Responses of type II pneumocyte antioxidant enzymes to normoxic and hyperoxic culture

Summary Cultured type II pneumocyte responses to in vitro normoxia (95% air: 5% CO₂) or hyperoxia (95% O₂:5% CO₂) were quantified. Normoxic culture (0 to 96 h) of rabbit type II cells resulted in enhanced cell-monolayer protein and DNA content. During this same time, cellular activities of superoxide dismutase (SOD), catalase, and glutathione peroxidase (GSH Px) decreased. Compared to cultures maintained in normoxia, hyperoxic exposure of cultures resulted in decreased cell-associated protein and DNA content. Exposure to hyperoxia also resulted in cytotoxicity as demonstrated by elevated cellular release of DNA, lactate dehydrogenase (LDH), and preincorporated 8-[¹⁴C]adenine. Cellular catalase and GSH Px activities in hyperoxic cells decreased similarly to normoxic controls. In contrast, cellular SOD activity in hyperoxic cells decreased less than in normoxic cultures. Cellular SOD activity in hyperoxic cultures, when normalized for cellular protein, but not DNA, was greater than normoxic values after 24 to 96 h of exposure. Unlike the decrease in cellular antioxidant enzymes during normoxic and hyperoxic culture, cellular LDH activity increased during both these exposures. Cellular LDH activity in 24 to 96 h hyperoxia-exposed cells increased to a lesser extent than normoxic controls. The extent of depression in LDH activity was dependent on whether the activity was normalized for cellular protein or DNA. Type II pneumocytes, which normally undergo hyperplasia and hypertrophy during hyperoxia in vivo, exhibited oxygen sensitivity in vitro. Exposure of type II cells to hyperoxia in vitro resulted in alterations in cellular SOD and LDH activities, but recognition of such changes were dependent on whether enzymatic activities were normalized for cellular DNA or protein. This work was supported by a grant from the Health Effects Institute, grant HL40458 from the National Institutes of Health, Bethesda, MD, and a grant from the American Lung Association, New York, NY.     

Superoxide dismutase mimetics elevate superoxide dismutase activity in vivo but do not retard aging in the nematode Caenorhabditis elegans

According to the oxidative damage theory a primary cause of aging is the accrual of molecular damage from reactive oxygen species (ROS), particularly superoxide and its derivatives. This predicts that treatments that reduce ROS levels should retard aging. Using the nematode Caenorhabditis elegans, we tested the effects on stress resistance and life span of treatment with EUK-8 and EUK-134, synthetic mimetics of the antioxidant enzyme superoxide dismutase (SOD), which neutralises superoxide. Treatment with SOD mimetics elevated in vivo SOD activity levels, particularly in mitochondria, where up to 5-fold increases in SOD activity were recorded. Treatment with exogenous SOD mimetics did not affect endogenous protein SOD levels. Where life span was reduced by the superoxide generators paraquat and plumbagin, EUK-8 treatment increased life span in a dose-dependent fashion. Yet in the absence of a superoxide generator, treatment with EUK-8 or EUK-134 did not increase life span, even at doses that were optimal for protection against pro-oxidants. Thus, an elevation of SOD activity levels sufficient to increase life span when it is limited by superoxide generators does not retard aging in the absence of superoxide generators. This suggests that C. elegans life span is not normally limited by levels of superoxide and its derivatives.     

Role of reactive oxygen species in LPS-induced production of prostaglandin E2 in microglia

