Effects Of Active Oxygen Species Research Articles

In vitro influence of plant extracts beyond taxonomy: a comparison ...

Detailed information on the in vivo activity of vegetal extracts on the immune system in birds and mammals, mainly the farmed ones, is helpful in improving their resilience and welfare. Therefore there is need to investigate the exact roles of active plant components in animals, mainly their influence on the immune system, which is crucial for antimicrobial resistance. The farming system, serving principally economic aims, overlooks the physiological support of several extremely stress-sensitive, short-lived species such as chickens or rabbits. Recovering from the damage caused by adverse microclimates or nutritional deficiencies in these species can be a prolonged process often standing beyond their economic lifespan. Additionally, the compromised condition makes them susceptible to various diseases, particularly microbial or parasitic infections which can easily take hold in the weakened state. Under these circumstances, it seems very important to select and study natural enhancers of the immune response. Setting up optimal therapeutic protocols and establishing the optimal age range and administration route boost their effects. The in vitro tests, performed mainly on isolated cell- or cell-line cultures, proved the beneficial influence of certain vegetal extracts, which diminished oxidative stress and tumor growth, as well as overcame the negative effects of active oxygen species, increasing the synthesis of cytokines. The study aimed a) to establish a potential dose-effect relationship for selected plant extracts, b) considering that belonging to either of groups Aves or Mammalia changes the response to these extracts. Blood samples taken from hens and rabbits were subjected to in vitro blast transformation tests (RPMI 1640 culture medium, 48 hours of incubation in a 5% CO2 atmosphere) to estimate the effects of alcoholic extracts of Calendula officinalis, Vaccinum myrtilus, Echinacea purpurea, and Hippophae rhamnoides at 1.5% and 6.5% concentrations by a colorimetric assay (orto-toluidine test). The results in hens indicated maximal stimulation indices, in a reverse dose-dependent manner, for Vaccinum myrtillus (81.93 ± 18.74% for 1.5% concentration) and Calendula officinalis (74.70 ± 22.15% for 1.5% concentration). In rabbits, all the extracts had stimulating effects, Calendula (80.72 ± 6.82% for 6.5% concentration) and Echinacea (68.67 ± 17.04% for 6.5% concentration) being more effective than Vaccinum (65.06 ± 35.78% at 1.5% concentration). Obviously, the in vitro effects of the extracts depended on the phylogeny of the animal and the plant of origin. Thus, the birds reacted to certain extracts in an inversely dose-dependent manner, while in rabbits, the effects were directly dose-dependent. The results highlight the importance of preliminary tests in selecting an appropriate vegetal extract for immune stimulation/modulation of a given species of animals.

IMPACT OF CHEMOTHERAPY ON PROOXIDANT-ANTIOXIDANT STATUS IN PATIENTS WITH MULTIPLE MYELOMA AND CONCOMITANT CORONARY ARTERY DISEASE

