Alain Favier

Increased lipid peroxidation in pregnant women after iron and vitamin C supplementation

Iron overload could promote the generation of free radicals and result in deleterious cellular damages. A physiological increase of oxidative stress has been observed in pregnancy. A routine iron supplement, especially a combined iron and vitamin C supplementation, without biological justifications (low hemoglobin [Hb] and iron stores) could therefore aggravate this oxidative risk. We investigated the effect of a daily combined iron supplementation (100 mg/d as fumarate) and vitamin C (500 mg/d as ascorbate) for the third trimester of pregnancy on lipid peroxidation (plasma TBARS), antioxidant micronutriments (Zn, Se, retinol, vitaminE, (β-carotene) and antioxidant metalloenzymes (RBC Cu-Zn SOD and Se-GPX). The iron-supplemented group (n=27) was compared to a control group (n=27), age and number of pregnancies matched. At delivery, all the women exhibited normal Hb and ferritin values. In the supplemented group, plasma iron level was higher than in the control group (26.90±5.52 mmol/L) and TBARs plasma levels were significantly enhanced (p<0.05) (3.62±0.36 vs 3.01±0.37 mmol/L). No significant changes were observed in plasma trace elements and red blood cell antioxidant metalloenzymes. Furthermore, the α-tocopherol plasma level was lowered in the iron-supplemented groups, suggesting an increased utilization of vitamin E.These data show that pharmalogical doses of iron, associated with high vitamin C intakes, can result in uncontrolled lipid peroxidation. This is predictive of adverse effects for the mother and the fetus. This study illustrates the potential harmful effects of iron supplementation when prescribed only on the assumption of anemia and not on the bases of biological criteria.

No Significant Effects of Lutein, Lycopene or β-Carotene Supplementation on Biological Markers of Oxidative Stress and LDL Oxidizability in Healthy Adult Subjects

Objective: The objective of this study was to determine the effect of individual carotenoid supplementation on biochemical indices of oxidative status in apparently healthy adult males. Methods:The study was a placebo controlled single blind study. Healthy male volunteers (n=175) were assigned to four groups. They received daily supplements of β-carotene (15 mg), lutein (15 mg), lycopene (15 mg) and placebo for three months. The effects of the supplementation on antioxidant status were monitored by plasma carotenoid, vitamin C and A levels, glutathione (GSH and GSSG) concentrations, protein SH groups, erythrocyte antioxidant enzyme activities (Cu-Zn SOD, Se-GSH-Px) and susceptibility of LDL to copper-induced oxidation. Results:β-carotene, lycopene and lutein supplementation led to significant plasma and LDL increases in each of these carotenoids, without modifications of other carotenoid levels in plasma or in LDL. The supplementation failed to enhance the resistance of LDL to oxidation or to modify the LDL polyunsaturated/saturated fatty acid ratio. Vitamin C, GSH, protein SH groups and antioxidant metalloenzyme activities were also unchanged. Conclusion: We did not observe beneficial or adverse effects of lutein, lycopene or β-carotene supplementation on biomarkers of oxidative stress. In apparently healthy subjects, carotenoid supplementation does not lead to significantly measurable improvement in antioxidant defenses.

Effect of increased fruit and vegetable intake on the susceptibility of lipoprotein to oxidation in smokers

