Accumulation and oxidative stress biomarkers in Japanese flounder larvae and juveniles under chronic cadmium exposure

This study investigated how Cd exposure affected oxidative biomarkers in Japanese flounder, Paralichthys olivaceus, at early life stages (ELS). Fish were exposed to waterborne Cd (0–48 µg L^− 1) from embryonic to juvenile stages for 80 days. Growth, Cd accumulation, activities of superoxide dismutase (SOD, EC 1.15.1.1), catalase (CAT, EC 1.11.1.6), glutathione S-transferase (GST, EC 2.5.1.18), and levels of glutathione (GSH) and lipid peroxidation (LPO) were investigated at three developmental stages. Flounder growth decreased and Cd accumulation increased with increasing Cd concentration. In metamorphosing larvae, CAT and SOD activities were inhibited and GSH level was elevated, while LPO was enhanced by increasing Cd concentrations. CAT and GST activities of settling larvae were inhibited but GSH level was elevated at high Cd concentrations. In juveniles, SOD activity and LPO level were increased but GST activity was inhibited as Cd concentration increased. Antioxidants in flounder at ELS were able to develop ductile responses to defend against oxidative stress, but LPO fatally occurred due to Cd exposure. These biochemical parameters could be used as effective oxidative biomarkers for evaluating Cd contamination and toxicity in marine environments: CAT, SOD, GSH, and LPO for metamorphosing stage; CAT, GSH, and GST for settling stage; and SOD, GST, and LPO for juvenile stage.     

Rye intercrop management in transplant tomatoes

Rye cv. Dańkowskie was seeded in September and desiccated with glyphosate in May of the following year. Transplants of tomato cv. Rumba were planted on plot covered with rye mulch and also on plot cultivated conventionally. Weed infestation on no-tilled plot was reduced significantly in comparison to conventional tillage during the first three weeks after planting of tomatoes. Temperature of no-tilled soil covered with rye mulch was lower during the whole period of vegetation. Total and marketable yield and size of tomato fruits were significantly bigger in conventional cultivation.     

Effect of copper exposure on GST activity and on the expression of four GSTs under oxidative stress condition in the monogonont rotifer, Brachionus koreanus

Glutathione S-transferases (GSTs; EC 2.5.1.18) are major enzymes that function in Phase II detoxification reactions by catalyzing the conjugation of reduced glutathione through cysteine thiol. In this study, we cloned and sequenced four GST genes from the monogonont rotifer Brachionus koreanus. The domain regions of four Bk-GSTs showed a high similarity to those of other species. In addition, to evaluate the potential of GST genes as an early warning signal for oxidative stress, we exposed sublethal concentrations of copper (Cu) to B. koreanus and measured glutathione (GSH) contents and several antioxidant enzymes such as glutathione S-transferase (GST), glutathione peroxidase (GPx; EC 1.11.1.9), and glutathione reductase (GR; EC 1.8.1.7). The reactive oxygen species (ROS) at 12 h and 24 h after copper exposure increased significantly. GSH contents however did not increase significantly and even it decreased at 0.24 mg/L at 12 h. The activities of several antioxidant enzymes, particularly GPx and GR, showed a dramatic increase in 0.24 mg/L of CuCl₂. Messenger RNAs of each Bk-GST showed different patterns of modulations according to GST types, and particularly, Bk-GST-omega, Bk-GST-sigma, and Bk-GST zeta genes were highly sensitive to Cu. These results indicate that Bk-GSTs, functioning as one of the enzymatic defense mechanisms particularly in the early stage of oxidative stress response, were induced by Cu exposure. This also suggests that these genes and related enzymes have a potential as biomarkers for a more sensitive initial stress response.     

