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The effect of coffee and cocoa on oxidative damage to macromolecules has been investigated in several studies, often with controversial results. This study aimed to investigate the effect of one-month consumption of different doses of coffee or cocoa-based products containing coffee on markers of DNA damage and lipid peroxidation in young healthy volunteers. Twenty-one volunteers were randomly assigned into a three-arm, crossover, randomized trial. Subjects were assigned to consume one of the three following treatments: one cup of espresso coffee/day (1C), three cups of espresso coffee/day (3C), and one cup of espresso coffee plus two cocoa-based products containing coffee (PC) twice per day for 1 month. At the end of each treatment, blood samples were collected for the analysis of endogenous and H2O2-induced DNA damage and DNA oxidation catabolites, while urines were used for the analysis of oxylipins. On the whole, four DNA catabolites (cyclic guanosine monophosphate (cGMP), 8-OH-2′-deoxy-guanosine, 8-OH-guanine, and 8-NO2-cGMP) were detected in plasma samples following the one-month intervention. No significant modulation of DNA and lipid damage markers was documented among groups, apart from an effect of time for DNA strand breaks and some markers of lipid peroxidation. In conclusion, the consumption of coffee and cocoa-based confectionery containing coffee was apparently not able to affect oxidative stress markers. More studies are encouraged to better explain the findings obtained and to understand the impact of different dosages of these products on specific target groups., This study was partially funded by Soremartec Italia S.r.l. (Alba, Italy). The sponsor approved the final trial protocol but was not involved in any stage of the study as well as in the drafting of the manuscript.

PMCID: PMC5666899; International audience; Argan oil is widely used in Morocco in traditional medicine. Its ability to treat cardiovascular diseases is well-established. However, nothing is known about its effects on neurodegenerative diseases, which are often associated with increased oxidative stress leading to lipid peroxidation and the formation of 7-ketocholesterol (7KC) resulting from cholesterol auto-oxidation. As 7KC induces oxidative stress, inflammation and cell death, it is important to identify compounds able to impair its harmful effects. These compounds may be either natural or synthetic molecules or mixtures of molecules such as oils. In this context: (i) the lipid profiles of dietary argan oils from Berkane and Agadir (Morocco) in fatty acids, phytosterols, tocopherols and polyphenols were determined by different chromatographic techniques; and (ii) their anti-oxidant and cytoprotective effects in 158N murine oligodendrocytes cultured with 7KC (25-50 mu M; 24 h) without and with argan oil (0.1% v/v) or -tocopherol (400 mu M, positive control) were evaluated with complementary techniques of cellular and molecular biology. Among the unsaturated fatty acids present in argan oils, oleate (C18:1 n-9) and linoleate (C18:1 n-6) were the most abundant; the highest quantities of saturated fatty acids were palmitate (C16:0) and stearate (C18:0). Several phytosterols were found, mainly schottenol and spinasterol (specific to argan oil), cycloartenol, beta-amyrin and citrostadienol alpha- and gamma-tocopherols were also present. Tyrosol and protocatechic acid were the only polyphenols detected. Argan and extra virgin olive oils have many compounds in common, principally oleate and linoleate, and tocopherols. Kit Radicaux Libres (KRL) and ferric reducing antioxidant power (FRAP) tests showed that argan and extra virgin olive oils have anti-oxidant properties. Argan oils were able to attenuate the cytotoxic effects of 7KC on 158N cells: loss of cell adhesion, cell growth inhibition, increased plasma membrane permeability, mitochondrial, peroxisomal and lysosomal dysfunction, and the induction of oxiapoptophagy (OXIdation + APOPTOsis + autoPHAGY). Altogether, our data obtained in 158N oligodendrocytes provide evidence that argan oil is able to counteract the toxic effects of 7KC on nerve cells, thus suggesting that some of its compounds could prevent or mitigate neurodegenerative diseases to the extent that they are able to cross the blood-brain barrier.

International audience; AIMS: The translocator protein (TSPO) is located on the outer mitochondrial membrane where it is responsible for the uptake of cholesterol into mitochondria of steroidogenic organs. TSPO is also present in the heart where its role remains uncertain. We recently showed that TSPO ligands reduced infarct size and improved mitochondrial functions after ischaemia-reperfusion. This study, thus, sought to determine whether cholesterol could play a role in the cardioprotective effect of TSPO ligands. METHODS AND RESULTS: In a model of 30 min coronary occlusion/15 min reperfusion in Wistar rat, we showed that reperfusion induced lipid peroxidation as demonstrated by the increase in conjugated diene and thiobarbituric acid reactive substance formation and altered mitochondrial function (decrease in oxidative phosphorylation and increase in the sensitivity of mitochondrial permeability transition pore opening) in ex-vivo isolated mitochondria. This was associated with an increase in mitochondrial cholesterol uptake (89.5 ± 12.2 vs. 39.9 ± 3.51 nmol/mg protein in controls, P < 0.01) and a subsequent strong generation of auto-oxidized oxysterols, i.e. 7α- and 7β-hydroxycholesterol, 7-ketocholesterol, cholesterol-5α,6α-epoxide, and 5β,6β-epoxide (+173, +149, +165, +165, and +193% vs. controls, respectively; P < 0.01). Administration of the selective TSPO ligand 4'-chlorodiazepam inhibited oxidative stress, improved mitochondrial function, and abolished both mitochondrial cholesterol accumulation and oxysterol production. This was also observed with the new TSPO ligand TRO40303. CONCLUSION: These data suggest that 4'-chlorodiazepam inhibits oxidative stress and oxysterol formation by reducing the accumulation of cholesterol in the mitochondrial matrix at reperfusion and prevents mitochondrial injury. This new and original mechanism may contribute to the cardioprotective properties of TSPO ligands.

International audience; The present study was undertaken to determine whether subacute treatment with aqueous extract of carob (Ceratonia siliqua L.) pods (AECPs) protects against ethanol (EtOH)-induced oxidative stress in rat liver. Animals were divided into four groups: control, carob, EtOH and EtOH+carob. Wistar rats were intraperitoneally pretreated with AECP (600mg/kg body weight (bw)) during 7days and intoxicated for 6h by acute oral administration of EtOH (6g/kg bw) 24h after the last injection. We found that acute administration of EtOH leads to hepatotoxicity as monitored by the increase in the levels of hepatic marker aspartate aminotransferase and alanine aminotransferase as well as hepatic tissue injury. EtOH also increased the formation of malondialdehyde in the liver, indicating an increase in lipid peroxidation and depletion of antioxidant enzyme activities as superoxide dismutase, catalase and glutathione peroxidase. Subacute carob pretreatment prevented all the alterations induced by EtOH and returned their levels to near normal. Importantly, we showed that acute alcohol increased hepatic and plasmatic hydrogen peroxide and free iron levels. The carob pretreatment reversed EtOH effects to near control levels. These data suggest that carob could have a beneficial effect in inhibiting the oxidative damage induced by acute EtOH administration and that its mode of action may involve an opposite effect on plasma and tissue-free iron accumulation. Indeed, carob can be offered as a food additive to protect against EtOH-induced oxidative damage.

ISI Document Delivery No.: 004QQTimes Cited: 0Cited Reference Count: 46Cited References: Pillon Nicolas J., 2011, CHEMICAL RESEARCH IN TOXICOLOGY, V24, P752Awada, Manar Soulage, Christophe O. Meynier, Anne Debard, Cyrille Plaisancie, Pascale Benoit, Berengere Picard, Gregory Loizon, Emmanuelle Chauvin, Marie-Agnes Estienne, Monique Peretti, Noel Guichardant, Michel Lagarde, Michel Genot, Claude Michalski, Marie-CarolineFrench National Research Agency (ANR)[ANR-08-ALIA-002]; ANRThis work was supported by the French National Research Agency (ANR) Grant ANR-08-ALIA-002, AGECANINOX project.M. Awada acknowledges her PhD grant from ANR. The authors thank Lucie Ribourg and Michele Viau for their work on oil mixtures. Patricia Daira is acknowledged for her help with the 4-hydroxy-2-alkenal assay. The authors thank Jean-Michel Chardigny for useful discussions. Isabelle Jouanin (Plateforme AXIOM-MetaToul, Toulouse) is acknowledged for the synthesis of internal standards to measure 4-hydroxyl-2-alkenals in fat. Yvonne Masson is acknowledged for revising the English-language manuscript.Amer soc biochemistry molecular biology incBethesda[Awada, Manar; Plaisancie, Pascale; Estienne, Monique; Michalski, Marie-Caroline] INRA, CarMeN Lab U1235, F-69621 Villeurbanne, France. [Awada, Manar; Soulage, Christophe O.; Guichardant, Michel; Lagarde, Michel; Michalski, Marie-Caroline] Inst Natl Sci Appl, IMBL, F-69621 Villeurbanne, France. [Meynier, Anne; Genot, Claude] INRA, Biopolymeres Interact Assemblages UR1268, F-44300 Nantes, France. [Debard, Cyrille; Chauvin, Marie-Agnes] INSERM, U1060, CarMeN Lab, F-69921 Oullins, France. [Benoit, Berengere; Picard, Gregory; Loizon, Emmanuelle; Peretti, Noel; Michalski, Marie-Caroline] Univ Lyon 1, F-69622 Villeurbanne, France.marie-caroline.michalski@insa-lyon.fr; International audience; Dietary intake of long-chain n-3 PUFA is now widely advised for public health and in medical practice. However, PUFA are highly prone to oxidation, producing potentially deleterious 4-hydroxy-2-alkenals. Even so, the impact of consuming oxidized n-3 PUFA on metabolic oxidative stress and inflammation is poorly described. We therefore studied such effects and hypothesized the involvement of the intestinal absorption of 4-hydroxy-2-hexenal (4-HHE), an oxidized n-3 PUFA end-product. In vivo, four groups of mice were fed for 8 weeks high-fat diets containing moderately oxidized or unoxidized n-3 PUFA. Other mice were orally administered 4-HHE and euthanized postprandially versus baseline mice. In vitro, human intestinal Caco-2/TC7 cells were incubated with 4-hydroxy-2-alkenals. Oxidized diets increased 4-HHE plasma levels in mice (up to 5-fold, P < 0.01) compared with unoxidized diets. Oxidized diets enhanced plasma inflammatory markers and activation of nuclear factor kappaB (NF-κB) in the small intestine along with decreasing Paneth cell number (up to -19% in the duodenum). Both in vivo and in vitro, intestinal absorption of 4-HHE was associated with formation of 4-HHE-protein adducts and increased expression of glutathione peroxidase 2 (GPx2) and glucose-regulated protein 78 (GRP78). Consumption of oxidized n-3 PUFA results in 4-HHE accumulation in blood after its intestinal absorption and triggers oxidative stress and inflammation in the upper intestine.

