Plumericin prevents intestinal inflammation and oxidative stress in vitro and in vivo.

Inflammatory bowel diseases (IBDs) are characterized by an inflammatory and oxidative stress condition in the intestinal tissue. In this study, we evaluated the effect of plumericin, one of the main bioactive components of Himatanthus sucuuba (Woodson) bark, on intestinal inflammation and oxidative stress, both in vitro and in vivo. The effect of plumericin (0.5-2 µM) in vitro was evaluated in rat intestinal epithelial cells (IEC-6) treated with lipopolysaccharides from E. coli (10 µg/mL) plus interferon-γ (10 U/mL). Moreover, a 2,4,6-dinitrobenzene sulfonic acid (DNBS)-induced colitis model was used to evaluate the anti-inflammatory and antioxidant activity of plumericin (3 mg/kg) in vivo. The results showed that plumericin significantly reduces intestinal inflammatory factors such as tumor necrosis factor-α, cyclooxygenase-2 and inducible nitric oxide synthase expression, and nitrotyrosine formation. Plumericin also inhibited nuclear factor-κB translocation, reactive oxygen species (ROS) release, and inflammasome activation. Moreover, plumericin activated the nuclear factor erythroid-derived 2 pathway in IEC-6. Using the DNBS-induced colitis model, a significant reduction in the weight loss and in the development of the macroscopic and histologic signs of colon injury, together with a reduced inflammatory and oxidative stress state, were observed in plumericin-treated mice. These results indicate that plumericin exerts a strong anti-inflammatory and antioxidant activity. Thus, it might be a candidate for the development of a new pharmacologic approach for IBDs treatment.

ß-Lactoglobulin Heptapeptide Reduces Oxidative Stress in Intestinal Epithelial Cells and Angiotensin II-Induced Vasoconstriction on Mouse Mesenteric Arteries by Induction of Nuclear Factor Erythroid 2-Related Factor 2 (Nrf2) Translocation.

Peptides derived from buffalo dairy products possess multiple healthy properties that cannot be exerted as long as they are encrypted in parent proteins. To evaluate the biological activities of encrypted peptide sequences from buffalo ricotta cheese, we performed a simulated gastrointestinal (GI) digestion. Chemical and pharmacological characterization of the digest led to the identification of a novel peptide endowed with antioxidant and antihypertensive action. The GI digest was fractionated by Semiprep-HPLC, and fractions were tested against reactive oxygen species (ROS) release in an H2O2-treated intestinal epithelial cell line. UHPLC-PDA-MS/MS analysis revealed the presence of an abundant ß-lactoglobulin peptide (BRP2) in the most active fraction. Pharmacological characterization of BRP2 highlighted its antioxidant activity, involving ROS reduction, nuclear factor erythroid 2-related factor 2 (Nrf2) activation, and cytoprotective enzyme expression. The bioavailability of BRP2 was evaluated in intestinal transport studies through a Caco-2 cell monolayer. Equal bidirectional transport and linear permeability indicate that BRP2 was absorbed mainly through passive diffusion. In addition to its local effects, the BRP2 administration on mouse mesenteric arteries was able to reduce the angiotensin II-induced vasoconstriction by the Nrf2 nuclear translocation, the reduction of the active form of Ras-related C3 botulinum toxin substrate 1 (Rac1), and the NADPH oxidase activity. These data further highlight the role of buffalo ricotta cheese-derived peptides against oxidative stress-related diseases and suggest their health-promoting potential.

Effect of Indoxyl Sulfate on the Repair and Intactness of Intestinal Epithelial Cells: Role of Reactive Oxygen Species' Release.

