945 resultados para Reactive species of oxygen


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Studies of DNA damage in gastric epithelial cells of Helicobacter pylori (H. pylori)-infected patients are conflicting, possibly due to different methods used for scoring DNA damage by Comet assay. Therefore, we compared the sensitivity of visual microscopic analysis (arbitrary units-scores and comets%) and image analysis system (tail moment), in the gastric epithelial cells from the antrum and corpus of 122 H. pylori-infected and 32 non-infected patients. The feasibility of cryopreserved peripheral blood lymphocytes and whole-blood cells for DNA damage biomonitoring was also investigated. In the antrum, the levels of DNA damage were significantly higher in H. pylori-infected patients with gastritis than in non-infected patients with normal mucosa, when evaluated by image analysis system, arbitrary units and comets%. In the corpus, the comets% was not sufficiently sensitive to detect the difference between H. pylori-infected patients with gastritis and non-infected patients with normal mucosa. The image analysis system was sensitive enough to detect differences between non-infected patients and H. pylori-infected patients with mild gastritis and between infected patients with moderate and severe gastritis, in both antrum, and corpus, while arbitrary units and comets% were unable to detect these differences. In cryopreserved peripheral blood lymphocytes, the levels of DNA damage (tail moment) were significantly higher in H. pylori-infected patients with moderate and severe gastritis than in non-infected patients. Overall, our results indicate that the image analysis system is more sensitive and adequate to measure the levels of DNA damage in gastric epithelial cells than the other methods assayed. (c) 2005 Elsevier B.V. All rights reserved.

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Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

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En el paradigma clásico, los efectos biológicos de la radiación ionizante se atribuyen al daño en el ADN inducido en cada célula irradiada. La demostración de efectos de vecindad causados por radiación ionizante (EVIR) ha generado un cambio profundo en la concepción actual de la radiobiología. Los EVIR son aquellos efectos causados por la radiación que se producen en células que no han sido irradiadas. Diversos avances técnicos, en particular el empleo de microhaces, han permitido estudiar los EVIR in vitro. Se conocen dos vías por las cuales las células irradiadas pueden comunicarse con las no irradiadas, a saber: mediante uniones especializadas (nexos) que comunican los citoplasmas de células adyacentes, y mediante la secreción de factores solubles al medio extracelular. Estos factores incluyen varias citokinas y especies reactivas del oxígeno y nitrógeno. Las vías de señalización en las células afectadas involucran en particular la activación de proteína kinasas activadas por mitógenos (MAPK) y del factor de transcripción NFciclooxigenasa 2, sintasa de óxido nítrico 2 y NAD(P)H oxidasa. Los EVIR pueden causar mutaciones puntuales y cambios epigenéticos. Los efectos sobre las vías de señalización pueden persistir indefinidamente e incluso transmitirse a la descendencia. Paradójicamente, en ciertas condiciones los EVIR pueden ser adaptativos, es decir que tornan a las células afectadas más resistentes a la radiación. La adaptación exige síntesis de proteínas y mejora la capacidad celular de reparar el ADN y resistir el estrés oxidativo. Los EVIR también se han demostrado in vivo. Por tanto, pueden tener implicaciones importantes en radioterapia, tanto para mejorar la eficacia terapéutica como para reducir la incidencia de efectos adversos. Asimismo, su mejor conocimiento puede influenciar las normas internacionales de radioprotección.

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Iron and oxidative stress have a regulatory interplay. During the oxidative burst, phagocytic cells produce free radicals such as hypochlorous acid (HOCl). Nevertheless, scarce studies evaluated the effect of either iron deficiency anemia (IDA) or anemia of chronic disease (ACD) on phagocyte function in the elderly. The aim of the present study was to determine the oxidative burst, phagocytosis, and nitric oxide ((aEuro cent)NO) and HOCl, reactive species produced by monocytes and neutrophils in elderly with ACD or IDA. Soluble transferrin receptor, serum ferritin, and soluble transferrin receptor/log ferritin (TfR-F) index determined the iron status. The study was constituted of 39 patients aged over 60 (28 women and 11 men) recruited from the Brazilian Public Health System. Oxidative burst fluorescence intensity per neutrophil in IDA group and HOCl generation in both ACD and IDA groups were found to be lower (p < 0.05). The percentages of neutrophils and monocytes expressing phagocytosis in ACD group were found to be higher (p < 0.05). There was an overproduction of (aEuro cent)NO from monocytes, whereas the fundamental generation of HOCl appeared to be lower. Phagocytosis, oxidative burst, and (aEuro cent)NO and HOCl production are involved in iron metabolism regulation in elderly patients with ACD and IDA.

