Modulation of copper toxicity-induced oxidative damage by excess supply of iron in maize plants

In this study, we examined the modulation of Cu toxicity-induced oxidative stress by excess supply of iron in Zea mays L. plants. Plants receiving excess of Cu (100 μM) showed decreased water potential and simultaneously showed wilting in the leaves. Later, the young leaves exhibited chlorosis and necrotic scorching of lamina. Excess of Cu suppressed growth, decreased concentration of chloroplastic pigments and fresh and dry weight of plants. The activities of peroxidase (EC 1.11.1.7; POD), ascorbate peroxidase (EC 1.11.1.11; APX) and superoxide dismutase (EC 1.15.1.1; SOD) were increased in plants supplied excess of Cu. However, activity of catalase (EC 1.11.1.6; CAT), was depressed in these plants. In gel activities of isoforms of POD, APX and SOD also revealed upregulation of these enzymes. Excess (500 μM)-Fe-supplemented Cu-stressed plants, however, looked better in their phenotypic appearance, had increased concentration of chloroplastic pigments, dry weight, and improved leaf tissue water status in comparison to the plants supplied excess of Cu. Moreover, activities of antioxidant enzymes including CAT were further enhanced and thiobarbituric acid reactive substance (TBARS) and H₂O₂ concentrations decreased in excess-Fe-supplemented Cu-stressed plants. In situ accumulation of H₂O₂, contrary to that of O₂ ^·− radical, increased in both leaf and roots of excess-Cu-stressed plants, but Cu-excess plants supplied with excess-Fe showed reduced accumulation H₂O₂ and little higher of O₂ ^·− in comparison to excess-Cu plants. It is, therefore, concluded that excess-Cu (100 μM) induces oxidative stress by increasing production of H₂O₂ despite of increased antioxidant protection and that the excess-Cu-induced oxidative damage is minimized by excess supply of Fe.     

Aluminum-induced oxidative stress in maize

The relation between Al-toxicity and oxidative stress was studied for two inbred lines of maize (Zea mays L.), Cat100-6 (Al-tolerant) and S1587-17 (Al-sensitive). Peroxidase (PX), catalase (CAT) and superoxide dismutase (SOD) activities were determined in root tips of both lines, exposed to different Al(3+) concentrations and times of exposure. No increases were observed in CAT activities in either line, although SOD and PX were found to be 1.7 and 2.0 times greater than initial levels, respectively, in sensitive maize treated with 36 microM of Al(3+) for 48 h. The results indicate that Al(3+) induces the dose- and time dependent formation of reactive oxygen species (ROS) and subsequent protein oxidation in S1587-17, although not in Cat100-6. After exposure to 36 microM of Al(3+) for 48 h, the formation of 20+/-2 nmol of carbonyls per mg of protein was observed in S1587-17. The onset of protein oxidation took place after the drop of the relative root growth observed in the sensitive line, indicating that oxidative stress is not the primary cause of root growth inhibition. The presence of Al(3+) did not induce lipid peroxidation in either lines, contrasting with the observations in other species. These results, in conjunction with the data presented in the literature, indicate that oxidative stress caused by Al may harm several components of the cell, depending on the plant species. Moreover, Al(3+) treatment and oxidative stress in the sensitive maize line induced cell death in root tip cells, an event revealed by the high chromatin fragmentation detected by TUNEL analysis.     

Copper-induced oxidative damage and enhanced antioxidant defenses in the root apex of maize cultivars differing in Cu tolerance