Chronic kidney disease (CKD) is characterized by an oxidative stress status, driving some CKD-associated complications, even at the gastrointestinal level. Indoxyl Sulfate (IS) is a protein-bound uremic toxin, poorly eliminated by dialysis. This toxin is able to affect the intestinal system, but its molecular mechanism/s in intestinal epithelial cells (IECs) remain poorly understood. This study's aim was to evaluate the effect of IS (31.2-250 µM) on oxidative stress in IEC-6 cells and on the intactness of IECs monolayers. Our results indicated that IS enhanced oxidative cell damage by inducing reactive oxygen species (ROS) release, reducing the antioxidant response and affecting Nuclear factor (erythroid-derived 2)-like 2 (Nrf2) nuclear translocation as well its related antioxidant enzymes. In the wound healing assay model, IS reduced IEC-6 migration, slightly impaired actin cytoskeleton rearrangement; this effect was associated with connexin 43 alteration. Moreover, we reported the effect of CKD patients' sera in IEC-6 cells. Our results indicated that patient sera induced ROS release in IEC-6 cells directly related to IS sera content and this effect was reduced by AST-120 serum treatment. Results highlighted the effect of IS in inducing oxidative stress in IECs and in impairing the intactness of the IECs cell monolayer, thus significantly contributing to CKD-associated intestinal alterations.

AST-120 Reduces Neuroinflammation Induced by Indoxyl Sulfate in Glial Cells.

Chronic kidney disease (CKD) involves multiple organ dysfunction, and the neurological complications that are often present in CKD patients support the idea of a crosstalk between the kidneys and the brain. Evidence suggests a possible role for products accumulating in these patients as uremic toxins in various CKD complications, including neurodegeneration. Indoxyl sulfate (IS), derived from tryptophan metabolism, is well-known as a uremic nephron-vascular toxin, and recent evidence suggests it also has a role in the immune response and in neurodegeneration. Inflammation has been associated with neurodegenerative diseases, as well as with CKD. In this study, we demonstrated that sera of CKD patients induced a significant inflammation in astrocyte cells which was proportional to IS sera concentrations, and that the IS adsorbent, AST-120, reduced this inflammatory response. These results indicated that, among the uremic toxins accumulating in serum of CKD patients, IS significantly contributed to astrocyte inflammation. Moreover, being also chronic inflammation associated with CKD, here we reported that IS further increased inflammation and oxidative stress in primary central nervous system (CNS) cells, via Nuclear Factor-κB (NF-κB) and Aryl hydrocarbon Receptor (AhR) activation, and induced neuron death. This study is a step towards elucidating IS as a potential pharmacological target in CKD patients.

Supplementation of Short-Chain Fatty Acid, Sodium Propionate, in Patients on Maintenance Hemodialysis: Beneficial Effects on Inflammatory Parameters and Gut-Derived Uremic Toxins, A Pilot Study (PLAN Study)

BACKGROUND: In end-stage renal disease (ESRD), gut-derived uremic toxins play a crucial role in the systemic inflammation and oxidative stress promoting the excess morbidity and mortality. The biochemical derangement is in part a consequence of an insufficient generation of short-chain fatty acids (SCFA) due to the dysbiosis of the gut and an insufficient consumption of the fermentable complex carbohydrates. AIM OF THE STUDY: The primary end-point was to evaluate the potential efficacy of SCFA (specifically, sodium propionate (SP)) for patients on maintenance hemodialysis (MHD) on systemic inflammation. Secondary end-points included potential attenuation of oxidative stress markers, insulin resistance and production of gut-derived uremic toxins indoxyl sulfate and p-cresol sulfate, as well as health status after SP supplementation. STUDY DESIGN: We performed a single-center non-randomized pilot study in 20 MHD patients. They received the food additive SP with a daily intake of 2 × 500 mg in the form of capsules for 12 weeks. Pre-dialysis blood samples were taken at the beginning, after six weeks and at the end of the administration period, as well as four weeks after withdrawal of the treatment. RESULTS: The subjects revealed a significant decline of inflammatory parameters C-reactive protein (-46%), interleukin IL-2 (-27%) and IL-17 (-15%). The inflammatory parameters IL-6 and IFN-gamma showed a mild non-significant reduction and the anti-inflammatory cytokine IL-10 increased significantly (+71%). While the concentration of bacterial endotoxins and TNF-α remained unchanged, the gut-derived uremic toxins, indoxyl sulfate (-30%) and p-cresyl sulfate (-50%), revealed a significant decline. The SP supplementation reduced the parameters of oxidative stress malondialdehyde (-32%) and glutathione peroxidase activity (-28%). The serum insulin levels dropped by 30% and the HOMA-index by 32%. The reduction of inflammatory parameters was associated with a lowering of ferritin and a significant increase in transferrin saturation (TSAT). Four weeks after the end of the treatment phase, all improved parameters deteriorated again. Evaluation of the psycho-physical performance with the short form 36 (SF-36) questionnaire showed an enhancement in the self-reported physical functioning, general health, vitality and mental health. The SP supplementation was well tolerated and without important side effects. No patient had left the study due to intolerance to the medication. The SP supplementation in MHD patients reduced pro-inflammatory parameters and oxidative stress and improved insulin resistance and iron metabolism. Furthermore, SP effectively lowered the important gut-derived uremic toxins indoxyl and p-cresol sulfate. These improvements were associated with a better quality of life. Further controlled studies are required in a larger cohort to evaluate the clinical outcome.

