS2R10 expression in the rat CC, where denatonium induces a strong SM relaxation per se and promotes the H2S- and NO-mediated inhibitory gaseous neurotransmission. Thus, TAS2R10 might represent a valuable therapeutic target in ED.

Proanthocyanidins: Impact on Gut Microbiota and Intestinal Action Mechanisms in the Prevention and Treatment of Metabolic Syndrome.

The metabolic syndrome (MS) is a cluster of risk factors, such as central obesity, hyperglycemia, dyslipidemia, and arterial hypertension, which increase the probability of causing premature mortality. The consumption of high-fat diets (HFD), normally referred to high-saturated fat diets, is a major driver of the rising incidence of MS. In fact, the altered interplay between HFD, microbiome, and the intestinal barrier is being considered as a possible origin of MS. Consumption of proanthocyanidins (PAs) has a beneficial effect against the metabolic disturbances in MS. However, there are no conclusive results in the literature about the efficacy of PAs in improving MS. This review allows a comprehensive validation of the diverse effects of the PAs on the intestinal dysfunction in HFD-induced MS, differentiating between preventive and therapeutic actions. Special emphasis is placed on the impact of PAs on the gut microbiota, providing a system to facilitate comparison between the studies. PAs can modulate the microbiome toward a healthy profile and strength barrier integrity. Nevertheless, to date, published clinical trials to verify preclinical findings are scarce. Finally, the preventive consumption of PAs in MS-associated dysbiosis and intestinal dysfunction induced by HFD seems more successful than the treatment strategy.

Metabolic regulation of (-)-epicatechin and the colonic metabolite 2,3-dihydroxybenzoic acid on the glucose uptake, lipid accumulation and insulin signalling in cardiac H9c2 cells.

Epicatechin (EC) and main colonic phenolic acids derived from flavonoid intake have been suggested to exert healthful effects, although their mechanism of action remains unknown. Heart damage is highly prevalent in metabolic diseases, and the failure of this organ is a major cause of death worldwide. In this study, the modulation of the energy metabolism and insulin signalling by the mentioned compounds in cardiac H9c2 cells was evaluated. Incubation of cells with EC (1-20 µM) and 2,3-dihydroxybenzoic acid (DHBA, 10 µM) reduced glucose uptake, and both compounds decreased lipid accumulation at concentrations higher than 0.5 µM. EC and DHBA also increased the tyrosine phosphorylated and total insulin receptor (IR) levels, and activated the phosphatidylinositol 3-kinase (PI3K)/protein kinase B (AKT) pathway in cardiac H9c2 cells. Interestingly, EC and DHBA did not modify glucose transporters (SGLT-1 and GLUT-1) levels, and increased GLUT-4 values. In addition, EC and DHBA decreased cluster of differentiation 36 (CD36) and fatty acid synthase (FAS) values, and enhanced carnitine palmitoyl transferase 1 (CPT1) and proliferator activated receptor α (PPARα) levels. By using specific inhibitors of AKT and 5'-AMP-activated protein kinase (AMPK), the participation of both proteins in EC- and DHBA-mediated regulation on glucose uptake and lipid accumulation was shown. Taken together, EC and DHBA modulate glucose uptake and lipid accumulation via AKT and AMPK, and reinforce the insulin signalling by activating key proteins of this pathway in H9c2 cells.

In vitro inhibition of phosphodiesterase type 4 enhances rat corpus cavernosum nerve-mediated relaxation induced by gasotransmitters.