The root apex is highly sensitive to many soil-derived stress factors. Copper (Cu), as a Fenton-type metal, may cause severe oxidative damage in plants at toxic concentrations. The aim of this study was to establish whether the apex is the primary site of Cu-induced oxidative stress and if so, whether there is a site-specific change in antioxidant defenses that can contribute to varietal differences in Cu tolerance. For this purposes, the influence of Cu excess on cell integrity and antioxidant defenses was investigated in two maize cultivars differing in Cu tolerance, Cu-tolerant cv. Oropesa and the Cu-sensitive cv. Orense. Three root zones were considered: 0–5 mm from the root apex (including root cap, meristem and transition zone), 5–10 mm (elongation zone) and 10–15 mm (maturation zone). The 24-h exposure to nominally 2 or 5 μM Cu (pCu7 or 6) confirmed the cultivar differences in Cu tolerance. Both cell membrane integrity, especially at the transition zone in the apex, and root elongation were considerably less damaged by elevated Cu in cv. Oropesa than in cv. Orense. Root tips of both cultivars accumulated similar Cu levels (analyzed after desorption of apoplastic Cu), but 5 μM Cu induced a higher increase of SOD activity (EC 1.15.1.1) in the 0–5 mm root tip region in Oropesa than in Orense. We conclude that this apical root tip zone is the most Cu-sensitive root part, but that the better performance of cv. Oropesa is not due to greater exclusion of Cu from the root apex. Further, the local increase of SOD activity in the root apex (0–5 mm) contributed to the maintenance of cell membrane integrity in the Cu-tolerant cv. Oropesa.     

Acephate induced oxidative damage in erythrocytes

The effect of oral administration of acephate (360 mg/kg body weight), for 15 days, daily, was investigated on the erythrocytes of male rats. Activities of acetyl cholinesterase and glucose-6-phosphate dehydrogenase decreased, while those of glutathione-s-transferase and glutathione reductase increased. Decreased glutathione content and increased lipid peroxidation suggest that there was increased oxidative stress in the erythrocytes of treated animals. Increased cholesterol/phospholipid ratio in the erythrocyte membranes and morphological changes in RBCs (scanning electron microscopy studies) were observed in acephate treated animals. The results clearly suggest that acephate induced oxidative stress in erythrocytes leads to morphological changes.     

PKC signaling in oxidative hepatic damage

Protein kinase C (PKC) is a family of isoenzymes differently involved in cell response to injury and many studies describe their role as "stress sensors". Oxidative stress is strictly involved in the pathogenesis of chronic liver diseases including alcohol- or drug-induced hepatotoxicity, iron overload, hepatitis and hepatocarcinoma development, but molecular mechanisms are not really defined. A crucial role of PKC as a redox sensitive signaling molecule has been widely accepted.     

The oxidative environment and protein damage

Proteins are a major target for oxidants as a result of their abundance in biological systems, and their high rate constants for reaction. Kinetic data for a number of radicals and non-radical oxidants (e.g. singlet oxygen and hypochlorous acid) are consistent with proteins consuming the majority of these species generated within cells. Oxidation can occur at both the protein backbone and on the amino acid side-chains, with the ratio of attack dependent on a number of factors. With some oxidants, damage is limited and specific to certain residues, whereas other species, such as the hydroxyl radical, give rise to widespread, relatively non-specific damage. Some of the major oxidation pathways, and products formed, are reviewed. The latter include reactive species, such as peroxides, which can induce further oxidation and chain reactions (within proteins, and via damage transfer to other molecules) and stable products. Particular emphasis is given to the oxidation of methionine residues, as this species is readily oxidised by a wide range of oxidants. Some side-chain oxidation products, including methionine sulfoxide, can be employed as sensitive, specific, markers of oxidative damage. The product profile can, in some cases, provide valuable information on the species involved; selected examples of this approach are discussed. Most protein damage is non-repairable, and has deleterious consequences on protein structure and function; methionine sulfoxide formation can however be reversed in some circumstances. The major fate of oxidised proteins is catabolism by proteosomal and lysosomal pathways, but some materials appear to be poorly degraded and accumulate within cells. The accumulation of such damaged material may contribute to a range of human pathologies.     

Flooding-induced changes in photosynthesis and oxidative status in maize plants

Maize plants (Zea mays L.) were subjected to soil flooding for 72, 96, and 120 h. A noticeable decrease in the rate of net photosynthesis (P_N) and the activity of ribulose-1,5-bisphosphate carboxylase (RuBPC, EC 4.1.1.39) were observed. The values of intercellular CO₂ concentrations (c_i) increased in all flooded plants without significant changes in stomatal conductance (g_s). The activity of phosphoenolpyruvate carboxylase (PEPC, EC 4.1.1.31) increased twofold 120 h after soil flooding. Flooding of maize plants led to a decrease in chlorophyll and protein levels and to slight increase of proline content. Flooded plants exhibited a large accumulation of leaf acidity. An increase in the values of some important parameters associated with oxidative stress, namely peroxides production, lipid peroxidation, and electrolyte leakage, confirmed the suggestion that root oxygen deficiency caused photooxidative damage in maize leaves.     