Objective: To evaluate the effect of an increased dietary intake of fruit and vegetables on susceptibility of LDL to oxidation in smokers and nonsmokers. Design: A descriptive and prospective study. Setting: Joseph Fourier University, Grenoble. Subjects: Volunteers were age and sex matched in the smoking and nonsmoking groups and were recruited by announcement. Interventions: Increased intake of fruits and vegetables for two weeks providing 30u2005mg/day of carotenoids. Main outcome measures: Circulating levels of beta-carotene, lutein, lycopene, α-tocopherol; susceptibility of LDL to oxidation. Cu-Zn superoxide dismutase (Cu-Zn SOD), and Se glutathione peroxidase (Se-GSH-Px) activities and reduced (GSH) and oxidized (GSSG) glutathione. Results: At entry (week 0: W0) smokers exhibited a lower plasma carotene level but the plasma parameters of oxidative stress and LDL oxidizability were not different from nonsmokers. After two weeks of increased intake of fruits and vegetables the circulating levels of carotenoids increased in smokers 23% and 11% in nonsmokers. At the same time the resistance of LDL to oxidation increased by 14% in smokers (P≤0.05) and by 28% in nonsmokers (P≤0.025). The mean whole bloodGSH level was higher in smokers at entry but returned to a concentration similar to nonsmokers at the end of the study. Conclusion: This pilot study indicates that an increased, carotenoid rich food intake through its inhibitory effect on the susceptibility of LDL to oxidation may be an interesting approach to reduce the risk of atherosclerosis both in smokers and nonsmokers. Sponsorship: This research has been supported by the European Union: AAIR project (AIR2-CT93-0888, DG 12SSMA).

Antioxidant Supplementation and Tapering Exercise Improve Exercise-Induced Antioxidant Response

Objective and Methods: The present controlled-training, double-blind study (supplemented, n = 7; placebo, n = 9) investigated whether taper training (TT) and antioxidant supplementation, i.e., 150 μg of selenium, 2000 IU of retinol, 120 mg of ascorbic acid and 30 IU of α-tocopherol, modulates antioxidant potential, redox status and oxidative damage occurrence both at rest and in response to exercise. Two weeks of TT followed four weeks of overloaded training. Dietary intakes were recorded. Before and after TT, triathletes did a duathlon consisting of 5-km run, 20-km bike and 5-km run. Biological studies were conducted at rest and after exercise. Results: Whatever the nutritional status, TT induced a decrease in resting blood reduced glutathione (GSH) concentration (p < 0.001), erythrocyte superoxide dismutase (SOD) activity (p < 0.0001) and plasma total antioxidant status (TAS) (p < 0.05). Only in the supplemented group (Su) with TT, did plasma glutathione peroxidase (GSH-Px) activity decrease (p < 0.05) and CD4+ cell concentration increase (p < 0.05). However, antioxidant supplementation increased plasma TAS increase in response to exercise and TT (p < 0.05). After exercise, TT also induced a lower decrease in blood reduced and oxidized (GSSG) glutathione (p < 0.01) in both groups, but TT had no effect on lipoperoxidation as estimated by plasma thiobarbituric reactive substances or on muscular damage occurrence estimated by plasma creatine kinase isoenzyme MB mass. Conclusion: During TT, antioxidant supplementation at nutritional doses reinforces antioxidant status response to exercise, with an effect on exercise-induced oxidative stress, and no effect on oxidative damage.

Induction of thioredoxin by ultraviolet-A radiation prevents oxidative-mediated cell death in human skin fibroblasts

The present study analyzes the expression of the thioredoxin/thioredoxin reductase (Trx/TR) system in UVA-irradiated human skin fibroblasts. Irradiation increases the intracellular level of Trx and a time-dependent increase of Trx mRNA is observed. Our data indicate that Trx translocates from the cytoplasm to the nucleus. In addition, UV exposure results in an increase in TR synthesis. In order to evaluate the function of Trx/TR system, we investigated the antioxidant role of Trx in transient transfected cells. The ROS accumulation in UVA irradiated cells was assessed using flow cytometry. A 3-fold decrease in ROS production was observed in transiently transfected fibroblasts. These results indicate that Trx acts as an antioxidant protein in UVA irradiated fibroblasts. As ROS are inducers of cell death, this raises the question as to whether Trx is able to protect cells from apoptosis and/or necrosis induced by UVA. Six hours after UVA-irradiation, 29.92% of cells were annexin-V positive. This population was significantly reduced in Trx-transfected cells (8.58%). Moreover, this work demonstrates that Trx prevents the loss of the membrane potential of the mitochondria, the depletion of cellular ATP content, and the loss of cell viability induced by irradiation.