Nickel-induced oxidative stress and the role of antioxidant defence in rice seedlings

Seedlings of rice (Oryza sativa L.) cv. Pant-12 grown in sand cultures containing 200 and 400 μM NiSO₄, showed a decrease in length and fresh weight of roots and shoots. Nickel was readily taken up by rice seedlings and the concentration was higher in roots than shoots. Nickel-treated seedlings showed increased rates of superoxide anion (O₂ ^•− ) production, elevated levels of H₂O₂ and thiobarbituric acid reactive substances (TBARS) demonstrating enhanced lipid peroxidation, and a decline in protein thiol levels indicative of increased protein oxidation compared to controls. With progressively higher Ni concentrations, non-protein thiol and ascorbate (AsA) increased, whereas the level of low-molecular-weight thiols (such as glutathione and hydroxyl-methyl glutathione), the ratio of these thiols to their corresponding disulphides, and the ratio of AsA to dehydroascorbic acid declined in the seedlings. Among the antioxidant enzymes studied, the activities of all isoforms of superoxide dismutase (Cu-Zn SOD, Mn SOD and Fe SOD), guaiacol peroxidases (GPX) and ascorbate peroxidase (APX) increased in Ni-treated seedlings, while no clear alteration in catalase activity was evident. Activity of the ascorbate-glutathione cycle enzymes monodehydroascorbate reductase (MDHAR), dehydroascorbate reductase (DHAR) and glutathione reductase (GR)—significantly increased in Ni-treated seedlings. However such increase was apparently insufficient to maintain the intracellular redox balance. Results suggest that Ni induces oxidative stress in rice plants, resulting in enhanced lipid peroxidation and decline in protein thiol levels, and that (hydroxyl-methyl) glutathione and AsA in conjunction with Cu-Zn SOD, GPX and APX are involved in stress response.     

The Fructose Transport Mechanism in Microsomal Membrane Vesicles from Rye Roots

In a fructose-containing medium in which rye root-microsomal membrane vesicles had reached the equilibrium of uptake of fructose, the presence of both Mg²⁺ and ATP caused the efflux of fructose from the vesicles. Among nucleotides examined, ATP caused the largest efflux of fructose. The efflux of fructose dependent on Mg²⁺ and ATP was quite insensitive to a protonophore, carbonylcyanide m-chlorophenylhydrazone (CCCP), which actually abolished MgATP-dependent proton accumulation in the vesicles, while it was largely inhibited by vanadate, which inhibits ATP-binding cassette transporters (ABCTs). The Michaelis-Menten constant (Km) of the efflux of fructose was 0.4 mM. It was observed that fructose stimulated the ATPase activity of the vesicles and that vanadate markedly decreased the fructose-stimulated ATPase activity. This revised version was published online in July 2006 with corrections to the Cover Date.     

Chromosomal Location of 46 New RAPD Markers in Rye (Secale Cereale L.)

The polymerase chain reaction (PCR) was used to locate RAPD markers using disomic wheat–rye addition lines in order to develop a set of molecular markers distributed on the seven rye chromosomes. We carried out RAPD amplifications on genomic DNA of wheat Chinese Spring (CS), rye Imperial (I), the amphiploid wheat–rye and the seven disomic wheat–rye addition lines (1R–7R) using 140 different 10-mer oligonucleotides. Forty six new RAPD markers were located on the seven rye chromosomes and all the disomic wheat–rye addition lines were identified on the basis of their amplification patterns. The number of RAPD bands located on 1R, 2R, 3R, 4R, 5R, 6R and 7R chromosomes were 5, 8, 11, 8, 8, 10 and 6, respectively. The seven wheat–rye addition lines can be distinguished using only the following three 10-mer oligonucleotides: OPA16, OPF19 and GEN3-605, the other RAPD primers being useful for this purpose. The use of these RAPDs as a source of molecular markers that could be linked to interesting genes or other important agronomic traits is discussed.     