International audience; Prolonged strenuous exercise (PSE) induces transient left ventricular (LV) dysfunction. Previous studies suggest that β-adrenergic pathway desensitization could be involved in this phenomenon, but it remains to be confirmed. Moreover, other underlying mechanisms involving oxidative stress have been recently proposed. The present study aimed to evaluate the involvement of both the β-adrenergic pathway and NADPH oxidase (Nox) enzyme-induced oxidative stress in myocardial dysfunction in rats following PSE. Rats were divided into 4 groups: controls (Ctrl), 4-h exercised on treadmill (PSE), and 2 groups in which Nox enzyme was inhibited with apocynin treatment (Ctrl APO and PSE APO, respectively). We evaluated cardiac function in vivo and ex vivo during basal conditions and isoproterenol stress. GSH/GSSG ratio, cardiac troponin I (cTnI) release, and lipid peroxidation (MDA) were evaluated. PSE induced a decrease in LV developed pressure, intrinsic myocardial contractility, and relaxation associated with an increase in plasma cTnI release. Our in vivo and ex vivo results demonstrated no differences in myocardial response to isoproterenol and of effective dose 50 between control and PSE rats. Interestingly, the LV dysfunction was reversed by apocynin treatment. Moreover, apocynin prevented cellular oxidation [GSH/GSSG ratio: PSE APO rats vs. PSE rats in arbitrary units (au): 1.98 ± 0.07 vs. 1.35 ± 0.10; P < 0.001]. However, no differences in MDA were observed between groups. These data suggest that myocardial dysfunction observed after PSE was not due to β-adrenergic receptor desensitization but could be due to a signaling oxidative stress from the Nox enzyme.

International audience; Acrolein is the most reactive aldehyde among the by-products of lipid peroxidation. Growing evidence indicates that acrolein may play an important role in the pathogenesis of Alzheimer's disease (AD). In AD, levels of acrolein are significantly higher in hippocampus and temporal cortex regions of the brain. However, little is known about its toxicity in neuronal cells. Using the neuroblastoma cell line SK-N-SH, our results show that acrolein is toxic from 10 μM, but its toxicity does not induce the activation of caspase-3 and DNA fragmentation. Protein carbonylation and 4-hydroxynonenal levels were increased after 0.5 hr and 1 hr of treatment, respectively. Furthermore acrolein induced a biphasic effect on glutathione levels with a rapid depletion followed by a progressive increase. We have further investigated the regulation of different redox signaling pathways. A treatment with 10 μM of acrolein for 30 min activated NFκB while this activation was suppressed after a 24 hrs of treatment. In contrast, Nrf2 was activated only after 24 hrs of acrolein exposure. Consequently, the expression of heme oxygenase-1 and γ-glutamyl-cysteine-synthase were elevated after 24 hrs of acrolein treatment. Sirt-1 was also upregulated after 24 hrs of acrolein treatment. The p66Shc and ERK1/2 proteins are known to be involved in oxidative stress. Acrolein, at 10 μM, induced the phosphorylation of p66Shc and ERK1/2 only after a short period of treatment. Collectively, these data strengthen the contribution of acrolein in the induction of oxidative stress as observed in mild cognitive impairment and in AD brain.

International audience; The antidiabetic drug metformin can diminish apoptosis induced by oxidative stress in endothelial cells and prevent vascular dysfunction even in nondiabetic patients. Here we tested whether it has a beneficial effect in a rat model of gentamicin toxicity. Mitochondrial analysis, respiration intensity, levels of reactive oxygen species, permeability transition, and cytochrome c release were assessed 3 and 6 days after gentamicin administration. Metformin treatment fully blocked gentamicin-mediated acute renal failure. This was accompanied by a lower activity of N-acetyl-beta-D-glucosaminidase, together with a decrease of lipid peroxidation and increase of antioxidant systems. Metformin also protected the kidney from histological damage 6 days after gentamicin administration. These in vivo markers of kidney dysfunction and their correction by metformin were complemented by in vitro studies of mitochondrial function. We found that gentamicin treatment depleted respiratory components (cytochrome c, NADH), probably due to the opening of mitochondrial transition pores. These injuries, partly mediated by a rise in reactive oxygen species from the electron transfer chain, were significantly decreased by metformin. Thus, our study suggests that pleiotropic effects of metformin can lessen gentamicin nephrotoxicity and improve mitochondrial homeostasis.

Thrombin inhibition protects against liver fibrosis. However, it is not known whether the thrombin profibrogenic effect is due to effects on blood coagulation or to signaling via protease-activated receptors (PARs). We took advantage of the lack of blood coagulation defects in PAR-1-knockout mice. Acute carbon tetrachloride (CCl4) toxicity was similar in wild-type (WT), PAR-1−/−, and PAR-1+/−mice as judged by aminotransferase levels, area of liver necrosis, and liver peroxidation measured by Fourier-transformed infrared spectroscopy. Fifteen mice/group received CCl4or its solvent for 6 wk (300 μl/kg, 3 times a week). Fibrosis area was increased 10-fold by CCl4treatment in WT mice. PAR-1 deficiency protected against fibrosis, with 36% and 56% decrease in PAR-1+/−and PAR-1−/−mice, respectively ( P < 0.001). Similar results were obtained for area of activated fibrogenic cells (64% and 79% decrease in PAR-1+/−and PAR-1−/−mice, respectively, P < 0.001). These findings were corroborated by measurements of type I collagen, matrix metalloproteinase-2, and PDGF-β receptor mRNA levels. There was also a significant decrease in T lymphocyte infiltration in PAR-1-deficient mice. Altogether, these results suggest that thrombin profibrogenic effects are independent of effects on blood coagulation and are instead due to direct effects on fibrogenic cells and possibly on T lymphocytes.

The purpose of this study was to examine the impacts of anthropogenic activity on the health status of intertidal clam populations of the Saguenay Fjord and the St. Lawrence Estuary (Québec, Canada). Clams were collected during low tide at sites subject to direct contamination and at sites far from human activity. Clams were analyzed for tributyltin and dibutyltin total levels and toxic stress (glutathione S-transferase, gonadal lipid peroxidation and DNA strand breaks), immunocompetence (phagocytic activity, hemocyte count and viability), reproduction (gonado-somatic index, gamete maturation, and vitellogenin-like proteins), energy status (temperature-dependent mitochondrial electron transport, and gonad lipids), and individual status (age, condition factor, and growth index). These responses were compared against population characteristics such as live clam density, number of empty shells, and sex ratio. The results show that clam density decreased with distance from the estuary (high salinity level) to upstream of the fjord (low salinity). There was no clear relationship between the number of empty shells and distance or site quality. Clam density values corrected against distance were significantly correlated with hemocyte viability, phagocytic activity, mitochondrial electron transport (MET), DNA damage in gonad, and temperature-dependent mitochondrial electron transport activity. A canonical analysis of the various groups of biomarkers revealed that population metrics were more strongly related with immunocompetence, followed by energy status and temperature-dependent mitochondrial electron transport activity. However, toxic stress biomarkers were strongly associated with energy status and reproduction. This was further confirmed by non-linear modeling using adaptive artificial neural networks (genetic selection and back propagation learning paradigms), where the following parameters were able to predict population parameters with

Nitrated-polycyclic aromatic hydrocarbons (nitro-PAHs) and diesel exhaust particle extracts (DEPE) induced apoptosis in Hepa1c1c7 cells with the following potency: 1,3-dinitropyrene (1,3-DNP)>1-nitropyrene (1-NP) >> DEPE >> 1,8-dinitropyrene (1,8-DNP). The compounds induced cyp1a1, and activated various intracellular signalling pathways related to apoptosis. The CYP inhibitor alpha-naphthoflavone strongly reduced 1,3-DNP-induced cell death, whereas cell death induced by 1-NP was rather increased. Toxic 1,3-DNP and 1-NP were found to induce a concentration-dependent lipid peroxidation. 1,3-DNP caused pro-apoptotic events, including increased phosphorylation and accumulation of p53 in the nucleus, cleavage of bid and of caspases 8 and 3, down-regulation of bcl-x(L) and phosphorylation of p38 and JNK MAPK. Furthermore, 1,3-DNP increased the activation of survival signals including phosphorylation of Akt and inactivation (phosphorylation) of pro-apoptotic bad. Although less potent, rather similar effects were observed following exposure to DEPE, compared to 1-NP. The most important finding was that the most mutagenic and carcinogenic compound tested, 1,8-DNP, induced little (if any) cell death, despite the fact that this compound seemed to give the most DNA damage as judged by DNA adduct formation, increased phosphorylation of p53 and accumulation of cells in S-phase. Immunocytochemical studies revealed that the p53 protein did not accumulate into the nucleus suggesting that 1,8-DNP inactivated the pro-apoptotic function of the p53 protein by a non-mutagenic event. These results suggest that after exposure to 1,8-DNP more cells may survive with DNA damage, thereby increasing its mutagenic and carcinogenic potential. The study has been supported in part by The Research Council of Norway, by Aurora grant (Egide) and Cancer Research UK; and Volker M. Arlt is a member of the European ECNIS (Environmental Cancer Risk, Nutrition and Individual Susceptibility) Network of Excellence. The technical assistance of Leni Johanne Ekeren and Trine Isaksen is very much appreciated. We also thank Dr. Trine Husøy for help with immunocytochemistry and Dr. Per E. Schwarze for support and critical reading of the manuscript.

