Teratogenicity of cobalt chloride in Xenopus laevis, assayed by the FETAX procedure.

The teratogenicity of cobalt chloride (CoCl2) was tested by the FETAX (Frog Embryo Teratogenesis Assay: Xenopus) procedure in the South African frog, Xenopus laevis. In five assays, beginning at 5 h post-fertilization, groups of Xenopus embryos were incubated for 96 h in media that contained CoCl2 at concentrations ranging from 1.8 x 10(-6) to 1.8 x 10(-2) mol/L; control groups were incubated in the same medium without added CoCl2. At 101 h post-fertilization, surviving embryos were counted, fixed in formalin, and examined by microscopy to score malformations and measure head-to-tail lengths. In control embryos, survival was greater than or equal to 95% and malformations were less than or equal to 5%. Malformations were found in greater than 99% of embryos exposed to Co2+ levels greater than or equal to 56 mumol/L. Co2+)-exposed embryos showed a concentration-related pattern of malformations, comprising gut malrotation, ocular anomalies, kinked tail, craniofacial dysplasia, cardiac deformities, and dermal blisters. Other concentration-dependent abnormalities, not categorized as malformations, included stunted growth, edema, ventral distention, and hypopigmentation. The median embryolethal concentration (LC50) of CoCl2 was 10.4 (SE +/- 0.4) mmol/L; the median teratogenic concentration (EC50) was 25 (SE +/- 2) mumol/L; the teratogenic index (TI = LC50/EC50) was 416 (SE +/- 13), indicating that CoCl2 is a potent teratogen for Xenopus laevis.     

Toxicological evaluation of lactose and chitosan delivered by inhalation

These days, inhalation constitutes a promising administration route for many drugs. However, this route exhibits unique limitations, and formulations aimed at pulmonary delivery should include as few as possible additives in order to maintain lung functionality. The purpose of this work was to investigate the safety of lactose and chitosan to the pulmonary tissue when delivered by inhalation. The study was carried out with 18 Wistar rats divided in three groups receiving distilled water, lactose or chitosan. A solution of each excipient was administered by inhalation at a dose of 20 mg. The lungs were excised and processed to determine several biochemical parameters used as toxicity biomarkers. Protein and carbonyl group content, lipid peroxidation, reduced and oxidized glutathione (GSSG), myeloperoxidase (MPO), cooper/zinc and manganese superoxide dismutase, catalase, glutathione S-transferase and glutathione peroxidase were determined. Results of myeloperoxidase activity and glutathione disulfide lung concentrations showed a relevant decrease for chitosan group compared to control: 4.67 +/- 2.27 versus 15.10 +/- 7.27 (P = 0.011) for MPO and 0.89 +/- 0.68 versus 2.02 +/- 0.22 (P = 0.014) for GSSG. The other parameters did not vary significantly among groups. Lactose and chitosan administered by inhalation failed to show toxic effects to the pulmonary tissue. A protective effect against oxidative stress might even be attributed to chitosan, since some biomarkers had values significantly lower than those observed in the control group when this product was inhaled. Nevertheless, caution must be taken regarding chemical composition and technological processes applied to incorporate these products during drug formulation, in particular for dry powder inhalators.     

Oxidative stress during energy impairment in mesencephalic cultures is not a downstream consequence of a secondary excitotoxicity

Past studies have shown that inhibiting energy metabolism with malonate in mesencephalic cultures damages neurons by mechanisms involving N-methyl-D-aspartate receptors and free radicals. Overstimulation of N-methyl-D-aspartate receptors is known to produce free radicals. This study was, therefore, carried out to determine if N-methyl-D-aspartate receptor activation triggered by energy impairment was a significant contributor to the oxidative stress generated during energy inhibition. Exposure of mesencephalic cultures to malonate for the minimal time required to produce toxicity, i.e. 6h, resulted in an increase in the efflux of both oxidized and reduced glutathione, and a decrease in tissue levels of reduced glutathione. In contrast, exposure to 1mM glutamate for 1h caused an increased efflux of reduced glutathione, but no changes in intra- or extracellular oxidized glutathione or intracellular reduced glutathione. Blocking N-methyl-D-aspartate receptors with MK-801 (0.5 microM) during malonate exposure did not modify malonate-induced alterations in glutathione status or free radical generation as monitored by dihydrochlorofluorescein diacetate and dihydrorhodamine 123 fluorescence. In contrast, the increase in dihydrorhodamine fluorescence caused by glutamate was completely blocked by MK-801. Reduction of tissue glutathione with a 24h pretreatment with 10 microM buthionine sulfoxamine, as shown previously, greatly potentiated malonate-induced toxicity to dopamine and GABA neurons, but had no potentiating effect on toxicity due to glutamate.The findings indicate that although oxidative stress mediates damage due either to energy deprivation or excitotoxicity, N-methyl-D-aspartate receptor over-stimulation does not contribute significantly to the oxidative stress that is incurred during malonate exposure.     

