Crop Enhancement of Cucumber Plants under Heat Stress by Shungite Carbon.

Heat stress negatively impacts plant growth and yield. The effects of carbon materials on plants in response to abiotic stress and antioxidant activity are poorly understood. In this study, we propose a new method for improving heat tolerance in cucumber (Cucumis sativus L.) using a natural carbon material, shungite, which can be easily mixed into any soil. We analyzed the phenotype and physiological changes in cucumber plants maintained at 35 °C or 40 °C for 1 week. Our results show that shungite-treated cucumber plants had a healthier phenotype, exhibiting dark green leaves, compared to the plants in the control soil group. Furthermore, in the shungite-treated plants, the monodehydroascorbate content (a marker of oxidative damage) of the leaf was 34% lower than that in the control group. In addition, scavengers against reactive oxygen species, such as superoxide dismutase, catalase, and peroxidase were significantly upregulated. These results indicate that the successive pre-treatment of soil with a low-cost natural carbon material can improve the tolerance of cucumber plants to heat stress, as well as improve the corresponding antioxidant activity.

12-Oxophytodienoic Acid Reductase 3 (OPR3) Functions as NADPH-Dependent α,ß-Ketoalkene Reductase in Detoxification and Monodehydroascorbate Reductase in Redox Homeostasis.

Arabidopsis (Arabidopsis thaliana) 12-oxophytodienoic acid reductase isoform 3 (OPR3) is involved in the synthesis of jasmonic acid (JA) by reducing the α,ß-unsaturated double bond of the cyclopentenone moiety in 12-oxophytodienoic acid (12-OPDA). Recent research revealed that JA synthesis is not strictly dependent on the peroxisomal OPR3. The ability of OPR3 to reduce trinitrotoluene suggests that the old yellow enzyme homolog OPR3 has additional functions. Here, we show that OPR3 catalyzes the reduction of a wide spectrum of electrophilic species that share a reactivity toward the major redox buffers glutathione (GSH) and ascorbate (ASC). Furthermore, we show that 12-OPDA reacts with ASC to form an ASC-12-OPDA adduct, but in addition OPR3 has the ability to regenerate ASC from monodehydroascorbate. The presented data characterize OPR3 as a bifunctional enzyme with NADPH-dependent α,ß-ketoalkene double-bond reductase and monodehydroascorbate reductase activities (MDHAR). opr3 mutants showed a slightly less-reduced ASC pool in leaves in line with the MDHAR activity of OPR3 in vitro. These functions link redox homeostasis as mediated by ASC and GSH with OPR3 activity and metabolism of reactive electrophilic species.

Is monodehydroascorbate reductase activity in leaf tissue critical for the maintenance of yield in tomato?

Ascorbate redox metabolism and growth have been shown to be linked and related to the activity of enzymes that produce or remove the radical monodehydroascorbate, the semi-oxidized form of ascorbate (ascorbate oxidase or peroxidase and monodehydroascorbate reductase respectively). Previous work in cherry tomato has revealed correlations between monodehydroascorbate reductase and ascorbate oxidase activity and fruit yield: decreased whole plant MDHAR activity decreases yield while decreased whole plant ascorbate oxidase activity increases yield under unfavourable environmental conditions. We aimed to investigate if similar effects on yield are obtained in a large-fruited variety of tomato, Moneymaker. Furthermore we wished to establish whether previously observed effects on yield in cherry tomato following changes in whole plant enzyme activity could be reproduced by reducing MDHAR activity in fruit only by using a fruit-specific promoter in cherry tomato (West Virginia 106). In Moneymaker, RNAi lines for monodehydroascorbate reductase did not show significant yield decrease compared to control lines when plants were grown under optimal or non-optimal conditions of carbon stress generated by mature leaf removal. In addition, we show that a decrease in monodehydroascorbate reductase activity in fruit of cherry tomato had no effect on yield compared to a reduction in whole-plant monodehydroascorbate reductase activity: we therefore show that whole plant MDHAR activity is necessary to maintain yield in cherry tomato, suggesting that the carbon source in autotrophic tissue is more important than fruit sink activity. The present data also revealed differences between cherry and large fruited tomato that could be linked to a source of genetic variability in the response to monodehydroascorbate metabolism in tomato: maybe the domestication of tomato towards large-fruited lines could have affected the importance of MDHAR in yield maintenance.