Introduction. Multiple myeloma, an oncopathology that remains challenging to treat, is characterized by a 54% 5-year survival rate despite advancements in modern treatment strategies. Numerous studies have substantiated the correlation between high prooxidant concentrations and the progression of multiple myeloma. This progression is often associated with alterations in antioxidant defense mechanisms. Patients with coronary artery disease also experience changes in their antioxidant defense that may enhance the deleterious effects of active oxygen species and elevate the risk of chemotherapy-related organotoxic effects.
 The aim of this study is to investigate the specificities of the prooxidant-antioxidant status in patients with multiple myeloma and concomitant coronary artery disease during chemotherapy.
 Materials and methods. 42 patients with multiple myeloma were examined, 22 (52,5%) of them had concomitant coronary artery disease. According to the coronary artery disease, patients with multiple myeloma were divided into two groups. All patients obtain standard chemotherapy schemes. Assessment of the patients was performed four times: before and after the 1st and 5th chemotherapy course. TBA-reactants and catalase activity were measured in blood serum.
 Results. Both in I and II group of patients, TBA-reactants concentration in blood serum was increased in 1.85 (р1<0,0001) and in 2.35 (р1<0,0001) times respectively compare to healthy people before chemotherapy. Although in the patients with concomitant coronary artery disease TBA-reactants concentration in blood serum was increased in 1,28 (р2<0,0001) times compare to I group of patients. Simultaneously in II group of patients catalase activity was decreased in 1.28 (р4 <0,0001) times compare to healthy people and in 1,37 (р5 <0,0001) times compare to I group patients. Before the second course of chemotherapy in II group of patients, TBA-reactants level was increased in 1.03 (р<0.05) times compared to the first examination and in 1.27 (р<0.05) times compared to I group. In II group of patients, catalase activity was decreased in 1.24 (р<0.05) times compared to the first examination and in 1.38 (р<0.001) times compared to the II group. In II group of patients level of TBA-reactants was decreased in 1,13 (p<0,01) times compare to the first examination, but it was in 1,17(р<0,05) higher than TBA-reactants level in I group. Simultaneously catalase activity was decreased in 1.4 (р<0.05) times compare to the first examination.
 Conclusions. The progression of multiple myeloma is marked by the development of a prooxidant-antioxidant imbalance. In both the I and II patient groups, the level of TBA-reactants increased significantly by 1.85 (p<0.0001) and 2.35 (p<0.0001) times, respectively, compared to the reference value. The presence of concomitant coronary artery disease in multiple myeloma patients led to a 1.28-fold elevation in TBA-reactants levels (p<0.0001) compared to those without cardiovascular diseases. Simultaneously, catalase activity decreased by 1.37 times (p<0.0001) compared to patients without cardiovascular diseases.
 During chemotherapy, a progressive increase in the prooxidant-antioxidant imbalance was observed in multiple myeloma patients. This was evidenced by a 1.4-fold decrease in catalase activity (p<0.05) compared to the initial examination.

Hydrothermal-treatment-based facile one-step preparation of K-promoted NOx adsorbents derived from hydrotalcite-like compounds

K-promoted mixed metal oxides were prepared from K2CO3-promoted Co-Mg-Al hydrotalcite-like compounds using a simple hydrothermal synthesis method. The introduced K2CO3 largely increased the NOx uptake by generating active oxygen species; however, the presence of an excessive amount of K2CO3 in the aggregated bulk phase suppressed the formation of active oxygen species. The developed NOx adsorbents presented excellent NOx uptake at low temperatures (0.566 and 2.63 mmol g−1 at 150 and 250 °C, respectively). In addition, the effects of active oxygen species on the NO oxidation and NOx adsorption ability were investigated. The amount of active oxygen species and the NOx adsorption performance at 150 °C were highly correlated. However, the NOx uptake at 250 °C gradually increased with the amount of K2CO3, although the K2CO3 species in the aggregated bulk phase inhibited the formation of active oxygen species. It was revealed that the K2CO3 in the bulk phase promoted the adsorption of NOx at 250 °C by directly trapping it. Overall, the introduced K2CO3 enhanced the NOx adsorption performance in two ways: (1) as a surface modifier to generate active oxygen species and (2) via the direct adsorption of NOx at high temperature.

Promotion effect of S and N co-addition on the catalytic performance of V2O5/TiO2 for NH3-SCR of NOX

The influence of S and N in the V2O5/TiO2 catalyst system on the activity was evaluated for the selective catalytic reduction (SCR) of NOX. The VTi1 (S/N/Ti = 3:1:100) with wide temperature window has the highest catalytic activity. The properties of the catalysts were characterized by physicochemical measurements, including X-ray diffraction, BET surface area, NH3 temperature-programmed desorption, H2 temperature-programmed reduction and O2 temperature-programmed desorption, and so on. The structure, surface area, Lewis acidity, reducibility and active sites of the catalyst were significantly influenced by adding S and N. The results of ultraviolet–visible spectroscopy and electron paramagnetic resonance indicated that the presence of the strong electron interactions between dispersed vanadia and TiO2 species and more active oxygen species on the surface of catalyst. This work provides new insights into the effect of active oxygen species on the catalytic conversion of the nitrogen oxides.