The circadian clock regulates rhythmic activation of the NRF2/glutathione-mediated antioxidant defense pathway to modulate pulmonary fibrosis

The disruption of the NRF2 (nuclear factor erythroid-derived 2-like 2)/glutathione-mediated antioxidant defense pathway is a critical step in the pathogenesis of several chronic pulmonary diseases and cancer. While the mechanism of NRF2 activation upon oxidative stress has been widely investigated, little is known about the endogenous signals that regulate the NRF2 pathway in lung physiology and pathology. Here we show that an E-box-mediated circadian rhythm of NRF2 protein is essential in regulating the rhythmic expression of antioxidant genes involved in glutathione redox homeostasis in the mouse lung. Using an in vivo bleomycin-induced lung fibrosis model, we reveal a clock “gated” pulmonary response to oxidative injury, with a more severe fibrotic effect when bleomycin was applied at a circadian nadir in NRF2 levels. Timed administration of sulforaphane, an NRF2 activator, significantly blocked this phenotype. Moreover, in the lungs of the arrhythmic Clock^Δ19 mice, the levels of NRF2 and the reduced glutathione are constitutively low, associated with increased protein oxidative damage and a spontaneous fibrotic-like pulmonary phenotype. Our findings reveal a pivotal role for the circadian control of the NRF2/glutathione pathway in combating oxidative/fibrotic lung damage, which might prompt new chronotherapeutic strategies for the treatment of human lung diseases, including idiopathic pulmonary fibrosis.     

Differential regulation of hydrogen peroxide and Fas-dependent apoptosis pathways by dehydroascorbate,the oxidized form of vitamin C

Dehydroascorbate (DHA), the oxidized form of vitamin C (ascorbate), enhanced antioxidant defenses of human T cells preferentially importing DHA over ascorbate. In itself, DHA did not affect cytosolic or mitochondrial reactive oxygen intermediate levels as monitored by flow cytometry using oxidation-sensitive fluorescent probes. DHA at 200-1,000 microM stimulated activity of pentose phosphate pathway enzymes glucose 6-phosphate dehydrogenase, 6-phosphogluconate dehydrogenase, and transaldolase, elevated intracellular glutathione levels, and inhibited H(2)O(2)-induced changes in mitochondrial transmembrane potential and cell death. With respect to the CD4 antigen, DHA selectively enhanced cell-surface expression of the Fas receptor and increased susceptibility of Jurkat and H9 human T cells to Fas-mediated cell death. The data identify DHA as a selective regulator of H(2)O(2)- and Fas-dependent apoptosis pathways.     

Toxicological evaluation of lactose and chitosan delivered by inhalation

These days, inhalation constitutes a promising administration route for many drugs. However, this route exhibits unique limitations, and formulations aimed at pulmonary delivery should include as few as possible additives in order to maintain lung functionality. The purpose of this work was to investigate the safety of lactose and chitosan to the pulmonary tissue when delivered by inhalation. The study was carried out with 18 Wistar rats divided in three groups receiving distilled water, lactose or chitosan. A solution of each excipient was administered by inhalation at a dose of 20 mg. The lungs were excised and processed to determine several biochemical parameters used as toxicity biomarkers. Protein and carbonyl group content, lipid peroxidation, reduced and oxidized glutathione (GSSG), myeloperoxidase (MPO), cooper/zinc and manganese superoxide dismutase, catalase, glutathione S-transferase and glutathione peroxidase were determined. Results of myeloperoxidase activity and glutathione disulfide lung concentrations showed a relevant decrease for chitosan group compared to control: 4.67 ± 2.27 versus 15.10 ± 7.27 (P = 0.011) for MPO and 0.89 ± 0.68 versus 2.02 ± 0.22 (P = 0.014) for GSSG. The other parameters did not vary significantly among groups. Lactose and chitosan administered by inhalation failed to show toxic effects to the pulmonary tissue. A protective effect against oxidative stress might even be attributed to chitosan, since some biomarkers had values significantly lower than those observed in the control group when this product was inhaled. Nevertheless, caution must be taken regarding chemical composition and technological processes applied to incorporate these products during drug formulation, in particular for dry powder inhalators.     