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Sera from patients infected with Taenia solium, Hymenolepis nana and Echinococcus granulosus were tested against homologous and heterologous parasite antigens using an ELISA assay, and a high degree of cross-reactivity was verified. To identify polypeptides responsible for this cross reactivity, the Enzyme Linked Immunoelectro Transfer Blot (EITB) was used. Sera from infected patients with T.solium, H.nana, and E.granulosus were assessed against crude, ammonium sulphate precipitated (TSASP), and lentil-lectin purified antigens of T.solium and crude antigens of.H.nana and E.granulosus. Several bands, recognized by sera from patients with T.solium, H.nana, and E.granulosus infections, were common to either two or all three cestodes. Unique reactive bands in H.nana were noted at 49 and 66 K-Da and in E.granulosus at 17-21 K-Da and at 27-32 K-Da. In the crude cysticercosis extract, a specific non glycoprotein band was present at 61-67 K-Da in addiction to specific glycoprotein bands of 50, 42, 24, 21, 18, 14, and 13 K-Da. None of the sera from patients with H.nana or E.granulosus infection cross reacted with these seven glycoprotein bands considered specific for T.solium infection.

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In the ecologically important arbuscular mycorrhizal fungi (AMF), Sod1 encodes a functional polypeptide that confers increased tolerance to oxidative stress and that is upregulated inside the roots during early steps of the symbiosis with host plants. It is still unclear whether its expression is directed at scavenging reactive oxygen species (ROS) produced by the host, if it plays a role in the fungus-host dialogue, or if it is a consequence of oxidative stress from the surrounding environment. All these possibilities are equally likely, and molecular variation at the Sod1 locus can possibly have adaptive implications for one or all of the three mentioned functions. In this paper, we analyzed the diversity of the Sod1 gene in six AMF species, as well as 14 Glomus intraradices isolates from a single natural population. By sequencing this locus, we identified a large amount of nucleotide and amino acid molecular diversity both among AMF species and individuals, suggesting a rapid divergence of its codons. The Sod1 gene was monomorphic within each isolate we analyzed, and quantitative PCR strongly suggest this locus is present as a single copy in G. intraradices. Maximum-likelihood analyses performed using a variety of models for codon evolution indicated that a number of amino acid sites most likely evolved under the regime of positive selection among AMF species. In addition, we found that some isolates of G. intraradices from a natural population harbor very divergent orthologous Sod1 sequences, and our analysis suggested that diversifying selection, rather than recombination, was responsible for the persistence of this molecular diversity within the AMF population.

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Background: The activation of hepatic stellate cells (HSCs) plays a pivotal role during liver injury because the resulting myofibroblasts (MFBs) are mainly responsible for connective tissue re-assembly. MFBs represent therefore cellular targets for anti-fibrotic therapy. In this study, we employed activated HSCs, termed M1-4HSCs, whose transdifferentiation to myofibroblastoid cells (named M-HTs) depends on transforming growth factor (TGF)-β. We analyzed the oxidative stress induced by TGF-β and examined cellular defense mechanisms upon transdifferentiation of HSCs to M-HTs. Results: We found reactive oxygen species (ROS) significantly upregulated in M1-4HSCs within 72 hours of TGF-β administration. In contrast, M-HTs harbored lower intracellular ROS content than M1-4HSCs, despite of elevated NADPH oxidase activity. These observations indicated an upregulation of cellular defense mechanisms in order to protect cells from harmful consequences caused by oxidative stress. In line with this hypothesis, superoxide dismutase activation provided the resistance to augmented radical production in M-HTs, and glutathione rather than catalase was responsible for intracellular hydrogen peroxide removal. Finally, the TGF-β/NADPH oxidase mediated ROS production correlated with the upregulation of AP-1 as well as platelet-derived growth factor receptor subunits, which points to important contributions in establishing antioxidant defense. Conclusion: The data provide evidence that TGF-β induces NADPH oxidase activity which causes radical production upon the transdifferentiation of activated HSCs to M-HTs. Myofibroblastoid cells are equipped with high levels of superoxide dismutase activity as well as glutathione to counterbalance NADPH oxidase dependent oxidative stress and to avoid cellular damage.

