Efthalia Kerasioti

Anti-inflammatory effects of a special carbohydrate-whey protein cake after exhaustive cycling in humans.

Intense exercise induces increased levels of pro-inflammatory and anti-inflammatory cytokines. Thus, the purpose of this study was to examine the effects of a special cake (consisting of carbohydrate to whey protein 3.5:1) vs. an isocaloric carbohydrate cake on inflammatory markers after exhaustive cycling in humans. Nine subjects received either the experimental or placebo cake in a counterbalanced fashion using a crossover, double-blind, repeated-measures design. They performed one trial involving a 2h exercise on a cycle ergometer at 60-65% VO2max followed by a 4h recovery and then a second trial involving an 1h exercise at 60-65% VO2max which was increased at 95% VO2max. Blood samples were collected pre-exercise, 30 min and 4h post-exercise, post-time Trial and 48 h post-time Trial. Cakes were consumed immediately post-exercise and every 1h for the next 3h. The results showed that consumption of the experimental cake reduced significantly (p<0.05), 4h post-exercise, the pro-inflammatory protein levels IL-6 and CRP compared to the control group by 50% and 46% respectively. Moreover, in the experimental cake group, the level of the anti-inflammatory cytokine IL-10 was higher by 118%, 4h post-exercise, compared to the control group but not statistically significant.

Antioxidant effects of whey protein on muscle C2C12 cells

In the present study, the in vitro scavenging activity of sheep whey protein against free radicals, as well as its reducing power were determined and compared with that of beef protein, soy protein and cow whey protein. Moreover, the possible protective effects of sheep whey protein from tert-butyl hydroperoxide (tBHP)-induced oxidative stress in muscle C2C12 cells were determined by assessing oxidative stress markers by flow cytometry and spectrophotometry. The results showed that sheep whey protein scavenged DPPH, ABTS(+) and OH radicals with IC50 values of 3.1, 4.1 and 1.8 mg of protein/ml. Moreover, the reducing power activity assessed with potassium ferricyanide of sheep whey protein was 1.3mg/ml. As regards to the antioxidant effects in muscle cell line, sheep whey protein at 0.78, 1.56, 3.12 and 6.24 mg of protein/ml increased GSH levels up to 138%, lowered TBARS levels up to 25% and decreased ROS levels up to 41.4%.

Effects of polyphenolic grape extract on the oxidative status of muscle and endothelial cells

A grape pomace extract enhanced antioxidant mechanisms in muscle and endothelial cells both in the absence and in the presence of oxidative stress-induced agent tert-butyl hydroperoxide (tBHP). In particular, muscle (C2C12) and endothelial (EA.hy926) cells were treated with the extract at noncytotoxic concentrations for 24 h, and the oxidative stress markers, total reactive oxygen species (ROS), glutathione (GSH), thiobarbituric reactive substances (TBARS), and protein carbonyl levels were assessed. The results showed that the grape extract treatment reduced significantly ROS, TBARS, and protein carbonyl levels and increased GSH in C2C12 cells, while it increased GSH and decreased protein carbonyl levels in EA.hy926 cells. In the presence of tBHP, the grape extract treatment in C2C12 cells reduced significantly ROS, TBARS, and protein carbonyls and increased GSH compared with tBHP alone treatment, while, in EA.hy926 cells, the extract decreased significantly TBARS and protein carbonyls but increased GSH. The antioxidant potency of the extract was different between muscle and endothelial cells suggesting that the antioxidant activity depends on cell type. Moreover, the antioxidant activity of the grape extract, in both cell lines, exerted, at least in part, through increase in GSH levels. The present work is the first to report the effects of grape extract shown for skeletal muscle cells.

Effect of a special carbohydrate-protein cake on oxidative stress markers after exhaustive cycling in humans.