Nutraceutical, Anti-Inflammatory, and Immune Modulatory Effects of ß-Glucan Isolated from Yeast.

ß-Glucan is a dietary fibre, found in many natural sources, and controls chronic metabolic diseases effectively. However, ß-glucan from the yeast has rarely been investigated. Objectively, conditions were optimized to isolate ß-glucan from the yeast (max. 66% yield); those optimized conditions included 1.0 M NaOH, pH 7.0, and 90°C. The purity and identity of the isolated ß-glucan were characterized through FT-IR, SEM, DSC, and physicofunctional properties. The obtained results from DSC revealed highly stable ß-glucan (m.p., 125°C) with antioxidant activity (TAC value 0.240 ± 0.0021 µg/mg, H2O2 scavenging 38%), which has promising bile acid binding 40.463% and glucose control (in vitro). In line with these results, we evaluated the in vivo anti-inflammatory potential, that is, myeloperoxidase activity and reduction in MDA and NO; protective effect on proteins and keeping viscosity within normal range exhibited improvement. Also, the in vivo cholesterol binding and reduction in the skin thickness by ß-glucan were highly encouraging. Finally, our results confirmed that yeast ß-glucan is effective against some of the inflammatory and oxidative stress markers studied in this investigation. In general, the effect of 4% ß-glucan was more noticeable versus 2% ß-glucan. Therefore, our results support the utilization of ß-glucan as a novel, economically cheap, and functional food ingredient.

Seasonality and toxins effects on oxidative/nitrosative metabolism in digestive glands of the bivalve Mytilus edulis platensis.

The hypothesis presented here is that oxidative and/or nitrosative metabolism in the bivalve Mytilus edulis platensis is altered by the presence of planktonic toxins. Digestive glands (DG) were isolated from specimens collected in the Argentinean Sea during summer, winter and spring (in the presence of harmful planktonic toxins). The labile iron pool content was not significantly different in DG from animals collected in summer and winter, but was 2.3-fold increased in samples from spring compared to summer collected mollusks. The 2',7' dichlorofluorescein diacetate (DCFH-DA) oxidation, ascorbyl radical/ascorbate and lipid radical/α-tocopherol content ratios showed no significant differences between samples collected in winter and summer. However, spring collected samples showed significantly higher DCFH-DA oxidation rate and oxidative ratios in comparison to DG from mollusks collected in summer. Superoxide dismutase activity decreased by 75% in winter, and 93% in spring, compared to samples collected in summer. Glutathione S-transferase activity decreased by 89% in winter, and 30% in spring, compared to samples collected in summer. Catalase activity in winter animals increased by 3.8-fold in comparison to summer values, with no differences between spring and summer collected mollusks. Nitrite plus nitrate content was not significantly different among samples collected in the three seasons, but nitric oxide content was 8.5- and 2.7-fold higher in samples from winter and spring collected mollusks than values obtained in summer, respectively. These results showed the lack of effects of climatic changes on the integrative oxidative indexes; however, under exposure to toxins, both oxidative and nitrosative metabolisms were affected.

Reduction of MDHAR activity in cherry tomato suppresses growth and yield and MDHAR activity is correlated with sugar levels under high light.

