Effect of the period of resting in elite judo athletes

The purpose of this study was to evaluate the effect of the resting period on hematological and copper-zinc-dependent antioxidant indices in Brazilian elite judo athletes (n=7). Venous blood samples were collected after 24-h and 5-d periods of resting following a competition, with an interval of 30 d between collections. Two months prior to and during the study, each athlete received an individualized adequate diet. Body composition was determined at both study periods. The following were analyzed: in whole blood, hemoglobin, hematocrit, red cell count, mean corpuscular volume, mean corpuscular hemoglobin, mean corpuscular hemoglobin concentration, red cell distribution width, and white cell count; in plasma, zinc, copper, iron, ceruloplasmin, and total iron-binding capacity; in erythrocytes, metallothionein, copper/zinc superoxide dismutase, and osmotic fragility. Dietary intake and body composition did not affect the biochemical measurements. A significant reduction in ceruloplasmin and superoxide dismutase activity was found after 5 d compared to 24 h of resting. A significant correlation between erythrocyte metallothionein and red cell distribution width was observed after 24 h of resting (r=−0.83, p=0.02) whereas positive correlations of metallothionein with hemoglobin, red cell count, and mean corpuscular hemoglobin concentration were observed after 5 d of resting (r≥0.76, p≤0.05). Our results suggest that a longer resting period favors homeostatic adjustments in the erythrocyte population and in the copper/zinc-dependent antioxidant system in elite judo athletes.     

Characterization of the superoxide dismutase SOD1 gene of Kluyveromyces marxianus L3 and improved production of SOD activity

Superoxide dismutase (SOD) activity is one major defense line against oxidative stress for all of the aerobic organisms, and industrial production of this enzyme is highly demanded. The Cu/Zn superoxide dismutase gene (KmSOD1) of Kluyveromyces marxianus L3 was cloned and characterized. The deduced KmSod1p protein shares 86% and 71% of identity with Kluyveromyces lactis and Saccharomyces cerevisiae Sod1p, respectively. The characteristic motifs and the amino acid residues involved in coordinating copper and zinc and in enzymatic function were conserved. To the aim of developing a microbial production of Cu/Zn superoxide dismutase, we engineered the K. marxianus L3 strain with the multicopy plasmid YG-KmSOD1 harboring the KmSOD1 gene. The production of KmSOD1p in K. marxianus L3 and K. marxianus L3 (pYG-KmSOD1) in response to different compositions of the culture medium was evaluated. The highest specific activity (472 U_SOD mg_prot ⁻¹) and the highest volumetric yield (8.8 × 10⁵ U_SOD l⁻¹) were obtained by the recombinant strain overexpressing KmSOD1 in the presence of Cu²⁺ and Zn²⁺ supplements to the culture media. The best performing culture conditions were positively applied to a laboratory scale fed-batch process reaching a volumetric yield of 1.4 × 10⁶ U_SOD l⁻¹. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

In yeast redistribution of Sod1 to the mitochondrial intermembrane space provides protection against respiration derived oxidative stress

The antioxidative enzyme copper-zinc superoxide dismutase (Sod1) is an important cellular defence system against reactive oxygen species (ROS). While the majority of this enzyme is localized to the cytosol, about 1% of the cellular Sod1 is present in the intermembrane space (IMS) of mitochondria. These amounts of mitochondrial Sod1 are increased for certain Sod1 mutants that are linked to the neurodegenerative disease amyotrophic lateral sclerosis (ALS). To date, only little is known about the physiological function of mitochondrial Sod1. Here, we use the model system Saccharomyces cerevisiae to generate cells in which Sod1 is exclusively localized to the IMS. We find that IMS-localized Sod1 can functionally substitute wild type Sod1 and that it even exceeds the protective capacity of wild type Sod1 under conditions of mitochondrial ROS stress. Moreover, we demonstrate that upon expression in yeast cells the common ALS-linked mutant Sod1^G93A becomes enriched in the mitochondrial fraction and provides an increased protection of cells from mitochondrial oxidative stress. Such an effect cannot be observed for the catalytically inactive mutant Sod1^G85R. Our observations suggest that the targeting of Sod1 to the mitochondrial IMS provides an increased protection against respiration-derived ROS.     