We determined the roles of reactive oxygen species (ROS) in the expression of cyclooxygenase-2 (COX-2) and the production of prostaglandin E2 (PGE2) in lipopolysaccharide (LPS)-activated microglia. LPS treatment increased intracellular ROS in rat microglia dose-dependently. Pre-treatment with superoxide dismutase (SOD)/catalase, or SOD/catalase mimetics that can scavenge intracellular ROS, significantly attenuated LPS-induced release in PGE2. Diphenylene iodonium (DPI), a non-specific NADPH oxidase inhibitor, decreased LPS-induced PGE2 production. In addition, microglia from NADPH oxidase-deficient mice produced less PGE2 than those from wild-type mice following LPS treatment. Furthermore, LPS-stimulated expression of COX-2 (determined by RT-PCR analysis of COX-2 mRNA and western blot for its protein) was significantly reduced by pre-treatment with SOD/catalase or SOD/catalase mimetics. SOD/catalase mimetics were more potent than SOD/catalase in reducing COX-2 expression and PGE2 production. As a comparison, scavenging ROS had no effect on LPS-induced nitric oxide production in microglia. These results suggest that ROS play a regulatory role in the expression of COX-2 and the subsequent production of PGE2 during the activation process of microglia. Thus, inhibiting NADPH oxidase activity and subsequent ROS generation in microglia can reduce COX-2 expression and PGE2 production. These findings suggest a potential therapeutic intervention strategy for the treatment of inflammation-mediated neurodegenerative diseases.     

Overexpression of glutathione reductase extends survival in transgenic Drosophila melanogaster under hyperoxia but not normoxia.

The purpose of this study was to test the hypothesis that overexpression of glutathione reductase in transgenic Drosophila melanogaster increases resistance to oxidative stress and retards the aging process. Transgenic flies were generated by microinjection and subsequent mobilization of a P element construct containing the genomic glutathione reductase gene of Drosophila, with 4 kb upstream and 1.5 kb downstream of the coding region. Transgenic animals stably overexpressed glutathione reductase by up to 100% throughout adult life and under continuous exposure to 100% oxygen or air. Under hyperoxic conditions, overexpressors had increased longevity, decreased accrual of protein carbonyls, and dramatically increased survival rates after recovery from a semi-lethal dose of 100% oxygen. Under normoxic conditions, overexpression of glutathione reductase had no effect on longevity, protein carbonyl content, reduced glutathione, or glutathione disulfide content, although the total consumption of oxygen was slightly decreased. Glutathione reductase activity does not appear to be a rate-limiting factor in anti-aging defenses under normoxic conditions, but it may become a limiting factor when the level of oxidative stress is elevated.     

Hyperoxia results in transient oxidative stress and an adaptive response by antioxidant enzymes in goldfish tissues

The effects of hyperoxia on the status of antioxidant defenses and markers of oxidative damage were evaluated in goldfish tissues. The levels of lipid peroxides, thiobarbituric acid reactive substances, carbonyl proteins and the activities of some antioxidant enzymes were measured in brain, liver, kidney and skeletal muscle of goldfish, Carassius auratus L., over a time course of 3-12 h of hyperoxia exposure followed by 12 or 36 h of normoxic recovery. Exposure to high oxygen resulted in an accumulation of protein carbonyls in tissues throughout hyperoxia and recovery whereas lipid peroxides and thiobarbituric acid reactive substances accumulated transiently under short-term hyperoxia stress (3-6 h) but were then strongly reduced. This suggests that hyperoxia stimulated an enhancement of defenses against lipid peroxidation or mechanisms for enhancing the catabolism of peroxidation products. The activities of principal antioxidant enzymes, superoxide dismutase and catalase, were not altered under hyperoxia but catalase increased during normoxic recovery; activities may rise in anticipation of further hyperoxic excursions. In most tissues, the activities of glutathione-utilizing enzymes (glutathione peroxidase, glutathione-S-transferase, glutathione reductase) as well as glucose-6-phosphate dehydrogenase, were not affected under hyperoxia but increased sharply during normoxic recovery. Correlations between some enzyme activities and oxidative stress markers were found, for example, an inverse correlation was seen between levels of thiobarbituric acid reactive substances and glutathione-S-transferase activity in liver and catalase and glucose-6-phosphate dehydrogenase in kidney. The results suggest that liver glutathione-S-transferase plays an important role in detoxifying end products of lipid peroxidation accumulated under hyperoxia stress.     

