Blood markers of oxidative stress predict weaning failure from mechanical ventilation.

Patients undergoing mechanical ventilation (MV) often experience respiratory muscle dysfunction, which complicates the weaning process. There is no simple means to predict or diagnose respiratory muscle dysfunction because diagnosis depends on measurements in muscle diaphragmatic fibre. As oxidative stress is a key mechanism contributing to MV-induced respiratory muscle dysfunction, the aim of this study was to determine if differences in blood measures of oxidative stress in patients who had success and failure in a spontaneous breathing trial (SBT) could be used to predict the outcome of MV. This was a prospective analysis of MV-dependent patients (≥72 hrs; n = 34) undergoing a standard weaning protocol. Clinical, laboratory and oxidative stress analyses were performed. Measurements were made on blood samples taken at three time-points: immediately before the trial, 30 min. into the trial in weaning success (WS) patients, or immediately before return to MV in weaning failure (WF) patients, and 6 hrs after the trial. We found that blood measures of oxidative stress distinguished patients who would experience WF from patients who would experience WS. Before SBT, WF patients presented higher oxidative damage in lipids and higher antioxidant levels and decreased nitric oxide concentrations. The observed differences in measures between WF and WS patients persisted throughout and after the weaning trial. In conclusion, WF may be predicted based on higher malondialdehyde, higher vitamin C and lower nitric oxide concentration in plasma.

Alpha-lipoic acid restores tear production in an animal model of dry eye.

The tear film comprises a major mechanism for protection of the ocular surface against harmful external agents. Disruption of tear production can lead to dry eye syndrome, causing damage ranging from mild discomfort to scarring of the ocular surface with irreversible vision impairment. The production of tears by the lacrimal gland is influenced by neuroendocrine, hormonal, and immunological factors. Reactive oxygen and nitrogen species play an important role in its regulation. We assessed the effects of oxidative stress on antioxidant defenses in the lacrimal gland and ocular surface in ovariectomized rats supplemented with n-3 polyunsaturated fatty acids (n-3 PUFA) and alpha-lipoic acid (ALP). We found that n-3 PUFA did not measurably influence oxidative stress, but ALP had site-specific pro-oxidant and antioxidant effects, and was an important influence on ocular surface dry eye improvement. As an index of oxidative damage to proteins and lipids, we measured levels of carbonyl and malondialdehyde (MDA), respectively. Enzymatic antioxidant defenses were measured as total superoxide dismutase (tSOD) and glutathione peroxidase (GPx), and non-enzymatic defenses were estimated by vitamin C, total glutathione, and indirect oxide nitric levels. PUFA and ALP treatment restored lacrimal production with resulting improvement in the dry eye Schirmer test in all supplemented groups. The results indicated that reactive oxygen species resulting from oxidative stress in the lacrimal gland did not play an important role in dry eye through reactive oxygen species; however, alpha-lipoic acid altered the metabolism of reactive nitrogen species, causing increased activity of lacrimal peroxidase and improved lacrimal production.

Oxidative stress in the kidney of reproductive female rats during aging.

Reproduction is a costly life process, and the reproductive investment by females appears to be greater than males in many species. We have analyzed the effects of reproductive investment during aging with respect to oxidative stress parameters in female Wistar rats. We measured the activity glutathione peroxidase, glutathione S-transferase, superoxide dismutase, consumption of hydrogen peroxide, protein carbonylation, lipid peroxidation, nitrite and nitrate levels, and Vitamin C (Vit. C) and E levels. We traced oxidative profiles at ages 3, 6, 12, and 24 months. Animals were grouped according to reproductive experience: experienced or naive with respect to reproductive activity. We measured aconitase activity and sex hormone levels. The naive animals exhibited an increase with respect to experienced in most parameters studied at 6 and 24 months, whereas experienced animals exhibited a similar increase at 3 and 12 months. At 6 months of age, during the period that would represent peak reproductive activity, naive animals showed higher levels of MDA, Vit. C, consumption of hydrogen peroxide and GPx, aconitase, and SOD activities. In naive elderly rats, we observed an increase in oxidative damage markers and an increase in enzymatic and non-enzymatic antioxidants, with the exception of consumption of hydrogen peroxide and Vit. C. In the long term, the reproductive investment was not sufficient to interfere with antioxidant capacity, and did not contribute to oxidative damage in kidneys of female Wistar rats.

An insight into the functional role of antioxidant and detoxification enzymes in adult Rhipicephalus microplus female ticks.