Acclimatization of micropropagated plantlets induces an antioxidative burst: a case study with <Emphasis Type="Italic">Ulmus minor</Emphasis> Mill.

In this article, the effects of increased light intensities on antioxidant metabolism during ex vitro establishment of Ulmus minor micropropagated plants are investigated. Three month old in vitro plants were acclimatized to ex vitro conditions in a climate chamber with two different light intensities, 200 μmol m⁻² s⁻¹ (high light, HL) and 100 μmol m⁻² s⁻¹ (low light, LL) during 40 days. Immediately after ex vitro transfer, the increase of both malondialdehyde (MDA) and electrolyte leakage in persistent leaves is indicative of oxidative stress. As the acclimatization continues, an upregulation of the superoxide dismutase (SOD), catalase (CAT), and glutathione reductase (GR) enzyme activities were also observed. Simultaneously, MDA content and membrane permeability stabilized, suggesting that the antioxidant enzymes decrease the deleterious effects of reactive oxygen species (ROS) generation. Unexpectedly, newly formed leaves presented a different pattern of antioxidative profile, with high levels of MDA and membrane leakage and low antioxidant enzyme activity. Despite these differences, both leaf types looked healthy (e.g. greenish, with no necrotic spots) during the whole acclimatization period. The results indicate that micropropagated U. minor plantlets develop an antioxidant enzyme system after ex vitro transfer and that, in general, LL treatment leads to lower oxidative stress. Moreover, new leaves tolerate higher levels of ROS without the need to activate the antioxidative pathway, which suggests that the environment at which leaves are exposed during its formation determinate their ability to tolerate ROS.     

Effect of oxidative stress on telomere maintenance in aortic smooth muscle cells

Reactive oxygen species (ROS) and telomere dysfunction are both associated with aging and the development of age-related diseases. Although there is evidence for a direct relationship between ROS and telomere dysfunction as well as an independent association of oxidative stress and telomere attrition with age-related disorders, there has not been sufficient exploration of how the interaction between oxidative stress and telomere function may contribute to the pathophysiology of cardiovascular diseases (CVD). To better understand the complex relationships between oxidative stress, telomerase biology and pathophysiology, we examined the telomere biology of aortic smooth muscle cells (ASMCs) isolated from mutant mouse models of oxidative stress. We discovered that telomere lengths were significantly shorter in ASMCs isolated from superoxide dismutase 2 heterozygous (Sod2^+/?) mice, which exhibit increased arterial stiffness with aging, and the observed telomere attrition occurred over time. Furthermore, the telomere erosion occurred even though telomerase activity increased. In contrast, telomeres remained stable in wild-type and superoxide dismutase 1 heterozygous (Sod1^+/?) mice, which do not exhibit CVD phenotypes. The data indicate that mitochondrial oxidative stress, in particular elevated superoxide levels and decreased hydrogen peroxide levels, induces telomere erosion in the ASMCs of the Sod2^+/? mice. This reduction in telomere length occurs despite an increase in telomerase activity and correlates with the onset of disease phenotype. Our results suggest that the oxidative stress caused by imbalance in mitochondrial ROS, from deficient SOD2 activity as a model for mitochondrial dysfunction results in telomere dysfunction, which may contribute to pathogenesis of CVD.     

