Antioxidant Scavenging Enzymes Research Articles

Inorganic Polyphosphates Regulate Hexokinase Activity and Reactive Oxygen Species Generation in Mitochondria of Rhipicephalus (Boophilus) microplus Embryo

The physiological roles of polyphosphates (poly P) recently found in arthropod mitochondria remain obscure. Here, the possible involvement of poly P with reactive oxygen species generation in mitochondria of Rhipicephalus microplus embryos was investigated. Mitochondrial hexokinase and scavenger antioxidant enzymes, such as superoxide dismutase, catalase, and glutathione reductase were assayed during embryogenesis of R. microplus. The influence of poly P3 and poly P15 were analyzed during the period of higher enzymatic activity during embryogenesis. Both poly Ps inhibited hexokinase activity by up to 90% and, interestingly, the mitochondrial membrane exopolyphosphatase activity was stimulated by the hexokinase reaction product, glucose-6-phosphate. Poly P increased hydrogen peroxide generation in mitochondria in a situation where mitochondrial hexokinase is also active. The superoxide dismutase, catalase and glutathione reductase activities were higher during embryo cellularization, at the end of embryogenesis and during embryo segmentation, respectively. All of the enzymes were stimulated by poly P3. However, superoxide dismutase was not affected by poly P15, catalase activity was stimulated only at high concentrations and glutathione reductase was the only enzyme that was stimulated in the same way by both poly Ps. Altogether, our results indicate that inorganic polyphosphate and mitochondrial membrane exopolyphosphatase regulation can be correlated with the generation of reactive oxygen species in the mitochondria of R. microplus embryos.

Assessment of Protective Antioxidant Mechanisms in some Ethno- Medicinally Important wild Edible Fruits of Odisha, India

Fruits and vegetables have now been documented as nutraceuticals or functional foods useful for health and medical benefits including prevention and treatment of diseases. With the background of ethno medicinal evidences and the view to utilize the wild fruit resources of Odisha, India as functional food enriched with antioxidants, 4 wild edible fruits were studied for in vitro radical scavenging activity and antioxidant enzymes such as Peroxidase, Catalase and Superoxide dismutase (SOD) following standard methods. It was found that the fruit with highest DPPH scavenging activity is Antidesma ghaesembilla (1020.6 AEAC mg/100 g dwt) and the lowest recorded in Morinda tinctoria (235 AEAC mg/100 g dwt). The highest FRAP value was recorded in Antidesma ghaesembilla (2114 μM AEAC/g dwt) and the fruit with lowest FRAP value was Careya arborea (538 μM AEAC/g dwt) Antidesma ghaesembilla showed the highest Peroxidase value of 1.12 OD/min/g tissue wt while the lowest was found in Morinda tinctoria (0.054 OD/min/g tissue wt). Catalase was found in high amounts in Antidesma ghaesembilla (5.4×104 IEU/g fresh tissue), the lowest value was observed in Dillenia pentagyna (1.2×104 IEU/g fresh tissue). Similarly for superoxide dismutase (SOD), the highest value was recorded in Morinda tinctoria (4.43 Δ OD/min/mg protein) and lowest in Careya arborea (1.12 Δ OD/min/mg protein). Current research reveals that these wild edible fruits, especially Antidesma ghaesembilla are rich source of antioxidants and can further be subjected to identification of individual compounds responsible for such high antioxidant activity.

Jujube preservation using chitosan film with nano-silicon dioxide

The effect of 1% chitosan film with 0.04% nano-silicon dioxide on the qualitative properties of harvested jujube under ambient temperature was investigated. After 32d, the red index, decay incidence, weight loss, and respiration rate of the coated jujubes were lower compared with those of the control. The lower phenylalnine ammonialyase activity and higher activities of scavenger antioxidant enzymes (i.e., superoxide dismutase, peroxidase, and catalase) of the coated jujubes can be attributed to the compound coating. Increased malonaldehyde in the coated jujubes was restrained. Composite coating had shown to be superior in preserving total flavonoid than chitosan coating alone. But no differences were observed in terms of vitamin C loss and total polyphenol content between composite coating and control. Coating jujubes with chitosan+nano-silicon dioxide will be a promising alternative if nano-silicon dioxide is allowed for food use.