Exercise has been associated with oxidative stress that is correlated with muscle fatigue and reduced exercise performance. The aim of this study was to examine the effects of a special cake (consisting of carbohydrate to whey protein 3.5:1) vs an isocaloric carbohydrate cake on biomarkers of oxidative stress in 9 males after exhaustive cycling. A randomized single-blind cross-over study was completed. They performed one trial involving a 2-h exercise on a cycle ergometer at 60-65% VO(2)max followed by a 4-h recovery and then a second trial involved an 1-h exercise at 60-65% VO(2)max which was increased at 95% VO(2)max (time trial). The subjects received 4 experimental or placebo cakes after the first trial (the first immediately after and then one every hour). Blood samples were collected at eight time intervals: pre-exercise, 30 min, 1.5 h and 4 h post-exercise, post time Trial, 1 h, 24 h and 48 h post time Trial. Thiobarbituric Acid Reactive Substances (TBARS), protein carbonyls, total antioxidant capacity (TAC), catalase and glutathione (GSH) were determined spectrophotometrically. The mean time to exhaustion did not differ upon cake consumption. Consumption of the special cake reduced TBARS significantly, but had no effect on other oxidative stress markers.

Grape pomace extract exerts antioxidant effects through an increase in GCS levels and GST activity in muscle and endothelial cells

In a previous study, we demonstrated that a grape pomace extract (GPE) exerted antioxidant activity in endothelial (EA.hy926) and muscle (C2C12) cells through an increase in glutathione (GSH) levels. In the present study, in order to elucidate the mechanisms responsible for the antioxidant activity of GPE, its effects on the expression of critical antioxidant enzymes, such as catalase (CAT), superoxide dismutase (SOD)1, heme oxygenase 1 (HO-1) and gamma-glutamylcysteine synthetase (GCS) were assessed in EA.hy926 and C2C12 cells. Moreover, the effects of GPE on CAT, SOD and glutathione S-transferase (GST) enzymatic activity were evaluated. For this purpose, the C2C12 and EA.hy926 cells were treated with GPE at low and non-cytotoxic concentrations (2.5 and 10 µg/ml for the C2C12 cells; 0.068 and 0.250 µg/ml for the EA.hy926 cells) for 3, 6, 12, 18 and 24 h. Following incubation, enzymatic expression and activity were assessed. The results revealed that treatment with GPE significantly increased GCS levels and GST activity in both the C2C12 and EA.hy926 cells. However, GPE significantly decreased CAT levels and activity, but only in the muscle cells, while it had no effect on CAT levels and activity in the endothelial cells. Moreover, treatment with GPE had no effect on HO-1 and SOD expression and activity in both cell lines. Therefore, the present results provide further evidence of the crucial role of GSH systems in the antioxidant effects exerted by GPE. Thus, GPE may prove to be effective for use as a food supplement for the treatment of oxidative stress-induced pathological conditions of the cardiovascular and skeletal muscle systems, particularly those associated with low GSH levels.

Antioxidant Effects of Sheep Whey Protein on Endothelial Cells

Excessive production of reactive oxygen species (ROS) may cause endothelial dysfunction and consequently vascular disease. In the present study, the possible protective effects of sheep whey protein (SWP) from tert-butyl hydroperoxide- (tBHP-) induced oxidative stress in endothelial cells (EA.hy926) were assessed using oxidative stress biomarkers. These oxidative stress biomarkers were glutathione (GSH) and ROS levels determined by flow cytometry. Moreover, thiobarbituric acid-reactive substances (TBARS), protein carbonyls (CARB), and oxidized glutathione (GSSG) were determined spectrophotometrically. The results showed that SWP at 0.78, 1.56, 3.12, and 6.24 mg of protein mL−1 increased GSH up to 141%, while it decreased GSSG to 46.7%, ROS to 58.5%, TBARS to 52.5%, and CARB to 49.0%. In conclusion, the present study demonstrated for the first time that SWP protected endothelial cells from oxidative stress. Thus, SWP may be used for developing food supplements or biofunctional foods to attenuate vascular disturbances associated with oxidative stress.