International audience; In recent years there has been enormous interest in researching oxidative stress. Reactive oxygen species (ROS) are derived from the metabolism of oxygen as by-products of cell respiration, and are continuously produced in all aerobic organisms. Oxidative stress occurs as a consequence of an imbalance between ROS production and the available antioxidant defence against them. Nowadays, a variety of diseases and degenerative processes such as cancer, Alzheimer’s and autoimmune diseases are mediated by oxidative stress. Heat stress was suggested to be an environmental factor responsible for stimulating ROS production because of similarities in responses observed following heat stress compared with that occurring following exposure to oxidative stress. This manuscript describes the main mitochondrial sources of ROS and the antioxidant defences involved to prevent oxidative damage in all the mitochondrial compartments. It also deals with discussions concerning the cytotoxic effect of heat stress, mitochondrial heat-induced alterations, as well as heat shock protein (HSP) expression as a defence mechanism.

Polyunsaturated fatty acids have been reported to enhance the cytotoxic activity of several anticancer drugs. In the present study, we observed that doxorubicin chemosensitization of breast cancer cell lines by docosahexaenoic acid (DHA, a long-chain omega-3 polyunsaturated fatty acid) was cell-line selective, affecting MDA-MB-231 and MCF-7 dox (a doxorubicin-resistant cell line) but not the parental MCF-7 cell line. DHA supplementation led to an increase in membrane phospholipid DHA level, but did not induce changes in intracellular [(14)C]doxorubicin accumulation. In MDA-MB-231, doxorubicin efficacy enhancement by DHA was linked to an increase in malondialdehyde level, a final product of lipid peroxidation. DHA elicited by itself a 3.7-fold malondialdehyde level increase, additive to that induced by doxorubicin. Addition of doxorubicin to DHA further increased the glutathione level, indicative of the generation of an oxidative stress. In contrast to MDA-MB-231, doxorubicin did not increase the malondialdehyde level in MCF-7, although DHA induced lipid peroxidation. Therefore in MCF-7, lipid peroxidation induced by DHA itself was not sufficient to trigger an oxidative stress and to subsequently increase sensitivity to doxorubicin. These data indicate that the differential effect of DHA among cells on drug toxicity results from a differential oxidative response to doxorubicin. Chemosensitization through fatty acids appears as a new promising adjuvant therapeutic paradigm, since omega-3 fatty acids are physiological molecules found in food and are nontoxic in vivo.

International audience; For a better understanding of the role of iron imbalance in neuropathology, a liposoluble iron complex (ferric hydroxyquinoline, FHQ) was injected into striatum of rats. The effects of two modalities of iron injections on brain damage, hydroxyl radical (*OH) production (assessed by the salicylate method coupled to microdialysis) and tissue reactive iron level (evaluated ex vivo by the propensity of the injected structure for lipid peroxidation) were examined. Rapid injection of FHQ (10 nmoles of 5 mM FHQ pH 3 solution over 1-min period) but not that of corresponding vehicle led to extensive damage associated with increased tissue free iron level in the injected region. Conversely, neither lesion nor free iron accumulation was observed after slow FHQ injection (10 nmoles of a 100 microM FHQ pH 7 solution over 1-h period) as compared to corresponding vehicle injection. Production of *OH was induced by slow FHQ injection but not by rapid FHQ injection, probably as a result of in vivo abolition of iron-induced *OH formation by acid pH. Indeed, rapid injection of FAC pH 7 (ferric ammonium citrate, 5 mM in saline) was associated with *OH formation whereas rapid injection of FAC pH 3 did not. Our results identify the rate of iron delivery to cells as an important determinant of iron toxicity and do not support a major role for extracellular *OH in damage associated with intracerebral iron injection.

ISI Document Delivery No.: 004QQTimes Cited: 0Cited Reference Count: 46Cited References: Pillon Nicolas J., 2011, CHEMICAL RESEARCH IN TOXICOLOGY, V24, P752Awada, Manar Soulage, Christophe O. Meynier, Anne Debard, Cyrille Plaisancie, Pascale Benoit, Berengere Picard, Gregory Loizon, Emmanuelle Chauvin, Marie-Agnes Estienne, Monique Peretti, Noel Guichardant, Michel Lagarde, Michel Genot, Claude Michalski, Marie-CarolineFrench National Research Agency (ANR)[ANR-08-ALIA-002]; ANRThis work was supported by the French National Research Agency (ANR) Grant ANR-08-ALIA-002, AGECANINOX project.M. Awada acknowledges her PhD grant from ANR. The authors thank Lucie Ribourg and Michele Viau for their work on oil mixtures. Patricia Daira is acknowledged for her help with the 4-hydroxy-2-alkenal assay. The authors thank Jean-Michel Chardigny for useful discussions. Isabelle Jouanin (Plateforme AXIOM-MetaToul, Toulouse) is acknowledged for the synthesis of internal standards to measure 4-hydroxyl-2-alkenals in fat. Yvonne Masson is acknowledged for revising the English-language manuscript.Amer soc biochemistry molecular biology incBethesda[Awada, Manar; Plaisancie, Pascale; Estienne, Monique; Michalski, Marie-Caroline] INRA, CarMeN Lab U1235, F-69621 Villeurbanne, France. [Awada, Manar; Soulage, Christophe O.; Guichardant, Michel; Lagarde, Michel; Michalski, Marie-Caroline] Inst Natl Sci Appl, IMBL, F-69621 Villeurbanne, France. [Meynier, Anne; Genot, Claude] INRA, Biopolymeres Interact Assemblages UR1268, F-44300 Nantes, France. [Debard, Cyrille; Chauvin, Marie-Agnes] INSERM, U1060, CarMeN Lab, F-69921 Oullins, France. [Benoit, Berengere; Picard, Gregory; Loizon, Emmanuelle; Peretti, Noel; Michalski, Marie-Caroline] Univ Lyon 1, F-69622 Villeurbanne, France.marie-caroline.michalski@insa-lyon.fr; International audience; Dietary intake of long-chain n-3 PUFA is now widely advised for public health and in medical practice. However, PUFA are highly prone to oxidation, producing potentially deleterious 4-hydroxy-2-alkenals. Even so, the impact of consuming oxidized n-3 PUFA on metabolic oxidative stress and inflammation is poorly described. We therefore studied such effects and hypothesized the involvement of the intestinal absorption of 4-hydroxy-2-hexenal (4-HHE), an oxidized n-3 PUFA end-product. In vivo, four groups of mice were fed for 8 weeks high-fat diets containing moderately oxidized or unoxidized n-3 PUFA. Other mice were orally administered 4-HHE and euthanized postprandially versus baseline mice. In vitro, human intestinal Caco-2/TC7 cells were incubated with 4-hydroxy-2-alkenals. Oxidized diets increased 4-HHE plasma levels in mice (up to 5-fold, P < 0.01) compared with unoxidized diets. Oxidized diets enhanced plasma inflammatory markers and activation of nuclear factor kappaB (NF-κB) in the small intestine along with decreasing Paneth cell number (up to -19% in the duodenum). Both in vivo and in vitro, intestinal absorption of 4-HHE was associated with formation of 4-HHE-protein adducts and increased expression of glutathione peroxidase 2 (GPx2) and glucose-regulated protein 78 (GRP78). Consumption of oxidized n-3 PUFA results in 4-HHE accumulation in blood after its intestinal absorption and triggers oxidative stress and inflammation in the upper intestine.