Effects of a combined micronutrient supplementation on maternal biological status and newborn anthropometrics measurements: a randomized double-blind, placebo-controlled trial in apparently healthy pregnant women

Objective: To investigate the possible beneficial effects of a micronutrient supplementation to apparently healthy pregnant women on maternal biological status and new born anthropometric characteristics.Setting: Departments of Obstetric of the University Hospital of Grenoble (France) and Lyon (France), Laboratoire of Biology of Oxidative Stress, UFR de Pharmacie. Grenoble (France).Study design: Double-blind, randomized placebo-controlled intervention trial.Subjects: A total of 100 apparently healthy pregnant women were recruited at 14±2 weeks of gestation to delivery. At the end, they were 65 women to follow out the study.Interventions: Daily consumption over gestation of a miconutrients supplement or placebo.Main outcome measures: Plasma micronutrient levels and oxidative stress parameters were measured in mothers at 14 and 38 weeks of gestation. New borns anthropometric characteristics were measured at delivery.Results: In the supplemented group, folic acid, vitamin C, E, B2, B6 and β-carotene levels were higher than in the placebo group. Oxidative stress parameters were not different between the groups. Birth weights were increased by 10% and the number of low newborn weights (<2700g) decreased significantly when the mother received the supplementation. Maternal plasma Zn levels were positively correlated to the newborn heights.Conclusion: A regular intake of a micronutrient supplement at nutritional dose may be sufficient to improve micronutrient status of apparently healthy pregnant women and could prevent low birth weight of newborn.Sponsorship: The supplement and placebo were supplied and the study was partially funded by BOIRON Pharmaceutical Company.

THIOLS AND SELENIUM : PROTECTIVE EFFECT ON HUMAN SKIN FIBROBLASTS EXPOSED TO UVA RADIATION

The sensitivity of human dermal fibroblasts to UVA radiation has been linked to a decrease in intracellular glutathione (GSH) levels. GSH (gamma-glutamyl-cysteinyl-glycine) is a radical scavenger and a cofactor for protective enzymes such as selenium-dependent GSH peroxidases. In this study, we examine the possibility of a cooperative interaction between three cysteine delivery systems and selenium in protecting human cultured fibroblast exposed to UVA radiation. Cells were irradiated (9, 15 and 20 J cm-2) following incubation with N-acetyl-cysteine (NAC, 5 mM), N-acetyl-homocysteine-thiolactone (citiolone (CIT), 1 mM) or L-2-oxothiazolidine-4-carboxylate (OTC, 1 mM). The modulation of the intracellular GSH levels by the addition of the different compounds was determined by enzymatic and separative methods. Cells were harvested for survival analysis by measuring the ability of the cell to adhere and proliferate. Treatments with NAC and CIT resulted in a significant rise in GSH levels compared with control cells, with protection against UVA radiation. OTC did not induce any rise in GSH level; nevertheless, the protective effect afforded by OTC is similar to that observed with NAC and CIT. Moreover, selenium (0.1 mg 1-1), as sodium selenite, significantly increased the protective efficiency of NAC and CIT, but not of OTC. Although the precise mechanism is not known, thiol molecules can inhibit the deleterious effects of UVA radiation. These results provide evidence that compounds capable of inducing GSH synthesis can act with selenium to protect cells against UVA damage.

Trace elements and cardioprotection: increasing endogenous glutathione peroxidase activity by oral selenium supplementation in rats limits reperfusion-induced arrhythmias.