Antioxidant Properties of Buffalo-Milk Dairy Products: A ß-Lg Peptide Released after Gastrointestinal Digestion of Buffalo Ricotta Cheese Reduces Oxidative Stress in Intestinal Epithelial Cells.

Redox signaling regulates different gastrointestinal (G.I.) epithelium functions. At the intestinal level, the loss of redox homeostasis in intestinal epithelial cells (IECs) is responsible for the pathogenesis and development of a wide diversity of G.I. disorders. Thus, the manipulation of oxidative stress in IECs could represent an important pharmacological target for different diseases. In this study, peptides released from in vitro gastro intestinal digestion of different buffalo-milk commercial dairy products were identified and evaluated for their bioactive properties. In particular, six G.I. digests of dairy products were tested in a model of oxidative stress for IECs. Among them, buffalo ricotta cheese was the most active and the presence of an abundant ß-lactoglobulin peptide (YVEELKPTPEGDL, f:60-72) was also revealed. The antioxidant potential of the identified peptide was also evaluated in a model of hydrogen peroxide (H2O2)-induced oxidative stress in the IEC-6 cell line. The peptide was able to reduce ROS release, while, on the other hand, it increased nuclear factor (erythroid-derived 2)-like 2 (Nrf2) activation and the expression of antioxidant cytoprotective factors, such as heme oxygenase 1 (HO-1), NAD(P)H:quinone oxidoreductase 1 (NQO1), and superoxide dismutase (SOD). These results indicate that buffalo ricotta cheese-isolated peptide could have potential in the treatment of some gastrointestinal disorders.

Astragalus membranaceus Extract Attenuates Inflammation and Oxidative Stress in Intestinal Epithelial Cells via NF-κB Activation and Nrf2 Response.

Astragalus membranaceus, dried root extract, also known as Astragali radix, is used in traditional Chinese medicine as a tonic remedy. Moreover, it has been reported that Astragalus membranaceus could attenuate intestinal inflammation; however, the underlying mechanism for its anti-inflammatory activity in intestinal epithelial cells (IECs) remains unclear. In this study, we evaluated Astragalus membranaceus extract (5-100 µg/mL) in a model of inflammation and oxidative stress for IECs. We showed that Astragalus membranaceus extract reduced the inflammatory response induced by lipopolysaccharide from E. coli (LPS) plus interferon-γ (IFN), decreasing tumor necrosis factor-α (TNF-α) release, cycloxygenase-2 (COX-2) and inducible nitric oxide synthase (iNOS) expression, nitrotyrosine formation, nuclear factor-κB (NF-κB) activation, and reactive oxygen species (ROS) release in the non-tumorigenic intestinal epithelial cell line (IEC-6). The antioxidant potential of Astragalus membranaceus extract was also evaluated in a model of hydrogen peroxide (H2O2)-induced oxidative stress in IEC-6, indicating that this extract reduced ROS release and increased nuclear factor (erythroid-derived 2)-like 2 (Nrf2) activation and the expression of antioxidant cytoprotective factors in these cells. The results contributed to clarify the mechanisms involved in Astragalus membranaceus extract-reduced inflammation and highlighted the potential use of this extract as an anti-inflammatory and antioxidant remedy for intestinal diseases.

Medicinal plants of northern Angola and their anti-inflammatory properties.