Expression of multiple copies of mitochondrially targeted catalase or genomic Mn superoxide dismutase transgenes does not extend the life span of Drosophila melanogaster

The simultaneous overexpression of multiple copies of Mn superoxide dismutase (SOD) and ectopic catalase (mtCat) transgenes in the mitochondria of the fruit fly, Drosophila melanogaster, was shown previously to diminish the life span. The hypothesis tested in this study was that this effect was due primarily to the presence of one or the other transgene. An alternative hypothesis was that both transgenes have additive, negative effects. Crosses were performed between five pairs of transgenic lines containing single-copy insertions of mtCat, Mn SOD, or P element vector control transgenes at unique loci, and the life spans of progeny containing two mtCat, Mn SOD, or vector insertions were determined. Increasing amounts of mitochondrial catalase activity tended to be associated with decreases in mean life span. Overexpression of two copies of the genomic Mn SOD transgene had no effect on life span. The results do not support the hypothesis that enhanced mitochondrial SOD or catalase activity promotes longevity in flies.     

Differential responses of juvenile and adult South African abalone (Haliotis midae Linnaeus) to low and high oxygen levels

Marine invertebrates have evolved multiple responses to naturally variable environmental oxygen, all aimed at either maintaining cellular oxygen homeostasis or limiting cellular damage during or after hypoxic or hyperoxic events. We assessed organismal (rates of oxygen consumption and ammonia excretion) and cellular (heat shock protein expression, anti-oxidant enzymes) responses of juvenile and adult abalone exposed to low (~ 83% of saturation), intermediate (~ 95% of saturation) and high (~ 115% of saturation) oxygen levels for one month. Using the Comet assay, we measured DNA damage to determine whether the observed trends in the protective responses were sufficient to prevent oxidative damage to cells. Juveniles were unaffected by moderately hypoxic and hyperoxic conditions. Elevated basal rates of superoxide dismutase, glutathione peroxidase and catalase were sufficient to prevent DNA fragmentation and protein damage. Adults, with their lower basal rate of anti-oxidant enzymes, had increased DNA damage under hypoxic and hyperoxic conditions, indicating that the antioxidant enzymes were unable to prevent oxidative damage under hypoxic and hyperoxic conditions. The apparent insensitivity of juvenile abalone to decreased and increased oxygen might be related to their life history and development in algal and diatom biofilms where they are exposed to extreme diurnal fluctuations in dissolved oxygen levels.     

Mitochondrial aldehyde dehydrogenase attenuates hyperoxia-induced cell death through activation of ERK/MAPK and PI3K-Akt pathways in lung epithelial cells

Oxygen toxicity is one of the major risk factors in the development of the chronic lung disease or bronchopulmonary dysplasia in premature infants. Using proteomic analysis, we discovered that mitochondrial aldehyde dehydrogenase (mtALDH or ALDH2) was downregulated in neonatal rat lung after hyperoxic exposure. To study the role of mtALDH in hyperoxic lung injury, we overexpressed mtALDH in human lung epithelial cells (A549) and found that mtALDH significantly reduced hyperoxia-induced cell death. Compared with control cells (Neo-A549), the necrotic cell death in mtALDH-overexpressing cells (mtALDH-A549) decreased from 25.3 to 6.5%, 50.5 to 9.1%, and 52.4 to 15.1% after 24-, 48-, and 72-h hyperoxic exposure, respectively. The levels of intracellular and mitochondria-derived reactive oxygen species (ROS) in mtALDH-A549 cells after hyperoxic exposure were significantly lowered compared with Neo-A549 cells. mtALDH overexpression significantly stimulated extracellular signal-regulated kinase (ERK) phosphorylation under normoxic and hyperoxic conditions. Inhibition of ERK phosphorylation partially eliminated the protective effect of mtALDH in hyperoxia-induced cell death, suggesting ERK activation by mtALDH conferred cellular resistance to hyperoxia. mtALDH overexpression augmented Akt phosphorylation and maintained the total Akt level in mtALDH-A549 cells under normoxic and hyperoxic conditions. Inhibition of phosphatidylinositol 3-kinase (PI3K) activation by LY294002 in mtALDH-A549 cells significantly increased necrotic cell death after hyperoxic exposure, indicating that PI3K-Akt activation by mtALDH played an important role in cell survival after hyperoxia. Taken together, these data demonstrate that mtALDH overexpression attenuates hyperoxia-induced cell death in lung epithelial cells through reduction of ROS, activation of ERK/MAPK, and PI3K-Akt cell survival signaling pathways.     