AIMS: Nitric oxide (NO) and hydrogen sulfide (H2S) are involved in nerve-mediated corpus cavernosum (CC) relaxation. Expression of phosphodiesterase type 5 (PDE5) and type 4 (PDE4), cyclic guanosine monophosphate (cGMP)- and cyclic adenosine monophosphate (cAMP)-specific, respectively, has been described and PDE5- and PDE4-inhibitors induce cavernous smooth muscle relaxation. Whereas the NO/cGMP signaling pathway is well established in penile erection, the cAMP-mediated mechanism is not fully elucidated. The aim of this study is to investigate the localization and the functional significance of PDE4 in rat CC tone regulation. MAIN METHODS: We performed immunohistochemistry for the detection of the PDE4A isoenzyme. Isometric tension recordings for roflumilast and tadalafil, PDE4 and PDE5 inhibitors, respectively, electrical field stimulation (EFS) and ß-adrenoceptor agonist isoproterenol and endogenous H2S production measurement. KEY FINDINGS: A marked PDE4A expression was detected mainly localized in the nerve cells of the cavernous smooth muscle. Furthermore, roflumilast and tadalafil exhibited strong corpus cavernous relaxations. Endogenous H2S production was decreased by NO and H2S synthase inhibitors and increased by roflumilast. Isoproterenol- and EFS-induced relaxations were increased by roflumilast. SIGNIFICANCE: These results indicate that PDE4A is mainly expressed within the nerves cells of the rat CC, where roflumilast induces a potent corpus cavernous relaxation per se and potentiates the response induced by ß-adrenoceptor activation. The fact that roflumilast enhances H2S production, as well as EFS-elicited responses suggests that PDE4 inhibitors modulate, in a positive feedback fashion, nerve-mediated relaxation induced by gasotransmitters, thus indicating a key role for neuronal PDE4 in penile erection.

Supplementation with a Cocoa-Carob Blend, Alone or in Combination with Metformin, Attenuates Diabetic Cardiomyopathy, Cardiac Oxidative Stress and Inflammation in Zucker Diabetic Rats.

Diabetic cardiomyopathy (DCM) is one of the main causes of mortality among diabetic patients, with oxidative stress and inflammation major contributors to its development. Dietary flavonoids show strong antioxidant and anti-inflammatory activities, although their potential additive outcomes in combination with antidiabetic drugs have been scarcely explored. The present study investigates the cardioprotective effects of a cocoa-carob blend (CCB) diet, rich in flavonoids, alone or in combination with metformin, in the development of DCM. Zucker diabetic fatty rats (ZDF) were fed with a CCB rich-diet or a control diet, with or without metformin for 12 weeks. Glucose homeostasis, cardiac structure and function, and oxidative and inflammatory biomarkers were analysed. CCB improved glucose homeostasis, and mitigated cardiac dysfunction, hypertrophy, and fibrosis in ZDF rats. Mechanistically, CCB counteracted oxidative stress in diabetic hearts by down-regulating NADPH oxidases, reducing reactive oxygen species (ROS) generation and modulating the sirtuin-1 (SIRT1)/ nuclear factor E2-related factor 2 (Nrf2) signalling pathway, overall improving antioxidant defence. Moreover, CCB suppressed inflammatory and fibrotic reactions by inhibiting nuclear factor kappa B (NFκB) and pro-inflammatory and pro-fibrotic cytokines. Noteworthy, several of these effects were further improved in combination with metformin. Our results demonstrate that CCB strongly prevents the cardiac remodelling and dysfunction observed in diabetic animals, highlighting its potential, alone or in adjuvant therapy, for treating DCM.

Functional Meat Products as Oxidative Stress Modulators: A Review.

High meat consumption has been associated with increased oxidative stress mainly due to the generation of oxidized compounds in the body, such as malondialdehyde, 4-hydroxy-nonenal, oxysterols, or protein carbonyls, which can induce oxidative damage. Meat products are excellent matrices for introducing different bioactive compounds, to obtain functional meat products aimed at minimizing the pro-oxidant effects associated with high meat consumption. Therefore, this review aims to summarize the concept and preparation of healthy and functional meat, which could benefit antioxidant status. Likewise, the key strategies regarding meat production and storage as well as ingredients used (e.g., minerals, polyphenols, fatty acids, walnuts) for developing these functional meats are detailed. Although most effort has been made to reduce the oxidation status of meat, newly emerging approaches also aim to improve the oxidation status of consumers of meat products. Thus, we will delve into the relation between functional meats and their health effects on consumers. In this review, animal trials and intervention studies are discussed, ascertaining the extent of functional meat products' properties (e.g., neutralizing reactive oxygen species formation and increasing the antioxidant response). The effects of functional meat products in the frame of diet-gene interactions are analyzed to 1) discover target subjects that would benefit from their consumption, and 2) understand the molecular mechanisms that ensure precision in the prevention and treatment of diseases, where high oxidative stress takes place. Long-term intervention-controlled studies, testing different types and amounts of functional meat, are also necessary to ascertain their positive impact on degenerative diseases.