Release of elicitors from rice blast spores under the action of reactive oxygen species

Effects of reactive oxygen species (ROS) on the release of putative elicitors from spores of rice blast causal fungus Magnaporthe grisea (Hebert) Barr were studied. While studying the influence of exogenous ROS, the spores were germinated for 5 h in the presence of 50 μM H₂O₂ and then treated with catalase to decompose hydrogen peroxide. The spore germination fluid was then boiled to inactivate catalase. When the resulting diffusate was applied onto rice (Oryza sativa L.) leaves, it caused necroses and stimulated superoxide (O₂⁻) production. Both effects were observed with the resistant rice cultivar but not with the cultivar susceptible to the fungal strain. The susceptible cultivar did not acquire resistance to challenge with fungal spores, which were applied one day after the treatment. The fractionation of the spore diffusate showed that both low- and high-molecular compounds (mol wt < 3 kD and >3 kD, respectively) should be present in combination to induce O₂⁻ production by leaves. The diffusates from spores germinated in water also caused necroses and stimulated O₂⁻ generation, though to a weaker extent than diffusates from spores germinated in H₂O₂. The effect of diffusates from spores germinated in water was abolished by catalase or superoxide dismutase added initially to the spore suspension. The results suggest that germinating spores of M. grisea are able to release elicitors and this ability depends on ROS formation by spores. Presumably, the yield of elicitors is increased additionally if fungus M. grisea is stressed or subjected to exogenous ROS. The described phenomena may be involved in incompatibility mechanisms.     

Light intensity determines the extent of mercury toxicity in the cyanobacterium Nostoc muscorum

The extent of mercury (Hg) toxicity in the heterocystous cyanobacterium Nostoc muscorum grown for 72 h in three different light intensities was tested for various physiological parameters viz. growth, pigment contents, photosynthesis, respiration, reactive oxygen species (ROS), malondialdehyde formation and antioxidants. A general reduction in growth and pigments, whole cell O₂-evolution, photosynthetic electron transport activities and ¹⁴CO₂-fixation was observed in a metal concentration–dependent manner, and this effect was more pronounced in high light (130 μmol photon m⁻² s⁻¹)–exposed cells as compared to low (10 μmol photon m⁻² s⁻¹) and normal (70 μmol photon m⁻² s⁻¹) light intensity–exposed cells; however, carotenoids and respiration showed reverse trend. Among photosynthetic electron transport activities, whole chain activity was found to be most sensitive in comparison with photosystem II (PS II) and photosystem I (PS I). Comparing the different photosynthetic processes, ¹⁴CO₂-fixation was most affected in cyanobacterial cells when exposed to Hg and different light intensities. After application of various exogenous electron donors, diphenyl carbazide was found to be more effective to restore PS II activity, suggesting that site of damage lies in between oxygen evolving complex and PS II. Level of oxidative stress (superoxide radical and lipid peroxidation) was maximum at 3.0 μM of Hg when coupled with high light intensity (except hydrogen peroxide). A dose-dependent increase in enzymatic antioxidants such as superoxide dismutase, peroxidase and catalase as well as non-enzymatic antioxidants such as proline, ascorbate, cysteine (except under high light intensity) and non-protein thiols [NP-SH] was observed, which further increased with the increase in light intensity. It was noticed that Hg intoxicates N. muscorum through ROS production, which is aggravated along with the increase in light intensity. Overall results suggest that the severity of the metal stress does increase with Hg concentrations but when coupled with light, it was the light intensity that determines the extent of Hg toxicity.     

Damaging mechanisms of chilling- and salt stress to <Emphasis Type="Italic">Arachis hypogaea</Emphasis> L. leaves