低氧-复氧胁迫对鲢抗氧化酶活性及Cu/Zn-SOD和Mn-SOD基因表达的影响

为探究低氧-复氧胁迫对鲢(Hypophthalmichthys molitrix)抗氧化酶活性及Cu/Zn-SOD和Mn-SOD基因表达的影响, 对鲢进行急性低氧、持续低氧及复氧实验, 进而分析血清、心脏和肝脏中不同抗氧化酶和SODs基因表达的变化特征。结果表明: 在急性低氧胁迫后, 血清中总抗氧化能力(T-AOC)、过氧化氢酶(CAT)和谷胱甘肽过氧化物酶(GSH-PX)活性随着氧浓度的降低均呈上升趋势, 但超氧化物歧化酶(SOD)活性呈先升后降的趋势。在持续低氧胁迫后, 血清中T-AOC和GSH-PX活性随着低氧胁迫时间的增加显著升高(P<0.05); 心脏中SOD活性显著高于常氧水平(P<0.05), 但Cu/Zn-SOD和Mn-SOD基因表达在低氧胁迫24h时显著低于常氧水平(P<0.05); 肝脏中SOD活性在低氧胁迫24h时显著高于常氧水平(P<0.05), 且Cu/Zn-SOD和Mn-SOD基因表达在低氧胁迫24h时也显著高于常氧水平(P<0.05)。复氧后, 血清、心脏和肝脏中T-AOC、SOD、CAT和GSH-PX活性均能恢复至常氧水平, 且心脏和肝脏中Cu/Zn-SOD和Mn-SOD基因表达的也能恢复至常氧水平, 但肝脏中Mn-SOD基因表达恢复至常氧水平较在心脏中所需时间更少。因而, 鲢可以通过调节抗氧化酶的活性来保护自身免受氧化应激造成的损伤。研究为解析低氧胁迫下鲢抗氧化应激机制提供了基础。     

抗氧化剂对皮肤角质细胞体外寿命的影响

考察了抗氧化剂对鼠角质细胞体外培养寿命的影响。实验发现在角质细胞的体外培养过程中添加抗氧化剂有利于延长细胞的寿命,其中效果最好的是巯基乙醇,其次为过氧化氢酶和SOD,但在体外培养过程中,角质细胞生长速率仍然逐渐下降。实验还发现,添加抗氧化剂可在一定程度上提高角质细胞的克隆形成率,减缓细胞衰老速率。同时,通过考察鼠表皮角质细胞衰老动力学,获得了对应于不同抗氧化剂的细胞衰老动力学常数。      

The effects of dissolved oxygen and aquatic surface respiration on the critical thermal maxima of three intermittent-stream fishes

Synopsis Critical thermal maxima (CTM) were determined for three species of fish in hypoxic (1.2 mg l^-1), normoxic (7 mg l^-1) and hyperoxic (12 mg l^-1) oxygen tensions. All fish were acclimated to 30°C under a regulated photoperiod of 12:12. During each CTM determination, one-half of the test fish were allowed free access to the surface (to permit aquatic surface respiration) while the remaining one-half were denied access to the surface. Under hypoxic conditions, CTMs of Fundulus notatus, Notropis lutrensis and Pimephales vigilax given access to the surface were significantly higher than those of conspecifics without access to the surface. With access to the surface, all species had significantly lower CTMs under hypoxic conditions than under normoxic and hyperoxic conditions. CTMs measured under normoxic and hyperoxic conditions were not different for any of the test species. Under all comparable oxygen tensions with surface access, F. notatus had the highest CTM, N. lutrensis intermediate and P. vigilax had the lowest CTM. The critical oxygen concentration for N. lutrensis during CTM determinations occurred between 1.2 and 2 mg l^-1.     