Preventive effect of cocoa flavanols against glucotoxicity-induced vascular inflammation in the arteria of diabetic rats and on the inflammatory process in TNF-α-stimulated endothelial cells.

Hyperglycaemia induces a vascular inflammatory process that is a critical event in cardiovascular disease in type 2 diabetes. Cocoa and its flavanols have been widely investigated for its antioxidant and anti-inflammatory properties, and several clinical and pre-clinical studies support their vascular benefits. However, the effects of cocoa flavanols on vascular inflammation in diabetes remains to be elucidated. Herein, we evaluated the anti-inflammatory effect of a cocoa-rich diet on the aortas of Zucker diabetic fatty (ZDF) rats. Moreover, the potential role of flavanol-derived colonic metabolites to modulate the adhesion and inflammatory processes were also evaluated using TNF-α-stimulated endothelial cells. Results demonstrate that cocoa attenuates the levels of phospho-p65-nuclear factor-kappaB (NF-κB) and the expression of inflammatory factors including intercellular adhesion molecule-1 (ICAM-1), vascular adhesion molecule-1 (VCAM-1) and inducible nitric oxide synthase in the aortas of ZDF rats. Experiments with endothelial cells further confirm that a mix of flavanol-derived colonic metabolites effectively down-regulate the levels of p-p65-NF-κB and the cell adhesion molecules ICAM-1 and VCAM-1, preventing thus the increase of monocyte-endothelial adhesion induced by TNF-α. These novel data provide the first evidence of the relevant role of cocoa and their flavanol-derived metabolites to avoid the development of endothelial inflammation and diabetic complications.

(-)-Epicatechin and the colonic metabolite 2,3-dihydroxybenzoic acid protect against high glucose and lipopolysaccharide-induced inflammation in renal proximal tubular cells through NOX-4/p38 signalling.

Chronic hyperglycaemia and inflammation are present in diabetes and both processes have been related to the pathogenesis of diabetic kidney disease. Epicatechin (EC) and main colonic phenolic acids derived from flavonoid intake, such as 2,3-dihydroxybenzoic acid (DHBA), 3,4-dihydroxyphenylacetic acid (DHPAA) and 3-hydroxyphenylpropionic acid (HPPA), have been suggested to exert beneficial effects in diabetes. This study was aimed at investigating whether the mentioned compounds could prevent inflammation in renal proximal tubular NRK-52E cells induced by high glucose and lipopolysaccharide (LPS). Pre-treatment of cells with EC and DHBA (5 µM) reverted the enhanced levels of pro-inflammatory cytokines, such as tumour necrosis factor-α (TNF-α), interleukin-6 (IL-6) and monocyte chemoattractant protein 1 (MCP-1), activated by high glucose and LPS. Additionally, EC and DHBA pre-incubation reduced the increased values of adhesion molecules, namely, intercellular cell adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1), as well as those of mitogen-activated protein kinases (MAPKs) [extracellular signal-regulated kinase (ERK), -c-jun N-terminal kinase (JNK) and -p38 protein kinase (p38)] activated by the high glucose and LPS challenge. Thus, in EC and DHBA pre-treated cells ICAM-1, p-ERK and p-JNK were returned to control values, and VCAM-1 and p-p38 levels were reduced by ∼20 and 25%, respectively, when compared to high glucose plus LPS-stimulated cells. Likewise, pre-treatment with EC and DHBA protected against high glucose plus LPS-triggered oxidative stress by preventing increased ROS and NADPH oxidase 4 (NOX-4) levels (∼25 and 45% reduction, respectively). By using specific inhibitors of p38 and NOX-4, the participation of both proteins in EC- and DHBA-mediated protection against inflammation and associated oxidative stress was shown. Taken together, EC and DHBA exert beneficial effects in renal proximal tubular cells, as they contribute to preventing the inflammatory-induced milieu and the accompanying redox imbalance, playing NOX-4/p38 a crucial role.