Ascorbate is oxidized into the radical monodehydroascorbate (MDHA) through ascorbate oxidase or peroxidase activity or non-enzymatically by reactive oxygen species. Regeneration of ascorbate from MDHA is ensured by the enzyme MDHA reductase (MDHAR). Previous work has shown that growth processes and yield can be altered by modifying the activity of enzymes that recycle ascorbate; therefore, we have studied similar processes in cherry tomato (Solanum lycopersium L.) under- or overexpressing MDHAR. Physiological and metabolic characterization of these lines was carried out under different light conditions or by manipulating the source-sink ratio. Independently of the light regime, slower early growth of all organs was observed in MDHAR silenced lines, decreasing final fruit yield. Photosynthesis was altered as was the accumulation of hexoses and sucrose in a light-dependent manner in plantlets. Sucrose accumulation was also repressed in young fruits and final yield of MDHAR silenced lines showed a stronger decrease under carbon limitation, and the phenotype was partially restored by reducing fruit load. Ascorbate and MDHA appear to be involved in control of growth and sugar metabolism in cherry tomato and the associated enzymes could be potential targets for yield improvement.

The Beneficial Effect of Cape Gooseberry Juice on Carbon Tetrachloride- Induced Neuronal Damage.

OBJECTIVE: Cape gooseberry (Physalis peruviana L.) belongs to the Solanaceae family. Physalis has many medicinal properties however, the beneficial effect of physalis in protecting against neurotoxins has not yet been evaluated. This experimental study investigated the protective effect of physalis juice against the oxidative damage induced by carbon tetrachloride (CCl4) in the rat brain. METHODS: The degrees of protection by physalis in brain tissues were evaluated by determining the brain levels of lipid peroxidation, nitric oxide, glutathione content and antioxidant enzyme activities (superoxide dismutase, catalase, glutathione-S-transferase, glutathione peroxidase and glutathione reductase), after CCl4) induction in the presence or absence of physalis. Adult male albino Wistar rats were divided into 4 groups, Group I served as the control group, Group II was intraperitoneally treated with 2 ml CCl4)/kg bwt for 12 weeks, Group III was supplemented with physalis juice via the drinking water for 12 weeks, Group IV was supplemented with physalis juice and was intraperitoneally injected weekly with CCl4). RESULTS: Treatment with CCl4) was significantly associated with a disturbance in the oxidative status in the brain tissues; this was marked by a significant (p<0.05) elevation in the lipid peroxidation and nitric oxide levels with a concomitant reduction in glutathione content compared to the control, along with a remarkable reduction in antioxidant enzymes. The administration of physalis along with CCl4) juice significantly (p<0.05) alleviated the changes in enzymatic antioxidant activity when compared to the CCl4) treated group. Furthermore, physalis juice supplemention inhibited apoptosis, as indicated by the increase of Bcl-2 immunoreactivity in brain tissue. CONCLUSION: Our results suggest that physalis juice could be effective in preventing neurotoxicity and the neuroprotective effect of physalis might be mediated via antioxidant and anti-apoptosis activities.

[Resveratrol alleviates oxidative injury from high glucose-induced human lens epithelial cells and its possible mechanism].