Endothelial NOS expression and ischemia-reperfusion in isolated working rat heart from hypoxic and hyperoxic conditions

Induction of endothelial nitric oxide synthase (eNOS) contributes to the mechanism of heart protection against ischemia-reperfusion damage. We analyzed the effects of hypoxia and hyperoxia on eNOS expression in isolated working rat hearts after ischemia-reperfusion damage. Adult male Wistar rats were submitted to chronic hypoxia (2 weeks) and hyperoxia (72 h). The hearts were submitted to 15 min of ischemia and reperfused for 60 min, then we evaluated hemodynamic parameters and creatine phosphokinase (CPK) release. eNOS expression was estimated by RT-PCR; enzyme localization was evaluated by immunohistochemistry and the eNOS protein levels were detected by Western blot. All hemodynamic parameters in hypoxic conditions were better with respect to other groups. The CPK release was lower in hypoxic (P<0.01) than in normoxic and hyperoxic conditions. The eNOS deposition was significantly higher in the hypoxic group versus the normoxic or hyperoxic groups. The eNOS protein and mRNA levels were increased by hypoxia versus both other groups. Chronic hypoxic exposure may decrease injury and increase eNOS protein and mRNA levels in heart subjected to ischemia-reperfusion.     

Effect of antioxidants on apoptosis and cytokine release in fetal rat Type II pneumocytes exposed to hyperoxia and nitric oxide

The response of the fetal rat Type II pneumocyte (FTIIP), the stem cell of the alveolar epithelium, to hyperoxia would be helpful to understand the effects of oxygen-induced injury to the developing lung. Our goals were to evaluate the effect of antioxidants (AO) on apoptosis and release of cytokines in freshly isolated FTIIP (day-19) in the presence of 95% O2 and/or nitric oxide (NO). There was increased apoptosis in FTIIP exposed to hyperoxia alone and in combination with NO; this was significantly attenuated (p < 0.01) in the presence of 3 AO, namely grape seed proanthocyanidin extract (GSPE), superoxide dismutase (SOD) and catalase. The anti-inflammatory cytokine IL-10 has been shown to have a role in ameliorating tissue damage owing to persistent inflammation. The release of IL-10 was significantly decreased (p < 0.01) in the presence of GSPE and catalase, compared to control. Addition of SOD led to increased IL-10 compared to GSPE or catalase (p < 0.01) or the combination of GSPE + SOD + catalase (p < or = 0.01). Thus, in our in vitro model of hyperoxic and NO mediated injury to FTIIP, protection from apoptotic cell death with the addition of AO was associated with varying levels of IL-10 release. Our data suggest that the use of SOD and/or IL-10 may decrease hyperoxic lung injury by decreasing apoptosis. Further studies are needed to understand the mode of protection from catalase and GSPE.     

Biochemical adaptation in rat liver in response to marginal oxygen toxicity

1. Hepatic glucose 6-phosphate dehydrogenase activity was increased in rats exposed to 5lb/in(2) (equivalent to 27000ft), 100% O(2) when compared with control animals in a 14.7lb/in(2) (sea level), air environment. Glyceraldehyde 3-phosphate dehydrogenase, isocitrate dehydrogenase, and succinate dehydrogenase were not affected by the 5lb/in(2), 100% O(2) environment. 2. Animals exposed to the hyperoxic environment consumed food, expired CO(2) and gained weight at the same rate as normoxic control animals. Additionally, blood glucose and liver glycogen concentrations were unchanged in the hyperoxic animals. The only readily apparent physiological difference in the hyperoxic animals was a decreased haematocrit. 3. The increase in glucose 6-phosphate dehydrogenase was eliminated by the injection of actinomycin D or cycloheximide. 4. Expiration of (14)CO(2) from [1-(14)C]glucose was approximately the same in hyperoxic and normoxic rats. However, (14)CO(2) expiration from [6-(14)C]glucose was markedly decreased in the animals exposed to the hyperoxic environment. 5. Calculations of the relative importance of the pentose phosphate pathway versus the tricarboxylic acid cycle plus glycolysis indicated that the livers from animals in the 5lb/in(2), 100% O(2) environment metabolized twice as much carbohydrate by way of the pentose phosphate pathway as did those from the sea-level air control animals. 6. In livers of rats exposed to 5lb/in(2), 100% O(2) the concentrations of pyruvate, citrate and 2-oxoglutarate were increased, that of isocitrate was slightly elevated, whereas the concentrations of succinate, fumarate and malate were decreased. 7. An inactivation of both tricarboxylic acid cycle lipoate-containing dehydrogenases, pyruvate and 2-oxoglutarate, under hyperoxic conditions is proposed. 8. The adaptive significance of the induction of glucose 6-phosphate dehydrogenase and the resultant production of NADPH under hyperoxic conditions is discussed.     