The effect of active oxygen species in nano-ZnCr2O4 spinel oxides for methane catalytic combustion

The nano-ZnCr2O4 spinel oxides was synthesized by a ethylene glycol mediated solvothermal method. Catalytic combustion of methane test showed that an excellent activity over nano-ZnCr2O4 with T10% = 300 °C and T90% = 400 °C. The results of X-ray diffraction (XRD), scanning electron microscopy (SEM), N2 adsorption-desorption measurements (BET) indicated that a uniform nano-ZnCr2O4 spinel oxides particles with the high surface area (96.2 m2 g−1) was successfully synthesized. Oxygen temperature programmed desorption (O2-TPD) profile revealed there were two obvious desorption of oxygen species from nano-ZnCr2O4 in the range of 300–400 °C and 500–700 °C. It was clear that the desorption temperature range of the first oxygen species coincided with the methane catalytic combustion temperature. X-ray photoelectron spectroscopy (XPS) analysis exhibited that Cr6+ was present in the lattice of ZnCr2O4 apart from Cr3+. High valence cations of chromium in crystal lattice probable caused the presence of interstitial oxygen species in the structure to maintain the electroneutrality. Additionally, Raman spectra proved that there is the interstitial oxygen species in the crystal lattice of ZnCr2O4. Therefore, the excellent catalytic activity for methane combustion was contributed to the flexible interstitial oxygen in the ZnCr2O4.

FEATURES OF FREE RADICAL OXIDATION AND ANTIOXIDANT PROTECTION IN CHILDREN WITH SUGAR DIABETES OF THE FIRST TYPE

Aim. Assessment of the state of lipid peroxidation, antioxidant defense and oxidative stress indicators on the model of functional activity of neutrophil granulocytes of blood in children with type 1 diabetes mellitus depending on the stage of endocrinopathy compensation. Materials and methods. The material of laboratory-diagnostic and clinical studies was the results of a survey of 38 children without endocrine pathology and 89 children with type 1 diabetes mellitus at various stages of disease compensation. The state of antioxidant protection was assessed taking into account the overall antioxidant activity, superoxide dismutase activity, retinol level, α-tocopherol, the content of reduced and oxidized glutathione. The intensity of lipoperoxidation processes was studied by the level of substrates with conjugated double bonds, diene conjugates, ketodienes, conjugated trienes, malonic dialdehyde, common lipids. Functional activity of neutrophil granulocytes was investigated by means of luminol-dependent chemiluminescence. Results. In children with type 1 diabetes mellitus, the intensification of lipid peroxidation processes and the activation of the antioxidant defense system, including variously directed changes in non-enzymatic mechanisms, are noted against the negative effect of active oxygen species. In children with decompensated type 1 diabetes mellitus, severe lipid metabolism disorders combined with oxidative stress indices increase the course of endocrine pathology, significantly increasing the likelihood of intravascular complications. Conclusion. The coordinated generation of active forms of oxygen and the processes of oxygen-dependent metabolism of blood cells in children with compensated type 1 diabetes mellitus indicates the development of the second stage (resistance) of oxidative stress. Reduction of reactive oxygen species production, reduced activation rate оxygendependent metabolism phagocytes incompleteness phagocytosis mechanisms, correlating with an increase in lesion area (destruction of insulin-producing β-cells) of the pancreas in children with decompensated type 1 diabetes, indicating the occurrence of the third step (debilitation) oxidative stress. Violations of metabolic pathways in children with type 1 diabetes are determined by the intensity of «respiratory burst» of neutrophil granulocytes in the «Lipid peroxidation – antioxidant protection».

活性酸素および紫外光曝露によるポリジメチルシロキサンの表面処理と細胞培養基板への応用

活性酸素および紫外光曝露によるポリジメチルシロキサンの表面処理と細胞培養基板への応用

Characterization of manganese superoxide dismutase from a marine cyanobacterium Leptolyngbya valderianaBDU20041