Ticks have developed physiological adaptations to transport, store, metabolize and secrete toxic components from the diet and environment. Different classes of enzymes are involved in these processes, however, the role of several of them is not yet characterized in Rhipicephalus microplus. In this context, this work investigated the action of antioxidant and detoxification enzymes, as well as the levels of essential cellular reductants in R. microplus partially engorged females (PEF) and fully engorged females (FEF). Results demonstrated that enzymes transcriptional levels and enzymatic activity from ovary and fat body were higher in PEF than in FEF, except for ovary Glutathione peroxidase (GPx), which was the only enzyme showing highest activity in the FEF stage. These results indicated a higher demand for antioxidant potential in these organs at the initial feeding phase than during egg-laying. In midgut, however, there was more variability in the transcriptional levels and activity of the different enzymes between the PEF and FEF phases. Similar NADPH levels were found in PEF and FEF phases, suggesting a remarkable capacity to maintain a regular supply of reducing power, despite the developmental changes and large intake of heme and iron. However, reduced glutathione (GSH) levels were variable between PEF and FEF when distinct organs were compared. Taken together, our data suggest a higher demand for reducing potential in FEF ticks. The silencing of catalase (CAT) or thioredoxin reductase (TRx) genes in females did not impair feeding, egg-laying capacity, or larvae hatching. CAT-silenced ticks had increased ovary peroxidase activity, a possible compensatory antioxidant mechanism. Altogether, the results shed light on the complexity of the antioxidant and detoxification enzyme system in ticks and its involvement in different physiological mechanisms.

Effects of lipoic acid and ω-3 long-chain polyunsaturated fatty acids on the kidney in the ovariectomized rat model of menopause.

OBJECTIVES: The loss of antioxidant protection from estrogen during menopause may lead to oxidative stress in the kidneys. Thus, antioxidant supplementation may potentially decrease the menopause-derived oxidative stress. The aim of this study was to evaluate the effect of α-lipoic acid (LA) and ω-3 long-chain polyunsaturated fatty acids on the redox profile of the kidneys in the ovariectomized rat model of menopause. METHODS: We assessed oxidative damage markers and antioxidant defenses in the kidneys of ovariectomized rats supplemented with LA, docosahexaenoic acid (DHA), and eicosapentaenoic acid (EPA). Animals received 3 mo of dietary supplementation. RESULTS: Ovariectomy did not increase the levels of the damage markers carbonyl and malondialdehyde. EPA supplementation increased carbonyl and malondialdehyde levels. Ovariectomy increased fumarase activity but did not affect the levels of vitamin C, glutathione, and glutathione S-transferase activity. LA, DHA, and EPA supplementation decreased fumarase activity, but increased the levels of vitamin C, glutathione, and glutathione S-transferase activity. Vitamin E, superoxide dismutase, glutathione peroxidase, and peroxide consumption were not affected by ovariectomy or supplementation. CONCLUSIONS: The results suggest that ovariectomy did not affect the redox profile in the kidneys. LA, DHA, and EPA supplementation increased certain endogenous antioxidants; however, EPA may have a prooxidant effect on the kidneys.

The effect of n-3 long-chain polyunsaturated fatty acids and lipoic acid on the heart in the ovariectomized rat model of menopause.

Menopause occurs as consequence of ovarian senescence that leads to a drop of oestrogen hormone. The decreased oestrogen levels combined with the impairment of the redox system may contribute to the increased risk of postmenopausal cardiovascular disease. Supplementation with antioxidants may be an alternative to reduce cardiovascular risk. The study evaluated the effect of dietary supplementation with docosahexaenoic acid (DHA), eicosapentaenoic acid (EPA), and α-lipoic acid (LA) for a period of 16 weeks on oxidative stress biomarkers in the hearts of ovariectomized 3-month-old rats. Ovariectomy did not increase the level of the damage markers malondialdehyde and carbonyl, and both were decreased by LA supplementation. Ovariectomy increased the levels of the endogenous antioxidants glutathione, vitamin C and H2O2 consumption, after restoration by DHA, EPA, and LA supplementation. Vitamin E, glutathione peroxidase, glutathione-S-transferase, and superoxide dismutase are not altered by ovariectomy. Lipid and protein damage are not increased after ovariectomy and a portion of the endogenous antioxidants concomitantly increased, suggesting that hearts may be protected by these antioxidants. DHA, EPA, and LA restored these endogenous antioxidants, showing that all evaluated supplements are effective in modulating the antioxidant redox system in the heart. LA showed additional effect on redox markers, decreasing lipid and protein damage markers.