Role of antioxidant scavenging enzymes and extracellular polysaccharide in pathogenicity of rice bacterial blight pathogen Xanthomonas oryzae pv. oryzae

In the present work, we studied the role of antioxidant scavenging enzymes of plant pathogenic bacteria: catalase, ascorbate peroxidase and a virulence factor; extracelluar polysaccharide production in determining the virulence of Xanthomonas oryzae pv. oryzae (Xoo) isolates and its differential reaction to rice cultivars. A varied level of antioxidant scavenging activity and exopolysaccharide production was observed among 34 isolates studied, and most of the Xoo isolates with higher catalase activity also exhibited higher ascorbate peroxidase activity. The maximum level of catalase (45 µM H 2 O 2 min -1 mg -1 protein), ascorbate peroxidase (29 µM ascorbate min -1 mg -1 proteins) activity and exopolysaccharide production (70 mg) was found in isolate Xoo32 which induced maximum lesion length on cultivar ‘Jaya’ upon clip inoculation in virulence assay. Among the 44 cultivars screened, cultivar ‘Jeerigesanna’ recorded least bacterial blight disease incidence, with 0.7 cm lesion length. The activity of catalase, ascorbate peroxidase and exopolysaccharide can be employed as bio-chemical markers in determining the virulence of Xoo under laboratory conditions. Key words : Paddy, plant pathogenic bacteria, antioxidant scavenging enzymes, exopolysaccharide, virulence, bio-chemical markers.

Effect of postharvest treatments and storage temperatures on the quality and shelf life of sweet pepper (Capsicum annum L.)

The effect of salicylic acid (SA) and calcium chloride (CaCl2) treatments on the physicochemical characteristics, softening and antioxidative enzyme activities of sweet pepper fruits packaged in plastic (polythene) bags and stored at 25°C and 10°C were evaluated at 0, 9 and 18 days of storage period. The activities of cell wall modifying enzymes such as polygalacturonase (PG), pectin methyl esterase (PME), cellulase, β-galactosidase and antioxidant enzymes like peroxidase (POD), catalase (CAT) and ascorbic acid oxidase (AAO) were elucidated in the fruits of experimental sets and they were compared with that of control set. Fruits treated with SA and CaCl2 showed a significant (p<0.05) delay in changes of weight, total soluble solids (TSS), pH, titrable acidity (TA) and shelf life compared to untreated control fruits. The data of the present study may be an indicative that the lower enzyme activities of PG, PME, cellulase, β-galactosidase in the treated peppers might have been associated with a high integrity of the cell membrane and few changes in the cell wall constituents, which contributed to high levels of crispness and firmness in the peppers during storage. The higher activities of scavenger antioxidant enzymes, including POD and CAT in the treated peppers at the 18 days storage period are probably due to the effect of SA and CaCl2. In comparison to the fruits of control set, all the treatments have shown positive effectiveness on the inhibition of AAO. Also the results of the present study suggest that by using SA (1mM and 2mM) and CaCl2 (1.5%), the ripening process can be delayed, and the shelf life of capsicum fruits (stored at 10°C) can be extended up to 71 days without any spoilage and off-flavour. Thus it may be concluded that the SA and CaCl2 treatments may aid in delaying the softening process, enhancing the keeping quality while retaining the nutritional quality of sweet peppers more than that of control fruits in both the storage conditions (25°C and 10°C).