BackgroundCyanobacteria are recognized as the primordial organisms to grace the earth with molecular oxygen ~3.5 billion years ago as a result of their oxygenic photosynthesis. This laid a selection pressure for the evolution of antioxidative defense mechanisms to alleviate the toxic effect of active oxygen species (AOS) in cyanobacteria. Superoxide dismutases (SODs) are metalloenzymes that are the first arsenal in defense mechanism against oxidative stress followed by an array of antioxidative system. Unlike other living organisms, cyanobacteria possess multiple isoforms of SOD. Hence, an attempt was made to demonstrate the oxidative stress tolerance ability of marine cyanobacterium, Leptolyngbya valderiana BDU 20041 and to PCR amplify and sequence the SOD gene, the central enzyme for alleviating stress.ResultL. valderiana BDU 20041, a filamentous, non-heterocystous marine cyanobacterium showed tolerance to the tested dye (C.I. Acid Black 1) which is evident by increased in biomass (i.e.) chlorophyll a. The other noticeable change was the total ROS production by culture dosed with dye compared to the control cultures. This prolonged incubation showed sustenance, implying that cyanobacteria maintain their antioxidant levels. The third significant feature was a two-fold increase in SOD activity of dye treated L. valderiana BDU20041 suggesting the role of SOD in alleviating oxidative stress via Asada-Halliwell pathway. Hence, the organism was PCR amplified for SOD gene resulting in an amplicon of 550 bp. The sequence analysis illustrated the presence of first three residues involved in motif; active site residues at H4, 58 and D141 along with highly conserved Mn specific residues. The isolated gene shared 63.8% homology with MnSOD of bacteria confirmed it as Mn isoform. This is the hitherto report on SOD gene from marine cyanobacterium, L. valderiana BDU20041 of Indian subcontinent.ConclusionGeneration of Reactive Oxygen Species (ROS) coupled with induction of SOD by marine cyanobacterium, L. valderiana BDU20041 was responsible for alleviating stress caused by an azo dye, C. I. Acid Black 1. The partial SOD gene has been sequenced and based on the active site, motif and metal specific residues; it has been identified as Mn metalloform.

Inflammatory biomarkers in coronary artery disease

Current evidence supports that inflammation is a major driving force underlying the initiation of coronary plaques, their unstable progression, and eventual disruption; patients with a more pronounced vascular inflammatory response have a poorer outcome. Biomarkers are generally considered to be proteins or enzymes - measured in serum, plasma, or blood - that provide independent diagnostic and prognostic value by reflecting an underlying disease state. In the case of coronary artery disease (CAD), inflammatory biomarkers, have been extensively investigated; more evidence exists for C-reactive protein (CRP). Using high sensitivity (hs) assays, epidemiologic data demonstrate an association between hs-CRP and risk for future cardiovascular morbidity and mortality among those at high risk or with documented CAD. Moreover, a series of prospective studies provide consistent data documenting that mild elevation of baseline levels of hs-CRP among apparently healthy individuals is associated with higher long-term risk for cardiovascular events. Yet, the predictive value of hs-CRP is found to be independent of traditional cardiovascular risk factors. Recent studies suggest that, besides CRP, other inflammatory biomarkers such as cytokines [interleukin (IL)-1, IL-6, IL-8, monocyte chemoattractant protein-1 (MCP-1)], soluble CD40 ligand, serum amyloid A (SAA), selectins (E-selectin, P-selectin), myeloperoxidase (MPO), matrix metalloproteinases (MMPs), cellular adhesion molecules [intercellular adhesion molecule 1 (ICAM-1), vascular adhesion molecule 1 (VCAM-1)], placental growth factor (PlGF) and A(2) phospholipases may have a potential role for the prediction of risk for developing CAD and may correlate with severity of CAD. Finally, indications suggest that the increased risk associated with inflammation may be modified with certain preventive therapies and biomarkers may help to identify the individuals who would benefit most from these interventions.

Polyphenols from Honeybush tea (Cyclopia intermedia).

The fermented leaves and stems of Cyclopia intermedia are used to brew Honeybush tea, a herbal tea indigenous to South Africa. The plant is also used to manufacture a sweet herbal infusion used for restorative properties such as soothing coughs and alleviating bronchial complaints including tuberculosis, pneumonia, and catarrh. It is claimed to have a low tannin content and no caffeine and contains various antioxidants. Continued investigations into the phenolic content of the leaves and stems of C. intermedia yielded tyrosol and a methoxy analogue, 2-[4-[O-alpha-apiofuranosyl-(1' '-->6')-beta-d-glucopyranosyloxy]phenyl]ethanol, 4-[O-alpha-apiofuranosyl-(1' '-->2')-beta-d-glucopyranosyloxy]benzaldehyde, five glycosylated flavonols, two isoflavones, four flavanones, two isoflavones, and two flavones. Structure elucidation was done by NMR, CD, and MS methods. Because flavonoids are presumed to contribute significantly toward the scavenging effects of active oxygen species, our results indicate that the tentative claimed health-promoting properties may be attributed to the presence of these and other phenolics in C. intermedia.