OBJECTIVE: To investigate the effects and its mechanism of resveratrol against human lens epithelial cells (LEC) apoptosis mediated by high glucose-induced oxidative injury. METHODS: An experimental study. LEC were cultured in different concentrations (5.5, 15.0, 25.0, 35.0, 45.0 mmol/L) of glucose medium or 25.0 mmol/L glucose medium at different time (0, 6, 12, 24, 48, 72 h), and established an interventional models of (5.0, 15.0, 25.0, 35.0 mg/L) resveratrol. Av-FITC-PI (annexin V-fluorescein isothiocyanate-propidium iodium) was used to detect apoptosis. The amount of ROS was calculated by flow cytometry. The expression of apoptosis of the protein Bcl-2 and Bax, iNOS, NF-κB, IκB and MnSOD were showed by Western blotting and the amount of oxidative damage marker MDA was explored by Spectrometers and Analytical Photometers. Differences between the two groups were evaluated by One-way ANOVA. RESULTS: The apoptosis of LEC induced by high glucose was time-and dose-dependent obviously. As the glucose concentration increased and duration prolonged, the expression of anti-apoptotic protein Bcl-2 was decreased and pro-apoptotic protein Bax was increased.Intracellular ROS and MDA induced by high glucose were increased significantly with dose-and time-dependence. Compared with 5.5 mmol/L group, ROS generation increased significantly in the concentration of 15.0 mmol/L (F = 14.06, P = 0.035), 25.0 mmol/L (F = 17.46, P = 0.000), 35.0 mmol/L (F = 16.58, P = 0.001), 45.0 mmol/L (F = 12.88, P = 0.000) and were statistically significant. Compared with 5.5 mmol/L glucose cultured group, ROS generation increased significantly at 6 h (F = 6.778, P = 0.014), 12 h (F = 6.551, P = 0.001), 24 h (F = 7.327, P = 0.001), 48 h (F = 10.84, P = 0.000), 72 h (F = 13.36, P = 0.000) in LEC cultured group by 25.0 mmol/L glucose and were statistically significant. Compared with 5.5 mmol/L group, the content of MDA were significantly increased in 15.0 mmol/L (F = 1.177, P = 0.035), 25.0 mmol/L (F = 1.704, P = 0.000), 35.0 mmol/L (F = 2.412, P = 0.001) and 45.0 mmol/L (F = 2.347, P = 0.000) glucose medium and were statistically significant. Compared with 5.5 mmol/L cultured group, the content of MDA were significantly increased at 6 h (F = 1.704, P = 0.014), 12 h (F = 5.676, P = 0.001), 24 h (F = 3.325, P = 0.001), 48 h (F = 6.669, P = 0.000), 72 h (F = 3.011, P = 0.000) in LEC cultured group by 25.0 mmol/L high glucose and were statistically significant. When resveratrol (5.0, 15.0, 25.0, 35.0 mg/L) was added to 25.0 mmol/L glucose medium, respectively, the apoptotic cells were decreased, the expression of pro-apoptotic protein Bax was decreased and anti-apoptotic protein Bcl-2 was increased.Intracellular ROS (compared with the basic concentration of 5.5 mmol/L, F values were 14.76, 7.018, 13.96, 4.733, 1.921, P values were 0.000, 0.000, 0.003, 0.086, 0.100 respectively) and MDA (compared with the basic concentration of 5.5 mmol/L, F values were 2.454, 1.108, 1.630, 1.563, 2.250, P values were 0.000, 0.001, 0.026, 0.068, 0.183 respectively) were decreased. MnSOD expression was increased, iNOS and NF-κB activation were inhibited. CONCLUSION: Resveratrol could alleviates oxidative injury from high glucose-induced LEC, and inhibited of iNOS-mediated oxidative damage through inhibiting the activities of NF-κB could be the mechanism of this effect.

Mechanisms of ascorbyl radical formation in human platelet-rich plasma.

Recently, many clinical reports have suggested that the ascorbyl free radical (Asc(∙)) can be treated as a noninvasive, reliable, real-time marker of oxidative stress, but its generation mechanisms in human blood have rarely been discussed. In this study, we used upstream substances, enzyme inhibitors, and free radical scavengers to delineate the mechanisms of Asc(∙) formation in human platelet-rich plasma (PRP). Our results show that the doublet signal was detected in PRP samples by using electron spin resonance, and the hyperfine splitting of the doublet signal was a(H) = 1.88 gauss and g-factor = 2.00627, which was determined to be the Asc(∙). We observed that the inhibitors of NADPH oxidase (NOX), cyclooxygenase (COX), lipoxygenase (LOX), cytochrome P450 (CYP450), mitochondria complex III, and nitric oxide synthase (NOS), but not xanthine oxidase, diminished the intensity of the Asc(∙) signal dose dependently. All enzyme inhibitors showed no obvious antioxidant activity during a Fenton reaction assay. In summary, the obtained data suggest that Asc(∙) formation is associated with NOX, COX, LOX, CYP450, eNOS, and mitochondria in human PRP.

Massive and long-lasting decrease in vitamin C plasma levels as a consequence of extracorporeal circulation.