Chemomodulatory potential of Asparagus adscendens against murine skin and forestomach papillomagenesis

We have evaluated the cancer chemopreventive efficacy of the roots of Asparagus adscendens, which have been used in the Indian traditional medicine system for a long time for the treatment of various ailments. For the first time, the effect of its different doses in a test diet was examined on 7,12-dimethylbenz(a)anthracene-induced skin and benzo(a)pyrene-induced forestomach papillomagenesis in mice. The effect of these test diets was also examined on drug-metabolizing phase I and phase II enzymes, antioxidant enzymes, reduced glutathione content, and peroxidative damage in mice. Results exhibited a significant reduction in the skin and the forestomach tumor incidence with respect to all the three (2, 4, and 6%, w/w) doses as compared with control. Among all the doses tested, 4% of test diet was most effective in protecting the animals against papillomagenesis. Further, the roots of A. adscendens inhibited phase I, and activated phase II system and antioxidant enzymes in the liver especially with 4% of test diet. The content of reduced glutathione was also significantly elevated whereas the peroxidative damage along with lactate dehydrogenase activity were reduced with all the three doses of the test diet. Together, these results suggest the cancer chemopreventive potential of A. adscendens, which could be mediated through drug-metabolizing phase I and phase II enzymes as well as free radical scavenging antioxidant enzymes.

Regulation of growth and metabolism in rice (Oryza sativa L.) by arsenic and its possible reversal by phosphate

The effect of arsenate with or without phosphate on the growth and metabolism in rice seedlings cv. MTU1010 was studied. In the test, cv. arsenic was more toxic for root growth, than for shoot growth, where root hairs were fewer and short, roots were characteristically stubby, brittle and root tips gradually turned brown. Arsenic caused damage to the root epidermal cells and aerenchymatous cortex. The level of total chlorophyll, chlorophyll-a, chlorophyll-b and fluorescence intensity were decreased in arsenic treated rice seedlings. Arsenic toxicity affected the activities of different antioxidant scavenging enzymes in the test seedlings. Activities of superoxide dismutase and ascorbic acid oxidase were increased, whereas catalase and catechol peroxidase activities were decreased by arsenic application. In these seedlings, the oxidative stress has been observed with arsenic treatments and the level of proline, H2O2 and malondialdehyde contents were increased. Joint application of phosphate with arsenic showed significant alterations on all parameters tested under the purview of arsenic treatment alone leading to better growth and metabolism in rice seedlings. Thus the use of phosphorus enriched fertilizers in arsenic contaminated soil may improve the production of healthy rice plants.

Effects of chitosan coating enriched with cinnamon oil on qualitative properties of sweet pepper ( Capsicum annuum L.)

Effect of chitosan–oil coating on qualitative properties of sweet pepper ( Capsicum annuum L.) stored at 8 °C for 35 days was investigated. The chitosan–oil coating treatment exhibited the best control effect on decay (below 5%). At the end of storage, samples treated with chitosan-oil coatings maintained good sensory acceptability, whereas the sensory quality of control samples became unacceptable. The higher activities of scavenger antioxidant enzymes, including superoxide dismutase (SOD), peroxidase (POD) and catalase (CAT), in treated peppers at the 35th day should be contributed to the chitosan–oil coating. Malondialdehyde (MDA) and electrolyte leakage contents in chitosan–oil-coated peppers were increased but were still lower than in control samples. Atomic force microscopy images showed that the surface of sweet pepper without coating treatment was rougher than that of peppers treated with chitosan–oil coating. Our study suggests that chitosan–oil coating might be a promising candidate for enhancing the keeping quality of sweet peppers.