Glutathione-dependent redox status of frataxin-deficient cells in a yeast model of Friedreich's ataxia

Friedreich's ataxia is a neurodegenerative disease caused by reduced expression of the mitochondrial protein frataxin. The main phenotypic features of frataxin-deficient human and yeast cells include iron accumulation in mitochondria, iron-sulphur cluster defects and high sensitivity to oxidative stress. Glutathione is a major protective agent against oxidative damage and glutathione-related systems participate in maintaining the cellular thiol/disulfide status and the reduced environment of the cell. Here, we present the first detailed biochemical study of the glutathione-dependent redox status of wild-type and frataxin-deficient cells in a yeast model of the disease. There were five times less total glutathione (GSH+GSSG) in frataxin-deficient cells, imbalanced GSH/GSSG pools and higher glutathione peroxidase activity. The pentose phosphate pathway was stimulated in frataxin-deficient cells, glucose-6-phosphate dehydrogenase activity was three times higher than in wild-type cells and this was coupled to a defect in the NADPH/NADP(+) pool. Moreover, analysis of gene expression confirms the adaptative response of mutant cells to stress conditions and we bring evidence for a strong relation between the glutathione-dependent redox status of the cells and iron homeostasis. Dynamic studies show that intracellular glutathione levels reflect an adaptation of cells to iron stress conditions, and allow to distinguish constitutive stress observed in frataxin-deficient cells from the acute response of wild-type cells. In conclusion, our findings provide evidence for an impairment of glutathione homeostasis in a yeast model of Friedreich's ataxia and identify glutathione as a valuable indicator of the redox status of frataxin-deficient cells.     

冬虫夏草对慢性阻塞性肺疾病模型鼠还原型谷胱甘肽-氧化型谷胱甘肽平衡和Thl—Th2型细胞因子的影响

目的探讨冬虫夏草对大鼠慢性阻塞性肺疾病（COPD）模型还原型谷胱甘肽（GSH）-氧化型谷胱甘肽（GSSG）失衡的干预作用及对Thl．Th2型细胞因子的影响。方法18只sD大鼠完全随机分为COPD对照组、冬虫夏草干预组和N-乙酰半胱氨酸（NAC）干预组（每组各6只）,分别在COPD模型制作中胃饲生理盐水、冬虫夏草和NAC,观察大鼠肺组织病理改变。检测冬虫夏草和NAC干预后支气管肺泡灌洗液（BALF）中Thl．Th2型细胞因子和巨噬细胞中GSH和GSH／GSSG变化。结果与COPD对照组（32＋13）相比,冬虫夏草干预组和NAC干预组的平均肺泡计数增加（49＋10,52＋14,P〈O．05）。冬虫夏草干预组和NAC干预组BALF中巨噬细胞GSH水平和GSH-GSSG较COPD对照组升高（t=3．06,t=3．24;t=2．36,t=2．82;均P〈0．05）。冬虫夏草干预组和NAC干预组BALF上清液中Thl型细胞因子IFN-y水平较COPD对照组升高（f=2．34,t=2．32,P〈0．05）。结论冬虫夏草可能通过提高COPD模型巨噬细胞内GSH水平,补充抗氧化水平而达到纠正Thl—Th2失衡的免疫调节。     

Effect of Methylsulfonylmethane on Paraquat-Induced Acute Lung and Liver Injury in Mice