ETHNOPHARMACOLOGICAL RELEVANCE: 32 plants, from which 30 are used in local traditional medicine - identified by interviews with the resident population - in the province of Uíge in northern Angola for the treatment of inflammation related disorders, were screened on different anti-inflammatory parameters. Three extracts were selected for a detailed ethnobotanical, pharmacological and phytochemical investigation based on their in vitro activity. AIM OF THE STUDY: We aimed to assess the in vitro anti-inflammatory activity of these plants and highlight the active principles of the three most promising candidates. MATERIALS AND METHODS: Plant material was collected in northern Angola during eight field trips from 2013 to 2015 based on data documented in 61 interviews with 142 local informants. 36 methanol (MeOH) extracts were prepared and tested at different concentrations (100, 50, 10µg/mL) to evaluate their inhibition on cyclooxygenase (COX)-2 expression and on nitric oxide (NO) release in lipopolysaccharide (LPS)-stimulated J774A.1 macrophages. Five selected extracts were additionally tested at the lower concentrations of 5, 2.5, and 1.25µg/mL and for their potential on inhibition of tumor necrosis factor-α (TNF-α) and interleukin 6 (IL-6) release. The major compounds of three of those five extracts were either identified by HPLC coupled with (tandem) mass spectrometry and comparison with data from literature or isolated from the respective extracts and confirmed by NMR experiments (one and two dimensional). RESULTS: 30 plant species with in total 161 citations were mentioned by the informants to have anti-inflammatory properties. The predominantly used plant part is the leaf (39%), followed by underground organs like roots and rhizomes (25%), bark (18%) as well as fruits and seeds with 15%. With 47%, decoction is the most frequent preparation form. A large number of the MeOH extracts showed promising activities in our preliminary screening for the inhibition of COX-2 expression and NO release. Five extracts with high activities in both assays showed also concentration dependent inhibition at lower concentrations and a decreased release of two additional pro-inflammatory mediators (IL-6 and TNF-α) vs. LPS. Three leaf extracts where chosen for a detailed investigation, which lead to the identification of several constituents: verbascoside and isoverbascoside (Acanthus montanus), geraniin, chebulagic acid and a large flavonoid fraction (Alchornea cordifolia) as well as the four flavonoids astilbin, isovitexin, isoorientin and swertisin (Chaetocarpus africanus). Their implication in the observed biological activity was proved by comparison with published data of these compounds in identical or similar pharmacological models. CONCLUSIONS: The indigenous use of these plants against inflammation related ailments could be - at least partly - verified by our in vitro models for many of the investigated extracts. Acanthus montanus and Alchornea cordifolia particularly stood out with their high activity in all four performed assays, which was in accordance with pharmacological studies of their major constituents in literature. In addition, this study was the first phytochemical investigation of Chaetocarpus africanus and first description of the occurrence of the ellagitannins geraniin and chebulagic acid in Alchornea cordifolia. These results support the traditional use and should encourage further investigations of medicinal plants of northern Angola.

Olive oil polyphenols extracts inhibit inflammatory markers in J774A.1 murine macrophages and scavenge free radicals.

Here we evaluate the olive oil antiradical and anti-inflammatory potential through its polyphenols extracts and examine the influence of olive maturity on olive oil quality properties, polyphenols composition and biological potentials. Samples have been obtained from minor Tunisian olive cultivars (Chemchali, Fouji and Zarrazi) at different maturity indices. Principal quality properties were evaluated and polyphenols analysis was carried out by Folin Ciocalteu reagent and HPLC-UV-MS. Antiradical activity was examined by DPPH and FRAP scavenging assays while J774A.1 murine macrophages were used to evaluate anti-inflammatory potential by analyzing NO production with Griess reagent method and iNOS and COX-2 expression by cytofluorimetric analysis. Our results revealed that quality characteristics, total phenol content, as well as phenolic compound concentrations were significantly affected by the olive maturity levels. On the other hand, the polyphenols extracts showed an interesting radical scavenging capacity and a potential ability to inhibit inflammatory markers at 90% for NO release and 75% for iNOS expression. Thus, our study establishes that olive oil through its polyphenols extracts has a substantial antiradical and anti-inflammatory potential. Likewise a lot of attention should be attributed to olive ripening level in order to decide the optimum harvesting time.