Antioxidant Enzyme Activity and Coronary Heart Disease: Meta-analyses of Observational Studies

Controversial data exist concerning the relation between the activities of scavenger antioxidant enzymes and coronary heart disease (CHD) risk. The authors report updated meta-analyses of studies assessing the activities of 3 antioxidant enzymes--glutathione peroxidase, superoxide dismutase, and catalase--and CHD risk. Computer-based and manual searches of the relevant literature from January 1966 to January 2008 were performed. Studies assessing glutathione peroxidase, superoxide dismutase, and catalase activities in cells or biologic fluids and clinical CHD outcomes were selected. Pooled odds ratios for CHD were calculated by using an inverse-variance-weighted random-effects model. Forty-two case-control studies and 3 prospective studies were included. The pooled odds ratios for CHD associated with a 1-standard-deviation increase in glutathione peroxidase, superoxide dismutase, and catalase activity levels were 0.51 (95% confidence interval: 0.35, 0.75), 0.48 (95% confidence interval: 0.32, 0.72), and 0.32 (95% confidence interval: 0.16, 0.61), respectively, with substantial between-study heterogeneity (I(2) > 90% for the 3 enzymes). These findings were remarkably robust in the sensitivity analysis. The authors' meta-analyses support an inverse association between circulating levels of superoxide dismutase, glutathione peroxidase, and catalase activities with CHD and emphasize the need for additional high-quality prospective studies.

Comparative Anticancer and Antioxidant Activities of Different Ingredients ofGinkgo bilobaExtract (EGb 761)

Flavonoid glycosides are the major constituents of Ginkgo biloba extract (EGb 761) and are well known to be an antioxidant for inhibiting tumor growth. Because it contains several flavonoid glycosides and other bioactive substances, the activities of EGb 761 against cancer in vivo and in vitro remain poorly understood. This situation prompted interest in the compounds for experimental study. We have investigated the anticancer effects of three analogues of EGb 761 samples on sarcoma 108 (S180)-bearing mice and leukemic 1210 (L1210) cell lines. We have also evaluated the changes of endogeneous antioxidant scavenging enzymes, including superoxide dismutase (SOD), glutathione (GST), lipid peroxidation (LPx), and catalase (CAT), in the blood of the S180-bearing mice. The EGb 761, EGb 761-H (containing mainly flavonoid aglycones and terpene trilactones), and EGb 761-DT-H (containing mainly flavonoid aglycones) samples exhibited cytotoxicity and inhibitory activity with IC (50) values of 46.36 +/- 2.43 microM, 10.27 +/- 0.88 microM, and 14.93 +/- 0.73 microM in L1210 cell-based assays, respectively. This resulted in 41.74 %, 60.72 %, and 63.76 % reductions in tumor weight after 10 days of treatment, respectively. In summary, the anticancer activity of EGb 761 can be improved by increasing the concentration of the aglycone form of the flavonoid. Terpene trilactones cannot exert the anticancer effects of flavonoids in vivo. Raising the levels of the free radical scavenger enzymes GST, SOD and CAT may be one of the involved anticancer mechanisms.

S29-04 Markers of oxidative stress in bipolar disorder and in schizophrenia

There has been strong evidence in recent years that oxygen free radicals may play an important role in the pathophysiology of schizophrenia and bipolar disorder. More recent evidence of altered antioxidant defenses in major mental illnesses has been accumulating. Much of the focus on antioxidant defense mechanism has been on the key scavenging antioxidant enzymes: Superoxide dismutase (SOD), Catalase (CAT) and Glutathione peroxidase (GSH-Px). Of these, SOD activity has been the most frequently examined.The imbalance between antioxidant and oxidant mechanisms increases what is called oxidative stress, when the oxygen free species react and could damage lipids, proteins, carbohydrates and DNA. Lipids are the main compound of cellular membrane and the oxygen free radicals reaction to them is called lipid peroxidation. Several plasma lipid peroxides, like MDA, are measured by Thiobarbituric acid reactive substances (TBARS). Increased levels of TBARS have been reported in the plasma of drug-free and medicated bipolar disorder and schizophrenic patients. It is hypothesized that psychotic symptoms seen in both disorders might be a consequence of peroxidative injury to membrane phospholipids. Moreover, neuronal functioning is affected by this injury either by changes in membrane fluidity or by alterations in membrane receptors, which can cause neurotransmitter uptake and release impairment and even cell death.