Methylsulfonylmethane (MSM) is a natural organosulfur compound that exhibits antioxidative and anti-inflammatory effects. This study was carried out to investigate the effect of MSM on paraquat (PQ)-induced acute lung and liver injury in mice. A single dose of PQ (50 mg/kg, i.p.) induced acute lung and liver toxicity. Mice were treated with MSM (500 mg/kg/day, i.p.) for 5 days. At the end of the experiment, animals were euthanized, and lung and liver tissues were collected for histological and biochemical analysis. Tissue samples were used to determine malondialdehyde (MDA), myeloperoxidase (MPO), catalase (CAT), superoxide dismutase (SOD), glutathione (GSH), and tumor necrosis factor-α (TNF-α) levels. Blood samples were used to measure plasma alanine transaminase (ALT), γ-glutamyl transferase (GGT), and alkaline phosphatase (ALP). Histological examination indicated that MSM decreased lung and liver damage caused by PQ. Biochemical results showed that MSM treatment significantly reduced tissue levels of MDA, MPO, and TNF-α, while increased the levels of SOD, CAT, and GSH compared with PQ group. MSM treatment also significantly reduced plasma levels of ALT, GGT, and ALP. These findings suggest that MSM as a natural product attenuates PQ-induced pulmonary and hepatic oxidative injury.     

Increased concentrations of glutathione in induced sputum of patients with mild or moderate allergic asthma.

BACKGROUND: Oxidative stress is involved in the pathogenesis of allergic inflammatory diseases, such as rhinitis and asthma. Glutathione is a vital intracellular and extracellular protective pulmonary antioxidant. It plays a key role in regulating oxidant-induced lung epithelial cell function and also in the control of proinflammatory processes. OBJECTIVE: To quantify oxidative stress in sputum of asthmatic patients compared with healthy subjects. METHODS: We quantified induced sputum supernatant concentrations of total and oxidized glutathione in 20 patients with mild asthma without inhaled corticosteroid treatment, 19 patients with moderate-persistent asthma treated with inhaled corticosteroids (median dose, 900 microg/d of beclomethasone equivalent), and 15 healthy, nonatopic, nonsmoking subjects. RESULTS: Total glutathione levels were significantly increased in mild and persistent asthma compared with healthy subjects [geometric mean (95% confidence interval), 9.2 microM (7.1-12 microM) and 8.7 microM (5.9-12.5 microM) vs 4 microM (2.7-6 microM); P = .039 and .042, respectively]. In contrast, there were no differences in total and oxidized glutathione levels between steroid-na?ve and steroid-treated asthmatic patients (P > .20 for all comparisons). Persistent, steroid-treated asthmatic patients had higher sputum counts of neutrophils than steroid-na?ve asthmatic patients [geometric mean (95% confidence interval), 35.6% (28.2%-42.7%) vs 17.7% (11.7%-27.1%), P = .04]. There was a positive correlation of total glutathione with sputum total cells (rho = 0.32, P = .02). CONCLUSIONS: Total glutathione is increased in induced sputum of patients with mild and moderate asthma. These data underline the contribution of oxidative stress to the pathogenesis of allergic asthmatic inflammation.     

Glucose‐6‐phosphate dehydrogenase inhibition attenuates acute lung injury through reduction in NADPH oxidase‐derived reactive oxygen species

Acute lung injury (ALI) is a heterogeneous disease with the hallmarks of alveolar capillary membrane injury, increased pulmonary oedema and pulmonary inflammation. The most common direct aetiological factor for ALI is usually parenchymal lung infection or haemorrhage. Reactive oxygen species (ROS) generated by nicotinamide adenine dinucleotide phosphate (NADPH) oxidase (NOX2) are thought to play an important role in the pathophysiology of ALI. Glucose‐6‐phosphate dehydrogenase (G6PD) plays an important role both in production of ROS as well as their removal through the supply of NADPH. However, how G6PD modulation affects NOX2‐mediated ROS in the airway epithelial cells (AECs) during acute lung injury has not been explored previously. Therefore, we investigated the effect of G6PD inhibitor, 6‐aminonicotinamide on G6PD activity, NOX2 expression, ROS production and enzymatic anti‐oxidants in AECs in a mouse model of ALI induced by lipopolysaccharide (LPS). ALI led to increased G6PD activity in the AECs with concomitant elevation of NOX2, ROS, SOD1 and nitrotyrosine. G6PD inhibitor led to reduction of LPS‐induced airway inflammation, bronchoalveolar lavage fluid protein concentration as well as NOX2‐derived ROS and subsequent oxidative stress. Conversely, ALI led to decreased glutathione reductase activity in AECs, which was normalized by G6PD inhibitor. These data show that activation of G6PD is associated with enhancement of oxidative inflammation in during ALI. Therefore, inhibition of G6PD might be a beneficial strategy during ALI to limit oxidative damage and ameliorate airway inflammation.     