Can Meat and Meat-Products Induce Oxidative Stress?

High meat and meat-products consumption has been related to degenerative diseases. In addition to their saturated fatty acids and cholesterol contents, oxidation products generated during their production, storage, digestion, and metabolization have been largely implicated. This review begins by summarizing the concept of meat and meat-products by the main international regulatory agencies while highlighting the nutritional importance of their consumption. The review also dials in the controversy of white/red meat classification and insists in the need of more accurate classification based on adequate scores. Since one of the negative arguments that meat receives comes from the association of its consumption with the increase in oxidative stress, main oxidation compounds (malondialdehyde, thermaloxidized compounds, 4-hydroxy-nonenal, oxysterols, or protein carbonyls) generated during its production, storage, and metabolization, are included as a central aspect of the work. The review includes future remarks addressed to study the effects meat consumption in the frame of diet-gene interactions, stressing the importance of knowing the genetic variables that make individuals more susceptible to a possible oxidative stress imbalance or antioxidant protection. The importance of consumed meat/meat-products in the frame of a personalized nutrition reach in plant-food is finally highlighted considering the importance of iron and plant biophenols on the microbiota abundance and plurality, which in turn affect several aspects of our physiology and metabolism.

Cocoa diet modulates gut microbiota composition and improves intestinal health in Zucker diabetic rats.

Cocoa supplementation improves glucose metabolism in Zucker diabetic fatty (ZDF) rats via multiple mechanisms. Furthermore, cocoa rich-diets modify the intestinal microbiota composition both in humans and rats in healthy conditions. Accordingly, we hypothesized that cocoa could interact with the gut microbiota (GM) in ZDF rats, contributing to their antidiabetic effects. Therefore, here we investigate the effect of cocoa intake on gut health and GM in ZDF diabetic rats. Male ZDF rats were fed with standard (ZDF-C) or 10% cocoa-rich diet (ZDF-Co) during 10 weeks. Zucker Lean animals (ZL) received the standard diet. Colon tissues were obtained to determine the barrier integrity and the inflammatory status of the intestine and faeces were analysed for microbial composition, short-chain fatty acids (SCFA) and lactate levels. We found that cocoa supplementation up-regulated the levels of the tight junction protein Zonula occludens-1 (ZO-1) and the mucin glycoprotein and reduced the expression of pro-inflammatory cytokines such as tumour necrosis factor-α (TNF-α), interleukin-6 (IL-6) and monocyte chemoattractant protein 1 (MCP-1) in the colon of ZDF diabetic animals. Additionally, cocoa modulated the microbial composition of the ZDF rats to values similar to those of the lean group. Importantly, cocoa treatment increased the relative abundance of acetate-producing bacteria such as Blautia and prevented the increase in the relative amount of lactate-producing bacteria (mainly Enterococcus and Lactobacillus genera) in ZDF diabetic animals. Accordingly, the total levels of SCFA (mainly acetate) increased significantly in the faeces of ZDF-Co diabetic rats. Finally, modified GM was closely associated with improved biochemical parameters related to glucose homeostasis and intestinal integrity and inflammation. These findings demonstrate for the first time that cocoa intake modifies intestinal bacteria composition towards a healthier microbial profile in diabetic animals and suggest that these changes could be associated with the improved glucose homeostasis and gut health induced by cocoa in ZDF diabetic rats.

Differential contribution of Nox1, Nox2 and Nox4 to kidney vascular oxidative stress and endothelial dysfunction in obesity.