Mild Therapeutic Hypothermia Increases Glutathione Levels in Postcardiac Arrest Patients.

Ischemia-reperfusion (I/R)-induced oxidative stress is one of the main mechanisms of tissue injury after cardiac arrest (CA). A decrease in antioxidant defenses may contribute to I/R injury. The present study aims to investigate the influence of mild therapeutic hypothermia (MTH) on levels of nonenzymatic antioxidants after CA. We investigated antioxidant levels at 6, 12, 36, and 72 hours after CA in central venous blood samples of patients admitted to intensive care. The sample consisted of 31 patients under controlled normothermia (36°C) and 11 patients treated with 24 hours of MTH (33°C). Erythrocyte glutathione (GSH) levels were elevated by MTH, increasing at 6, 12, 36, and 72 hours after CA in hypothermic patients (mean GSH levels in normothermic patients: 6 hours = 73.89, 12 hours = 56.45, 36 hours = 56.46, 72 hours = 61.80 vs. hypothermic patients: 6 hours = 176.89, 12 hours = 198.78, 36 hours = 186.96, and 72 hours = 173.68 µmol/g of protein). Vitamin C levels decreased significantly at 6 and 12 hours after CA in hypothermic patients (median vitamin C levels in normothermic patients: 6 hours = 7.53, 12 hours = 9.40, 36 hours = 8.56, and 72 hours = 8.51 vs. hypothermic patients: 6 hours = 5.46, 12 hours = 5.44, 36 hours = 6.10, and 72 hours = 5.89 mmol/L), coinciding with the period of therapeutic hypothermia. Vitamin E and nitric oxide levels were not altered by hypothermic treatment. These findings suggest that MTH alters nonenzymatic antioxidants differently, decreasing circulating vitamin C levels during treatment; however, MTH elevates GSH levels, possibly protecting tissues from I/R injury after CA.

Therapeutic Hypothermia Reduces Oxidative Damage and Alters Antioxidant Defenses after Cardiac Arrest.

After cardiac arrest, organ damage consequent to ischemia-reperfusion has been attributed to oxidative stress. Mild therapeutic hypothermia has been applied to reduce this damage, and it may reduce oxidative damage as well. This study aimed to compare oxidative damage and antioxidant defenses in patients treated with controlled normothermia versus mild therapeutic hypothermia during postcardiac arrest syndrome. The sample consisted of 31 patients under controlled normothermia (36°C) and 11 patients treated with 24 h mild therapeutic hypothermia (33°C), victims of in- or out-of-hospital cardiac arrest. Parameters were assessed at 6, 12, 36, and 72 h after cardiac arrest in the central venous blood samples. Hypothermic and normothermic patients had similar S100B levels, a biomarker of brain injury. Xanthine oxidase activity is similar between hypothermic and normothermic patients; however, it decreases posthypothermia treatment. Xanthine oxidase activity is positively correlated with lactate and S100B and inversely correlated with pH, calcium, and sodium levels. Hypothermia reduces malondialdehyde and protein carbonyl levels, markers of oxidative damage. Concomitantly, hypothermia increases the activity of erythrocyte antioxidant enzymes superoxide dismutase, glutathione peroxidase, and glutathione S-transferase while decreasing the activity of serum paraoxonase-1. These findings suggest that mild therapeutic hypothermia reduces oxidative damage and alters antioxidant defenses in postcardiac arrest patients.

Serum S100B protein is increased in fasting rats.

BACKGROUND: S100B is a calcium-binding protein expressed and secreted by astrocytes; serum and cerebrospinal fluid (CSF) S100B elevation has been proposed as an index of brain damage. However, other tissues are shown to produce this protein and the clinical significance of serum S100B elevation has been discussed. METHODS: We investigated the levels of serum and CSF S100B in fasting Wistar rats. Animals were divided into two groups, control and fasting for 48 h, and S100B levels in serum and CSF were determined by ELISA. S100B secretion in dissociated epididymal fat cells was investigated in the presence of epinephrine. RESULTS: We observed a significant >2-fold increase of S100B levels in serum of fasting rats, without changes in its CSF content. Moreover, we demonstrated in vitro epinephrine stimulated S100B release from fat cells. CONCLUSIONS: Present results reinforce that extracerebral sources of S100B, particularly adipocytes, contribute to its serum levels and support the idea that caution is needed when interpreting serum S100B increase as a clinical marker of brain damage.

Food restriction during pregnancy alters brain's antioxidant network in dams and their offspring.