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The development of cancer in humans and animals is a multistep process. The complex series of cellular and molecular changes participating in cancer development are mediated by a diversity of endogenous and exogenous stimuli. One type of endogenous damage is that arising from intermediates of oxygen (dioxygen) reduction - oxygen-free radicals (OFR), which attacks not only the bases but also the deoxyribosyl backbone of DNA. Thanks to improvements in analytical techniques, a major achievement in the understanding of carcinogenesis in the past two decades has been the identification and quantification of various adducts of OFR with DNA. OFR are also known to attack other cellular components such as lipids, leaving behind reactive species that in turn can couple to DNA bases. Endogenous DNA lesions are genotoxic and induce mutations. The most extensively studied lesion is the formation of 8-OH-dG. This lesion is important because it is relatively easily formed and is mutagenic and therefore is a potential biomarker of carcinogenesis. Mutations that may arise from formation of 8-OH-dG involve GC. TA transversions. In view of these findings, OFR are considered as an important class of carcinogens. The effect of OFR is balanced by the antioxidant action of non-enzymatic antioxidants as well as antioxidant enzymes. Non-enzymatic antioxidants involve vitamin C, vitamin E, carotenoids (CAR), selenium and others. However, under certain conditions, some antioxidants can also exhibit a pro-oxidant mechanism of action. For example, beta-carotene at high concentration and with increased partial pressure of dioxygen is known to behave as a pro-oxidant. Some concerns have also been raised over the potentially deleterious transition metal ion-mediated (iron, copper) pro-oxidant effect of vitamin C. Clinical studies mapping the effect of preventive antioxidants have shown surprisingly little or no effect on cancer incidence. The epidemiological trials together with in vitro experiments suggest that the optimal approach is to reduce endogenous and exogenous sources of oxidative stress, rather than increase intake of anti-oxidants. In this review, we highlight some major achievements in the study of DNA damage caused by OFR and the role in carcinogenesis played by oxidatively damaged DNA. The protective effect of antioxidants against free radicals is also discussed.

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The alpha-aminoketone 1,4-diamino-2-butanone (DAB), a putrescine analogue, is highly toxic to various microorganisms, including Trypanosoma cruzi. However, little is known about the molecular mechanisms underlying DAB`s cytotoxic properties. We report here that DAB (pK(a) 7.5 and 9.5) undergoes aerobic oxidation in phosphate buffer, pH 7.4, at 37 degrees C, catalyzed by Fe(II) and Cu(II) ions yielding NH(4)(+) ion, H(2)O(2), and 4-amino-2-oxobutanal (oxoDAB). OxoDAB, like methylglyoxal and other alpha-oxoaldehydes, is expected to cause protein aggregation and nucleobase lesions. Propagation of DAB oxidation by superoxide radical was confirmed by the inhibitory effect of added SOD (50 U ml(-1)) and stimulatory effect of xanthine/xanthine oxidase, a source of superoxide radical. EPR spin trapping studies with 5,5-dimethyl-1-pyrroline-1-oxide (DMPO) revealed an adduct attributable to DMPO-HO(center dot), and those with alpha-(4-pyridyl-1-oxide)-N-tert-butylnitrone or 3,5-dibromo-4-nitrosobenzenesulfonic acid, a six-line adduct assignable to a DAB(center dot) resonant enoyl radical adduct. Added horse spleen ferritin (HoSF) and bovine apo-transferrin underwent oxidative changes in tryptophan residues in the presence of 1.0-10 mM DAB. Iron release from HoSF was observed as well. Assays performed with fluorescein-encapsulated liposomes of cardiolipin and phosphatidylcholine (20:80) incubated with DAB resulted in extensive lipid peroxidation and consequent vesicle permeabilization. DAB (0-10 mM) administration to cultured LLC-MK2 epithelial cells caused a decline in cell viability, which was inhibited by preaddition of either catalase (4.5 mu M) or aminoguanidine (25 mM). Our findings support the hypothesis that DAB toxicity to several pathogenic microorganisms previously described may involve not only reported inhibition of polyamine metabolism but also DAB pro-oxidant activity. (C) 2011 Elsevier Inc. All rights reserved.