OBJECTIVE: The use of cardiopulmonary bypass (CPB) is suggested to induce oxidative stress, reflected by an imbalance between prooxidant and antioxidant substances. The majority of studies published have either focused on only one aspect (prooxidant or antioxidant side) or covered only a short observation period. Therefore, the aim of this study was to investigate the long-term effects of CPB on the balance of prooxidative markers and antioxidant substances in one single group of patients, being able to estimate the degree of oxidative stress. METHODS: Blood samples were taken from 29 patients undergoing cardiovascular surgery beginning the day before surgery through postoperative day 6 (discharge). Plasma concentrations of vitamins C (total ascorbic acid) and E and malondialdehyde were measured by high-performance liquid chromatography. Plasma levels of ascorbyl free radical were determined using electron paramagnetic resonance spectroscopy. RESULTS: The study showed a significant decrease in vitamin C plasma levels during CPB without any recovery of vitamin C up to the time of discharge. Furthermore, CPB induced a significant increase in malondialdehyde plasma concentrations immediately after unclamping, accompanied by a significant increase in the ascorbyl free radical to total ascorbic acid ratio. The latter stayed elevated until the end of observation. CONCLUSIONS: Our findings indicate that the oxidative stress event after CPB can be divided into two phases: Immediately after reperfusion, a massive oxidative stress occurs, reflected by the increase in malondialdehyde. During convalescence, there must be an ongoing situation of oxidative stress, especially in the water-soluble compartment, leading to the consumption of vitamin C. Because the main antioxidant substance, vitamin C, did not increase again over the entire observation period, supplementation should be given consideration.

TaMDHAR4, a monodehydroascorbate reductase gene participates in the interactions between wheat and Puccinia striiformis f. sp. tritici.

Reactive oxygen species (ROS) in plants are induced in various cellular compartments upon pathogen infection and act as an early signal during plant-pathogen interactions. Monodehydroascorbate reductase (MDHAR) is involved in plant disease resistance through the regulation of the ROS level via the ascorbate-glutathione (AsA-GSH) cycle. In this study, TaMDHAR4 was firstly isolated from wheat cultivar Suwon 11, and this protein exhibits high similarity to MDHAR proteins from other plant species. Bioinformatics analyses indicated that TaMDHAR4 contains typical structural features, such as mPTS-like sequences in the C-terminal extension and trans-membrane domain followed by five basic arginine residues (-RKRRR), which predicted that this protein may be localized in the peroxisome. qRT-PCR analyses demonstrated that TaMDHAR4 could be induced by various exogenous hormones, such as ABA, MeJA, and ETH. TaMDHAR4 is sharply down-regulated at 12 and 18 hpi only in wheat leaves challenged with Puccinia striiformis f. sp. tritici (Pst) race CYR23 and induced at 48 hpi with both Pst races CYR23 and CYR31. SOD and APX injection analyses demonstrated that TaMDHAR4 may be involved in the interaction between wheat and Pst through the regulation of its expression. Moreover, the knockdown of TaMDHAR4 through virus-induced gene silencing (VIGS) enhanced the wheat resistance to Pst by inhibiting sporulation in the compatible interaction. Histological observations also demonstrated that silenced wheat resulted in an increased proportion of necrotic area at the infection sites and suppressed Pst hypha elongation. The study provided novel insights into the molecular functions of TaMDHAR4 during plant-pathogen interactions.

New evaluation of fetal oxidative stress: measurement of the umbilical cord blood dimethyl sulfate-induced ascorbyl-free radical by an electron spin resonance method.