Very long chain alkylresorcinols accumulate in the intracuticular wax of rye (Secale cereale L.) leaves near the tissue surface

Alkylresorcinols (ARs) are bioactive compounds occurring in many members of the Poaceae, likely at or near the surface of various organs. Here, we investigated AR localization within the cuticular wax layers of rye (Secale cereale) leaves. The total wax mixture from both sides of the leaves was found to contain primary alcohols (71%), alkyl esters (11%), aldehydes (5%), and small amounts (<3%) of alkanes, steroids, secondary alcohols, fatty acids and unknowns. A homologous series of ARs (3%) was identified by GC-MS and comparison with a synthetic standard of nonadecylresorcinol. The alkyl side chains of the wax ARs contained odd numbers of carbons ranging from C19 to C27, with a prevalence of C21, C23 and C25. Waxes from both sides of the leaf, analyzed separately in a second experiment, comprised the same compound classes in similar relative amounts and with similar homolog patterns. Finally, the epicuticular and intracuticular wax layers were sampled separately from the abaxial side of the leaf. While ARs accounted for 2% of the intracuticular wax, they were not detectable in the epicuticular wax. The intracuticular wax was also slightly enriched in steroids, whereas the epicuticular layer contained more primary alcohols. All other wax constituents were distributed evenly between both wax layers.     

Induction of apoptosis in lymphoid cells by thiol-mediated oxidative stress

Summary In previous studies, oxidants such as hydrogen peroxide (H₂O₂) or hydroperoxy fatty acids were shown to induce apoptosis in the CEM human T cell line as demonstrated by the cleavage of cellular DNA into a 180-base pair ladder. Oxidant-induced DNA fragmentation was detectable within 3 h and inhibitable by various antioxidants. In the present study, apoptosis is shown to also be induced by the addition of low doses (0.1–3 mM) of N-acetyl-L-cysteine (NAC), reduced glutathione (GSH) or cysteine. By contrast, higher concentrations (10 mM) of the same thiols displayed a paradoxical lack of toxicity. Thiol-induced apoptosis was completely prevented by the addition of BAPTA-AM, an intracellular calcium chelator, or by simultaneous treatment with 5 mM pyruvate which forms a thiazolidine complex with sulfhydryl compounds. Catalase or glutathione peroxidase, but not Superoxide dismutase, protected the cells from thiol-induced apoptosis demonstrating a role for H₂O₂. The ability of thiol compounds to either evoke or prevent oxidative stress implies a unique role for these agents in the control of apoptosis in lymphoid cells.     

Susceptibility to oxidative stress and modulated expression of antioxidant genes in the copper-exposed polychaete Perinereis nuntia

To identify and evaluate potentially useful biomarkers for oxidative stress as early warning indices in the polychaete, Perinereis nuntia, we exposed P. nuntia to copper (Cu) and measured several biomarker enzymes (glutathione S-transferase; GST, glutathione peroxidase; GPx, Metallothionein-like protein; MTLPs, and catalase; CAT) and genes (Pn-GSTs, Pn-CAT, and Pn-MT) with a cellular oxidative index, reactive oxygen species (ROS) level. Accumulated Cu concentrations in P. nuntia increased in a time-dependent manner. Intracellular ROS reached high levels 6h after exposure in P. nuntia with an increase of GST activity and glutathione (GSH) content. Particularly, GSH in polychaetes showed a positive correlation with Cu contents accumulated in P. nuntia. Messenger RNA expressions of GST sigma and GST omega showed relatively high expressions at 50 μg/L of Cu exposure, even though the moderate increase of rest of GST isoforms was also observed. Also regarding long-term exposure, we reared P. nuntia in sediments for 15 days, and found that there was an obvious increase of Pn-GSTs, Pn-CAT, and Pn-MT genes with elevated concentrations of Cu and Cd in polychaete body, compared to initial levels, suggesting that P. nuntia in sediment was affected by metals as well as by other organic pollutants to induce oxidative stress genes and enzymes. These findings suggest that oxidative stress is a potential modulator of defense system of P. nuntia. Several potential biomarker genes are available as early warning signals for environmental biomonitoring.     