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New technologies and sterilization agents for heat-sensitive materials are under intense investigation. Plasma sterilization, an atoxic low-temperature substitute for conventional sterilization, uses various gases that are activated by an electrical discharge, generating reactive species that promote lethality in microorganisms. Here, assays were performed using pure O-2 and O-2 + H2O2 mixture gas plasmas against a standard load of Bacillus atrophaeus spores inoculated on glass carriers inside PVC catheters. The sterilization efficiency was studied as a function of plasma system (reactive ion etching or inductively coupled plasma), biological monitor lumen diameter, gas, radio frequency power, and sub-lethal exposition time. After sterilization, the biological monitors were disassembled and the surviving bacteria were grown in trypticase soy broth using the most probable number technique. Plasma antimicrobial activity depended on the catheter's internal diameter and radio frequency powers. The N-2 + H2O2 mixture exhibited higher microbial efficacy than pure N-2 in both plasma systems.

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The effects of a moderate electrical stimulation on superoxide and nitric oxide production by primary cultured skeletal muscle cells were evaluated. The involvement of the main sites of these reactive species production and the relationship between superoxide and nitric oxide production were also examined. Production of superoxide was evaluated by cytochrome c reduction and dihydroethidium oxidation assays. Electrical stimulation increased superoxide production after 1?h incubation. A xanthine oxidase inhibitor caused a partial decrease of superoxide generation and a significant amount of mitochondria-derived superoxide was also observed. Nitric oxide production was assessed by nitrite measurement and by using 4,5-diaminofluorescein diacetate (DAF-2-DA) assay. Using both methods an increased production of nitric oxide was obtained after electrical stimulation, which was also able to induce an increase of iNOS content and NF-?B activation. The participation of superoxide in nitric oxide production was investigated by incubating cells with DAF-2-DA in the presence or absence of electrical stimulation, a superoxide generator system (xanthinexanthine oxidase), a mixture of NOS inhibitors and SOD-PEG. Our data show that the induction of muscle contraction by a moderate electrical stimulation protocol led to an increased nitric oxide production that can be controlled by superoxide generation. The cross talk between these reactive species likely plays a role in exercise-induced maintenance and adaptation by regulating muscular glucose metabolism, force of contraction, fatigue, and antioxidant systems activities. J. Cell. Physiol. 227: 25112518, 2012. (c) 2011 Wiley Periodicals, Inc.

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Analysis of human serum reactivities to antigenic components of soluble Taenia solium metacestode proteins showed the predominant presence of determinants shared by T. solium, Echinococcus multilocularis and E. granulosus. Two polypeptides were demonstrated by SDS-PAGE and Western blot or enzyme-linked immunoelectrotransfer blot (EITB) assay to bind serum and CSF antibodies only from T. solium cysticercosis patients. These species-specific antigenic polypeptides focused between pH 4.6 and 3.9 after resolution by isoelectric focusing followed by EITB. The high species-specificity demonstrated by the present techniques offers the opportunity to confirm serologically an infection by T. solium metacestode.

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The stable isotope composition of one epifaunal and three infaunal benthic foraminiferal species of a sediment core from 1800 m water depth of the western Arabian Sea was determined to evaluate deepwater oxygenation, organic matter remineralization, and early diagenetic processes during the past 190,000 years. The d18O records reveal species-specific metabolic effects, susceptibility to changes in carbonate ion concentration, and supralysoclinal calcite dissolution. The foraminiferal d13C records reveal changes in the stable carbon isotope gradients of pore water dissolved inorganic carbon (d13CDIC) and in the microhabitat depth of infaunal species. Maximum d13CDIC offsets between bottom and pore waters ranged between mean values of 0.8 and 1.2% corresponding to estimates of deepwater oxygen concentration between approximately 1 and 2.7 ml/l. Intervals of improved deepwater oxygenation coincided with high benthic foraminiferal diversity and indicate the admixture of well-oxygenated deepwater masses during interglacials. During interglacial maxima the d13C difference between epifauna and shallow infauna indicates highest organic matter remineralization rates at times of maximum organic matter fluxes.