To investigate damaging mechanisms of chilling and salt stress to peanut (Arachis hypogaea L.) leaves, LuHua 14 was used in the present work upon exposure to chilling temperature (4°C) accompanied by high irradiance (1,200 μmol m⁻² s⁻¹) (CH), salt stress accompanied by high irradiance (1,200 μmol m⁻² s⁻¹) (SH), and high-irradiance stress (1,200 μmol m⁻² s⁻¹) at room temperature (25°C) (NH), respectively. Additionally, plants under low irradiance (100 μmol m⁻² s⁻¹) at room temperature (25°C) were used as control plants (CK). Relative to CK and NH treatments, both the maximal photochemical efficiency of PSII (F_v/F_m) and the absorbance at 820 nm decreased greatly in peanut leaves under CH and SH stress, which indicated that severe photoinhibition occurred in peanut leaves under such conditions. Initial fluorescence (F_o), 1 − q_P and nonphotochemical quenching (NPQ) in peanut leaves significantly increased under CH- and SH stress. Additionally, the activity of superoxide dismutase (SOD), one of the key enzymes of water-water cycle, decreased greatly, the accumulation of malondialdehyde (MDA) and membrane permeability increased. These results suggested that damages to peanut photosystems might be related to the accumulation of reactive oxygen species (ROS) induced by excess energy, and the water-water cycle could not dissipate energy efficiently under the stress of CH and SH, which caused the accumulation of ROS greatly. CH and SH had similar damaging effects on peanut photosystems, except that CH has more severe effects. All the results showed that CH- and SH stress has similar damaging site and mechanisms in peanut leaves.     

Extracellular Superoxide Dismutase Protects Histoplasma Yeast Cells from Host-Derived Oxidative Stress

In order to establish infections within the mammalian host, pathogens must protect themselves against toxic reactive oxygen species produced by phagocytes of the immune system. The fungal pathogen Histoplasma capsulatum infects both neutrophils and macrophages but the mechanisms enabling Histoplasma yeasts to survive in these phagocytes have not been fully elucidated. We show that Histoplasma yeasts produce a superoxide dismutase (Sod3) and direct it to the extracellular environment via N-terminal and C-terminal signals which promote its secretion and association with the yeast cell surface. This localization permits Sod3 to protect yeasts specifically from exogenous superoxide whereas amelioration of endogenous reactive oxygen depends on intracellular dismutases such as Sod1. While infection of resting macrophages by Histoplasma does not stimulate the phagocyte oxidative burst, interaction with polymorphonuclear leukocytes (PMNs) and cytokine-activated macrophages triggers production of reactive oxygen species (ROS). Histoplasma yeasts producing Sod3 survive co-incubation with these phagocytes but yeasts lacking Sod3 are rapidly eliminated through oxidative killing similar to the effect of phagocytes on Candida albicans yeasts. The protection provided by Sod3 against host-derived ROS extends in vivo. Without Sod3, Histoplasma yeasts are attenuated in their ability to establish respiratory infections and are rapidly cleared with the onset of adaptive immunity. The virulence of Sod3-deficient yeasts is restored in murine hosts unable to produce superoxide due to loss of the NADPH-oxidase function. These results demonstrate that phagocyte-produced ROS contributes to the immune response to Histoplasma and that Sod3 facilitates Histoplasma pathogenesis by detoxifying host-derived reactive oxygen thereby enabling Histoplasma survival.     

Adaptive changes in structure of skeletal muscles from adult <Emphasis Type="Italic">Sod1</Emphasis> homozygous knockout mice

Cu/Zn superoxide dismutase (SOD1), which is localized cytoplasmically and in the mitochondrial intermembrane space, is an enzyme that is critically important for superoxide free-radical elimination. Compared with age-matched wild-type littermates (Sod1 ^+/+ ), SOD1 homozygous knockout (Sod1 ^-/- ) mice have smaller body masses, heart and skeletal muscle masses, and muscle cross-sectional areas. At the light-microscopic level, cross sections of skeletal muscles from Sod1 ^-/- mice show no gross structural abnormalities. Following the staining of muscles of Sod1 ^-/- mice for succinate dehydrogenase (SDH) enzymatic activity, a grouping of SDH-positive fibers has been observed. Immunostaining for neural cell adhesion marker in the gastrocnemius muscle of Sod1 ^-/- mice has revealed a small number of atrophic denervated muscle fibers. No denervated fibers are observed in extensor digitorum longus (EDL), tibialis anterior, or plantaris muscles. An increase in mRNA expression levels of myogenin and acetylcholine receptor alpha has been detected in muscles in Sod1 ^-/- mice, but no changes in MyoD expression occur. Compared with fast oxidative fibers in EDL muscles of Sod1 ^+/+ mice, those of Sod1 ^-/- mice show increased accumulations of sub-sarcolemmal mitochondria. We conclude that the lack of SOD1 in adult Sod1 ^-/- mice does not result in extensive denervation of skeletal muscle fibers, although the distribution of fiber types is modified, and that fast oxidative fibers develop alterations in the amount and spatial distribution of sub-sarcolemmal mitochondria. This study was supported by NIA grant PO1-AG20591, by the Nathan Shock Center Contractility Core (NIA grant P30-AG13283), and by a Nathan Shock Center Pilot Award (to T. Kostrominova).     