International audience; OBJECTIVES: Within the past decade, biochemical markers have emerged as attractive tools to assess pulmonary arterial hypertension (PAH) prognosis, being noninvasive and easily repeatable.The objective of this study was to determine whether biomarkers measured at initial diagnostic right-sided heart catheterization predict 3-year all-cause mortality for incident cases of PAH independently of clinical and hemodynamic parameters. METHODS: Patients with incident PAH were enrolled between December 2003 and April 2006 in six centers from the French Network on Pulmonary Hypertension and followed for 3 years.Venous blood samples were taken during right-sided heart catheterization, and analyses were centralized. RESULTS: Among 110 enrolled patients, 11 underwent lung or heart/lung transplantation, and 27 died during follow-up. The Kaplan-Meier estimates of survival were 91%, 78%, and 75% at 1, 2, and 3 years, respectively. Plasma big endothelin-1 (hazard ratio [HR] per 1-SD increase, 1.48; 95% CI,1.14-1.92), serum troponin T . 0.01 mg/L (HR, 2.35; 95% CI, 1.05-5.29), and urinary F 2 -isoprostanes(15-F2t -isoprostane) (HR per 1-SD increase, 1.76; 95% CI, 1.31-2.36) were associated with increased unadjusted hazard of death. In multivariate analysis adjusting for patient characteristics, the level of urinary F 2 -isoprostanes was the only biomarker that remained independently associated with increased hazard of death (HR per 1-SD increase, 1.82; 95% CI, 1.28-2.60). CONCLUSIONS: This study shows that levels of urinary F 2 -isoprostane, a biomarker of lipid peroxidation,quantified at initial diagnostic right-sided heart catheterization are independently associated with mortality in a cohort of patients with incident PAH.

International audience; Bisphenol A (BPA) is an endocrine-disrupting chemical that leaches from polycarbonate plastics that consequently leads to low-dose human exposure. In addition to its known xenoendocrine action, BPA exerts a wide variety of metabolic effects, but no data are available on its actions on the functions of liver mitochondrial. To assess these effects, HepG2 cells were exposed to BPA (10(-4)-10(-12)M) and physiological parameters were measured by flow cytometry. We demonstrated a significant mitochondrial dysfunction including ROS production, ΔΨ(M) hyperpolarization, lipid accumulation, lipoperoxidation and the release of pro-inflammatory cytokines. In conclusion, we showed that low concentrations of BPA promote lipid accumulation in hepatic cells triggered by disturbances in mitochondrial function, alterations in lipid metabolism and by inflammation that can therefore contribute to steatosis.

ISI Document Delivery No.: 004QQTimes Cited: 0Cited Reference Count: 46Cited References: Pillon Nicolas J., 2011, CHEMICAL RESEARCH IN TOXICOLOGY, V24, P752Awada, Manar Soulage, Christophe O. Meynier, Anne Debard, Cyrille Plaisancie, Pascale Benoit, Berengere Picard, Gregory Loizon, Emmanuelle Chauvin, Marie-Agnes Estienne, Monique Peretti, Noel Guichardant, Michel Lagarde, Michel Genot, Claude Michalski, Marie-CarolineFrench National Research Agency (ANR)[ANR-08-ALIA-002]; ANRThis work was supported by the French National Research Agency (ANR) Grant ANR-08-ALIA-002, AGECANINOX project.M. Awada acknowledges her PhD grant from ANR. The authors thank Lucie Ribourg and Michele Viau for their work on oil mixtures. Patricia Daira is acknowledged for her help with the 4-hydroxy-2-alkenal assay. The authors thank Jean-Michel Chardigny for useful discussions. Isabelle Jouanin (Plateforme AXIOM-MetaToul, Toulouse) is acknowledged for the synthesis of internal standards to measure 4-hydroxyl-2-alkenals in fat. Yvonne Masson is acknowledged for revising the English-language manuscript.Amer soc biochemistry molecular biology incBethesda[Awada, Manar; Plaisancie, Pascale; Estienne, Monique; Michalski, Marie-Caroline] INRA, CarMeN Lab U1235, F-69621 Villeurbanne, France. [Awada, Manar; Soulage, Christophe O.; Guichardant, Michel; Lagarde, Michel; Michalski, Marie-Caroline] Inst Natl Sci Appl, IMBL, F-69621 Villeurbanne, France. [Meynier, Anne; Genot, Claude] INRA, Biopolymeres Interact Assemblages UR1268, F-44300 Nantes, France. [Debard, Cyrille; Chauvin, Marie-Agnes] INSERM, U1060, CarMeN Lab, F-69921 Oullins, France. [Benoit, Berengere; Picard, Gregory; Loizon, Emmanuelle; Peretti, Noel; Michalski, Marie-Caroline] Univ Lyon 1, F-69622 Villeurbanne, France.marie-caroline.michalski@insa-lyon.fr; International audience; Dietary intake of long-chain n-3 PUFA is now widely advised for public health and in medical practice. However, PUFA are highly prone to oxidation, producing potentially deleterious 4-hydroxy-2-alkenals. Even so, the impact of consuming oxidized n-3 PUFA on metabolic oxidative stress and inflammation is poorly described. We therefore studied such effects and hypothesized the involvement of the intestinal absorption of 4-hydroxy-2-hexenal (4-HHE), an oxidized n-3 PUFA end-product. In vivo, four groups of mice were fed for 8 weeks high-fat diets containing moderately oxidized or unoxidized n-3 PUFA. Other mice were orally administered 4-HHE and euthanized postprandially versus baseline mice. In vitro, human intestinal Caco-2/TC7 cells were incubated with 4-hydroxy-2-alkenals. Oxidized diets increased 4-HHE plasma levels in mice (up to 5-fold, P < 0.01) compared with unoxidized diets. Oxidized diets enhanced plasma inflammatory markers and activation of nuclear factor kappaB (NF-κB) in the small intestine along with decreasing Paneth cell number (up to -19% in the duodenum). Both in vivo and in vitro, intestinal absorption of 4-HHE was associated with formation of 4-HHE-protein adducts and increased expression of glutathione peroxidase 2 (GPx2) and glucose-regulated protein 78 (GRP78). Consumption of oxidized n-3 PUFA results in 4-HHE accumulation in blood after its intestinal absorption and triggers oxidative stress and inflammation in the upper intestine.

International audience; Ischemia/reperfusion injury occurring during liver transplantation is mainly due to the generation of reactive oxygen species (ROS) upon revascularization. Thus, delivery of antioxidant enzymes might reduce the deleterious effects of ROS and improve liver graft initial function. Mangafodipir trisodium (MnDPDP), a contrast agent currently used in magnetic resonance imaging of the liver, has been shown to be endowed with powerful antioxidant properties. We hypothesized that MnDPDP could have a protective effect against liver ischemia reperfusion injury when administrated to the donor prior to harvesting. Livers from Sprague Dawley rats pretreated or not with MnDPDP were harvested and subsequently preserved for 24 h in Celsior® solution at 4°C. Organs were then perfused ex vivo for 120 min at 37°C with Krebs Henseleit solution. In MnDPDP (5 µmol/kg) group, we observed that ATP content was significantly higher at the end of the cold preservation period relative to untreated group. After reperfusion, livers from MnDPDP-treated rats showed better tissue integrity, less hepatocellular and endothelial cell injury. This was accompanied by larger amounts of bile production and higher ATP recovery as compared to untreated livers. The protective effect of MnDPDP was associated with a significant decrease of lipid peroxidation, mitochondrial damage, and apoptosis. Interestingly, MnDPDP-pretreated livers exhibited activation of Nfr2 and HIF-1α pathways resulting in a higher catalase and HO-1 activities. MnDPDP also increased total nitric oxide (NO) production which derived from higher expression of constitutive NO synthase and lower expression of inducible NO synthase. In conclusion, our results show that donor pretreatment with MnDPDP protects the rat liver graft from cold ischemia/reperfusion injury and demonstrate for the first time the potential interest of this molecule in the field of organ preservation. Since MnDPDP is safely used in liver imaging, this preservation strategy holds great promise for translation to clinical liver transplantation.

AIR:EAU+SPI:CGU; In order to compare the disinfection potential of photocatalysis and photochemistry, the effects of these two processes on bacteria in water were investigated under exposure to UV-A and UV-C. The well-known bacterial model Escherichia coli (E. coli) was used as the experimental organism. Radiation exposure was produced with an HPK 125 W lamp and the standard TiO2 Degussa P-25 was used as the photocatalyst. Firstly, the impact of photocatalysis and photochemistry on the cultivability of bacterial cells was investigated. UV-A radiation resulted in low deleterious effects on bacterial cultivability but generated colonies of size smaller than average. UV-C photocatalysis demonstrated a greater efficiency than UV-A photocatalysis in altering bacterial cultivability. From a cultivability point of view only, UV-C radiation appeared to be the most deleterious treatment. A rapid epifluorescence staining method using the LIVE/DEAD Bacterial Viability Kit was then used to assess the modifications in bacterial membrane permeability. UV-A radiation did not induce any alterations in bacterial permeability for 420 min of exposure whereas only a few minutes of exposure to UV-C radiation, with the same total radiance intensity, induced total loss of permeability. Moreover, after 20 and 60 min of exposure to UV-C and UV-A photocatalysis respectively, all bacteria lost their membrane integrity, suggesting that the bacterial envelope is the primary target of reactive oxygen species (ROS) generated at the surface of TiO2 photocatalyst. These results were further confirmed by the formation of malondialdehyde (MDA) during the photocatalytic inactivation of bacterial cells and suggest that destruction of the cell envelope is a key step in the bactericidal action of photocatalysis. The oxidation of bacterial membrane lipids was also correlated with the monitoring of carboxylic acids, which can be considered as representatives of lipid per-oxidation by-products. Finally, damages to bacterial morphology induced by UV-C photocatalysis and photochemistry were investigated through Scanning electron microscopy (SEM). Bacterial cells were observed on microscopy pictures at exposure durations corresponding to a loss of cultivability. After 90 min of exposure to UV-C radiation, bacterial cells showed little alteration of their outer membrane whereas they suffered deep deleterious damages under UV-C photocatalysis exposure. (c) 2012 Elsevier Ltd. All rights reserved.