Development of an assay to assess genotoxicity by particulate matter extract

The current study describes a method for assessing the oxidative potential of common environmental stressors (ambient air particulate matter), using a plasmid relaxation assay where the extract caused single-strand breaks, easily visualised through electrophoresis. This assay utilises a miniscule amount (11 µg) of particulate matter (PM) extract compared to other, cell-based methods (~3,000 µg). The negative impact of air pollution on human health has been extensively recognised. Among the air pollutants, PM plays an eminent role, as reflected in the broad scientific interest. PM toxicity highly depends on its composition (metals and organic compounds), which in turn has been linked to multiple health effects (such as cardiorespiratory diseases and cancer) through multiple toxicity mechanisms; the induction of oxidative stress is considered a major mechanism among these. In this study, the PM levels, oxidative potential, cytotoxicity and genotoxicity of PM in the region of Larissa, Greece were examined using the plasmid relaxation assay. Finally, coffee extracts from different varieties, derived from both green and roasted seeds, were examined for their ability to inhibit PM-induced DNA damage. These extracts also exerted an inhibitory effect on xanthine oxidase and catalase, but had no effect against superoxide dismutase. Overall, this study highlights the importance of assays for assessing the oxidative potential of widespread environmental stressors (PM), as well as the antioxidant capacity of beverages and food items, with the highlight being the development of a plasmid relaxation assay to assess the genotoxicity caused by PM using only a miniscule amount.

Effects of sheep/goat whey protein dietary supplementation on the redox status of rats

The purpose of the present study is to estimate the effects of sheep/goat whey protein dietary supplementation on the redox status of blood and tissues of rats. Twelve male Wistar rats were divided into the control group (standard commercial diet) and whey group [standard commercial diet + sheep/goat whey protein (1 g kg b.w/day)] (6 rats/group). The animals were maintainted on their respective diet for 28 days. At the end of the experimental period, reduced glutathione, catalase activity, total antioxidant capacity, thiobarbituric reactive substances, protein carbonyls and the decomposition rate of H2O2 were measured in blood and tissues of rats. According to the results, the rats fed with the sheep/goat whey protein exhibited improved antioxidant status and decreased free radical-induced toxic effects on lipids and proteins. Specifically, in blood, GSH and CAT levels were significantly increased while TBARS and protein carbonyl levels were significantly decreased compared to the control group. Regarding the effects on tissues, it was observed that GSH levels were significantly increased in small intestine, quadriceps muscle, pancreas and lung tissue compared to the control group. The decomposition rate of H2O2 was significantly decreased in liver, brain and quadriceps muscle, but was significantly increased in spleen tissue compared to the control group. TBARS levels were significantly decreased in liver, brain, quadriceps muscle, pancreas, lung and spleen tissue compared to the control group. Finally, protein carbonyl levels were significantly decreased in brain, small intestine, kidney, pancreas and spleen tissue compared to the control group. Thus, the present findings show the beneficial effects of sheep/goat whey protein, a by-product of cheese manufacturing, on the redox status in an in vivo model.

Six months exposure to a real life mixture of 13 chemicals' below individual NOAELs induced non monotonic sex-dependent biochemical and redox status changes in rats

This study assessed the potential adverse health effects of long-term low-dose exposure to chemical mixtures simulating complex real-life human exposures. Four groups of Sprague Dawley rats were administered mixtures containing carbaryl, dimethoate, glyphosate, methomyl, methyl parathion, triadimefon, aspartame, sodium benzoate, calcium disodium ethylene diamine tetra-acetate, ethylparaben, butylparaben, bisphenol A, and acacia gum at doses of 0, 0.25, 1 or 5 times the respective Toxicological Reference Values (TRV): acceptable daily intake (ADI) or tolerable daily intake (TDI) in a 24 weeks toxicity study. Body weight gain, feed and water consumption were evaluated weekly. At 24 weeks blood was collected and biochemistry parameters and redox status markers were assessed. Adverse effects were observed on body weight gain and in hepatotoxic parameters such as the total bilirubin, alanine aminotransferase (ALT) and alkaline phosphatase (ALP), especially in low dose and affecting mainly male rats. The low dose group showed increased catalase activity both in females and males, whereas the high dose group exhibited decreased protein carbonyl and total antioxidant capacity (TAC) levels in both sex groups. Non-monotonic effects and adaptive responses on liver function tests and redox status, leading to non-linear dose-responses curves, are probably produced by modulation of different mechanisms.

Tissue specific effects of feeds supplemented with grape pomace or olive oil mill wastewater on detoxification enzymes in sheep

Graphical abstract