Dietary restriction increases life span and protects distinct organisms against a series of diseases, among which, those related to oxidative stress, like neurodegenerative diseases. Interferences in the maternal environment are known to reprogram the offspring metabolism response, impacting in the risk of chronic diseases development in adulthood. We aimed to assess the effects of 40% food restriction on reactive species levels, enzymatic and non-enzymatic antioxidant defenses, and oxidative damage parameters in the cerebellum and total cerebral cortex of pregnant rats and their offspring. Dams and pups showed oxidative modulation caused by food restriction in both structures. Dichlorofluorescein oxidation, reflecting reactive species levels, was reduced in the cerebellum of dams and offspring, while the cerebral cortex was not affected. Decreased mitochondrial superoxide levels were found in the cerebellum and cerebral cortex of pups, while nitric oxide was increased in the cortex. We also measured the activities of important antioxidant enzymes responsible by reactive oxygen species elimination. Superoxide dismutase activity was increased in the cerebellum of dams and in both structures of pups, while it was decreased in dams' cerebral cortex. Both brain structures were affected concerning to catalase, glutathione peroxidase, and glutaredoxin activities, which were reduced in pups and dams. Non-enzymatic defenses were decreased in pups, while dams showed an adaptive pattern in the cerebellum and no alteration in the cerebral cortex. Even though the results suggest increased oxidative status, lipids and proteins were not oxidatively affected. Our data suggest that intrauterine food restriction may disrupt oxidative status, impairing the antioxidant network.

Induction of ROS generation by fluconazole in Candida glabrata: activation of antioxidant enzymes and oxidative DNA damage.

In this study, we assessed the generation of reactive oxygen species (ROS) induced by subinhibitory concentration of fluconazole in susceptible and resistant Candida glabrata strains at stationary growth phase and measured their oxidative responses parameters: glutathione peroxidase (GPx), superoxide dismutase (SOD), glutathione-S-transferase (GST), consumption of hydrogen peroxide, and total glutathione, as well as oxidative damage in lipids, proteins, and DNA. Data showed that fluconazole increased generation of ROS and GPx and SOD enzymatic activity in treated cells; however, these enzymatic activities did not differ between resistant and susceptible strains. Susceptible strains exhibited higher GST activity than resistant, and when susceptible cells were treated with fluconazole, GST activity decreased. Fluconazole treatment cause oxidative damage only in DNA. There are a possible participation of ROS, as organic peroxides and O2(•-), in antifungal mechanism of fluconazole, which results in higher GPx and SOD enzymatic activities and oxidative DNA damage in C. glabrata.

Treatment of oxidative stress in brain of ovariectomized rats with omega-3 and lipoic acid.

SCOPE: Postmenopausal women are often affected by a group of metabolic disorders related to oxidative stress. Alternative treatments that can improve the quality of life of these women have been the subject of recent studies. The objective of this study was to evaluate the response to oxidative stress in the brains of rats following ovariectomy, and to determine enzymatic and nonenzymatic antioxidant responses when the animals received 3 months of dietary supplementation. METHODS AND RESULTS: Ovariectomy produced changes in antioxidant profiles characterized by reductions in glutathione S-transferase activity, H2 O2 consumption, superoxide dismutase activity, and vitamin C levels and increases in protein carbonylation. Docosahexaenoic fatty acid (DHA) supplementation restored these parameters to normal values and increased values of other antioxidants (glutathione peroxidase and total glutathione). However, DHA supplementation also increased protein carbonylation and lipid peroxidation. Eicosapentaenoic acid supplementation produced no changes in antioxidants, but decreased lipid peroxidation. Lipoic acid supplementation increased consumption of H2 O2 and decreased protein carbonylation and lipid peroxidation. CONCLUSION: These results suggest that the antioxidant response to omega-3 varies in different tissues, and in this study DHA treatment had a prooxidant effect in the brain. Lipoic acid treatment, on the other hand, had a protective effect, reducing markers of oxidative damage.

Oxidative stress in the kidney of reproductive male rats during aging.