Mitochondrial dysfunction and oxidative damage in parkin-deficient mice

Loss-of-function mutations in parkin are the predominant cause of familial Parkinson's disease. We previously reported that parkin-/- mice exhibit nigrostriatal deficits in the absence of nigral degeneration. Parkin has been shown to function as an E3 ubiquitin ligase. Loss of parkin function, therefore, has been hypothesized to cause nigral degeneration via an aberrant accumulation of its substrates. Here we employed a proteomic approach to determine whether loss of parkin function results in alterations in abundance and/or modification of proteins in the ventral midbrain of parkin-/- mice. Two-dimensional gel electrophoresis followed by mass spectrometry revealed decreased abundance of a number of proteins involved in mitochondrial function or oxidative stress. Consistent with reductions in several subunits of complexes I and IV, functional assays showed reductions in respiratory capacity of striatal mitochondria isolated from parkin-/- mice. Electron microscopic analysis revealed no gross morphological abnormalities in striatal mitochondria of parkin-/- mice. In addition, parkin-/- mice showed a delayed rate of weight gain, suggesting broader metabolic abnormalities. Accompanying these deficits in mitochondrial function, parkin-/- mice also exhibited decreased levels of proteins involved in protection from oxidative stress. Consistent with these findings, parkin-/- mice showed decreased serum antioxidant capacity and increased protein and lipid peroxidation. The combination of proteomic, genetic, and physiological analyses reveal an essential role for parkin in the regulation of mitochondrial function and provide the first direct evidence of mitochondrial dysfunction and oxidative damage in the absence of nigral degeneration in a genetic mouse model of Parkinson's disease.     

Increased mitochondrial oxidative damage and oxidative DNA damage contributes to the neurodegenerative process in sporadic amyotrophic lateral sclerosis

To investigate the possibility that mitochondrial oxidative damage or oxidative DNA damage or both contribute to the neurodegenerative process of sporadic amyotrophic lateral sclerosis (sALS), this study used high-performance liquid chromatography with an electrochemical detector to measure the concentrations of the reduced and oxidized forms of coenzyme Q10 (CoQ10) and 8-hydroxy-2'-deoxyguanosine (8-OHdG) in the cerebrospinal fluid (CSF) of 17 patients with sALS and 17 age-matched controls with no neurological diseases. The percentage of oxidized CoQ10 in the CSF of sALS patients was greater than that in the CSF of controls (p<0.002) and was negatively correlated with the duration of illness (rho=-0.61, p<0.01). The concentration of 8-OHdG in the CSF of sALS patients was greater than that in the CSF of controls (p<0.005) and was positively correlated with the duration of illness (rho=0.53, p<0.005). The percentage of oxidized CoQ10 was correlated with the concentrations of 8-OHdG in the CSF of sALS patients (rho=-0.53, p<0.05). These results suggest that both mitochondrial oxidative damage and oxidative DNA damage play important roles in the pathogenesis of sporadic amyotrophic lateral sclerosis.     

Effect of abscisic acid on active oxygen species, antioxidative defence system and oxidative damage in leaves of maize seedlings.