Enhanced tolerance to oxidative stress in transgenic tobacco plants expressing three antioxidant enzymes in chloroplasts

The effect of simultaneous expression of genes encoding three antioxidant enzymes, copper zinc superoxide dismutase (CuZnSOD, EC 1.15.1.1), ascorbate peroxidase (APX, EC 1.11.1.11), and dehydroascorbate (DHA) reductase (DHAR, EC 1.8.5.1), in the chloroplasts of tobacco plants was investigated under oxidative stress conditions. In previous studies, transgenic tobacco plants expressing both CuZnSOD and APX in chloroplast (CA plants), or DHAR in chloroplast showed enhanced tolerance to oxidative stresses, such as paraquat and salt. In this study, in order to develop transgenic plants that were more resistant to oxidative stress, we introduced the gene encoding DHAR into CA transgenic plants. Mature leaves of transgenic plants expressing all three antioxidant genes (CAD plants) had approximately 1.6–2.1 times higher DHAR activity, and higher ratios of reduced ascorbate (AsA) to DHA, and oxidized glutathione (GSSG) to reduced glutathione (GSH) compared to CA plants. CAD plants were more resistant to paraquat-induced stress, exhibiting only 18.1% reduction in membrane damage relative to CA plants. In addition, seedlings of CAD plants had enhanced tolerance to NaCI (100 mM) compared to CA plants. These results indicate that the simultaneous expression of multiple antioxidant enzymes, such as CuZnSOD, APX, and DHAR, in chloroplasts is more effective than single or double expression for developing transgenic plants with enhanced tolerance to multiple environmental stresses.     

Ribulose-1,5-Bisphosphate Carboxylase/Oxygenase Content and Activity in Wheat, Rye and Triticale

Photosynthetic parameters were measured in triticale and its parents wheat and rye. Soluble protein content in leaves, ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) content per fresh mass, total chlorophyll content, biomass yield, leaf area, leaf mass and specific leaf mass were higher but Rubisco content expressed as percentage of soluble protein, carboxylase activity, photosynthetic rate and stomatal conductance were significantly lower in rye than in wheat. Native-PAGE of Rubisco revealed that rye carboxylase was different from that of wheat. The difference was not related to either the small or large subunit of Rubisco but, may be, to the ionic and/or other properties of the Rubisco protein moiety. Triticale Rubisco was similar to wheat. For most of the studied physiological parameters, triticale showed much more similarity with wheat than with rye.     

Alteration of mitochondrial protein complexes in relation to metabolic regulation under short-term oxidative stress in Arabidopsis seedlings

Plants reconfigure their metabolic network under stress conditions. Changes of mitochondrial metabolism such as tricarboxylic acid (TCA) cycle and amino acid metabolism are reported in Arabidopsis roots but the exact molecular basis underlying this remains unknown. We here hypothesise the reassembly of enzyme protein complexes to be a molecular mechanism for metabolic regulation and tried in the present study to find out mitochondrial protein complexes which change their composition under oxidative stress by the combinatorial approach of proteomics and metabolomics. Arabidopsis seedlings were treated with menadione to induce oxidative stress. The inhibition of several TCA cycle enzymes and the oxidised NADPH pool indicated the onset of oxidative stress. In blue native/SDS-PAGE analysis of mitochondrial protein complexes the intensities of 18 spots increased and those of 13 spots decreased in menadione treated samples suggesting these proteins associate with, or dissociate from, protein complexes. Some spots were identified as metabolic enzymes related to central carbon metabolism such as malic enzyme, glyceraldehyde-3-phosphate dehydrogenase, monodehydroascorbate reductase and alanine aminotransferase. The change in spot intensity was not directly correlated to the total enzyme activity and mRNA level of the corresponding enzyme but closely related to the metabolite profile, suggesting the metabolism is regulated under oxidative stress at a higher level than translation. These results are somewhat preliminary but suggest the regulation of the TCA cycle, glycolysis, ascorbate and amino acid metabolism by reassembly of plant enzyme complexes.     

Alterations in root lipid peroxidation and antioxidative responses in two rice cultivars under NaCl-salinity stress