Sudden infant death syndrome: oxidative stress

In studies of oxidative stress in sudden infant death syndrome (SIDS) there were two major findings: (1) During normal post-natal development, there was a gradual decline in the number of Cu/Zn superoxide dismutase (SOD) and glutathione peroxidase (GSHPx) immunoreactive neurons in the hippocampus and parahippocampus gyrus in the brain; (2) The total number of immunoreactive neurons was elevated in SIDS victims compared to age-matched controls in infants 6 months of age and under (1). SOD and neuronal aging and degeneration in the hippocampus and neocortex were features of SIDS, Alzheimer's disease and Down's syndrome. In the SIDS study of infants from 3-6 months of age, the elevation of SOD in SIDS victims was significant, whereas no significant elevation of GSHPx was detected. An imbalance between SOD and GSHPx was said to be crucial in the prevention of toxicity of free radicals (1). Zinc-deficient cells cannot up-regulate gene expression of the scavenger enzymes SOD and GSHPx in cells exposed to high levels of superoxide and hydrogen peroxide (2). GSHPx coupled to reduced nicotine adenine diphosphate (NADPH) regenerating systems via glutathione reductase is virtually able to guarantee an effective protection of biological structures against oxidative attack (22). When the capacity of the cell to regenerate GSH is exceeded - primarily due to an insufficient supply of NADPH-oxidised glutathione (GSSG) is released from the cell and protein synthesis turns off (20). We hypothesize that the increased incidence of aging and neuronal death and increased incidence of SOD and GSHPx reactive neurons in early post-natal development indicates an increased up-regulation of gene expression of scavenger enzymes during high exposure to oxidative stress after birth. GSH-dependent peroxide metabolism is linked to the pentose phosphate shunt via NADPH-dependent glutathione reductase (GR). GSHPx is a selenium containing enzyme which together with catalase (CAT) SOD and vitamin E protects cells in the free radical chain. Zinc upregulates gene expression of these antioxidants.     

In vitro stability of a novel compliant poly(carbonate-urea)urethane to oxidative and hydrolytic stress.

Poly(ester)urethane and poly(ether)urethane vascular grafts fail in vivo because of hydrolytic and oxidative degradative mechanisms. Studies have shown that poly(carbonate)urethanes have enhanced resistance. There is still a need for a viable, nonrigid, small-diameter, synthetic vascular graft. In this study, we sought to confirm this by exposing a novel formulation of compliant poly(carbonate-urea)urethane (CPU) manufactured by an innovative process, resulting in a stress-free. Small-diameter prosthesis, and a conventional poly(ether)urethane Pulse-Tec graft known to readily undergo oxidation in a variety of degradative solutions, and we assessed them for the development of oxidative and hydrolytic degradation, changes in elastic properties, and chemical stability. To simulate the in vivo environment, we used buffered solutions of phospholipase A(2) and cholesterol esterase; solutions of H(2)O(2)/CoCl(2), t-butyl peroxide/CoCl(2) (t-but/CoCl(2)), and glutathione/t-butyl peroxide/CoCl(2) (Glut/t-but/CoCl(2)); and plasma fractions I-IV, which were derived from fresh human plasma centrifuged in poly(ethylene glycol). To act as a negative control, both graft types were incubated in distilled water. Samples of both graft types (100 mm with a 5.0-mm inner diameter) were incubated in these solutions at 37 degrees C for 70 days before environmental scanning electron microscopy, radial tensile strength and quality control, gel permeation chromatography, and in vitro compliance assessments were performed. Oxidative degradation was ascertained from significant changes in molecular weight with respect to a control on all Pulse-Tec grafts treated with t-but/CoCl(2), Glut/t-but/CoCl(2), and plasma fractions I-III. Pulse-Tec grafts exposed to the H(2)O(2)/CoCl(2) mixture had significantly greater compliance than controls incubated in distilled water (p < 0.001 at 50 mmHg). No changes in molecular weight with respect to the control were observed for the CPU samples; only those immersed in t-but/CoCl(2) and Glut/t-but/CoCl(2) showed an 11% increase in molecular weight to 108,000. Only CPU grafts treated with the Glut/t-but/CoCl(2) mixture exhibited significantly greater compliance (p < 0.05 at 50 mmHg). Overall, results from this study indicate that CPU presents a far greater chemical stability than poly(ether)-urethane grafts do.     