International audience; HIV-infected patients and transplanted patients share similar immunosuppressed status. Recent insights gained through the field of heart transplantation may help to clarify the role of reactive oxygen species in HIV-infected patients.

International audience; The calcineurin-inhibitors (CNIs) cyclosporine (CsA) and tacrolimus (TAC) remain the pillars of modern immunosuppression regimens used in solid organ transplantation. Nephrotoxicity is an adverse effect that limits their successful use. The precise molecular mechanisms underlying this nephrotoxicity remain unclear. Using SILAC together with LC-MALDI-TOF/TOF, we investigated the CNIs-induced proteomic perturbations in renal cells. Among the 495 proteins quantifiable in both forward and reverse SILAC, 69 displayed CsA-induced perturbations: proteins involved in ER-stress/protein folding, apoptosis, metabolism/transport or cytoskeleton pathways were up-regulated, while cyclophilin B as well as nuclear and RNA-processing proteins were down-regulated. Co-administration of CsA with the antioxidant N-acetylcysteine significantly decreased lipid peroxidation and also partially corrected the CsA-induced unfolded protein response. TAC toxicity profile was apparently different from that of CsA, especially without perturbation of cyclophilins A and B, up-regulation of ER-chaperones nor down-regulation of a number of nuclear proteins. These results provide a new insight and are consistent with recent data regarding the molecular mechanisms of CNIs-induced nephrotoxicity. Our findings offer new directions for future research aiming to identify specific biomarkers of CsA nephrotoxicity.

International audience; Although carbon tetrachloride (CCl(4))-induced acute and chronic hepatotoxicity have been extensively studied, little is known about the very early in vivo effects of this organic solvent on oxidative stress and mitochondrial function. In this study, mice were treated with CCl(4) (1.5 ml/kg ie 2.38 g/kg) and parameters related to liver damage, lipid peroxidation, stress/defense and mitochondria were studied 3 h later. Some CCl(4)-intoxicated mice were also pretreated with the cytochrome P450 2E1 inhibitor diethyldithiocarbamate or the antioxidants Trolox C and dehydroepiandrosterone. CCl(4) induced a moderate elevation of aminotransferases, swelling of centrilobular hepatocytes, lipid peroxidation, reduction of cytochrome P4502E1 mRNA levels and a massive increase in mRNA expression of heme oxygenase-1 and heat shock protein 70. Moreover, CCl(4) intoxication induced a severe decrease of mitochondrial respiratory chain complex IV activity, mitochondrial DNA depletion and damage as well as ultrastructural alterations. Whereas DDTC totally or partially prevented all these hepatic toxic events, both antioxidants protected only against liver lipid peroxidation and mitochondrial damage. Taken together, our results suggest that lipid peroxidation is primarily implicated in CCl(4)-induced early mitochondrial injury. However, lipid peroxidation-independent mechanisms seem to be involved in CCl(4)-induced early hepatocyte swelling and changes in expression of stress/defense-related genes. Antioxidant therapy may not be an efficient strategy to block early liver damage after CCl(4) intoxication.

International audience; AIMS/HYPOTHESIS: This study assessed oxidative stress in LDL from obese patients with the metabolic syndrome and compared it with that in LDL from type 2 diabetic patients or control volunteers. It also determined the effect on platelets of LDL from the three groups. METHODS: The profiles of lipids, fatty acids and fatty acid oxidation products were determined in LDL isolated from plasma of patients with the metabolic syndrome, patients with type 2 diabetes and volunteers (n = 10 per group). The effects of LDL from the participant groups on the platelet arachidonic acid signalling cascade and aggregation were investigated. RESULTS: Compared with LDL from control volunteers, LDL from obese metabolic syndrome and type 2 diabetic patients had lower cholesteryl ester, higher triacylglycerol and lower ethanolamine plasmalogen levels. Proportions of linoleic acid were decreased in phosphatidylcholine and cholesteryl esters in LDL from both patient groups. Among the markers of lipid peroxidation, oxidation products of linoleic acid (hydroxy-octadecadienoic acids) and malondialdehyde were increased by 59% and twofold, respectively in LDL from metabolic syndrome and type 2 diabetic patients. LDL from metabolic syndrome and type 2 diabetic patients were equally potent in activating the platelet arachidonic acid signalling cascade through increased phosphorylation of p38 mitogen-activated protein kinase and cytosolic phospholipase A(2), and through increased thromboxane B(2) formation. LDL from patients with the metabolic syndrome and type 2 diabetes potentiated platelet aggregation by threefold and 3.5-fold respectively, whereas control LDL had no activating effects on platelets. CONCLUSIONS/INTERPRETATION: The metabolic syndrome in obese patients, without or with diabetes, is associated with increased oxidative stress in LDL, which triggers platelet activation.

International audience; The effect of meat consumption on cancer risk is a controversial issue. However, recent meta-analyses show that high consumers of cured meats and red meat are at increased risk of colorectal cancer. This increase is significant but modest (20-30%). Current WCRF-AICR recommendations are to eat no more than 500 g per week of red meat, and to avoid processed meat. Moreover, our studies show that beef meat and cured pork meat promote colon carcinogenesis in rats. The major promoter in meat is heme iron, via N-nitrosation or fat peroxidation. Dietary additives can suppress the toxic effects of heme iron. For instance, promotion of colon carcinogenesis in rats by cooked, nitrite-treated and oxidized high-heme cured meat was suppressed by dietary calcium and by α-tocopherol, and a study in volunteers supported these protective effects in humans. These additives, and others still under study, could provide an acceptable way to prevent colorectal cancer.

International audience; Dispersant use in nearshore areas is likely to increase the exposure of aquatic organisms to petroleum. To measure the toxicity of this controversial response technique, golden grey mullets (Liza aurata) were exposed to mechanically dispersed oil, chemically dispersed oil, dispersant alone in seawater, water-soluble fraction of oil and to seawater as a control treatment. Several biomarkers were assessed in the gills (enzymatic antioxidant activities, glutathione content, lipid peroxidation) and in the gallbladder (polycylic aromatic hydrocarbons metabolites). The significant differences between chemically dispersed oil and water soluble fraction of oil highlight the environmental risk to disperse an oil slick when containment and recovery can be conducted. The lack of significance between chemically and mechanically dispersed oil suggests that dispersant application is no more toxic than the natural dispersion of the oil slick. The results of this study are of interest in order to establish dispersant use policies in nearshore areas.

International audience; The role of hepatocyte apoptosis in the physiopathology of obstructive cholestasis is still controversial. Although some data have strongly suggested that hepatocellular cholestatic injury is due to Fas-mediated hepatocyte apoptosis, some others concluded that necrosis, rather than apoptosis, represents the main type of hepatocyte death in chronic cholestasis. Moreover, it has also been suggested that the reduced liver injury observed in the absence of Fas receptor after bile duct ligation was not due to lower hepatocyte apoptosis but to the indirect role of this receptor in non-hepatocytic cells such as cholangiocytes and inflammatory cells. The aim of this work was therefore to determine whether a protection against cell death limited to hepatocytes could be sufficient to reduce liver injury and delay cholestatic fibrosis. With this purpose, we performed bile duct ligation in transgenic mice overexpressing Bcl-2 in hepatocytes and in wild-type littermates. We found that, compared with necrosis, apoptosis was negligible in this model. Our results also showed that hepatocyte Bcl-2 expression protected hepatocytes against liver injury only in the early steps of the disease. This protection was correlated with reduced mitochondrial dysfunction and lipid peroxidation. However, in contrast to Fas receptor-deficient lpr mice, fibrosis progression was not hampered and liver inflammatory response was not reduced by Bcl-2 overexpression. These results therefore comfort the hypothesis that Fas-mediated apoptotic hepatocyte pathway is not a significant contributing factor to the clinical features observed in cholestasis. Moreover, in the absence of a blunted inflammatory response in transgenic mice, Bcl-2 protection against hepatocyte mitochondrial dysfunction and lipid peroxidation was not sufficient to block fibrosis progression.

International audience; In the last decade, a lot of patents have been filled regarding molecular biology and functions of cellular membranes. The membrane bilayer model has evolved from a static, passive, homogeneous barrier to a highly dynamic, asymmetric, heterogeneous structure composed of distinct domains. Changes in membrane fluidity and composition of microdomains have been proven to be involved in the regulation of many important physiological signaling pathways. Recently, several xenobiotics, including various drugs and environmental pollutants, have been reported to change plasma membrane characteristics, thereby altering cell physiology. Interestingly, it has been suggested that a cross talk between chemical-induced cellular membrane effects and DNA damages may be important for the final mutation outcome of genotoxic chemicals. Thus, effects on plasma membrane remodeling may give additional mechanistic explanations to how certain chemicals exert their carcinogenic effect. With respect to such effects, recent patents suggest to focus on plasma membrane and its components like caveolin-1 for cancer screening and chemotherapy. Here, we review the effects of environmental toxicants on cellular plasma membrane structure and function, and further describe possible implication for health and disease.