OBJECTIVE: The aim is to evaluate intrapartum fetal oxidative stress in real-time by umbilical cord blood dimethyl sulfate (DMSO)-induced ascorbyl-free radical (AFR) measured by an electron spin resonance (ESR) method. METHODS: Seventy-five mothers delivering at gestational age after 37 weeks were recruited. They were divided into three groups: spontaneous vaginal birth (n = 27), elective cesarean section (CS) (n = 34), and emergency CS due to non-reassuring fetal status (n = 14). Umbilical artery (UA) and venous (UV) cord blood gas analysis was performed. Serum levels of DMSO-induced AFR (AFR/DMSO) that reflect vitamin C concentrations were measured by ESR spectroscopy. RESULTS: Blood gas analysis showed no significant differences among the groups. UA-AFR/DMSO level of elective CS group was significantly lower compared with spontaneous delivery group (0.32 ± 0.12 versus 0.46 ± 0.14, p < 0.005). Emergency CS group showed significantly lower levels of UA-AFR/DMSO compared with elective CS group (0.25 ± 0.11 versus 0.32 ± 0.12, p < 0.005). UV-AFR/DMSO levels had no significant difference among the groups. CONCLUSIONS: It is suggested that fetal cord blood AFR/DMSO is a sensitive marker to assess fetal oxidative stress during delivery.

Altered apoplastic ascorbate redox state in tobacco plants via ascorbate oxidase overexpression results in delayed dark-induced senescence in detached leaves.

Ascorbate oxidase (AO) is an apoplastic enzyme that uses oxygen to catalyse the oxidation of ascorbate (AA) to dehydroascorbate (DHA) via the unstable radical monodehydroascorbate (MDHA). Here, we report that transgenic tobacco plants (Nicotiana tabacum L. cv. Xanthi) with an in vivo lowered apoplastic AA redox state through increased AO expression demonstrate signs of delayed dark-induced senescence compared with wild-type plants, as shown by chlorophyll loss assay. In situ localization of hydrogen peroxide (H2O2) suggests that, although transgenic plants have higher constitutive levels of H2O2 under normal growth conditions, imposed dark-induced senescence results in smaller induction levels of H2O2, an observation which correlates with increased antioxidant enzyme activities and an induction in the expression of AA recycling genes compared with that in wild-type plants. Our current findings, combined with previous studies which showed the contribution of AO in the regulation of AA redox state, suggest that the reduction in AA redox state in the leaf apoplast of these transgenic plants results in an increase in the endogenous levels of H2O2, which provides a form of 'acquired tolerance' to oxidative stress imposed by dark-induced senescence.

Antioxidant/oxidant status and cardiac function in bradykinin B(1)- and B(2)-receptor null mice.

Kinin-vasoactive peptides activate two G-protein-coupled receptors (R), B(1)R (inducible) and B(2)R (constitutive). Their complex role in cardiovascular diseases could be related to differential actions on oxidative stress. This study investigated impacts of B(1)R or B(2)R gene deletion in mice on the cardiac function and plasma antioxidant and oxidant status. Echocardiography-Doppler was performed in B(1)R (B(1)R(-/-)) and B(2)R (B(2)R(-/-)) deficient and wild type (WT) adult male mice. No functional alteration was observed in B(2)R(-/-) hearts. B(1)R(-/-) mice had significantly lowered fractional shortening and increased isovolumetric contraction time. The diastolic E and A waves velocity ratio was similar in all mice groups. Thus B(1)R(-/-) mice provide a model of moderate systolic dysfunction, whereas B(2)R(-/-) mice displayed a normal cardiac phenotype. Plasma antioxidant capacity (ORAC) was significantly decreased in both B(1)R(-/-) and B(2)R(-/-) mice whereas the vitamin C levels were decreased in B(2)R(-/-) mice only. Plasma ascorbyl free radical was significantly higher in B(1)R(-/-) compared to WT and B(2)R(-/-) mice. Therefore, the oxidative stress index, ascorbyl free radical to vitamin C ratio, was increased in both B(1)R(-/-) and B(2)R(-/-) mice. Hence, B(1)R and B(2)R deficiency are associated with increased oxidative stress, but there is a differential imbalance between free radical production and antioxidant defense. The interrelationship between the differential B(1)R and B(2)R roles in oxidative stress and cardiovascular diseases remain to be investigated.

[Study on effects and mechanism of traditional Mongolian medicine wuweifengshi capsule on adjuvant arthritis in rats].