Reproduction alters the male physiology. We performed a comprehensive examination of oxidative stress in the kidneys of male rats with (experienced) or without (naïve) reproductive activity during aging. Oxidative stress was assessed by measuring the activity of catalase, glutathione peroxidase, glutathione S-transferase, and superoxide dismutase, and by measuring protein carbonylation, lipid peroxidation, nitrite and nitrate levels, vitamin C levels, and glutathione (total, reduced, and oxidized forms) levels, and metabolism was accessed by aconitase activity in kidney tissue, as well as testosterone and estradiol levels in serum. Reproductively active animals exhibited increased testosterone levels and altered metabolism. Aging affects tissues and organs and contributes to their functional decline. Elderly naïve rats showed high nitrite and nitrate levels. The experienced rats had less damage in elderly ages, probably because they had higher antioxidant amount and antioxidant enzyme activities at earlier ages, which would have avoided oxidative damage seen in naïve group, and because of the metabolism decline. Glutathione increase in naïve elder rats probably was induced for direct protection against oxidative damage and indirect protection by higher glutathione peroxidase and glutathione S-transferase activities. Linear regression shows that lipid peroxidation levels explained vitamin C levels (B standardized value of 0.42), indicating that vitamin C was properly produced or recruited into kidneys to combat lipid peroxidation. Catalase activity reflected the protein carbonylation and lipid peroxidation levels (B standardized values of 0.28 and 0.48). These results add comprehensive data regarding changes in oxidative stress during aging, and suggest an explanation for the costs of reproduction.

Carbonyl groups: Bridging the gap between sleep disordered breathing and coronary artery disease.

Sleep disordered breathing (SDB) is related to coronary artery disease (CAD), but the mechanisms are uncertain. SDB is characterized by periods of intermittent hypoxia and free radical formation. This study tested the hypothesis that carbonylation can be the link between SDB and CAD. It included 14 cases with CAD and 33 controls with <50% coronary narrowing. CAD cases have higher erythrocyte carbonyl levels than controls (p = 0.012). Positive correlation was observed between apnea-hypopnea index (AHI) and erythrocyte carbonyl concentration (rho = 0.310; p = 0.027). To predict CAD, including as regressors: AHI, erythrocyte carbonyl, gender, age and body mass index, the significant variables in the Poisson multiple regression model were AHI and erythrocytes carbonyl. An increase of 1 pmol/gHb in erythrocyte carbonyl levels increases by 1.8% the risk of CAD and one unit of AHI increases by 3.8% the risk of CAD. The present findings represent the first evidence in humans that SDB may cause CAD through protein carbonylation.

Alpha-lipoic acid modifies oxidative stress parameters in sickle cell trait subjects and sickle cell patients.

BACKGROUND & AIMS: Oxidative stress plays a crucial role in the sickle cell disease. Alpha-lipoic acid (ALA) is a potent antioxidant that is employed in the treatment of several diseases. The objective of this study was to test the ALA effect in the sickle cell disease (SCD) treatment. METHODS: Sixty subjects were selected and divided into groups according to the hemoglobin profile: AA (normal), AS (SCD trait subject) and SS (SCD patient). Patients were randomized into a placebo-controlled trial and treated with either ALA (200 mg) or vehicle. Blood samples were collected before supplementation and after 3 months of treatment. Catalase (CAT), superoxide dismutase (SOD), glutathione peroxidase (GPx) activities and total antioxidant status (TAS) were evaluated as measure of antioxidant defense. Lipid and protein damages were quantified by malondialdehyde (MDA) and carbonyl assays, respectively. RESULTS: CAT activity significantly increased in the AS group after ALA treatment and GPx activity presented significant decrease in all groups. SOD activity was not different in any group. Data on MDA and carbonyl levels showed significant reduction in the AA group with ALA treatment. TAS decreased in the same group. CONCLUSION: ALA treatment protected AA individuals from oxidative damage to lipids and proteins. In SCD subjects, the dose applied was not effective to prevent the oxidative damage.

Stress protein response and catalase activity in freshwater planarian Dugesia (Girardia) schubarti exposed to copper.

The hsp60 expression pattern and catalase activity in the freshwater planarian Dugesia schubarti exposed to copper under laboratory conditions were investigated. In the hsp60 induction experiments, planarians were exposed to a range of copper concentrations (0-960 microgCu/L) for 4 or 24h, to concentrations of 50 or 100 microgCu/L for 2, 4, 8, and 24h at 19 degrees C, and to heat shock at 27 degrees C for 24h. The concentrations of hsp60 in whole-body homogenates were determined immunochemically by Western blotting. Stress protein induction was detected only after 24h treatment at 27 degrees C. The tissue concentration of hsp60 remained unaltered in Cu-exposed planarians under the experimental conditions used. Catalase activity was significantly induced at concentrations of 40, 80, and 160 microgCu/L after 24h exposure. Our results suggest that catalase levels in planarians could represent biomarkers of interest for the estimation of copper effects in freshwater ecosystems.