International audience; Retinopathy of prematurity (ROP), an ocular disease characterized by the onset of vascular abnormalities in the developing retina, is the major cause of visual impairment and blindness in premature neonates. ROP is a complex condition in which various factors participate at different stages of the disease leading to microvascular degeneration followed by neovascularization, which in turn predisposes to retinal detachment. Current ablative therapies (cryotherapy and laser photocoagulation) used in the clinic for the treatment of ROP have limitations and patients can still have long-term effects even after successful treatment. New treatment modalities are still emerging. The most promising are the therapies directed against VEGF; more recently the use of preventive dietary supplementation with ω-3 polyunsaturated fatty acid may also be promising. Other than pharmacologic and nutritional approaches, cell-based strategies for vascular repair are likely to arise from advances in regenerative medicine using stem cells. In addition to all of these, a greater understanding of other factors involved in regulating pathologic retinal angiogenesis continues to emerge, suggesting potential targets for therapeutic approaches. This review summarizes an update on the current state of knowledge on ROP from our and other laboratories, with particular focus on the role of nitro-oxidative stress and notably trans-arachidonic acids in microvascular degeneration, semaphorin 3 operating as vasorepulsive molecules in the avascular hypoxic retina and in turn impairing revascularization, succinate and its receptor GPR91 in neuron-mediated retinal neovascularization, and ω-3 lipids as modulators of preretinal neovascularization.

International audience; Lipid peroxidation leads to the formation of a number of by-products including acrolein. In brain from patients with Alzheimer's disease (AD), acrolein was found to be elevated in vulnerable regions. Astrocytes contribute to a variety of neuronal functions but the toxicity of acrolein in astroglial cells remains unknown. Using the rat primary astroglial cells, our results show that acrolein is toxic from 15 μM. Acrolein induced a biphasic effect on glutathione (GSH) levels with a depletion after 30 min of treatment followed by a progressive increase 24 hrs after exposure while the expression of γ-glutamyl-cysteine-synthase (γ-GCS) was induced. Protein carbonyls levels were significantly higher with all tested concentrations of acrolein. We have further investigated the effect of acrolein on the regulation of different redox-sensitive signaling pathways. A treatment with 20 μM of acrolein for 30 min activated NF-κB, Nrf2, and heme oxygenase-1 while after 24 hrs of exposure, their induction was observed with the subtoxic and toxic concentrations of acrolein except for NF-κB. Sirt-1 was also up-regulated after 24 hrs of exposure with acrolein. Acrolein also induced the phosphorylation of p66shc and of ERK1/2 after 30 min of treatment. Our results provide evidence that acrolein is a potent inducer of redox-sensitive pathways in astrocytes with a differential regulation after a short or a long term period of exposure to overcome cell death. Considering the crucial role of astrocytes in the brain, these results demonstrated that acrolein could disrupt neuronal functions and synaptic homeostasis by provoking dysfunctional or loss of astrocytes.

International audience; AIMS/HYPOTHESIS: This study assessed oxidative stress in LDL from obese patients with the metabolic syndrome and compared it with that in LDL from type 2 diabetic patients or control volunteers. It also determined the effect on platelets of LDL from the three groups. METHODS: The profiles of lipids, fatty acids and fatty acid oxidation products were determined in LDL isolated from plasma of patients with the metabolic syndrome, patients with type 2 diabetes and volunteers (n = 10 per group). The effects of LDL from the participant groups on the platelet arachidonic acid signalling cascade and aggregation were investigated. RESULTS: Compared with LDL from control volunteers, LDL from obese metabolic syndrome and type 2 diabetic patients had lower cholesteryl ester, higher triacylglycerol and lower ethanolamine plasmalogen levels. Proportions of linoleic acid were decreased in phosphatidylcholine and cholesteryl esters in LDL from both patient groups. Among the markers of lipid peroxidation, oxidation products of linoleic acid (hydroxy-octadecadienoic acids) and malondialdehyde were increased by 59% and twofold, respectively in LDL from metabolic syndrome and type 2 diabetic patients. LDL from metabolic syndrome and type 2 diabetic patients were equally potent in activating the platelet arachidonic acid signalling cascade through increased phosphorylation of p38 mitogen-activated protein kinase and cytosolic phospholipase A(2), and through increased thromboxane B(2) formation. LDL from patients with the metabolic syndrome and type 2 diabetes potentiated platelet aggregation by threefold and 3.5-fold respectively, whereas control LDL had no activating effects on platelets. CONCLUSIONS/INTERPRETATION: The metabolic syndrome in obese patients, without or with diabetes, is associated with increased oxidative stress in LDL, which triggers platelet activation.

International audience; Lipid peroxidation (LPO) product accumulation in human tissues is a major cause of tissular and cellular dysfunction that plays a major role in ageing and most age-related and oxidative stress-related diseases. The current evidence for the implication of LPO in pathological processes is discussed in this review. New data and literature review are provided evaluating the role of LPO in the pathophysiology of ageing and classically oxidative stress-linked diseases, such as neurodegenerative diseases, diabetes and atherosclerosis (the main cause of cardiovascular complications). Striking evidences implicating LPO in foetal vascular dysfunction occurring in pre-eclampsia, in renal and liver diseases, as well as their role as cause and consequence to cancer development are addressed.

hal-00680938, v1; International audience; Fat oxidation (FO) is optimized during low- to moderate-intensity exercise in lean and obese subjects, whereas high-intensity exercise induces preferential FO during the recovery period. After food intake during the postexercise period, it is unknown if FO differs according to the intensity exercise in overweight subjects. Fat oxidation was thus evaluated in overweight men after low- and high-intensity exercise during the recovery period before and after food intake as well as during a control session. Ten healthy, sedentary, overweight men (age, 27.9 +/- 5.6 years; body mass index, 27.8 +/- 1.3 kg m(-2); maximal oxygen consumption, 37 +/- 3.9 mL min(-1) kg(-1)) exercised on a cycloergometer (energy expenditure = 300 kcal) at 35% (E35) or 70% (E70) maximal oxygen consumption or rested (Cont). The subjects were fed 30 minutes after the exercise with 300 kcal (1256 kJ) more energy in the exercise sessions than in the Cont session. Respiratory quotient and FO were calculated by indirect calorimetry. Blood samples were analyzed to measure plasma glycerol, nonesterified fatty acid, glucose, and insulin. During exercise, mean respiratory quotient was lower (P < .05) and FO was higher (P < .01) in the E35 than in the E70 session (FO [in mg min(-1)]: E35 = 290 +/- 12, E70 = 256 +/- 38, and Cont = 131 +/- 7). Conversely, FO was higher in the E70 than in both the E35 session and the Cont session during the immediate recovery as well as during the postprandial recovery period (P = .005 for all; FO from the end of the exercise to the end of the session [in grams]: E70 = 45.7 +/- 8.9, E35 = 38.2 +/- 6.8, and Cont = 36.0 +/- 4.3). Blood parameters did not differ between the 3 sessions but changed according to the absorption of the nutrients. In overweight subjects, high-intensity exercise increased FO during the postexercise period even after food intake compared with the low-intensity exercise and the control session.

Background Increasing environmental and occupational exposures to nanoparticles (NPs) warrant deeper insight into the toxicological mechanisms induced by these materials. The present study was designed to characterize the cell death induced by carbon black (CB) and titanium dioxide (TiO2) NPs in bronchial epithelial cells (16HBE14o- cell line and primary cells) and to investigate the implicated molecular pathways. Results Detailed time course studies revealed that both CB (13 nm) and TiO2(15 nm) NP exposed cells exhibit typical morphological (decreased cell size, membrane blebbing, peripheral chromatin condensation, apoptotic body formation) and biochemical (caspase activation and DNA fragmentation) features of apoptotic cell death. A decrease in mitochondrial membrane potential, activation of Bax and release of cytochrome c from mitochondria were only observed in case of CB NPs whereas lipid peroxidation, lysosomal membrane destabilization and cathepsin B release were observed during the apoptotic process induced by TiO2 NPs. Furthermore, ROS production was observed after exposure to CB and TiO2 but hydrogen peroxide (H2O2) production was only involved in apoptosis induction by CB NPs. Conclusions Both CB and TiO2 NPs induce apoptotic cell death in bronchial epithelial cells. CB NPs induce apoptosis by a ROS dependent mitochondrial pathway whereas TiO2 NPs induce cell death through lysosomal membrane destabilization and lipid peroxidation. Although the final outcome is similar (apoptosis), the molecular pathways activated by NPs differ depending upon the chemical nature of the NPs.

International audience; This paper deals with the effects of exercise training on oxygen consumption (MO(2)) and ROS metabolism in the red muscle of trained and untrained female silver eels. Their critical swimming speed (U(crit)) was determined before and after a 4-day training (10h of swimming at 70% of U(crit) and 14 h at 50%, every day). The U(crit) of trained eels increased significantly (by about 7%). The in vitro MO(2) and ROS production by the red fibres were higher (not significant) in trained than in untrained eels, but the ROS production/MO(2) ratio was alike in both groups. The antioxidant-enzyme activities and lipoperoxidation index in trained eels were both lower than those of the untrained ones. These biochemical changes related to the increase in U(crit) suggest that such a training session could maintained or even increased aerobic power of the red muscle without deleterious impact by ROS. These regulations could play a role in the eel's swimming performance efficiency.

International audience; OBJECTIVE: To determine the effect of supplementation with increasing doses of docosahexaenoic acid (DHA), as the only n-3 polyunsaturated fatty acid (PUFA), on low-density lipoprotein (LDL) redox status and oxidizability. METHODS: Twelve healthy men aged 53-65 years ingested consecutive doses of DHA (200, 400, 800 and 1600 mg/day), each dose for two weeks. RESULTS: The proportions of DHA increased dose-dependently in LDL phospholipids and cholesteryl esters, even after two weeks of supplementation with 200mg/day DHA. The daily intake of 200, 400 and 800 mg DHA resulted in increased alpha-tocopherol concentrations, decreased MDA concentrations, and a longer lag time for copper-induced LDL oxidation. Supplementation with 1600 mg/day DHA had no effect on the above parameters. In plasma, concentrations of 4-hydroxy-hexenal, specifically derived from the peroxidation of n-3 fatty acids, significantly increased after 800 and 1600 mg DHA, representing 0.01% of plasma n-3 PUFAs, while 4-hydroxy-nonenal concentrations, derived from the peroxidation of n-6 fatty acids, did not change. CONCLUSION: Our results clearly show that an intake of 200-800 mg/day DHA may have protective and antioxidant effects on LDL and could represent optimal doses for cardiovascular disease prevention in a healthy population.