OBJECTIVE: To study the effects and immunoregulation mechanism of the traditional Mongolian medicine Wuweifengshi capsule on adjuvant arthritis (AA). METHOD: Wister rats were divided into several groups: normal group, AA model group, Wuweifengshi capsule groups (with low, moderate, high dose of 0.2, 0.4, 0.8 g x kg(-1) x d(-1) respectively), and Zhonglun-5 group (original dose of 1.68 g x kg(-1) x d(-1)). The edema degree, the level of IL-1beta, TNF-alpha, PGE2, NO and MDA and the activity of SOD in serum were detected. Through cell culture, the effects of the medicine on AA rat's splenic cell's multiplication capacity were studied. The influence of celiac macrophage cell culture fluid of AA rats' on C57BL/6J mice thymic cell multiplication capacity under the medicine was evaluated. RESULT: Wuweifengshi capsule showed an inhibiting function on the level of IL-1beta, TNF-alpha, PGE2, NO and increased the activity of SOD in serum, but showed no significant influence on MDA. It also inhibited the AA rat's splenic cell's multiplication capacity and the influence of celiac macrophage cell culture fluid of AA rat's on C57BL/6J mice thymic cell multiplication capacity. CONCLUSION: The anti-AA effect of Wuweifengshi capsule is possibly due to its inhibition of relevant cytokines and its adjustment of corresponding enzyme's activity and immunization organ's cell multiplication capacity.

Electron transfer reactions of candidate tumor suppressor 101F6 protein, a cytochrome b561 homologue, with ascorbate and monodehydroascorbate radical.

The candidate tumor suppressor 101F6 protein is a homologue of adrenal chromaffin granule cytochrome b561, which is involved in the electron transfer from cytosolic ascorbate to intravesicular monodehydroascorbate radical. Since the tumor suppressor activity of 101F6 was enhanced in the presence of ascorbate, it was suggested that 101F6 might utilize a similar transmembrane electron transfer reaction. Detailed kinetic analyses were conducted on the detergent-solubilized recombinant human 101F6 for its electron transfer reactions with ascorbate and monodehydroascorbate radical by stopped-flow and pulse radiolysis techniques. The reduction of oxidized 101F6 with ascorbate was found to be independent of pH in contrast to those observed for chromaffin granule and Zea mays cytochromes b561 in which both cytochromes exhibited very slow rates at pH 5.0 but faster at pH 6.0 and 7.0. The absence of the inhibition for the electron acceptance from ascorbate upon the treatment with diethyl pyrocarbonate suggested that 101F6 might not utilize a "concerted proton/electron transfer mechanism". The second-order rate constant for the electron donation from the ascorbate-reduced 101F6 to the pulse-generated monodehydroascorbate radical was found to be 5.0 × 10(7) M(-1 )s(-1), about 2-fold faster than that of bovine chromaffin granule cytochrome b561 and about five times faster than that of Zea mays cytochrome b561, suggesting that human 101F6 is very effective for regenerating ascorbate from monodehydroascorbate radical in cells. Present observations suggest that 101F6 employs distinct electron transfer mechanisms on both sides of the membranes from those of other members of cytochrome b561 protein family.

Electronic paramagnetic resonance (EPR) for the study of ascorbyl radical and lipid radicals in marine organisms.

Electron paramagnetic resonance (EPR) spectroscopy detects the presence of radicals of biological interest, such as ascorbyl radical (A(•)) and lipid radicals. A(•) is easily detectable by EPR even in aqueous solution at room-temperature. Under oxidative conditions leading to changes in total ascorbate (AH(-)) content, the A(•)/AH(-) ratio could be used to estimate early oxidative stress in the hydrophilic milieu. This methodology was applied to a wide range of aquatic systems including algae, sea urchin, limpets, bivalves and fish, under physiological and oxidative stress conditions as well. The A(•)/AH(-) ratio reflected the state of one part of the oxidative defense system and provided an early and simple diagnosis of environmental stressing conditions. Oxidative damage to lipids was assessed by the EPR-sensitive adduct formation that correlates well with cell membrane damage with no interference from other biological compounds. Probe instability, tissue metabolism, and lack of spin specificity are drawback factors for employing EPR for in vivo determination of free radicals. However, the dependability of this technique, mostly by combining it with other biochemical strategies, enhances the value of these procedures as contributors to the knowledge of oxidative condition in aquatic organisms.