Copper deficient rat heart can compensate for doxorubicin-induced oxidant stress

The hypothesis that copper (Cu) alters drug metabolizing enzymes and functions as an antioxidant nutrient in doxorubicin cardiotoxicity was tested. Male Sprague-Dawley rats were fed Cu adequate (⁺Cu; 5 mg Cu/kg of diet), marginally Cu deficient (MCu; 1.2 mg Cu/kg of diet), or severely Cu deficient (⁻Cu; 0.5 mg Cu/kg of diet) diets for 6 wk. Doxorubicin (1, 2, or 4 mg/kg body wt) or saline were administered intraperitoneally 1 time/wk for 4 wk. Compared to control hearts, Cu, Zn superoxide dismutase activity was decreased by 9% in MCu rats and by 21–40% in⁻Cu rats. Glutathione peroxidase activity was elevated 5–15% in⁻Cu rats. Doxorubicin administration increased heart Cu, Zn superoxide dismutase activity in⁺Cu and⁻Cu rats 18 h after the last of 4 injections, but not 18 h after 1 injection. There was no synergism between doxorubicin and Cu deficiency on lipid peroxidation, plasma creatine phosphokinase, cardiac hypertrophy, electrocardiographic abnormalities, or morphological changes. Heart glutathione S-transferase activity was decreased by Cu deficiency, and like Cu, Zn superoxide dismutase activity, returned to normal in⁻Cu rats given doxorubicin. Thus, the Cu deficient rat heart may be able to compensate for doxorubicin-induced oxidant stress by increasing the activity of Cu,Zn superoxide dismutase and glutathione S-transferase.     

Metabolism of reactive oxygen species in cotton cytoplasmic male sterility and its restoration

To elucidate reactive oxygen species (ROS) metabolism of cotton cytoplasmic male sterility and the effects of restorer gene on the metabolism of ROS, the metabolism changes in the production and scavenging of ROS and gene expression related to ROS-scavenging enzymes were investigated in the anther mitochondria of CMS line, maintainer line and hybrid F₁. During the abortion preliminary stage (sporogenous cell division stage), anthers of CMS line had a little higher superoxide (O₂ ⁻) production rate and hydrogen peroxide (H₂O₂) and malondialdehyde (MDA) contents than those of maintainer or hybrid F_1. Simultaneously, a little higher ROS contents might serve as a signal to increase the activity of superoxide dismutase (SOD) in anthers of CMS line to reduce the ROS damage to the anther development. But at the abortion peak (pollen mother cell meiosis stage), anthers of CMS line had extraordinarily higher ROS contents and lower ROS-scavenging enzymic activities compared with the hybrid F₁, during which the ROS contents and ROS-scavenging enzymic activities in hybrid F₁ were approximate to those of maintainer line. The expression of Mn-sod and apx mRNA in anther of CMS line was obviously inhibited when ROS produced with a great deal during anther abortion, however the gene expression in hybrid F₁ kept normal with the maintainer. Excessive accumulation of O₂^·−, H₂O₂ and MDA, significant reduction of ROS-scavenging enzymic activities and lower gene expression level of ROS-scavenging enzyme were coinstantaneous with male cells death in anthers of CMS line. But when the restorer gene was transferred into CMS line, excessive production of ROS could be eliminated in the anthers of hybrid F₁. The restorer gene likely plays an important role in keeping the dynamic balance between the production and elimination of ROS.     