The comparative alterations of short term NaCl stress and recovery on growth, water relations, ionic composition, lipid peroxidation and antioxidants in roots of two rice cultivars differing in salt tolerance were studied. Exposed for 24 h to increasing (50, 100 and 150 mmol l⁻¹) concentrations of NaCl, roots of 12D Oryza sativa L. cv. Lunishree and cv. Begunbitchi decreased in fresh weight, dry weight and relative water content. Increased Na⁺ and decreased K⁺ ion were determined at increasing NaCl concentrations. Both peroxide content and lipid peroxidation measured in terms of MDA level increased and the ratio was higher in Begunbitchi compared to Lunishree. Recovered roots showed lower peroxide and MDA content. Ascorbate and glutathione contents increased in the stressed and recovered roots of Lunishree, but decreased in Begunbitchi with increasing NaCl concentrations. Although SOD, CAT and GR activities decreased in the stressed roots, CAT activity also increased in recovered roots of both the cultivars. The POX activity increased in stressed and recovered roots of both Lunishree and Begunbitchi. Higher free radicals scavenging capacity and more efficient protection mechanism of Lunishree against salt stress, as revealed by the lower level of lipid peroxidation and improved plant water status as well as activities of some of the antioxidants, suggest that significant cultivar differences in response to salt stress in rice are closely related to differences in the activities of antioxidants and ion content. Another possible conclusion is that improved tolerance to salt stress may be accomplished by increased capacity of antioxidative system.     

Diabetes,oxidative stress,and antioxidants: a review

Increasing evidence in both experimental and clinical studies suggests that oxidative stress plays a major role in the pathogenesis of both types of diabetes mellitus. Free radicals are formed disproportionately in diabetes by glucose oxidation, nonenzymatic glycation of proteins, and the subsequent oxidative degradation of glycated proteins. Abnormally high levels of free radicals and the simultaneous decline of antioxidant defense mechanisms can lead to damage of cellular organelles and enzymes, increased lipid peroxidation, and development of insulin resistance. These consequences of oxidative stress can promote the development of complications of diabetes mellitus. Changes in oxidative stress biomarkers, including superoxide dismutase, catalase, glutathione reductase, glutathione peroxidase, glutathione levels, vitamins, lipid peroxidation, nitrite concentration, nonenzymatic glycosylated proteins, and hyperglycemia in diabetes, and their consequences, are discussed in this review. In vivo studies of the effects of various conventional and alternative drugs on these biomarkers are surveyed. There is a need to continue to explore the relationship between free radicals, diabetes, and its complications, and to elucidate the mechanisms by which increased oxidative stress accelerates the development of diabetic complications, in an effort to expand treatment options.     

Protective effect of Operculina turpethum against 7,12-dimethyl benz(a)anthracene induced oxidative stress with reference to breast cancer in experimental rats

Reactive oxygen species (ROS) directly or indirectly involves in multistage process of carcinogenesis. Antioxidant activity of methanolic extract of Operculina turpethum stems (MEOT) on 7,12 dimethylbenz(a)anthracene (DMBA) induced breast cancer was investigated in female Sprague-Dawley rats. Changes in the levels of lipid peroxidation and antioxidants system was evaluated in addition to tumour development. Twenty four female rats were divided into four groups: control, DMBA, DMBA + MEOT and MEOT. In the DMBA group, rats were intragastrically administered with 20 mg of DMBA using corn oil as vehicle. Animals of DMBA + MEOT group received a single dose of 20 mg of DMBA dissolved in corn oil intragastrically followed by O. turpethum extract (100 mg/kg body weight), while MEOT group received O. turpethum extract (100 mg/kg body weight) intragastrically daily for a period of 45 days. After the experimental period of 45 days, oxidative stress parameters were assessed in serum, liver and breast of both control and experimental groups. In addition to this, tumour weight of breast was also assessed. A significant increase in lipid peroxidation levels were observed in the tested samples of cancer induced rats while the activities of enzymic antioxidants such as superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx) and non-enzymic antioxidants like glutathione (GSH), ascorbic acid (Vitamin C) and α-tocopherol (Vitamin E) were decreased in cancer-bearing animals when compared to control animals. A significant (P < 0.05) increase in the tumour weight was observed in the breast of DMBA group and the breast tumour weight decreased significantly (P < 0.05) in the DMBA + MEOT groups. Oral administration of MEOT remarkably reduced the lipid peroxidation activity and increased the antioxidants level in drug treated animals and decreased the tumour weight significantly (P < 0.05). This result suggests that MEOT shows antioxidant activity and play a protective role against DMBA induced breast cancer.