Effect of active oxygen species on the productivity of torularhodin by Rhodotorula glutinis No. 21

The effect of active oxygen species on the productivity of torularhodin, an effective antioxidant, by soil yeast, Rhodotorula glutinis no. 21, was examined. Methylene blue, methyviologen and AAPH [2,2′-azobis(2-amidinopropane) dihydrochloride] were used as generators of singlet oxygen, superoxide anion radicals and peroxy radicals, respectively. All of them indicated effectiveness at a dose of 1.0 x 10 −10 to 3.0 x 10 −6 M. Addition of these generators to the culture medium had almost no influence on the biosynthesis of β-carotene, whereas it had marked enhancement on those of torulene and torularhodin. Production of uric acid by xanthineoxidase remained unchanged at a torularhodin concentration of up to 100 μM. This result suggests that torularhodin does not directly affect the productivity of superoxide anions. It has been proved that torularhodin has a more potent effect on the scavenging of peroxyl radicals and inhibits substrate degradation by singlet oxygen more effectively than β-carotene does. Continuous addition of methylene blue enhanced the torularhodin accumulation more markedly than single addition.

Effect of Active Oxygen Species on the Productivity of Torularhodin by Rhodotorula glutinis No. 21

The effect of active oxygen species on the productivity of torularhodin, an effective antioxidant, by soil yeast, Rhodotorula glutinis no. 21, was examined. Methylene blue, methyviologen and AAPH [2,2'-azobis(2-amidinopropane) dihydrochloride] were used as generators of singlet oxygen, superoxide anion radicals and peroxy radicals, respectively. All of them indicated effectiveness at a dose of 1.0 x 10(-10) to 3.0 x 10(-6) M. Addition of these generators to the culture medium had almost no influence on the biosynthesis of beta-carotene, whereas it had marked enhancement on those of torulene and torularhodin. Production of uric acid by xanthineoxidase remained unchanged at a torularhodin concentration of up to 100 microM. This result suggests that torularhodin does not directly affect the productivity of superoxide anions. It has been proved that torularhodin has a more potent effect on the scavenging of peroxyl radicals and inhibits substrate degradation by singlet oxygen more effectively than beta-carotene does. Continuous addition of methylene blue enhanced the torularhodin accumulation more markedly than single addition.

Synergistic effect of active oxygen species and alginate on chitinase production by Wasabia japonica cells and its application

The specific chitinase productivity of a Wasabia japonica cell suspension culture under pure oxygen aeration was 3.8 times higher than that of a suspension culture aerated with ordinary air. During aeration with pure oxygen, both oxygen consumption by the cells and the H 2O 2 concentration in the medium increased. Addition of H 2O 2 to the cultivation medium also promoted the specific chitinase productivity. H 2O 2 could pass freely through the cell membrane. It was assumed that the excess oxygen was converted into active oxygen species such as H 2O 2, and that the promotion of chitinase production was probably due to the generated active oxygen species. Addition of alginate oligomer (AO, an endogenous elicitor-like substance) to cultures aerated with pure oxygen or supplemented with H 2O 2 resulted in synergistic increases in chitinase production. Based on these results, the development of a simple and efficient chitinase production system was investigated. Cells were immobilized in alginate gel (instead of adding AO to the medium) and cultivated in a medium containing H 2O 2. The specific chitinase productivity increased to the levels observed in the suspension culture system. During repeated batch cultivation of immobilized cells, the chitinase production remained stable for three repeated batches. When immobilized protoplasts were cultivated in a medium containing H 2O 2, there was 7-fold increase in chitinase production compared with that of immobilized cells.

Effects of active oxygen species and methyl jasmonate on expression of the gene for a wound-inducible 1-aminocyclopropane-1-carboxylate synthase in winter squash ( Cucurbita maxima )