Cadmium and zinc-mediated oxidative burst in tobacco BY-2 cell suspension cultures

Generation of reactive oxygen species (ROS) constitutes an important first reaction under many stress conditions in plants. We demonstrate that Nicotiana tabacum L. cv. Bright Yellow 2 (TBY-2) cells in suspension cultures, generate superoxide radical and hydrogen peroxide upon treatment with cadmium and zinc. Addition of catalase and N,N-diethyldithiocarbamate (DDC) decreased the level of H₂O₂, whereas superoxide dismutase (SOD) induced a slight increase of the H₂O₂ production. The effects of catalase, DDC and SOD on the heavy metal-induced ROS production indicate that it occurs outside of the cells, and that at least part of the hydrogen peroxide is produced by dismutation of the superoxide radical (O₂ ^·−). The effect of pretreatment of the cell cultures with commonly used mammalian NADPH oxidase inhibitors was also tested. Strong inhibitions of cadmium and zinc-mediated ROS production were obtained with the flavoprotein inhibitors—diphenylene iodonium (DPI) and quinacrine and with an inhibitor of b-type cytochromes—imidazol. Membrane permeable-N-ethyl maleimide (NEM) and iodoacetate, and membrane non-permeable thiol reagents—para-chloromercuribenzoic acid (pCMBS) also inhibited the ROS production. These results suggested that the enzyme responsible for cadmium and zinc-induced ROS production in tobacco cells contains a flavocytochrome. They also show the importance of intra- and extracellular thiol groups in the observed stress reaction. The induction of ROS production with heavy metals showed properties comparable to the elicitor-induced oxidative burst in other plant cells.     

Chill-induced inhibition of photosynthesis was alleviated by 24-epibrassinolide pretreatment in cucumber during chilling and subsequent recovery

To investigate whether brassinosteroids (BRs) could be used to alleviate chill-induced inhibition of photosynthesis in cucumber (Cucumis sativus L) during chilling and subsequent recovery, the effects of exogenously applied 24-epibrassinolide (EBR) on gas exchange, chlorophyll fluorescence parameters, and antioxidant enzyme activity were studied. Cucumber plants were exposed to chilling under low light (12/8°C and 100 μmol m⁻² s⁻¹ PPFD) for 3 days and then recovered under normal temperature and high irradiance (28/18°C and 600 μmol m⁻² s⁻¹ PPFD) for 6 days. Chilling significantly decreased the net photosynthetic rate (P _N) and stomatal conductance (g _s), and increased rate of O₂ ^·− formation and H₂O₂ and malondialdehyde (MDA) content in cucumber leaves, but did not influence the optimal quantum yield of PSII (F_v/F_m). Chilling also decreased the effective quantum yield of PSII photochemistry (Φ_PSII) and photochemical quenching (q_P), but induced an increase in nonphotochemical quenching (NPQ), and the activities of superoxide dismutase (SOD) and ascorbate peroxidase (APX). High irradiance (600 μmol m⁻² s⁻¹) further aggravated the decrease in P _N, g _s, Φ_PSII and q_P, and enhanced the increase in reactive oxygen species (ROS) generation and accumulation in the first day of recovery after chilling. However, high irradiance induced a sharp decrease in F_v/F_m and NPQ, as well as the activities of SOD and APX on the first day of recovery. EBR pretreatment significantly alleviated chill-induced inhibition of photosynthesis during chilling stress and subsequent recovery period, which was mainly due to significant increases in g _s, Φ_PSII, q_P and NPQ. EBR pretreatment also reduced ROS generation and accumulation, and increased the activities of SOD and APX during chilling and subsequent recovery. Those results suggest that EBR pretreatment alleviates the chill reduction in photosynthesis and accelerated the recovery rate mainly by increasing of the stomatal conductance, the efficiency of utilization and dissipation of leaf absorbed light, and the activity of the ROS scavenging system during chilling and subsequent recovery period.     