Mitochondrial oxidative stress in the lungs of cystic fibrosis transmembrane conductance regulator protein mutant mice

Cystic fibrosis is a fatal genetic disorder involving dysfunction of the cystic fibrosis transmembrane regulator protein (CFTR) resulting in progressive respiratory failure. Previous studies indicate that CFTR regulates cellular glutathione (GSH) transport and that dysfunctional CFTR is associated with chronic pulmonary oxidative stress. The cause and the source of this oxidative stress remain unknown. The current study examines the role of the mitochondria in CFTR-mediated pulmonary oxidative stress. Mitochondrial GSH levels and markers of DNA and protein oxidation were assessed in the lung mitochondria from CFTR-knockout mice. In addition, in vitro models using human CFTR-sufficient and -deficient lung epithelial cells were also employed. Mitochondrial GSH levels were found to be decreased up to 85% in CFTR-knockout mice, and 43% in human lung epithelial cells deficient in CFTR. A concomitant 29% increase in the oxidation of mitochondrial DNA, and a 30% loss of aconitase activity confirmed the existence of a mitochondrial oxidative stress. Flow cytometry revealed significantly elevated levels of cellular reactive oxygen species (ROS) in CFTR-deficient human lung cells. These studies suggest that dysfunctional CFTR leads to an increase in the level of ROS and mitochondrial oxidative stress. This oxidative stress, however, appears to be a consequence of lower mitochondrial GSH levels and not increased oxidation of GSH. Further studies are needed to determine how CFTR deficiency contributes to mitochondrial oxidative stress and the role this plays in CFTR-mediated lung pathophysiology.     

低氧诱导的小窝蛋白-1上调参与人肺腺癌细胞A549迁移和侵袭

 目的:探讨低氧诱导剂二氯化钴（CoCl2）对小窝蛋白-1（Cav-1）生成的调节作用及后者对人肺腺癌A549细胞迁移、侵袭的影响。方法:检测肺癌患者伴恶性胸水（MPE）和结核性胸膜炎患者胸水（TBPE）中Cav-1和缺氧诱导因子(HIF)-1α浓度,比较两者相关性;以CoCl2和（或）HIF-1α抑制剂YC-1作用于A549细胞ELISA法检测细胞上清Cav-1和HIF-1α浓度;分别采用细胞划痕实验及Transwell小室侵袭实验研究CoCl2刺激表达的Cav-1对A549细胞迁移和侵袭的影响。结果:MPE中Cav-1和HIF-1α浓度明显高于TBPE,两组患者胸水中Cav-1与HIF-1α均呈正相关。CoCl2浓度和时间依赖性诱导A549细胞Cav-1和HIF-1α生成,200 μmol/L或24 h达到峰值;浓度>200 μmol/L或作用时间超过24 h则呈现浓度或时间依赖性抑制。HIF-1α抑制剂YC-1浓度依赖性抑制HIF-1α和Cav-1生成。CoCl2浓度依赖性增强A549细胞迁移和侵袭,200 μmol/L作用最强;YC-1对上述过程产生抑制效应。结论: 肺癌患者胸腔积液中Cav-1浓度升高,低氧诱导Cav-1生成的变化可能参与了A549细胞迁移和侵袭,HIF-1α可能对Cav-1生成发挥影响。     

Hydrogen sulfide protects HT22 neuronal cells from oxidative stress

Hydrogen sulfide (H2S) is a neuromodulator in the brain and a relaxant for smooth muscle. H2S protects primary cortical neurons from oxidative stress by increasing the intracellular concentrations of glutathione, the major antioxidant in cells. However, changes in glutathione alone are not sufficient to account for full protection in all types of nerve cells. H2S is here shown to protect an immortalized mouse hippocampal cell line from oxidative glutamate toxicity by activating ATP-dependent K+ (KATP) and Cl- channels, in addition to increasing the levels of glutathione. The present study therefore identifies a novel pathway for H2S protection from oxidative stress.     

Cytophotometric assay of cytochrome oxidase, lactate dehydrogenase and glucose-6-phosphate dehydrogenase activities in human peroxidized spermatozoa.