The Food Contaminants Nivalenol and Deoxynivalenol Induce Inflammation in Intestinal Epithelial Cells by Regulating Reactive Oxygen Species Release.

Fusarium mycotoxins are fungal metabolites whose ability to affect cereal grains as multi-contaminants is progressively increasing. The trichothecene mycotoxins nivalenol (NIV) and deoxynivalenol (DON) are often found in almost all agricultural commodities worldwide. They are able to affect animal and human health, including at the intestinal level. In this study, NIV, both alone and in combination with DON, induced inflammation and increased the inflammatory response induced by lipopolysaccharide (LPS) plus Interferon-γ (IFN) in the non-tumorigenic intestinal epithelial cell line (IEC-6). The inflammatory response induced by NIV and DON involves tumor necrosis factor-α (TNF-α) production, inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) expression, nitrotyrosine formation, reactive oxygen species (ROS) release, Nuclear Factor-κB (NF-κB), Nuclear factor (erythroid-derived 2)-like 2 (Nrf2) and inflammasome activation. The pro-inflammatory effect was strongly induced by NIV and by the mycotoxin mixture, when compared to DON alone. Mechanistic studies indicate a pivotal role for ROS in the observed pro-inflammatory effects induced by mycotoxins. In this study, the interactions between NIV and DON point out the importance of their food co-contamination, further highlighting the risk assessment process that is of growing concern.

Anti-Inflammatory and Anti-Oxidant Potential of the Root Extract and Constituents of Doronicum austriacum.

BACKGROUND: Doronicum austriacum Jacq., Asteraceae, is a plant which is used in traditional alpine medicine. Historical sources describe the medical use of the root, but up until now only a few studies evaluated its pharmacological properties. The evaluation of the dichloromethane extract, and its major compounds for their anti-inflammatory and anti-oxidant potential was performed in macrophages J774A.1 and C6 astrocytes. Nitric oxide (NO) and reactive oxygen species (ROS) release, as well as nitrotyrosine formation, were evaluated. Moreover, in order to evaluate the potential anti-proliferative activity, under the same experimental conditions, 3-(4,5-dimethyltiazol-2yl)-2,5-phenyl-2H-tetrazolium bromide (MTT) assay was also performed. Our results indicate that Doronicum austriacum has a significant effect in inhibiting both pro-inflammatory and pro-oxidative mediators. All isolated compounds were able to significantly inhibit NO and ROS release both in macrophage and in astrocytes cells, even if the effect was more pronounced in macrophage. In particular, among the tested compounds, 6,12-dihydroxy-(-)-2S-tremetone exerted stronger activity. Both extract and single compounds did not affect cellular viability. This study provides evidence for the pharmacological anti-inflammatory and anti-oxidant potential of Doronicum austriacum extract. These effects could be due to the activity of its major constituents and subsequent identification of benzofurans as a promising compound class to combat inflammation and related diseases.

Indoxyl Sulfate Affects Glial Function Increasing Oxidative Stress and Neuroinflammation in Chronic Kidney Disease: Interaction between Astrocytes and Microglia.

Indoxyl sulfate (IS) is a protein-bound uremic toxin resulting from the metabolism of dietary tryptophan which accumulates in patients with impaired renal function, such as chronic kidney disease (CKD). IS is a well-known nephrovascular toxin but little is known about its effects on central nervous system (CNS) cells. Considering the growing interest in the field of CNS comorbidities in CKD, we studied the effect of IS on CNS cells. IS (15-60 µM) treatment in C6 astrocyte cells increased reactive oxygen species release and decreased nuclear factor (erythroid-derived 2)-like 2 (Nrf2) activation, and heme oxygenase-1 (HO-1) and NAD(P)H dehydrogenase quinone 1 expression. Moreover, IS increased Aryl hydrocarbon Receptor (AhR) and Nuclear Factor-kB (NF-kB) activation in these cells. Similiar observations were made in primary mouse astrocytes and mixed glial cells. Inducible nitric oxide synthase and cyclooxygenase-2 (COX-2) expression, tumor necrosis factor-α and interleukin-6 release and nitrotyrosine formation were increased by IS (15-60 µM) in primary mouse astrocytes and mixed glial cells. IS increased AhR and NF-kB nuclear translocation and reduced Nrf2 translocation and HO-1 expression in primary glial cells. In addition, IS induced cell death in neurons in a dose dependent fashion. Injection of IS (800 mg/kg, i.p.) into mice induced histological changes and increased COX-2 expression and nitrotyrosine formation in thebrain tissue. Taken together, our results show a significant contribution of IS in generating a neurotoxic enviroment and it could also have a potential role in neurodegeneration. IS could be considered also a potential therapeutical target for CKD-associated neurodegenerative complications.