Oxidative stress-associated endothelial dysfunction is a key pathogenic factor underlying the microvascular complications of metabolic disease. NADPH oxidase (Nox) is a major source of oxidative stress in diabetic nephropathy and chronic kidney disease, despite Nox4 and Nox2 have been identified as relevant sources of vasodilator endothelial H2O2.The present study was sought to investigate the role of Nox enzymes in renal vascular oxidative stress and endothelial dysfunction in a rat model of genetic obesity. Endothelial function was assessed in intrarenal arteries of obese Zucker rats (OZR) and their counterparts lean Zucker rats (LZR) mounted in microvascular myographs, and superoxide (O2.-) and H2O2 production were measured. Impaired endothelium-dependent relaxations to acetylcholine (ACh) were associated to augmented O2.- generation, but neither ROS scavengers nor the Nox inhibitor apocynin significantly improved these relaxant responses in renal arteries of OZR. Whereas NO contribution to endothelial relaxations was blunted, catalase-sensitive non-NO non-prostanoid relaxations were enhanced in obese rats. Interestingly, NADPH-dependent O2.- production was augmented while NADPH-dependent H2O2 generation was reduced, and cytosolic and mitochondrial SOD were up-regulated in kidney of obese rats. Nox4 was down-regulated in renal arteries and Nox4-dependent H2O2 generation and endothelial relaxation were reduced in OZR. Up-regulation of both Nox2 and Nox1 was associated with augmented O2.- production but reduced H2O2 generation and blunted endothelial Nox2-derived H2O2-mediated in obese rats. Moreover, increased Nox1-derived O2.- contributed to renal endothelial dysfunction in OZR. In summary, the current data support a main role for Nox1-derived O2.- in kidney vascular oxidative stress and renal endothelial dysfunction in obesity, while reduced endothelial Nox4 expression associated to decreased H2O2 generation and H2O2-mediated vasodilatation might hinder Nox4 protective renal effects thus contributing to kidney injury. This suggests that effective therapies to counteract oxidative stress and prevent microvascular complications must identify the specific Nox subunits involved in metabolic disease.

Cocoa ameliorates renal injury in Zucker diabetic fatty rats by preventing oxidative stress, apoptosis and inactivation of autophagy.

Redox balance, autophagy and apoptosis are main processes involved in the development of diabetic nephropathy. Epidemiological and animal studies suggest that cocoa might reduce the risk of diabetic complications. However, the molecular mechanisms responsible for these potential preventive activities and whether cocoa exerts beneficial effects on dysregulated signalling pathways involved in cellular antioxidant defence, autophagy and apoptosis in the diabetic kidney remain largely unknown. Therefore, this work investigated the effect of a cocoa-rich diet on the mentioned processes in the renal cortex of Zucker Diabetic Fatty (ZDF) rats. Male ZDF rats were fed either a control or cocoa-rich diet (10%), and Zucker lean animals received the control diet (10-20 weeks-of-life). ZDF rats fed with cocoa decreased body weight and glucose and insulin levels, and improved renal function. Cocoa intake further prevented the enhanced renal cortical oxidative stress in diabetic rats by regulating the antioxidant defence system and close-related proteins to cytoprotection and cell response; thus, cocoa diminished oxidative markers (reactive oxygen species and carbonyl groups) and NADPH-oxidase-4 levels, and restored key enzymatic antioxidant activities (superoxide dismutase and catalase), nuclear-erythroid-2-related factor-2, and ERK-MAPK levels, as well as sirtuin-1/5'-AMP-activated-protein kinase signalling. Moreover, in ZDF rats cocoa-rich diet contributed to alleviation of the renal cortical injury through autophagy activation (p62 upregulation, and downregulation of beclin-1 and LC3), and inhibition of apoptosis (Bcl-xL stimulation and suppression of Bax and caspases-9 and -3). These findings provide the first in vivo evidence on the molecular mechanisms of cocoa to circumvent renal cortical damage that involve improvement of antioxidant competences, stimulation of autophagy and suppression of apoptosis in ZDF rats.

Bladder Dysfunction in an Obese Zucker Rat: The Role of TRPA1 Channels, Oxidative Stress, and Hydrogen Sulfide.