Leaves of maize (Zea mays L.) seedlings were supplied with different concentrations of abscisic acid (ABA). Its effects on the levels of superoxide radical (O(2)(-)), hydrogen peroxide (H(2)O(2)) and the content of catalytic Fe, the activities of several antioxidative enzymes such as superoxide dismutase (SOD), catalase (CAT), ascorbate peroxidase (APX) and glutathione reductase (GR), the contents of several non-enzymatic antioxidants such as ascorbate (ASC), reduced glutathione (GSH), alpha-tocopherol (alpha-TOC) and carotenoid (CAR), and the degrees of the oxidative damage to the membrane lipids and proteins were examined. Treatment with 10 and 100 microM ABA significantly increased the levels of O(2)(-) and H(2)O(2), followed by an increase in activities of SOD, CAT, APX and GR, and the contents of ASC, GSH, alpha-TOC and CAR in a dose- and time-dependent pattern in leaves of maize seedlings. An oxidative damage expressed as lipid peroxidation, protein oxidation, and plasma membrane leakage did not occur except for a slight increase with 100 microM ABA treatment for 24 h. Treatment with 1,000 microM ABA led to a more abundant generation of O(2)(-) and H(2)O(2) and a significant increase in the content of catalytic Fe, which is critical for H(2)O(2)-dependent hydroxyl radical production. The activities of these antioxidative enzymes and the contents of alpha-TOC and CAR were still maintained at a higher level, but no longer further enhanced when compared with the treatment of 100 microM ABA. The contents of ASC and GSH had no changes in leaves treated with 1,000 microM ABA. These results indicate that treatment with low concentrations of ABA (10 to 100 microM) induced an antioxidative defence response against oxidative damage, but a high concentration of ABA (1,000 microM) induced an excessive generation of AOS and led to an oxidative damage in plant cells.     

Spermine and putrescine enhance oxidative stress tolerance in maize leaves

The protective effects of spermine (SPM) and putrescine (PUT) against paraquat (PQ), a herbicide in agriculture and oxidative stress inducer, were investigated in the leaves of maize. Maize leaves were pretreated to SPM and PUT at concentrations of 0.2 and 1 mM and treated with PQ afterwards. Pretreatment with 1 mM of SPM and PUT significantly prevented the losses in chlorophyll and carotenoid levels induced by PQ. Ascorbic acid content in the leaves pretreated with both polyamines was found to be higher than those of the leaves pretreated with water. Also, pretreatment with SPM and PUT was determined to have some effects on the activities of superoxide dismutase (SOD) and peroxidase (POD). 1 mM of SPM increased SOD activity, but PUT has no significant effect on SOD activity. On the other hand, POD activity was recorded to increase slightly in response to both concentrations of SPM and 1 mM of PUT. The results showed that such polyamine pretreated plants may become more tolerant to oxidative stress due to increases in the antioxidative enzymes and antioxidants.     

Resveratrol ameliorates oxidative DNA damage and protects against acrylamide-induced oxidative stress in rats

Acrylamide (ACR), used in many fields from industrial manufacturing to laboratory personnel work is also formed during the heating process through interactions of amino acids. Therefore ACR poses a significant risk to human health. This study aimed to elucidate whether resveratrol (RVT) treatment could modulate ACR-induced oxidative DNA damage and oxidative changes in rat brain, lung, liver, kidney and testes tissues. Rats were divided into four groups as control (C); RVT (30 mg/kg i.p. dissolved in 0.9% NaCl), ACR (40 mg/kg i.p.) and RVT + ACR groups. After 10 days rats were decapitated and tissues were excised. 8-hydroxydeoxyguanosine (8-OHdG) is a biomarker of oxidative DNA damage. 8-OHdG content in the extracted DNA solution was determined by enzyme-linked immunosorbent assay method. Malondialdehyde (MDA), glutathione (GSH) levels and myeloperoxidase activity (MPO) were determined in tissues, while oxidant-induced tissue fibrosis was determined by collagen contents. Serum enzyme activities, cytokine levels, leukocyte apoptosis were assayed in plasma. As an indicator of oxidative DNA damage, 8-OHdG levels significantly increased in ACR group and this was reversed significantly by RVT treatment. In ACR group, GSH levels decreased significantly while the MDA levels, MPO activity and collagen content increased in the tissues suggesting oxidative organ damage. In RVT-treated ACR group, oxidant responses reversed significantly. Serum enzyme activities, cytokine levels and leukocyte late apoptosis which increased following ACR administration, decreased with RVT treatment. Therefore supplementing with RVT can be useful in individuals at risk of ACR toxicity.     