Anti-Inflammatory, Antioxidant and Crystallographic Studies of N-Palmitoyl-ethanol Amine (PEA) Derivatives.

N-Palmitoyl-ethanolamine (PEA) is an anti-inflammatory component of egg yolk that is usually employed for the prevention of respiratory apparatus virus infection and then frequently used for its efficient anti-inflammatory and analgesic effects in experimental models of visceral, neuropathic, and inflammatory diseases. Nevertheless, data of its use in animal or human therapy are still scarce and further studies are needed. Herein, we report the biological evaluation of a small library of N-palmitoyl-ethanolamine analogues or derivatives, characterized by a protected acid function (either as palmitoyl amides or hexadecyl esters), useful to decrease their hydrolysis rate in vitro and prolong their biological activity. Two of these compounds-namely phenyl-carbamic acid hexadecyl ester (4) and 2-methyl-pentadecanoic acid (4-nitro-phenyl)-amide (5)-have shown good anti-inflammatory and antioxidant properties, without affecting the viability of J774A.1 macrophages. Finally, crystals suitable for X-ray analysis of compound 4 have been obtained, and its solved crystal structure is here reported. Our outcomes may be helpful for a rational drug design based on new PEA analogues/derivatives with improved biological properties.

Bioavailable Citrus sinensis Extract: Polyphenolic Composition and Biological Activity.

Citrus plants contain large amounts of flavonoids with beneficial effects on human health. In the present study, the antioxidant and anti-inflammatory potential of bioavailable polyphenols from Citrus sinensis was evaluated in vitro and ex vivo, using the murine macrophages cell line J774A.1 and primary peritoneal macrophages. Following simulated gastro-intestinal digestion, the in vitro bioavailability of Citrus sinensis polyphenolic extract was assessed using the human cell line Caco-2 grown as monolayers on a transwell membrane. Data demonstrated a relative permeation of its compounds (8.3%). Thus, the antioxidant and anti-inflammatory effect of polyphenolic Citrus sinensis fraction (Cs) was compared to the bioavailable one (CsB). Results revealed that Citrus extract were able to reduce macrophages pro-inflammatory mediators, including nitric oxide, iNOS, COX-2 and different cytokines. Moreover, the effect of Citrus sinensis polyphenols was associated with antioxidant effects, such as a reduction of reactive oxygen species (ROS) and heme-oxygenase-1 (HO-1) increased expression. Our results provide evidence that the bioavailable polyphenolic constituents of the Citrussinensis extract accumulate prevalently at intestinal level and could reach systemic circulation exerting their effect. The bioavailable fraction showed a higher anti-inflammatory and antioxidant potential compared to the initial extract, thus highlighting its potential nutraceutical value.

Anti-inflammatory and antioxidant activity of polyphenolic extracts from Lactuca sativa (var. Maravilla de Verano) under different farming methods.