Sperm head defects and disturbances in spermatozoal chromatin and DNA integrities in idiopathic infertile subjects: Association with cigarette smoking

Objectives To evaluate sperm chromatin and DNA integrities in idiopathic infertile men and determine the possible association(s) of cigarette smoking on oxidative stress markers, antioxidant capacity and semen quality. Subjects and methods Semen samples from men referring to the andrology laboratory were categorized into 3 groups: fertile non-smokers ( n = 16), infertile non-smokers ( n = 36), and infertile smokers ( n = 34). Semen analysis was performed according to WHO criteria. The percentage of sperm DNA fragmentation index (%DFI) and the percentage of sperm with abnormally high DNA stainability (HDS%; immature spermatozoa) were determined by SCSA using the metachromatic properties of acridine orange. Lipid peroxidation, superoxide dismutase (SOD), catalase (CAT) and reduced glutathione (GSH) levels in seminal plasma and spermatozoa were measured by spectrophotometric assays. Results The classical semen parameters were negatively correlated with lipid peroxidation in spermatozoa; motility and morphology were negatively correlated with %DFI ( p < 0.05). HDS% was also negatively correlated with above markers except for morphology ( r = − 0.352, p = 0.081). DFI% and HDS% were significantly higher in the infertile smokers group than in infertile non-smokers ( p = 0.032; p = 0.001 respectively). Cigarette smoking was significantly associated with DFI%, HDS%, TBARS and the fraction of “round-headed” sperm ( r = 0.796, p = 0.0001; r = 0.371, p = 0.033; r = 0.606, r = 0.591, p = 0.001 respectively), and decreased SOD levels ( r = − 0.545). Conclusion DFI%, HDS% and round-head sperms are increased in idiopathic infertile men; this increase is associated with cigarette smoking. These defects may be attributed to increased oxidative stress and insufficient scavenging antioxidant enzymes in the seminal fluid of infertile patients.

Antioxidant potential by arabinoxylan rice bran, MGN-3/biobran, represents a mechanism for its oncostatic effect against murine solid Ehrlich carcinoma

We have recently examined the oncolytic effect of arabinoxylan rice bran, MGN-3/biobran, against solid Ehrlich carcinoma (SEC)-bearing mice via immune-modulation and apoptosis [N.K. Badr El-din, E. Noaman, M. Ghoneum, In vivo tumor inhibitory effects of nutritional rice bran supplement MGN-3/biobran on Ehrlich carcinoma-bearing mice, Nutr. Cancer 60 (2) (2008) 235–244]. In the present study, we examined the antioxidant system as another possible mechanism through which MGN-3 exerts its oncostatic potential. Female albino mice were inoculated intramuscularly in the right thigh with Ehrlich ascites carcinoma (EAC) cells. MGN-3 (25 mg/kg body weight) was injected intraperitoneally (i.p.) six times a week for 25 days into mice at either day 4 or day 11 post-EAC cell inoculation. Tumor growth, lipid peroxidation (LPx), glutathione (GSH) contents, the activity of the antioxidant scavenger enzymes, and alterations in gene expression were examined. MGN-3 efficiently suppressed the growth of tumors, which was associated with normalization of the LPx levels and augmentation of GSH contents. MGN-3 enhanced the activity of the endogenous antioxidant scavenging enzymes – superoxide dismutase (SOD), glutathione peroxidase (GPx), catalase (CAT) and glutathione- S-transferase (GST) – in blood, liver, and tumor tissue. Similarly it up-regulated the expression of GPx, SOD1 and CAT mRNA in the liver. The effect of MGN-3 was more pronounced when treated early, at day 4 of tumor cell inoculation, as compared to later treatment at 11 days. In conclusion, MGN-3-induced oncostatic activity by modulating lipid peroxidation, augmenting the antioxidant defense system and protecting against oxidative stress.