The production of ethylene by mesocarp tissue of winter squash is induced by mechanical wounding. We isolated a cDNA fragment (WSACS2) from the wounded mesocarp tissue of winter-squash (Cucurbita maxima Duch.) fruit by the reverse transcriptase-polymerase chain reaction using primers that corresponded to two conserved amino acid sequences found in 1-aminocyclopropane-1-carboxylate (ACC) synthases. The sequence of WSACS2 cDNA was almost identical to that of the cDNA for a wound-inducible ACC synthase (CM-ACS1) reported by N. Nakajima et␣al. 1990, Plant Cell Physiol 31: 1021–1029, with only one nucleotide difference. The expression of the WSACS2 transcript in tissue disks was examined by Northern blotting analysis. The accumulation of WSACS2 mRNA in the wounded tissue disks was inhibited by treatment with diphenyleneiodonium, which is known as an inhibitor of superoxide-generating NAD(P)H oxidases. The addition of xanthine and xanthine oxidase, which form a superoxide-generating system, strongly induced the accumulation of the WSACS2 transcript within 30 min and the level of the transcript fell thereafter. We measured levels of active oxygen species that were generated in the tissue disks by using chemiluminescence reagents and found that the chemiluminescence signal became apparent within a few seconds after wounding. The synthesis of the WSACS2 transcript was slightly inhibited by inhibitors of lipoxygenase, which is involved in the synthesis of jasmonic acid. Methyl jasmonate stimulated the synthesis of the WSACS2 transcript in the tissue disks.

Effect of active oxygen species on intimal proliferation in rat aorta after arterial injury.

Proliferation and migration of vascular smooth-muscle cells (VSMCs) are essential events in neointimal hyperplasia. Recent findings that active oxygen species induce pro-oncogene expression, and stimulate VSMC DNA synthesis and cell division, suggest that active oxygen species may play an important role in intimal proliferation after arterial injury. To determine how the redox state of the artery was altered by injury, the levels of thiobarbituric-acid-reactive substances (TBARs, markers of lipid oxidation) and glutathione peroxidase (GSH-PX, the enzyme responsible for the production of glutathione, a major intracellular antioxidant) were measured in the rat aorta after balloon injury. There was an inverse relationship between the level of TBARs, which increased significantly to a maximum 17% greater than normal at 10 days after injury (p<0.05, n=7), and the activity of GSH-PX, which decreased significantly to a minimum 36.5% less than normal at 10 days after injury (p<0.05, n=7). To determine whether maintaining a more reduced vessel environment would inhibit intimal proliferation, L-cysteine was administered by intraperitoneal injection from 3 days before to 14 days after balloon injury. At 14 days after the arterial injury, the aorta was harvested for histological and morphometric measurements of TBARs and GSH-PX. At 7 and 14 days after balloon injury, the aorta was harvested for [3H]thymidine incorporation studies. Efficacy of therapy was demonstrated by a significant decrease in the level of TBARs and an increase in the activity of GSH-PX (p<0.05 vs. the control group, n=7). In the L-cysteine group, all parameters of intimal proliferation were significantly reduced compared to the controls, including the intima:media ratio (0.091 vs. 0.253, p<0.05); the cross-sectional area of the neointima (0.0563 compared to 0.140 mm2, p<0.05), coverage of the internal elastic lamina by the neointima (23.44 compared to 43.27%, p<0.05), and [3H]thymidine incorporation (at 7 days 240.58 vs 350.68 cpm/mg tissue, p<0.05; at 14 days 181.71 vs. 275.30 cpm/mg tissue, p<0.05). These results demonstrate dynamic alterations in the vessel redox state after arterial injury, and suggest that maintaining a more reduced environment (e.g. administration of L-cysteine) will reduce intimal proliferation after arterial injury.

Biological effects of active oxygen on an X-ray-sensitive mutant mouse cell line (SL3-147).

The biological effects of active oxygen species were examined in a mutant mouse cell line (SL3-147) that is deficient in the repair of DNA double-strand breaks. This mutant cell line shows different sensitivities to X-rays, hydrogen peroxide, paraquat and menadione when compared to the wild-type cell line (LTA). SL3-147 was more sensitive to X-rays, hydrogen peroxide and paraquat, but was less sensitive to menadione in side by side comparisons to LTA cells. The greater number of DNA double-strand breaks in SL3-147 appears to account for the line's greater sensitivity to X-rays and paraquat. DNA damage other than double-strand breaks or injury to non-DNA targets, however, is responsible for the differences between LTA and SL3-147 in their sensitivities to hydrogen peroxide and menadione.

The loss of in vivo activity of recombinant human erythropoietin by active oxygen species.