Sodium nitroprusside induces mild oxidative stress in Saccharomyces cerevisiae

Nitric oxide is known to be a messenger in animals and plants. It may act either as a pro-oxidant or antioxidant. In the present work, the yeast Saccharomyces cerevisiae was treated under aerobic conditions with the nitric oxide donor, sodium nitroprusside (SNP), at concentrations of 1, 5 and 10 mM. The activities of antioxidant enzymes as well as concentrations of protein carbonyls and cellular thiols were measured. Yeast incubation with SNP increased the activities of catalase and superoxide dismutase. Cycloheximide, an inhibitor of translation, blocked SNP-induced catalase activation, but not SOD activation. Incubation with SNP increased the activity of peroxisomal catalase, whereas cytosolic catalase was not affected. SNP treatment inactivated aconitase in a dose-dependent manner. Surprisingly, in cells incubated with 1 mM SNP, the levels of low-molecular weight thiols were significantly higher, whereas the concentrations of protein carbonyl groups were lower than those in untreated cells. The incubation of yeast cells either with decomposed SNP or with SNP under anaerobic conditions did not result in SOD and catalase activation. It is suggested, that under aerobic conditions, the SNP effects are connected with induction of mild oxidative/nitrosative stress.     

Effects of sodium valproate and oxygen on the CD-1 mouse fetus

This study reports the effects of valproic acid (VA) on the CD-1 mouse fetus when the drug is administered continuously via osmotic minipumps at human therapeutic drug plasma levels. Two VA-filled Alzet osmotic minipumps were implanted subcutaneously on gestation day 5 for continuous exposure of a total daily dosage of 850 mg/kg on gestation days 5-12. Dams were then exposed continuously to either normoxic (21% oxygen), hyperoxic (50% oxygen), or hypoxic (12% oxygen) controlled environments during gestation days 5-12, in order to determine if hyperoxic maternal conditions offered a protective environment for the fetus, and conversely, if hypoxia exacerbated teratogenicity. Dams were sacrificed on gestation day 18, and litter and fetal data were collected. It was determined in separate groups under normoxic conditions that the osmotic minipump system maintained VA plasma levels corresponding to human therapeutic levels. Sodium valproate was found to induce developmental toxicity in the CD-1 mouse fetus at human therapeutic drug plasma levels. Fetal weights were reduced, and the number of resorptions, deaths, and hematomas was increased. While hypoxia exacerbated the toxic effect on the fetus, hyperoxia failed to ameliorate the outcome.     

Expression of manganese superoxide dismutase is not altered in transgenic mice with elevated level of copper-zinc superoxide dismutase

In evaluating the relative expression of copper-zinc and manganese superoxide dismutase (CuZnSOD and MnSOD) in vivo in states like Down syndrome in which one dismutase is present at increased levels, we measured activities of both enzymes, in tissues of control and transgenic mice constitutively expressing increased levels of CuZnSOD, during exposure to normal and elevated oxygen tensions. Using SOD gel electrophoresis assay, CuZnSOD and MnSOD activities of brain, lung, heart, kidney, and liver from mice exposed to either normal (21%) or elevated (>99% oxygen, 630 torr) oxygen tensions for 120 h were compared. Whereas CuZnSOD activity was elevated in tissues of transgenic relative to control mice under both normoxic or hyperoxic conditions, MnSOD activities in organs of transgenic mice were remarkably similar to those of controls under both conditions. To confirm the accuracy of this method in quantitating MnSOD relative to CuZnSOD expression, two other methods were utilized. In lung, which is the organ exposed to the highest oxygen tension during ambient hyperoxia, a sensitive, specific ELISA for MnSOD was used. Again, MnSOD protein was not different in transgenic relative to control mice during exposure to air or hyperoxia. In addition, lung MnSOD protein was not changed significantly by exposure to hyperoxia in either group. In kidney, a mitochondrion-rich organ, SOD assay, before and after inactivation of CuZnSOD with diethyldithiocarbamate, was used. MnSOD activity was not different in organs from air-exposed transgenic relative to control mice. The data indicated that expression of MnSOD in vivo was not affected by overexpression of the CuZnSOD and, therefore, the two enzymes are probably regulated independently.     