The dynamics of nutrient utilization and growth of apple root stock ‘M9 EMLA’ in temporary versus continuous immersion bioreactors

The present study investigated the dynamics of nutrient utilization and various growth and physiological parameters during in vitro proliferation of apple root stock ‘M9 EMLA’ in two different bioreactor systems, i.e. temporary and continuous immersions. Individual shoots obtained from temporary immersion system had higher dry mass and were of better quality than those obtained from continuous immersion. In continuous immersion bioreactor, apple shoots appeared to utilize more nutrients from liquid culture medium than that from temporary immersion. The shoot growth was limited by the availability of phosphate and nitrogen in continuous immersion system. The shoots produced in temporary immersion bioreactor showed higher photosynthetic rate, maximum quantum yield of photosystem-II and slow but steady rate of nutrient absorption, indicating the occurrence of higher photomixotrophic metabolism. The study also showed that high level of antioxidant scavenging enzymes in shoots grown in continuous immersion system induced physiological changes to foster adaptation to stresses.

Free radical scavenging capacity and antioxidant enzyme activity in deerberry ( Vaccinium stamineum L.)

Fruit from three genotypes (‘B-76’, ‘B-59’ and ‘SHF-3A’) of deerberry [ Vaccinium stamineum L.] were evaluated for fruit quality, total anthocyanin and phenolic contents, antioxidants, antioxidant capacity, and antioxidant enzyme activity. The fruit soluble solids, titratable acids, total anthocyanins, and total phenolic contents varied with genotypes. Cyanidin 3-galactoside and cyanidin 3-arabinoside were the two predominant anthocyanins. Resveratrol was also found in deerberries. Among the three genotypes, ‘B-76’ had higher amount of anthocyanins, phenolic compounds and resveratrol than ‘B-59’ and ‘SHF-3A’. Deerberries contained potent free radical scavenging activities for 2,2-Di (4- tert-octylphenyl)-1-picrylhydrazyl (DPPH ), 2,2′-azinobis(3-ethylbenzothiazoline-6-sulfonic acid) diammonium salt (ABTS + ), peroxyl radical (ROO ), superoxide radicals ( O 2 - ), hydrogen peroxide (H 2O 2), hydroxyl radicals ( OH), and singlet oxygen ( 1O 2) radicals and also had high activities of antioxidant enzymes including glutathione-peroxidase (GSH-POD), glutathione reductase (GR), superoxide dismutase (SOD), ascorbate peroxidase (AsA-POD), guaiacol peroxidase (G-POD), monodehydroascorbate radical reductase (MDAR), dehydroascorbate reductase (DHAR), glutathione reductase (GR)] and non-enzyme antioxidants [ascorbic acid (ASA) and reduced glutathione (GSH)]. Antioxidant capacities were highly correlated to antioxidant enzymes activities. Among the three genotypes, ‘B-76’ had the highest level of antioxidants and antioxidant enzyme activity.

Oxidative stress plays a role in diabetes-induced bladder dysfunction in a rat model

ObjectivesTo evaluate the oxidative status of the bladder 8 weeks after diabetes induction. Oxidative stress has recently been implicated in the pathogenesis of diabetes complications, but its role in diabetic cystopathy has not been studied. MethodsSprague-Dawley rats were divided into three groups: control (n = 11), diuretic control (5% sucrose drink; n = 6), and streptozotocin-induced diabetic group (n = 14). Eight weeks later, the bladders were dissected. We measured the antioxidant scavenging enzymes (catalase and superoxide dismutase)-like activity and the levels of the thiobarbituric acid reactive substances, as a marker of lipid peroxidation. We also examined the levels of inducible nitric oxide synthase and apoptosis in the bladders. ResultsWe found a statistically significant reduction in the catalase-like activity in the bladders from the diabetic group compared with the other groups (P = 0.017, diabetic versus control); the difference in the superoxide dismutase-like activity was not statistically significant among the groups. The thiobarbituric acid reactive substances levels were significantly greater in the diabetic compared with other groups (131.9 ± 47.5, 46.7 ± 17.9, and 60.9 ± 25.4 nmol/mg protein in the diabetic, control, and diuretic group, respectively, P = 0.006, diabetic versus control). Immunohistochemical and apoptosis studies showed a statistically significant increased number of inducible nitric oxide synthase-positive cells and apoptotic cells in the diabetic bladder smooth muscle cells (P <0.001). ConclusionsOur findings showed that oxidative stress occurred in the bladders of the STZ-diabetic rats and was not mediated by diuresis. The oxidative damage of the smooth muscle cells may be a contributory factor in diabetic cystopathy.