BACKGROUND: Besides their nutritional value, vegetables are a source of health-promoting compounds, such as polyphenols, and their content can be influenced by the particular farming method. In this study polyphenolic extracts from Lactuca sativa (var. Maravilla de verano) plants cultivated with different farming methods were chemically characterised and tested in vitro and ex vivo inflammation models. RESULTS: The tested extacts (250-2.5 µg mL(-1) ) were able to reduce both the inflammatory and oxidative stress in LPS-stimulated J774A.1 murine monocyte macrophage cells, by lowering the release of nitric oxide (NO) and reactive oxygen species (ROS) and promoting nuclear translocation of nuclear factor (erythroid-derived 2)-like 2; (Nrf2) and nuclear factor-κB (NF-κB). In this regard, quantitative profiles revealed different amounts of polyphenols, in particular quercetin levels were higher in plants under mineral fertilised treatment. Those extract showed an enhanced anti-inflammatory and antioxidant activity. CONCLUSION: Our data showed the anti-inflammatory and antioxidant potential of Maravilla de Verano polyphenolic extracts. The effect of farming methods on polyphenolic levels was highlighted. The higher reduction of inflammatory mediators release in extracts from plants cultivated under mineral fertilisation treatment was correlated to the higher amount of quercetin. These results can be useful for both nutraceutical or agronomic purposes. © 2016 Society of Chemical Industry.

Muscle stimulation in elderly patients with CKD and sarcopenia.

OBJECTIVES: Aim of our study was to assess the potential effects of high-tone external muscle stimulation (HTEMS) on improvement of endothelial dysfunction (ED) and kidney damage in elderly patients with chronic kidney disease (CKD), sarcopenia and/or serious physical disability with a high Multidisciplinary Prognostic Index (MPI). METHODS: We enrolled 12 consecutive CKD patients with MPI > 0,66 from January 1st, 2008 to December 31st, 2014. Six patients underwent a 2-hours HTEMS during the first day (group A) and the other six patients (group B) underwent a sham experiment with HTEMS without power supply. After 24 hours, patients of group A were shifted to group B and vice-versa. Nitrite/nitrate (NOx), endotheline-1 (ET-1) and urine creatinine concentration were measured in all patients. RESULTS: During HTEMS urine amount increased by 22% (p=0.049), so did urine creatinine that increased by 40%, (p=0.034) and creatinine clearance that increased by 26% (p=0.041). There was no statistical difference in urine nitrogen (that raised by 11%, p=0.526), urine sodium (that reduced by 42%, p=0.121) and urine potassium levels (p=0,491). At the same time, NOx changed from 44.15.1 to 38.45.3 M/L after 1 hour, to 36.44.8 M/L after 2 hours, to 41.15.7 M/L after 3 hours and to 46,95.0 M/L after 4 hours (p=0.008) during HTEMS, while it did not vary during the sham section of the experiment, respectively 43.66.1 M/L , 436.4 M/L, 42.85.5 M/L, 434.7 M/L, and 42.85.8 M/L (p=0.992). CONCLUSION: Our study showed that HTEMS may improve microcirculation and, through this mechanism, may reduce kidney damage in elderly patients with CKD and severe muscle atrophy.

Anti-inflammatory activity of horseradish (Armoracia rusticana) root extracts in LPS-stimulated macrophages.

Horseradish (Armoracia rusticana) is a perennial crop belonging to the Brassicaceae family. Horseradish root is used as a condiment due to its extremely pungent flavour, deriving from the high content of glucosinolates and their breakdown products such as isothiocyanates and other sulfur compounds. Horseradish also has a long history in ethnomedicine. In this study the anti-inflammatory potential of three accessions of Armoracia rusticana on lipopolysaccharide from E. coli treated J774A.1 murine macrophages was evaluated. Our results demonstrate that Armoracia rusticana reduced nitric oxide, tumor necrosis factor-α and interleukin-6 release and nitric oxide synthase and cyclooxygenase-2 expression in macrophages, acting on nuclear transcription factor NF-κB p65 activation. Moreover Armoracia rusticana reduced reactive oxygen species release and increased heme-oxygenase-1 expression, thus contributing to the cytoprotective cellular effect during inflammation.

Nivalenol induces oxidative stress and increases deoxynivalenol pro-oxidant effect in intestinal epithelial cells.