The effects of active oxygen species on the in vivo activity of recombinant human erythropoietin (EPO) treated by Fenton system, xanthine (X) plus xanthine oxidase (XO) system and hydrogen peroxide (H2O2) has been studied by means of counting the increase in number of hemolyser-resistant cells (HRCs) in EPO-injected mice. The results showed that both Fenton and X plus XO systems caused a significant reduction of the activity in proportion to the concentration of generated active oxygen species. Meanwhile, the treatment of EPO with H2O2 alone resulted in a relatively slight reduction of the activity. Electrophoretic studies on the structure of EPO revealed that its main protein band on sodium dodecyl sulfate-polyacrylamide gel (SDS-PAGE) disappeared in proportion with the extent of exposure to active oxygen generating systems. Both Fenton and X plus XO systems caused a significant loss of fluorescence in the pyridylamino (PA-) sugar chain in proportion to the concentration of generated active oxygen species, and no degradation products in the sugar chain part of the PA-sugar chain were detected. This showed that aromatic groups in EPO were sensitive to attack by active oxygen species. These results provide evidence that hydroxyl radical and other active oxygen species have a potential to react with EPO, leading to a reduction of its in vivo activity.

Genetic and biochemical analysis of glutathione-S-transferase in the oxygen defense system of Drosophila melanogaster.

Aerobic organisms possess an array of enzymatic defense mechanisms against the toxic effects of active oxygen species. These include CuZn superoxide dismutase (CuZn SOD), catalase (CAT), and glutathione peroxidase (GPOX). Insects, however, lack an independent GPOX enzyme and instead rely on the activity of the more general detoxification enzyme, glutathione-S-transferase (GST), to carry out a peroxidase function. We report here the developmental profile of GST in Drosophila melanogaster and show that GST is induced by paraquat, a known free-radical generating agent. We also report that glutathione (GSH) depletion induced by administration of buthionine sulfoximine (BSO) selectively reduces the viability of mutants lacking CuZnSOD. By measuring GST specific activity in flies carrying deficiencies for the 87B region, we confirm an earlier report that this region contains active GST-encoding genes. Finally, through a biochemical analysis of representative alleles of known lethal complementation gene. The implications of these findings to the role of GSH and GST in D. melanogaster oxygen defense are discussed.

Adenosine Contributes to Neutrophil-mediated Loss of Myocardial Function in Post-ischemic Guinea-pig Hearts

Tissue injury associated with myocardial ischemia is assumed to largely result from the toxic effects of active oxygen species generated by accumulated polymorphonuclear leukocytes (PMNs). Recent reports have indicated that adenosine can interfere with the PMN function in vitro. The potential of adenosine to influence PMN-mediated myocardial tissue injury was assessed using a model of ischemia-reperfusion injury developed in the isolated working guinea-pig heart perfused with homologous PMNs. After an initial work phase, hearts were subjected to 30 min low-flow ischemia (1 ml/min) in the absence and presence of PMNs. Work was resumed after 15 min reperfusion in a non-working mode (Langendorff). Adenosine in the coronary effluent reached a maximum of 0.2 μM during low-flow ischemia. Recoveries of external heart work and cardiac output were reduced from about 80% to about 40% by PMNs. Infusion of adenosine deaminase (ADA, 5 U/ml), theophylline (50 μM) or the selective A 1-antagonist dipropyl-8-cyclopentylxanthine (0.1 μM) prevented this effect. Furthermore, application of adenosine (0.1 μM) in combination with PMNs also resulted in a loss of pump function, even in the absence of a direct ischemic stimulus. The data indicate that adenosine contributes to post-ischemic, PMN-mediated damage in the isolate working guinea-pig heart model by a receptor-mediated action.

Glutathione, a regulator of cell functions.

Glutathione(GSH)has various important roles in maintaining cell functions. It is a major non-protein tissue thiol, supporting the redox potential of cells, regulating enzyme activities, processing endo-or exogenous substances and protecting cells from deleterious effects of active oxygen species and free radicals. GSH has been shown to be a regulator of gluconeogenesis, glycolysis or cholesterol biosynthesis, and, more recently, a component of several autacoids. Many reactive xenobiotics bind to SH-group of GSH and are transported out of the cells. Elevation of circular or tissue GSH levels is often effective for preventing the development of toxic and pathological processes. Various attempts have been made to explore the defense action of GSH, but our studies have led to an interesting finding that prostaglandin A2 (PGA 2) promotes biosynthesis of GSH and protects cells from oxidative stress. The finding suggests that PGA2 may be involved in regulating the physiological functions of cells by affecting the GSH status. The extension of work on the GSH status should reveal answers to the elaborate mysteries of bioorganisms.