Decreased IL-7 responsiveness is related to oxidative stress in HIV disease.

HIV disease results in decreased IL-7 receptor expression and IL-7 responsiveness in T cells. To explore mechanisms of these deficiencies, we compared CD127 expression and IL-7 induction of P-STAT5 in T cells from HIV-infected persons with serum concentrations of cytokines (IL-7, IL-6 and IL-15), markers of microbial translocation (sCD14 and LPS), and with an indicator of oxidative stress (malondialdehyde (MDA) adducts). CD127 expression was directly related to IL-7 responsiveness in most CD8+ T cell subsets but not in CD4+ T cells from HIV-infected persons. MDA adducts were increased in serum of HIV-infected patients and were inversely related to IL-7 responsiveness in CD8+ T cells and in central memory CD4+ T cells. Incubation of T cells from healthy controls with hydrogen peroxide resulted in impairments in IL-7 induction of P-STAT5. These findings suggest that oxidative stress that is characteristic of HIV disease could contribute to impairments in IL-7 responsiveness and disrupt T cell homeostasis.

A kinetic approach to assess oxidative metabolism related features in the bivalve Mya arenaria.

Electron paramagnetic resonance uses the resonant microwave radiation absorption of paramagnetic substances to detect highly reactive and, therefore, short-lived oxygen and nitrogen centered radicals. Previously, steady state concentrations of nitric oxide, ascorbyl radical (A·) and the labile iron pool (LIP) were determined in digestive gland of freshly collected animals from the North Sea bivalve Mya arenaria. The application of a simple kinetic analysis of these data based on elemental reactions allowed us to estimate the steady state concentrations of superoxide anion, the rate of A· disappearance and the content of unsaturated lipids. This analysis applied to a marine invertebrate opens the possibility of a mechanistic understanding of the complexity of free radical and LIP interactions in a metabolically slow, cold water organism under unstressed conditions. This data can be further used as a basis to assess the cellular response to stress in a simple system as the bivalve M. arenaria that can then be compared to cells of higher organisms.

New evaluation of fetal oxidative stress: measurement of the umbilical cord blood dimethyl sulfate-induced ascorbyl free radical by an electron spin resonance method.

OBJECTIVE: The aim is to evaluate intrapartum fetal oxidative stress in real-time by umbilical cord blood dimethyl sulfate (DMSO)-induced ascorbyl free radical (AFR) measured by an electron spin resonance (ESR) method. METHODS: 75 mothers delivering at gestational age after 37 weeks were recruited. They were divided into three groups: spontaneous vaginal birth (n = 27), elective cesarean section (CS) (n = 34), and emergency CS due to non-reassuring fetal status (n = 14). Umbilical artery (UA) and venous (UV) cord blood gas analysis was performed. Serum levels of DMSO-induced AFR (AFR/DMSO) that reflect vitamin C concentrations, was measured by ESR spectroscopy. RESULTS: Blood gas analysis showed no significant differences among the groups. UA-AFR/DMSO level of elective CS group was significantly lower compared with spontaneous delivery group (0.32 ± 0.12 vs. 0.46 ± 0.14, p < 0.005). Emergency CS group showed significantly lower levels of UA-AFR/DMSO compared with elective CS group (0.25 ± 0.11 vs. 0.32 ± 0.12, p < 0.005). UV-AFR/DMSO levels had no significant difference among the groups. CONCLUSIONS: It is suggested that fetal cord blood AFR/DMSO is a sensitive marker to assess fetal oxidative stress during delivery.