Parameters of Metabolic Syndrome and Its Relationship with Zincemia and Activities of Superoxide Dismutase and Glutathione Peroxidase in Obese Women

Alterations in antioxidant defense in obese people with metabolic syndrome can contribute to oxidative stress. This study assessed the relationship between the parameters of metabolic syndrome and the zincemia, activity of superoxide dismutase, and glutathione peroxidase enzymes in obese women. Seventy-three premenopausal women, aged between 20 and 50 years, were divided into two groups: case group, composed of obese (n = 37), and control group, composed of no obese (n = 36). Analyses of zinc intake, parameters of metabolic syndrome, plasma, and erythrocyte zinc, and activities of superoxide dismutase and glutathione peroxidase were carried out. The mean values of body mass index of obese women and control group were 34.5 ± 3.4 and 21.7 ± 1.9 kg/m², respectively (p < 0.05). In the study, body mass index, waist circumference, and zinc intake were higher in obese women than control group (p < 0.05). The plasma zinc and activity of superoxide dismutase did not show significant differences between obese and controls (p > 0.05). The values of erythrocyte zinc was 36.4 ± 15.0 μg/gHb and 45.4 ± 14.3 μg/gHb and of glutathione peroxidase was 46.4 ± 19.4 U/gHb and 36.7 ± 13.6 U/gHb in obese women and controls, respectively (p < 0.05). The study shows that there are alterations in biochemical parameters of zinc in obese women, with low zinc concentrations in erythrocytes. Regression analysis demonstrates that the erythrocyte zinc and activity of superoxide dismutase enzyme is influenced by components of the metabolic syndrome, and the plasmatic glucose, body mass index, and waist circumference have a negative correlation with this enzyme.     

Partial attenuation of cytotoxicity and apoptosis by SOD1 in ischemic renal epithelial cells

Reactive oxygen species (ROS) contribute significantly to apoptosis in renal ischemia-reperfusion (IR) injury, however the exact mechanisms are not well understood. We used novel lentiviral vectors to over-express superoxide dismutase 1 (SOD1) in proximal tubular epithelial (LLC-PK₁) cells and determined effects of SOD1 following ATP depletion-recovery, used as a model to simulate renal IR. SOD1 over-expression partially protected against cytotoxicity (P < 0.001) and decreased superoxide (O₂ ^•−) in ATP depleted cells. The ATP depletion-mediated increase in nuclear fragmentation, an index of apoptosis and activation of caspase-3 was also partially blocked by SOD1 (P < 0.05). However, SOD1 over-expression was insufficient to completely attenuate caspase-3, indicating that ROS other than cytoplasmic O₂ ^•− are involved in ATP depletion mediated injury. To test the contribution of hydrogen peroxide, a subset of enhanced green fluorescent protein (EGFP) and SOD1 (serum free and injured) cells were treated with polyethylene glycol-catalase (PEG-catalase). As expected there was 50% reduction in cytotoxicity and caspase-3 in SOD1 cells compared to EGFP cells; catalase treatment decreased both indices by an additional 28% following ATP depletion. To test the role of mitochondrial derived superoxide, we also treated a subset of LLC-PK₁ cells with the mitochondrial antioxidant, MitoTEMPO. Treatment with MitoTEMPO also decreased ATP depletion induced cytotoxicity in LLC-PK₁ cells in a dose dependant manner. These studies indicate that both SOD1 dependent and independent pathways are integral in protection against ATP depletion-recovery mediated cytotoxicity and apoptosis, however more studies are needed to delineate the signaling mechanisms involved.     