PURPOSE: This study investigates whether functionality and/or expression changes of transient receptor potential vanilloid 1 (TRPV1) and transient receptor potential ankyrin 1 (TRPA1) channels, oxidative stress, and hydrogen sulfide (H2S) are involved in the bladder dysfunction from an insulin-resistant obese Zucker rat (OZR). MATERIALS AND METHODS: Detrusor smooth muscle (DSM) samples from the OZR and their respective controls, a lean Zucker rat (LZR), were processed for immunohistochemistry for studying the expression of TRPA1 and TRPV1 and the H2S synthase cystathionine beta-synthase (CBS) and cysthathionine-γ-lyase (CSE). Isometric force recordings to assess the effects of TRPA1 agonists and antagonists on DSM contractility and measurement of oxidative stress and H2S production were also performed. RESULTS: Neuronal TRPA1 expression was increased in the OZR bladder. Electrical field stimulation- (EFS-) elicited contraction was reduced in the OZR bladder. In both LZR and OZR, TRPA1 activation failed to modify DSM basal tension but enhanced EFS contraction; this response is inhibited by the TRPA1 blockade. In the OZR bladder, reactive oxygen species, malondialdehyde, and protein carbonyl contents were increased and antioxidant enzyme activities (superoxide dismutase, catalase, GR, and GPx) were diminished. CSE expression and CSE-generated H2S production were also reduced in the OZR. Both TRPV1 and CBS expressions were not changed in the OZR. CONCLUSIONS: These results suggest that an increased expression and functionality of TRPA1, an augmented oxidative stress, and a downregulation of the CSE/H2S pathway are involved in the impairment of nerve-evoked DSM contraction from the OZR.

Dietary Cocoa Prevents Aortic Remodeling and Vascular Oxidative Stress in Diabetic Rats.

SCOPE: The aim of the present study is to investigate the potential protective effect of a cocoa-rich diet on functional and structural vascular alterations in diabetes and the mechanism involved. METHODS AND RESULTS: Male Zucker diabetic fatty (ZDF) rats are fed on a standard (ZDF-C) or cocoa-rich diet (ZDF-Co) from week 10 to 20 of life. Diabetic ZDF-C rats showed increased blood pressure and enhanced aortic stiffness, as demonstrated by the increased pulse pressure and the augmented aortic medial thickness with loss and disruption of elastic fibres. Interestingly, cocoa intake strongly avoided all these adverse effects and reduced aortic oxidative stress. Mechanistically, cocoa diet prevented sirtuin-1 (SIRT-1) depletion and increased NADPH oxidases (NOXs) and reactive oxygen species production as well as reduced active nuclear factor E2 related factor 2 (Nrf2) and their antioxidant products. CONCLUSION: The results demonstrate for the first time that a cocoa-rich diet strongly prevents aortic stiffening and remodeling in diabetic animals and avoids aortic oxidative stress. It is suggested that this effect could be mediated via its effects on SIRT-1, NOXs, and Nrf2.

Can Carob-Fruit-Extract-Enriched Meat Improve the Lipoprotein Profile, VLDL-Oxidation, and LDL Receptor Levels Induced by an Atherogenic Diet in STZ-NAD-Diabetic Rats?

Carob fruit extract (CFE) has shown remarkable in vitro antioxidant properties and reduces postprandial hyperglycemia and hyperlipidemia in healthy animals. Development of functional meat products that contain bioactive components are presented as a great nutritional strategy. Until now, the effect of the consumption of restructured meat enriched with CFE in a murine model of diabetes has not been investigated. The objective of this study was to evaluate the effect on glycemia, lipemia, lipoprotein profile, Ldlr, arylesterase (AE), and very low-density lipoproteins (VLDL) and liver oxidation in streptozotocin-nicotinamide (STZ-NAD) growing Wistar diabetic rats fed restructured meat in the frame of a high cholesterol/high saturated-fat diet. In the present study, three groups (D, ED and DE) were fed cholesterol-enriched (1.4% cholesterol and 0.2% cholic acid) and high saturated-fat diets (50% of total energy from fats and 20.4% from saturated fatty acids). Rats were subjected to a STZ-NAD administration at the 3rd week. Group D did not receive CFE, while ED and DE rat groups received CFE before and after the diabetic induction, respectively. After eight weeks, D rats showed hyperglycemia and hypercholesterolemia, an increased amount cholesterol-enriched VLDL (ß-VLDL), IDL and LDL particles and triglyceride-enriched HDL. ED and DE partially blocked the hypercholesterolemic induction with respect to D group (p < 0.001) and improved glycemia, cholesterol levels, lipoprotein profile, Ldlr, plasma AE activity and liver oxidation (p < 0.001). Fecal fat, moisture and excretion were higher while dietary digestibility was lower in ED and DE vs. D counterparts (p < 0.001). In conclusion, CFE-enriched meat shows, for the first time, hypoglycemic and hypolipidemic effects in STZ-NAD animals fed high cholesterol/high saturated-fat diets. Likewise, it manages to reverse possible diabetes lipoprotein alterations if CFE-enriched meat is consumed before pathology development or improves said modifications if Type 2 Diabetes Mellitus is already established.