Repair of oxidative DNA damage

Cellular genomes suffer extensive damage from exogenous agents and reactive oxygen species formed during normal metabolism. The MutT homologs (MutT/MTH) remove oxidized nucleotide precursors so that they cannot be incorporated into DNA during replication. Among many repair pathways, the base excision repair (BER) pathway is the most important cellular protection mechanism responding to oxidative DNA damage. The 8-oxoG glycosylases (Fpg or MutM/OGG) and the MutY homologs (MutY/MYH) glycosylases along with MutT/MTH protect cells from the mutagenic effects of 8-oxoG, the most stable and deleterious product known caused by oxidative damage to DNA. The key enzymes in the BER process are DNA glycosylases, which remove different damaged bases by cleavage of the N-glycosylic bonds between the bases and the deoxyribose moieties of the nucleotide residues. Biochemical and structural studies have demonstrated the substrate recognition and reaction mechanism of BER enzymes. Cocrystal structures of strated the substrate recognition and reaction mechanism of BER enzymes. Cocrystal structures of several glycosylases show that the substrate base flips out of the sharply bent DNA helix and the minor groove is widened to be accessed by the glycosylases. To complete the repair after glycosylase action, the apurinic/apyrimidinic (AP) site is further processed by an incision step, DNA synthesis, an excision step, and DNA ligation through two alternative pathways. The short-patch BER (1-nucleotide patch size) and long-patch BER (2–6-nucleotide patch size) pathways need AP endonuclease to generate a 3′ hydroxyl group but require different sets of enzymes for DNA synthesis and ligation. Protein-protein interactions have been reported among the enzymes involved in BER. It is possible that the successive players in the repair pathway are assembled in a complex to perform concerted actions. The BER pathways are proposed to protect cells and organisms from mutagenesis and carcinogenesis.     

Retinal degeneration from oxidative damage

Paraquat, a herbicide that generates reactive oxygen species, has been used to probe the oxidative defense systems of lung. In this study, we investigated the effects of paraquat in the retina. There was no significant decline in electroretinogram (ERG) a- or b-wave amplitudes after intravitreous injection of 1 mul of 0.5 mM paraquat in C57BL/6 mice, but loss of ERG function occurred after injection of 0.75 or 1 mM paraquat. Histology in paraquat-injected eyes showed condensation of chromatin and thinning of the inner and outer nuclear layers indicating cell death, and terminal deoxynucleotidyl transferase-mediated duTP-biotinide end labeling demonstrated that one mechanism of cell death was apoptosis. Fluorescence in the retina and retinal pigmented epithelium after intraocular injection of paraquat followed by perfusion with hydroethidine indicated high levels of superoxide radicals, and oxidative damage was demonstrated by staining for acrolein and enzyme-linked immunosorbent assay for carbonyl adducts. Paraquat-induced damage to the outer nuclear layer was greater in BALB/c mice than in C57BL/6 mice, suggesting strain differences in the oxidative defense system of photoreceptors and/or other modifier genes. Intravitreous injection of paraquat provides a new model of oxidative damage-induced retinal degeneration that is likely to be useful for testing new antioxidant treatments.     

Endogenous oxidative damage of mtDNA

Almost a decade ago, based on analytical measurements of the oxidative DNA adduct 8-oxo-deoxyguanosine (oxo8dG), it was reported that mitochondrial DNA suffers greater endogenous oxidative damage than nuclear DNA. The subsequent discovery that somatic deletions of mitochondrial DNA occur in humans, and that they do so to the greatest extent in metabolically active tissues, strengthened the hypothesis that mitochondrial DNA is particularly susceptible to endogenous oxidative attack. This hypothesis was (and is) appealing for a number of reasons. Nevertheless, solid direct support for the hypothesis is lacking. Since the initial measurements, attempts to repeat the observation of greater oxidation of mitochondrial DNA have resulted in a range of measurements that spans over four orders of magnitude. Moreover, this range includes values that are as low as published values for nuclear DNA. In the last 2 years or so, it has become apparent that the quantification of oxidative DNA adducts is prone to artifactual oxidation. We have reported that the analysis of small quantities of DNA may be particularly susceptible to such interference. Because yields of mitochondrial DNA are generally low, a systematic artifact associated with low quantities of DNA may have elevated the apparent level of adduct oxo8dG in mitochondrial DNA relative to nuclear DNA in some studies. Whatever the cause for the experimental variation, the huge disparity between published measurements of oxidative damage makes it impossible to conclude that mitochondrial DNA suffers greater oxidation than nuclear DNA. Despite the present confusion, however, there are reasons to hypothesize that this is indeed the case. We briefly describe methods being developed by a number of workers that are likely to surmount current obstacles and allow the hypothesis to be tested definitively.     