Protective role of sanumgerman against γ-irradiation–induced oxidative stress in Ehrlich carcinoma-bearing mice

This study was conducted to evaluate the potency of organogermanium sanumgerman (germanium lactate citrate) as a protective agent against γ-irradiation–induced toxicity, which may be extended to affect normal organs in mice bearing solid Ehrlich tumor. Another aim was to evaluate the antitumor effect of this agent, alone or in combination with irradiation, by measuring the change in tumor volume. Female Swiss albino mice were injected intraperitoneally with sanumgerman 1 day before tumor inoculation, then once every other day for a total of seven injections during 13 days. The last injection was given 20 minutes before exposure to a single dose of whole body γ-irradiation at a dose level of 6.5 Gy. Lipid peroxidation (LPx), glutathione (GSH) levels, and activity of the antioxidant scavenger enzymes glutathione reductase, glutathione peroxidase, glucose 6-phosphate dehydrogenase, superoxide dismutase, and catalase were estimated in blood, liver, and spleen at days 1 and 7 postirradiation. Ehrlich carcinoma–bearing mice, whether exposed to irradiation or not, showed marked elevation in LPx level accompanied by a decline in GSH content and in the activity of related antioxidant enzymes that was more apparent at day 7 postirradiation. Pretreatment with sanumgerman potentially reversed the investigated parameters in nonirradiated, tumor-bearing mice. Additionally, it prevented LPx, and ameliorated to a great extent GSH content and the activity of antioxidant scavenger enzymes in blood, liver, and spleen of mice exposed to γ-irradiation. Moreover, sanumgerman significantly suppressed the growth of tumor and efficiently produced a synergistic effect with irradiation in reducing tumor volume. Therefore, this agent may be a good candidate for use with irradiation to enhance tumor cell killing without affecting normal organs of the tumor host.

Yap1 and Skn7 Control Two Specialized Oxidative Stress Response Regulons in Yeast

Yap1 and Skn7 are two yeast transcriptional regulators that co-operate to activate thioredoxin (TRX2) and thioredoxin reductase (TRR1) in response to redox stress signals. Although they are both important for resistance to H2O2, only Yap1 is important for cadmium resistance, whereas Skn7 has a negative effect upon this response. The respective roles of Yap1 and Skn7 in the induction of defense genes by H2O2 were analyzed by two-dimensional gel electrophoresis. Yap1 controls a large oxidative stress response regulon of at least 32 proteins. Fifteen of these proteins also require the presence of Skn7 for their induction by H2O2. Although about half of the Yap1 target genes do not contain a consensus Yap1 recognition motif, the control of one such gene, TSA1, involves the binding of Yap1 and Skn7 to its promoter in vitro. The co-operative control of the oxidative stress response by Yap1 and Skn7 delineates two gene subsets. Remarkably, these two gene subsets separate antioxidant scavenging enzymes from the metabolic pathways regenerating the main cellular reducing power, glutathione and NADPH. Such a specialization may explain, at least in part, the dissociated function of Yap1 and Skn7 in H2O2 and cadmium resistance.

Endogenous antioxidant enzymes and glutathione S-transferase in protection of mesothelioma cells against hydrogen peroxide and epirubicin toxicity.