Human spermatozoa contain appreciable amounts of intracellular glutathione, which has a protective function against peroxidative degradation of spermatozoal polyunsaturated fatty acids by the NADPH-dependent glutathione peroxidase/reductase enzymatic system. The glutathione system provides a basic defense against peroxidative damage, without which the superoxide dismutase system would dominate. Since oxidative damage is said to include enzyme leakage and changes in metabolism, cytochrome oxidase and lactate dehydrogenase activities were used as indicators of the energy metabolism in unwashed and washed human spermatozoa during lipid peroxidation. Lipid peroxidation was induced by aerobic incubation of sperms in the presence of sodium ascorbate and ferrous sulphate. In addition, since NADPH concentrations influence the concentration of reduced glutathione, we studied glucose-6-phosphate dehydrogenase activity as an indicator of pentose phosphate shunt activity, the main source of NADPH. Microdensitometric measurements of the three enzymes were made by a Vickers M85a scanning microdensitometer. We found that the lipid peroxidation process greatly affects the 3 enzymatic activities examined and that seminal plasma protects against the extensive deleterious effects of lipid peroxidation.     

H2S protects against methionine-induced oxidative stress in brain endothelial cells

Homocysteine (Hcy) causes cerebrovascular dysfunction by inducing oxidative stress. However, to date, there are no strategies to prevent Hcy-induced oxidative damage. Hcy is an H2S precursor formed from methionine (Met) metabolism. We aimed to investigate whether H2S ameliorated Met-induced oxidative stress in mouse brain endothelial cells (bEnd3). The bEnd3 cells were exposed to Met treatment in the presence or absence of NaHS (donor of H2S). Met-induced cell toxicity increased the levels of free radicals in a concentration-dependent manner. Met increased NADPH-oxidase-4 (NOX-4) expression and mitigated thioredxion-1(Trx-1) expression. Pretreatment of bEnd3 with NaHS (0.05 mM) attenuated the production of free radicals in the presence of Met and protected the cells from oxidative damage. Furthermore, NaHS enhanced inhibitory effects of apocynin, N-acetyl-l-cysteine (NAC), reduced glutathione (GSH), catalase (CAT), superoxide dismutase (SOD), Nomega-nitro-l-arginine methyl ester (L-NAME) on ROS production and redox enzymes levels induced by Met. In conclusion, the administration of H2S protected the cells from oxidative stress induced by hyperhomocysteinemia (HHcy), which suggested that NaHS/H2S may have therapeutic potential against Met-induced oxidative stress.     

Systemic oxidative and antioxidative status in Chinese patients with asthma

BACKGROUND: Patients with asthma generate an increased amount of reactive oxygen species from peripheral blood cells. Reactive oxygen species produce many of the pathophysiologic changes associated with asthma and may contribute to its pathogenesis. OBJECTIVE: We investigated changes in antioxidant enzyme activities and oxidized glutathione (glutathione disulfide; GSSG) levels in erythrocytes from a group of healthy control Chinese subjects (n=135) and patients with asthma (n=106). METHODS: Baseline pulmonary function was measured for all subjects. Antioxidant status was evaluated by measuring erythrocyte superoxide dismutase, catalase, and glutathione peroxidase activities. Oxidative stress was also measured in terms of GSSG in erythrocytes with a kinetic microassay. RESULTS: Patients with asthma had significantly increased erythrocyte superoxide dismutase and catalase activities compared with controls (61.10 +/- 1.30 U/g hemoglobin [Hb] vs 55.51 +/- 1.82 U/g Hb [P=.018] and 0.0637 +/- 0.0021 U/g Hb vs 0.0257 +/- 0.0120 U/g Hb [P <.001] for the asthma and control groups, respectively). Conversely, erythrocyte glutathione peroxidase activity decreased (44.21 +/- 1.33 mU/g Hb vs 50.07 +/- 1.39 mU/g Hb for the asthma and control groups, respectively; P=.003). Patients with asthma also had significantly higher GSSG levels in erythrocyte hemolysates compared with controls (167.40 +/- 2.93 micromol/L vs 44.98 +/- 0.44 micromol/L for the asthma and control groups, respectively; P <.001), indicating increased oxidative stress. CONCLUSIONS: Asthma is accompanied by an alteration in systemic antioxidant status due to possible oxidative stress in this disease.