Origanum majorana Essential Oil Inhalation during Neurofeedback Training Reduces Saliva Myeloperoxidase Activity at Session-1 in Bruxistic Patients.

Introduction: Bruxism affects teeth and provokes sleep alterations. We evaluated whether saliva Myeloperoxidase (MPO) activity could be reduced after 21 neurofeedback training (NO) sessions in Origanum majorana (AE) bruxistic-treated patients (n = 12 patients, 120 saliva samples). The term divergence evaluates cerebral activity, which was compared between bruxistic patients with/without this essential oil exposure during 21 NO training sessions (n = 12, n = 120 saliva samples). The nasal filter used allow us to vehiculizate this Origanum majorana essential oil in patients. MPO activity was measured in six training NO sessions (Session S1, 6, 12, 18, 21). We included a total of 104 patients and 500 saliva samples. Results: there was a fast reduction in MPO activity after NO session-1 in bruxistic patients; divergence (an index of NO training brain efficacy) is the difference in cerebral activity found between pre-training and post-training levels. Thus, Divergence can fluctuate during NO training before reaching a final calm state after many sessions (21). Repeated NO training lead to habituation in so far as cerebral activity as well as MPO activity after 21 training sessions. Origanum majorana essential oil modulates cerebral activity at certain training sessions in bruxistic patients. Stress levels were reduced on the perceived stress scores (PSS: Cohen Scale) after 21 NO sessions than for those bruxistic without NO training sessions. Conclusions: MPO activity could predict stress level in bruxistic patients and repeated NO reduced their stress level; Origanum majorana essential oil enhanced these anxiolytic effects.

Hydrogen peroxide derived from NADPH oxidase 4- and 2 contributes to the endothelium-dependent vasodilatation of intrarenal arteries.

The role of NADPH oxidase (Nox)-derived reactive oxygen species in kidney vascular function has extensively been investigated in the harmful context of oxidative stress in diabetes and obesity-associated kidney disease. Since hydrogen peroxide (H2O2) has recently been involved in the non-nitric oxide (NO) non-prostanoid relaxations of intrarenal arteries, the present study was sought to investigate whether NADPH oxidases may be functional sources of vasodilator H2O2 in the kidney and to assess their role in the endothelium-dependent relaxations of human and rat intrarenal arteries. Renal interlobar arteries isolated from the kidney of renal tumor patients who underwent nephrectomy, and from the kidney of Wistar rats, were mounted in microvascular myographs to assess function. Superoxide (O2.-) and H2O2 production was measured by chemiluminescence and Amplex Red fluorescence, and Nox2 and Nox4 enzymes were detected by Western blotting and by double inmunolabeling along with eNOS. Nox2 and Nox4 proteins were expressed in the endothelium of renal arterioles and glomeruli co-localized with eNOS, levels of expression of both enzymes being higher in the cortex than in isolated arteries. Pharmacological inhibition of Nox with apocynin and of CYP 2C epoxygenases with sulfaphenazol, but not of the NO synthase (NOS), reduced renal NADPH-stimulated O2.- and H2O2 production. Under conditions of cyclooxygenase and NOS blockade, acetylcholine induced endothelium-dependent relaxations that were blunted by the non-selective Nox inhibitor apocynin and by the Nox2 or the Nox1/4 inhibitors gp91ds-tat and GKT136901, respectively. Acetylcholine stimulated H2O2 production that was reduced by gp91ds-tat and by GKT136901. These results suggest the specific involvement of Nox4 and Nox2 subunits as physiologically relevant endothelial sources of H2O2 generation that contribute to the endothelium-dependent vasodilatation of renal arteries and therefore have a protective role in kidney vasculature.