Cadmium modulates NADPH oxidase activity and expression in sunflower leaves

The production of reactive oxygen species (ROS) and the ways by which ROS are generated are very important facts related to heavy metal toxicity in plants. In this work, superoxide anion (O₂ ^·−) generation diminished in cadmium treated sunflower (Helianthus annuus L.) leaf discs, and this reduction was time and Cd-concentration dependent. In line with these findings, we observed that NADPH-dependent oxidase activity was significantly inhibited by 0.1 and 0.5 mM Cd²⁺ treatments and the expression of the NADPH oxidase putative gene related to O₂ ^·− synthesis in sunflower leaves was 83 % inhibited by 0.1 mM CdCl₂ and almost completely depleted by 0.5 mM CdCl₂.     

Response of Chara globularis and Hydrodictyon reticulatum to lead pollution: their survival, bioaccumulation, and defense

The responses of the pioneer submerged macroalga (Chara globularis) and the rapidly spreading floating macroalga (Hydrodictyon reticulatum) to high levels of lead (40, 80, and 160 mg L⁻¹) at pH 7.14 were studied. Growth rate, Pb bioaccumulation, and physiological response of plants were measured after 5 and 15 days exposure. Both macroalgae efficiently postponed the deposition process of Pb from water column to soil. The Pb bioaccumulation in C. globularis was concentration- and time-dependent increase during the experiment and the maximum bioaccumulation activity was about 3,650 mg Pb kg⁻¹ DW in 160 mg L⁻¹ Pb at pH 7.14 after 15 days, whereas H. reticulatum showed saturable bioaccumulation in 5 days and the maximum was approximately 4,000 mg Pb kg⁻¹ DW; in addition, H. reticulatum exhibited higher tolerance to Pb pollution than C. globularis. The results also showed that the antioxidant defense systems of both tested macroalgae were overwhelmed under high Pb levels with superoxide radical and malondiadehyde levels increasing significantly. The antioxidant enzymes, superoxide dismutase, catalase, and guaiacol peroxidase activities were inhibited severely increasing Pb levels and exposure time. These results indicate that the pioneer species C. globularis would have difficulty growing in a habitat polluted by Pb >40 mg L⁻¹and the rapidly spreading H. reticulatum may not grow in an environment polluted by >80 mg L⁻¹ Pb. Because Pb levels in most water bodies are lower than 40 mg L⁻¹, both C. globularis and H. reticulatum can be considered for phytoremediation of Pb pollution.     

Salicylic Acid-induced Nitric Oxide and ROS Generation Stimulate Ginsenoside Accumulation in Panax ginseng Roots

We evaluated the involvement of nitric oxide (NO) in salicylic acid (SA)-induced accumulation of ginsenoside in adventitious roots of Panax ginseng and its mediation by reactive oxygen species (ROS). Related effects of SA on components of the antioxidant system were also sought. Adventitious roots of P. ginseng were grown in suspension culture for 3 weeks in MS medium and treated over 5 days with SA (100 μM) alone, SA in combination with the NO scavenger 2-phenyl-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide (PTIO), or PTIO alone. Nitric oxide, the superoxide anion (O₂^·−), H₂O₂, nitrite, nonprotein thiol, and ascorbate were monitored together with ginsenoside, NADPH oxidase activity, and several antioxidant enzymes. Salicylic acid did not inhibit root growth but induced accumulation of ginsenoside, lipid peroxidation, and generation of NO and O₂^·−. It also enhanced activities of NADPH oxidase, superoxide dismutase, catalase, and peroxidase, including ascorbate peroxidase. These effects were suppressed by PTIO. Salicylic acid also decreased glutathione reductase activity. Inclusion of PTIO with SA decreased the activity of glutathione reductase further. Treatment with SA plus PTIO also decreased nonprotein thiol and ascorbate contents but caused nitrite to overaccumulate. Salicylic acid applied to adventitious roots in culture induced accumulation of ginsenoside in an NO-dependent manner that was mediated by the associated increases in O₂^·−, which gave other antioxidant responses that were dependent on NO.