Mycotoxins are secondary fungal metabolites often found as contaminants in almost all agricultural commodities worldwide, and the consumption of food or feed contaminated by mycotoxins represents a major risk for human and animal health. Reactive oxygen species are normal products of cellular metabolism. However, disproportionate generation of reactive oxygen species poses a serious problem to bodily homeostasis and causes oxidative tissue damage. In this study we analyzed the effect of two trichothecenes mycotoxins: nivalenol and deoxynivalenol, alone and in combination, on oxidative stress in the non-tumorigenic intestinal epithelial cell line IEC-6. Our results indicate the pro-oxidant nivalenol effect in IEC-6, the stronger pro-oxidant effect of nivalenol when compared to deoxynivalenol and, interestingly, that nivalenol increases deoxynivalenol pro-oxidative effects. Mechanistic studies indicate that the observed effects were mediated by NADPH oxidase, calcium homeostasis alteration, NF-kB and Nrf2 pathways activation and by iNOS and nitrotyrosine formation. The toxicological interaction by nivalenol and deoxynivalenol reported in this study in IEC-6, points out the importance of the toxic effect of these mycotoxins, mostly in combination, further highlighting the risk assessment process of these toxins that are of growing concern.

Evaluation of anti-inflammatory activity and fast UHPLC-DAD-IT-TOF profiling of polyphenolic compounds extracted from green lettuce (Lactuca sativa L.; var. Maravilla de Verano).

Fresh cut vegetables represent a widely consumed food worldwide. Among these, lettuce (Lactuca sativa L.) is one of the most popular on the market. The growing interest for this "healthy" food is related to the content of bioactive compounds, especially polyphenols, that show many beneficial effects. In this study, we report the anti-inflammatory and antioxidant potential of polyphenols extracted from lettuce (var. Maravilla de Verano), in J774A.1 macrophages stimulated with Escherichia coli lipopolysaccharide (LPS). Lettuce extract significantly decreased reactive oxygen species, nitric oxide release, inducible nitric oxide synthase and cycloxygenase-2 expression. A detailed quali/quantitative profiling of the polyphenolic content was carried out, obtaining fast separation (10 min), good retention time and peak area repeatability, (RSD% 0.80 and 8.68, respectively) as well as linearity (R(2)⩾ 0.999) and mass accuracy (⩽ 5 ppm). Our results show the importance in the diet of this cheap and popular food for his healthy properties.

High-tone external muscle stimulation in patients with acute kidney injury (AKI): beneficial effects on NO metabolism, asymmetric dimethylarginine, and endothelin-1.

OBJECTIVES: The aim of this study was to assess potential effects of high-tone external muscle stimulation (HTEMS) on parameters of endothelial dysfunction (ED) in patients with acute kidney injury (AKI). BACKGROUND: The bad outcome of AKI patients is markedly influenced by ED, microinflammation, oxidative stress and protein hypercatabolism. Recently, we have shown that intradialytic application of HTMS was associated with a faster resolution of AKI. Here, we investigated in the same cohort of patients whether parameters of ED such as nitric oxide (NO), asymmetric-dimethylarginine (ADMA), and endothelin 1 (ET-1) are modulated by HTEMS as compared to non-HTEMS-treated AKI patients. METHODS: In a post-hoc study we analyzed plasma samples of the 34 AKI patients stage 5, of whom 17 underwent intradialytic HTEMS treatment while the other 17 served as AKI dialysis controls. Measurements included plasma nitrate and nitrite (NOx), ADMA, ET-1 and were performed before and on days 3, 7, 14, 21, and 28 after start of daily dialysis. Additional 16 healthy volunteers served as controls. RESULTS: Initially, in both AKI groups NOx levels were markedly lower and ADMA and ET-1 levels were higher compared to the healthy controls. After initiation of daily hemodialysis the HTEMS group showed a faster improvement of NOx and ET-1 (after 1 week) and ADMA levels (after 2 weeks) compared to the No- HTEMS group. After 2 weeks, all parameters of the HTEMS group were not different from healthy controls, while the No-HTEMSAKI group needed 3 - 4 weeks. CONCLUSION: Our findings suggest for the first time that in AKI patients, application of HTEMS is associated with a faster normalization of lowered NOx and elevated ADMA and ET-1 plasma levels. We hypothesize that the more rapid amelioration of these parameters in the HTEMS group contributed to the accelerated recovery of AKI. With regard to the small study groups with different causes of AKI, investigations in a greater number of AKI patients is required.