We have previously shown that cultured malignant mesothelioma cells contain elevated manganese superoxide dismutase (MnSOD) mRNA levels and activities compared with non-malignant mesothelial cells. As many cytotoxic drugs generate both superoxide and hydrogen peroxide, we assessed the relative significance of catalase and the glutathione redox cycle, as well as glutathione S-transferase (GST), in protecting these cells against hydrogen peroxide and epirubicin toxicity. Mesothelioma cell lines containing high (M38K cells) and low (M14K cells) MnSOD, and non-malignant MeT-5A mesothelial cells were selected for the study. M38K cells were the most resistant of these three cell types to hydrogen peroxide (0.1-0.5 mM, 4 h) and epirubicin (0.1-0.5 microg ml(-1), 48 h) as judged by lactate dehydrogenase (LDH) release and by high-energy nucleotide (ATP, ADP, AMP) depletion. Total glutathione was higher in M38K cells (63.8 +/- 20.3 nnmol mg(-1) protein) than in M14K (25.2 +/- 8.2 nmol mg[-1]) or MeT-5A cells (23.5 +/- 4.5 nmol mg[-1]). Furthermore, GST specific activity was higher in M38K cells (111.3 +/- 15.8 U mg[-1]) than in M14K cells (77.4 +/- 6.6 U mg[-1]) or in MeT-5A cells (68.8 +/- 7.6 U mg[-1]). Western blotting indicated the presence of GST-pi in all these cells, the reactivity again being highest in M38K cells. Depletion of glutathione by buthionine sulphoximine and inhibition of catalase by aminotriazole enhanced hydrogen peroxide toxicity in all cell types, while only the depletion of glutathione increased epirubicin toxicity. We conclude that simultaneous induction of multiple antioxidant enzymes can occur in human mesothelioma cells. In addition to the high MnSOD activity, hydrogen peroxide scavenging antioxidant enzymes, glutathione and GST can partly explain the high hydrogen peroxide and epirubicin resistance of these cells in vitro.

Non-optimal growth temperatures and antioxidants in the leaves of Sorghum bicolor (L.) Moench. II. Short-term acclimation

Summary In pursuing previous studies on long-term acclimation to non-optimal, non-stressing growth temperatures (Badiani et al., 1993 b; Paolacci et al., 1997; Fusari et al., in press), the foliar antioxidant status and the photosynthetic capacity were evaluated in two sorghum [ Sorghum bicolor (L.) Moench.] cultivars of different agroclimatic provenance, namely Aralba and ICSV 112, which were grown at the near-optimal temperature of 27 ± 0.3 °C and then gradually (1°Ch −1 ) and transiently (up to 120 h) exposed to suboptimal, 17 ± 0.4 °C, or supraoptimal, 37 ± 0.1 °C, temperatures, under moderate light intensity and ad libitum water and mineral nutrition. Comparative analysis of short-term and long-term responses to non-optimal growth temperatures suggested that: i ) even realistic, i.e. limited and gradual, upward or downward shifts from the normative growth temperature, incapable of causing evident stress symptoms, might per se enhance the formation of reactive oxygen species whose effect, albeit not drastic, appear to be more noxious during the early stages of the exposure; ii ) this disthermia-driven oxidative burst triggers almost immediate and extensive changes in all of the major antioxidant metabolites and scavenging enzymes. This could be aimed at preparing plant tissues in case moderate disthermia flows into authentic temperature stress; iii ) certain short-term antioxidant changes persist beyond the relief of the disthermic regimes; and iv ) however, in case the exposure to non-optimal growth temperatures becomes chronic, long-term adjustment processes take place, consisting of increases in protective pigments, ascorbic acid and glutathione, but without the involvement of antioxidant enzymes. Such a strategy might be aimed at keeping disthermia-dependent oxidative stress under control at the lowest possible price in terms of metabolic resources.