International audience; Chelation therapy is thought to not only remove contaminating metals but also to decrease free radical production. However, in standard ethylene diamine tetracetic acid (EDTA) chelation therapy, high doses of vitamin C with potential pro-oxidant effects are often added to the chelation solution. The authors demonstrated previously that the intravenous administration of the standard chelation cocktail, containing high amounts of vitamin C, resulted in an acute transitory pro-oxidant burst that should be avoided in the treatment of pathologies at risk of increased oxidative stress such as diabetes and cardiovascular disease. The current study was designed to determine the acute and chronic biochemical effects of chelation therapy on accepted clinical, antioxidant variables. An EDTA chelation cocktail not containing ascorbic acid was administered to six adult patients for five weeks (10 sessions of chelation therapy); antioxidant indicators were monitored. Immediately after the initial chelation session, in contrast with the data previously reported with the standard cocktail containing high doses of vitamin C, none of the oxidative stress markers were adversely modified. After five weeks, plasma peroxide levels, monitored by malondialdehyde, decreased by 20 percent, and DNA damage, monitored by formamidopyrimidine-DNA glycosylase (Fpg) sensitive sites, decreased by 22 percent. Remaining antioxidant-related variables did not change. In summary, this study demonstrates that multiple sessions of EDTA chelation therapy in combination with vitamins and minerals, but without added ascorbic acid, decreases oxidative stress. These results should be beneficial in the treatment of diseases associated with increased oxidative stress such as diabetes and cardiovascular diseases.

International audience; OBJECTIVE: The aim of the study was to investigate whether zinc supplementation affects antioxidant status in European middle-aged and elderly people. DESIGN: Multicentre prospective intervention study, randomized, double-blind, placebo-control. SETTING: France (Clermont-Ferrand/Theix, and Grenoble), Italy (Rome), Northern Ireland (Coleraine). SUBJECTS: A total of 387 healthy middle-aged (55-70 yrs) and free-living older aged (70-85 yrs) subjects were randomly allocated to three groups: 0, 15 or 30 mg zinc gluconate/d in addition to usual dietary intake during 6 months. METHODS: Oxidative stress status was evaluated by measurement of protein oxidation (plasma thiol groups), lipid peroxidation (plasma thio-barbituric acid reactants, TBARS), whole blood glutathione levels, erythrocyte copper/zinc superoxide dismutase activity and plasma antioxidant status (ferric reducing antioxidant power assay), at baseline and after 3 and 6 months. RESULTS: Zinc supplementation did not alter oxidative stress markers and antioxidant defenses in elderly, after 3 or 6 months, except an increase in Cu/Zn superoxide dismutase activity. CONCLUSIONS: In apparently healthy free living elderly people, a single zinc supplementation had no effects on oxidative stress status.

International audience; In an experimental food chain, Wistar rats were fed cadmium (Cd) in an inorganic (CdCl(2)) or organic (mainly associated with metallothionein from Helix aspersa snail viscera) form. After 1 month of exposure to 100 microg inorganic Cd g(-1) in food, an induction of metallothionein was observed in all target tissues. In liver, glutathione peroxidase (GSH-Px) activity decreased and alanine aminotransferase (ALAT) activity increased, suggesting that Cd causes hepatotoxicity. However, lipid peroxidation as well as catalase and caspase 3 (a marker of apoptosis) activities were not modified. At a rather low exposure (2.5 microg Cd g(-1)), metallothionein level in the kidney was found to be the most sensitive biomarker of exposure for both Cd forms. In the small intestine of rats ingesting inorganic Cd, metallothionein expression was significantly higher than that observed for rats fed organic Cd. Present results allowed proposing a simple design to assess the effect of a chemical in a trophic transfer approach.

Background The interconversion of two important energy metabolites, 3-hydroxybutyrate and acetoacetate (the major ketone bodies), is catalyzed by D-3-hydroxybutyrate dehydrogenase (BDH1: EC 1.1.1.30), a NAD+-dependent enzyme. The eukaryotic enzyme is bound to the mitochondrial inner membrane and harbors a unique lecithin-dependent activity. Here, we report an advanced purification method of the mammalian BDH applied to the liver enzyme from jerboa (Jaculus orientalis), a hibernating rodent adapted to extreme diet and environmental conditions. Results Purifying BDH from jerboa liver overcomes its low specific activity in mitochondria for further biochemical characterization of the enzyme. This new procedure is based on the use of polyclonal antibodies raised against BDH from bacterial Pseudomonas aeruginosa. This study improves the procedure for purification of both soluble microbial and mammalian membrane-bound BDH. Even though the Jaculus orientalis genome has not yet been sequenced, for the first time a D-3-hydroxybutyrate dehydrogenase cDNA from jerboa was cloned and sequenced. Conclusion This study applies immunoaffinity chromatography to purify BDH, the membrane-bound and lipid-dependent enzyme, as a 31 kDa single polypeptide chain. In addition, bacterial BDH isolation was achieved in a two-step purification procedure, improving the knowledge of an enzyme involved in the lipid metabolism of a unique hibernating mammal. Sequence alignment revealed conserved putative amino acids for possible NAD+ interaction.

Tumor protein 53-induced nuclear protein 1 (TP53INP1) is an antiproliferative and proapoptotic protein involved in cell stress response. To address its physiological roles in colorectal cancer and colitis, we generated and tested the susceptibility of Trp53inp1-deficient mice to the development of colorectal tumors induced by injection of the carcinogen azoxymethane followed by dextran sulfate sodium (DSS)-induced chronic colitis. Trp53inp1-deficient mice showed an increased incidence and multiplicity of tumors compared to those of wild-type (WT) mice. Furthermore, acute colitis induced by DSS treatment was more severe in Trp53inp1-deficient mice than in WT mice. Treatment with the antioxidant N-acetylcysteine prevented colitis and colitis-associated tumorigenesis more efficiently in WT mice than in Trp53inp1-deficient mice, suggesting a higher oxidative load in the latter. Consistently, we demonstrated by electron spin resonance and spin trapping that colons derived from deficient mice produced more free radicals than those of the WT during colitis and that the basal blood level of the antioxidant ascorbate was decreased in Trp53inp1-deficient mice. Collectively, these results indicate that the oxidative load is higher in Trp53inp1-deficient mice than in WT mice, generating a more-severe DSS-induced colitis, which favors development of colorectal tumors in Trp53inp1-deficient mice. Therefore, TP53INP1 is a potential target for the prevention of colorectal cancer in patients with inflammatory bowel disease.

regular heavy ethanol consumption has been associated with a type of nonischemic dilated cardiomyopathy termed “alcoholic cardiomyopathy” (ACM), which is clinically expressed as an impairment of left ventricular function (nonsymptomatic stage) and occasionally as overt heart failure (symptomatic

International audience; Exogenous treatment with monosialoganglioside GM1 has been described to afford protection against different apoptotic insults. However, the underlying mechanisms remain to be determined. In this study, we focused on the effect of GM1 on the apoptotic cascade induced by benzo[a]pyrene (B[a]P) in rat hepatic F258 epithelial cells. We first demonstrated that a co-treatment with GM1 (80 microM) reduced B[a]P (50 nM)-induced apoptosis as evidenced by a decrease of both cell population exhibiting nuclear fragmentation and caspase 3 cleavage and activity. We next showed that the p53 phosphorylation and nuclear translocation as well as the intracellular alkalinization related to Na+/H+ exchanger 1 (NHE1) activation, two early events of the apoptosis induced by B[a]P, were not inhibited by GM1. In contrast, the late mitochondria-dependent acidification elicited by B[a]P was inhibited by GM1 co-treatment, and an inhibition of the oxidative stress was also observed. Because GM1 has been shown to reduce the low-molecular weight iron content related to ethanol-induced oxidative stress, we finally investigated the involvement of iron under our conditions. Using the two iron chelators deferiprone and desferrioxamine, we clearly showed that iron played an important role in B[a]P-induced apoptosis in F258 cells, and that B[a]P-treatment resulted in a significant GM1-sensitive increase in (55)Fe uptake. In conclusion, our results indicate that exogenous GM1 partly prevents B[a]P-induced apoptosis by interfering with mitochondria-related intracellular acidification and iron transport.