Oxyradicals have been implicated as a possible cause of reperfusion-arrhythmias (RA). However, the use of diverse exogenous oxyradical scavengers designed to reduce RA has given contradictory results. The aim of the present study was to determine whether enhancing the activity of the main endogenous enzyme involved in peroxide elimination in cardiac cells, namely glutathione peroxidase, may limit RA in isolated heart preparations by increasing their antioxidant status. For this purpose, a group of 15 male Wistar rats received a selenium enriched diet for ten weeks (1.5 mg Se/kg diet). Control animals (n = 15) received a standard diet containing 0.05 mg Se/kg diet. The incidence of early ventricular arrhythmias was investigated during the reperfusion period following 10 min regional ischemia induced ex-vivo by left coronary artery ligation. Our results show that selenium-supplementation significantly increased the global selenium status of the animals. In the isolated heart preparations, the selenium supplementation induced a significant reduction of the severity of RA as assessed by the arrhythmia score and the limitation of the incidence of both ventricular tachycardia (control: 91% vs selenium: 36%, p < 0.05) and irreversible ventricular fibrillation (control: 45% vs selenium: 0%, p < 0.05). These effects were associated with a significant increase in cardiac mitochondrial and cytosolic glutathione peroxidase activities in both the left and the right ventricles. These results illustrate the potential protective effect of selenium against ischemia-reperfusion injury and suggest that peroxides might play a key role in the genesis of some aspects of the reperfusion syndrome.

Effect of an acute zinc depletion on rat lipoprotein distribution and peroxidation

The aim of this study was to determine the extent to which zinc depletion leads to lipoprotein modifications by measuring both lipoprotein-fraction distribution and peroxidation in zinc-depleted rats.The animals were divided into three groups and fed for 8 wk a zinc-adequate diet (100 ppm) ad libitum (AL), a zinc-deficient diet (0.2 ppm) ad libitum (ZD), or a zinc-adequate diet according to the pair feeding method (PF).Trace-element status, tissular lipids, and lipoprotein-fraction study were performed. The MDA production by the lipoprotein fraction was measured before and after induced peroxidation.Cholesterol and phospholipids were increased in ZD rats. An important increase of VLDL and IDL was observed and a significant enhanced production of MDA by the LDL was related to zinc deficiency. From this observation, we may conclude that LDL fractions of ZD rats are more susceptible to induced oxidative damage.These results suggest that in zinc deficiency, the lipoprotein fragility is an aggravating factor of peroxidation and the dyslipoproteinemia may lead to an atherogenic risk.

Age-dependent changes in myocardial susceptibility to zero flow ischemia and reperfusion in isolated perfused rat hearts: relation to antioxidant status

Ageing is associated with an increase in myocardial susceptibility to ischemia and a decrease in post-ischemic recovery of function. In the present study, we have examined the effects of ageing on (i) myocardial ischemic contracture, (ii) the reperfusion syndrome and lipid peroxidation upon reperfusion, and (iii) the activity of enzymes involved in reactive oxygen species elimination. Hearts from male Wistar rats aged 4 (adults), 16 (old) or 24 months (senescent) were subjected to 20-min zero flow ischemia and 30-min reperfusion ex vivo. Cardiac activity of superoxide dismutase, catalase, and glutathione peroxidase, as well as cardiac content of thiobarbituric acid reactants were assessed in frozen heart samples. The effects of ageing on ischemic contracture of the sarcomeres were assessed on electromicrographs of tissues taken at the end of ischemia. In our experimental conditions, ischemic contracture of the sarcomeres increased progressively during ageing. In contrast, the severity of the reperfusion syndrome increased between 4 and 16 months of age, and then decreased up to 24 months of age. We propose that the peak of susceptibility of the myocardium to reperfusion observed during moderate ageing might be related to a decrease in the ability of cardiomyocytes to dismutate hydrogen peroxide as suggested by the observed decrease in catalase activity. Finally, the better resistance to the reperfusion syndrome exhibited by senescent rats compared to old rats might be due to a natural selection of a subpopulation of rats which is particularly resistant to oxidative stress.