Coccidian infection causes oxidative damage in greenfinches

The main tenet of immunoecology is that individual variation in immune responsiveness is caused by the costs of immune responses to the hosts. Oxidative damage resulting from the excessive production of reactive oxygen species during immune response is hypothesized to form one of such costs. We tested this hypothesis in experimental coccidian infection model in greenfinches Carduelis chloris. Administration of isosporan coccidians to experimental birds did not affect indices of antioxidant protection (TAC and OXY), plasma triglyceride and carotenoid levels or body mass, indicating that pathological consequences of infection were generally mild. Infected birds had on average 8% higher levels of plasma malondialdehyde (MDA, a toxic end-product of lipid peroxidation) than un-infected birds. The birds that had highest MDA levels subsequent to experimental infection experienced the highest decrease in infection intensity. This observation is consistent with the idea that oxidative stress is a causative agent in the control of coccidiosis and supports the concept of oxidative costs of immune responses and parasite resistance. The finding that oxidative damage accompanies even the mild infection with a common parasite highlights the relevance of oxidative stress biology for the immunoecological research.     

Induction of oxidative DNA damage in anaerobes

We compared oxidative DNA damage in strictly anaerobic Prevotella melaninogenica, aerotolerant anaerobic Bacteroides fragilis, and facultative anaerobic Salmonella typhimurium after exposure to O2 or H2O2. Using HPLC with electrochemical detection, we measured 8-hydroxydeoxyguanosine (8OHdG) as a damage marker. O2 induced 8OHdG in P. melaninogenica but not in B. fragilis, which shows catalase activity, or in S. typhimurium. In P. melaninogenica, with catalase, O2 induced less 8OHdG; superoxide dismutase had no effect; with glucose and glucose oxidase, O2 induced more 8OHdG. H2O2 also markedly increased 8OHdG. O2 was suggested to induce 8OHdG through H2O2. O2 or H2O2 decreased survival only in P. melaninogenica. Highly sensitive to oxidative stress, P. melaninogenica could prove useful for investigating oxidative DNA damage.     

Predation risk causes oxidative damage in prey

While there is increasing interest in non-consumptive effects of predators on prey, physiological effects are understudied. While physiological stress responses play a crucial role in preparing escape responses, the increased metabolic rates and shunting of energy away from other body functions, including antioxidant defence, may generate costs in terms of increased oxidative stress. Here, we test whether predation risk increases oxidative damage in Enallagma cyathigerum damselfly larvae. Under predation risk, larvae showed higher lipid peroxidation, which was associated with lower levels of superoxide dismutase, a major antioxidant enzyme in insects, and higher superoxide anion concentrations, a potent reactive oxygen species. The mechanisms underlying oxidative damage are likely to be due to the shunting of energy away from antioxidant defence and to an increased metabolic rate, suggesting that the observed increased oxidative damage under predation risk may be widespread. Given the potentially severe fitness consequences of oxidative damage, this largely overlooked non-consumptive effect of predators may be contributing significantly to prey population dynamics.     

Sickle cell membranes and oxidative damage.

Sickle erythrocytes and their membranes are susceptible to endogenous free-radical-mediated oxidative damage which correlates with the proportion of irreversibly sickled cells. The suppression of incubation-induced oxidative stress by antioxidants, free radical scavengers and an iron chelator suggest that oxidation products of membrane-bound haemoglobin contribute towards the pathology of the disease.