National audience; Intestinal hexose absorption and gluconeogenesis have been studied in relation to refeeding after two different fasting phases: a long period of protein sparing during which energy expenditure is derived from lipid oxidation (phase II), and a later phase characterized by a rise in plasma corticosterone triggering protein catabolism (phase III). Such a switch in body fuel uses, leading to changes in body reserves and gluconeogenic precursors, could modulate intestinal gluconeogenesis and glucose transport. The gene and protein levels, and the cellular localization of the sodium-glucose cotransporter SGLT1, and of GLUT5 and GLUT2, as well as that of the key gluconeogenic enzymes phosphoenolpyruvate carboxykinase (PEPCK) and glucose-6-phosphatase (Glc6Pase) were measured. PEPCK and Glc6Pase activities were also determined. In phase III fasted rats, SGLT1 was up-regulated and intestinal glucose uptake rates were higher than in phase II fasted and fed rats. PEPCK and Glc6Pase mRNA, protein levels and activities also increased in phase III. GLUT5 and GLUT2 were down-regulated throughout the fast, but increased after refeeding, with GLUT2 recruited to the apical membrane. The increase in SGLT1 expression during phase III may allow glucose absorption at low concentrations as soon as food is available. Furthermore, an increased epithelial permeability due to fasting may induce a paracellular movement of glucose. In the absence of intestinal GLUT2 during fasting, Glc6Pase could be involved in glucose release to the bloodstream via membrane trafficking. Finally, refeeding triggered GLUT2 and GLUT5 synthesis and apical recruitment of GLUT2, to absorb larger amounts of hexoses.

There is currently great interest in the development of methods to analyze intracellular redox state and the cellular damages generated by oxidative stress. General methods for analyzing reactive oxygen species and glutathione level are presented together with more recently developed protocols to analyze the consequences of oxidative stress on the oxidation of macromolecules. Finally, techniques to study modalities of constitutive expression of Hsp27 in mammalian cells are considered as well as methods used to determine the protective activity of this small heat shock protein against the deleterious effects induced by oxidative stress.

Although it is well established that symptomatology, morbidity and death following scorpion envenomation are due to increases in neurotransmitter release secondary to toxins binding to voltage-sensitive sodium channels, the mechanism by which venom action is involved in damaging heart, liver, lungs and kidneys remains unclear. We hypothesized that scorpion toxins could induce the generation of high levels of free radicals responsible for membrane damage in organs targeted by venom action. We have investigated lipid peroxidation in different organs, through the evaluation of thiobarbituric acid reactive substances (TBARS), after experimental envenomation of rats by toxic fractions of Androctonus australis Hector venom. We have shown that scorpion toxins cause considerable lipid peroxidation in most vital organs. We also evaluated the protective effects of antioxidants in mice injected with lethal doses of toxins. Among the drugs tested, N-acetylcysteine (NAC) was effective in protecting the mice when injected prior to toxin application. However, the free radical scavenging properties of NAC seem less implicated in these protective effects than its ability to increase the fluidity of bronchial secretions. We therefore conclude that free radical generation only plays a minor role in the toxicity of scorpion venom.

International audience; Red meat intake is associated with colon cancer risk. Puzzlingly, meat does not promote carcinogenesis in rat studies. However, we demonstrated previously that dietary heme promotes aberrant crypt foci (ACF) formation in rats given a low-calcium diet. Here, we tested the hypothesis that heme-rich meats promote colon carcinogenesis in rats treated with azoxymethane and fed low-calcium diets (0.8 g/kg). Three meat-based diets were formulated to contain varying concentrations of heme by the addition of raw chicken (low heme), beef (medium heme), or black pudding (blood sausage; high heme). The no-heme control diet was supplemented with ferric citrate and the heme control diet with hemoglobin to match iron and heme concentrations in the beef diet, respectively. After 100 d, colons were scored for ACF and mucin-depleted foci (MDF). Fecal water was assayed for lipoperoxides and cytotoxicity. Only diets with heme promoted the formation of MDF, but all meat diets promoted ACF formation. The number of MDF/colon was 0.55 +/- 0.68 in controls, but 1.2 +/- 0.6 (P = 0.13), 1.9 +/- 1.4 (P < 0.01), and 3.0 +/- 1.2 (P < 0.001) in chicken-, beef-, and black pudding-fed rats. MDF promotion by the high-heme black pudding diet was greater than that by the medium-heme beef diet. The number of ACF/colon was 72 +/- 16 in controls, but 91 +/- 18, 100 +/- 13, and 103 +/- 14 in chicken-, beef-, and black pudding-fed rats (all P < 0.001). ACF and MDF did not differ between rats fed the beef diet and those fed the heme control diet. MDF promotion was correlated with high fecal water lipoperoxides and cytotoxicity (r = 0.65, P < 0.01). This is the first study to show the promotion of experimental carcinogenesis by dietary meat and the association with heme intake.

International audience; We have previously shown, as have other authors, that trans-resveratrol (E-resveratrol, 3,4,5-trihydroxy-E-stilbene) reduces reactive oxygen species (ROS) generation of mitochondria freshly isolated from healthy rat brains and that it also counteracts the effect of uncouplers (CCCP) on mitochondrial respiration and oxidative phosphorylation. Two main mechanisms have been shown: firstly, a scavenger effect toward O2- and secondly inhibition of complex III ROS generation. We now report on the effects of resveratrol in a pathological model that mimics the ischemia followed by the reperfusion process which may occur in the human brain. Isolated brain mitochondria were submitted first to hypoxia then to reoxygenation. The aim of this study was to determine the extent of mitochondrial damage induced by this experimental model, to demonstrate which mitochondrial functions were altered and to quantify the extent to which they were prevented by resveratrol. Resveratrol was either added to mitochondria freshly isolated from healthy rat brains or was injected by subcutaneous chronically implanted pumps (0.5, 2 and 10 mg/kg/day for 7 days). The rats were then sacrificed and mitochondria were extracted from brains. To evaluate the respective effects of hypoxia and reoxygenation on mitochondrial functions and the relevant effects of resveratrol, this drug was added (first protocol) either before the complete process (i.e., hypoxia and reoxygenation), or after anoxia before reoxygenation. We found that resveratrol prevented alterations of mitochondrial functions. This substance partly counteracted the decrease in respiratory control and the increase in ROS generation. It fully inhibited the alteration of membrane fluidity and the mitochondrial step of the apoptotic process (evidenced by cytochrome c release and membrane potential collapse). The effects of resveratrol were concentration-dependent (in vitro) or dose-dependent (ex vivo, second protocol). They were not significantly different when the drug was added before or after hypoxia, which suggests that in this model, reoxygenation was the most deleterious process and the stage at which resveratrol was most effective.

International audience; High intake of red meat, but not of white meat, is associated with an increased risk of colon cancer. However, red meat does not promote cancer in rodents. Haemin, added to low-calcium diets, increases colonic proliferation, and haemoglobin, added to high-fat diets, increases the colon tumour incidence in rats, an effect possibly due to peroxyl radicals. We thus speculated that haem might be the promoting agent in meat, and that prevention strategies could use calcium and antioxidants. These hypotheses were tested in rats at the aberrant crypt foci (ACF) stage at 100 days. F344 rats (n = 124) were given an injection of azoxymethane and were then randomized to 11 groups fed with low-calcium (20 micro mol/g) AIN76-based diets, containing 5% safflower oil. Haemin (0.25, 0.5 and 1.5 micro mol/g) or haemoglobin (1.5 and 3 micro mol haem/g) was added to five experimental diets, compared with a control diet without haem. Three other high-haemin diets (1.5 micro mol/g) were supplemented with calcium (250 micro mol/g), antioxidant butylated hydroxyanisole and rutin (0.05% each), and olive oil, which replaced safflower oil. Faecal water was assayed for lipid peroxidation by thiobarbituric acid reactive substances (TBARs) test, and for cytolytic activity. Haemin strikingly increased the ACF size, dose-dependently, from 2.6 to 11.4 crypts/ACF (all P < 0.001). The high-haemin diet also increased the number of ACF per colon (P < 0.001). Promotion was associated with increased faecal water TBARs and cytotoxicity. Calcium, olive oil and antioxidants each inhibited the haemin-induced ACF promotion, and normalized the faecal TBARs and cytotoxicity. The haemoglobin diets increased the number of ACF and faecal TBARs, but not the ACF size or the faecal cytotoxicity. In conclusion, dietary haemin is the most potent known ACF promoter. Haemoglobin is also a potent promoter of colorectal carcinogenesis. The results suggest that myoglobin in red meat could promote colon cancer. Diets high in calcium, or in oxidation-resistant fats, may prevent the possible cancer-promoting effect of red meat.

Chronic inactivity may not be the sole factor involved in the myopathy of chronic obstructive pulmonary disease (COPD) patients. One hypothesis is that exercise-induced oxidative stress that leads to muscle alterations may also be involved. This study investigated whether exercise localised to a peripheral muscle group would induce oxidative stress in COPD patients. Eleven COPD patients (FEV 1 1.15±0.4 L (mean±sd)) and 12 healthy age-matched subjects with a similar low quantity of physical activity performed endurance exercise localised to a peripheral muscle group, the quadriceps of the dominant leg. The authors measured plasma levels of thiobarbituric reactive substances (TBARs) as an index of oxidative stress, the release in superoxide anion (O 2 · − ) by stimulated phagocytes as an oxidant, and blood vitamin E as one antioxidant. Quadriceps endurance was significantly lower in the COPD patients compared with healthy subjects (136±16 s versus 385±69 s (mean±sem), respectively). A significant increase in TBARs 6 h after quadriceps exercise was only found in the COPD patients. In addition, significantly higher O 2 · − release and lower blood vitamin E levels were found in COPD patients than in controls at rest. This blood vitamin E level was significantly correlated with the resting level of plasma TBARs in the COPD patients. This study mainly showed that quadriceps exercise induced systemic oxidative stress in chronic obstructive pulmonary disease patients and that vitamin E levels were decreased in these patients at rest. The exact relevance of these findings to chronic obstructive pulmonary disease myopathy needs to be elucidated.