Growth on ethanol results in co-ordinated Saccharomyces cerevisiae response to inactivation of genes encoding superoxide dismutases

Superoxide dismutase (SOD) is an essential enzyme protecting cells against oxidative stress. However, its specific role under different conditions is not clear. To study the possible role of SOD in the cell during respiration, Saccharomyces cerevisiae single and double mutants with inactivated SOD1 and/or SOD2 genes growing on ethanol as an energy and carbon source were used. Activities of antioxidant and associated enzymes as well as the level of protein carbonyls were measured. SOD activity was significantly higher in a Mn-SOD deficient strain than that in the wild-type parental strain, but significantly lower in a Cu, Zn-SOD mutant. A strong positive correlation between SOD and catalase activities (R(2) = 0.99) shows possible protection of catalase by SOD from inactivation in vivo and/or decrease in catalase activity because of lower H(2)O(2) formation in the mutant cells. SOD deficiency resulted in a malate dehydrogenase activity increase, whereas glucose-6-phosphate dehydrogenase (G6PDH) activity was lower in SOD-deficient strains. Linear and non-linear positive correlations between SOD and isocitrate dehydrogenase activities are discussed. No changes in the activity of glutathione reductase and protein carbonyl levels support the idea that SOD-deficient cells are not exposed to strong oxidative stress during exponential growth of yeast cultures on ethanol.     

Nucleotides and the adenylate energy charge as indicators of stress in rainbow trout (Oncorhynchus mykiss) subjected to a range of dissolved oxygen concentrations

Liver nucleotides (ATP, ADP, AMP, IMP), the adenylate energy charge (AEC), total adenylate concentration (TA), and IMP-load were used as measures of stress in rainbow trout (Oncorhynchus mykiss) acclimated to normoxic (10.0 mg/l), hypoxic (6.5 mg/l), and supersaturated (13.0 mg/l) dissolved oxygen concentrations and subjected to a challenge by confinement. Liver ATP (783.0 nmol/g) was significantly different in the normoxic fish compared to either hyperoxic (447.7 nmol/g) or hypoxic (402.0 nmol/g) fish at the end of the confinement. Within 6.0 hr in the confinement, liver AEC in the normoxic fish increased significantly (0.58) compared to hypoxic (0.42) and hyperoxic fish (0.42). Similarly, the IMP-load in normoxic fish (0.16) decreased to near prestress levels by 6.0 hr in confinement compared to either the hypoxic (0.31) or hyperoxic (0.30) fish. Nucleotides in liver were significantly affected by the dissolved oxygen treatments and the confinement stress in contrast to the muscle nucleotides which were not.     

Physiological significance of catalase and glutathione peroxidases,and in vivo peroxidation,in selected tissues of the toadDiscoglossus pictus (Amphibia) during acclimation to normobaric hyperoxia

1. Various parameters related to oxidative stress were measured in adult Discoglossus pictus acclimated for 15 days to either normoxia or hyperoxia (PO2 = 710 mmHg). 2. Total weight of the toads and total and relative wet weight of liver, kidneys, lungs and heart were not changed by hyperoxic acclimation. 3. In vivo tissue peroxidation increased in lung, decreased in skeletal muscle, and was not changed in liver, kidney, heart and skin after hyperoxic exposure. 4. Hyperoxic acclimation increased catalase activities in the lung, liver, kidney and heart but not in skeletal muscle and skin. 5. Liver showed higher GSH-peroxidase activity with cumene-OOH than with H2O2 as substrate, whereas lung, skeletal muscle and skin presented similar GSH-peroxidase activities with both substrates. 6. GSH-peroxidase activities did not change between hyperoxic and normoxic animals in liver, lung, skeletal muscle and skin. 7. These results show that catalase, not GSH-peroxidase, is the principal H2O2 detoxifying enzyme involved in the adaptation of D. pictus to hyperoxia.