Molecular epidemiology of norovirus from patients with acute gastroenteritis in northwestern Spain.

The high incidence of norovirus (NoV) infections seems to be related to the emergence of new variants that evolved by genetic drift of the capsid gene. In this work, that represents a first effort to describe the molecular epidemiology of NoV in the northwest of Spain, a total of eight different NoV genotypes (GII.1, GII.3, GII.4, GII.6, GII.7, GII.12, GII.13, GII.14) were detected. The major genotypes observed were GII.4 (45·42%) and GII.14 (34·9%), being detected in all age groups. In addition, and although most of GII.4 sequences belonged to 2006b (7·2%) and 2010 (50·35%) variants, the presence of new NoV variants was observed. Phylogenetic analysis revealed that a high number of GII.4 sequences (35·24%) could be assigned to the newly emerging Sydney 2012 variant, even during late 2010. The high prevalence of NoV GII.14 observed in this study may indicate the emergence of this genotype in Spain.

Genotyping of hepatitis A virus detected in bivalve shellfish in Galicia (NW Spain).

Hepatitis A virus (HAV) represents a significant public health problem due to its high persistence in the environment and its transmission through contaminated water and food. Bivalve shellfish are filter feeders that can bioaccumulate human pathogens found in contaminated waters, their consumption being a potential cause of hepatitis A outbreaks. In this work, cultured and wild bivalve shellfish from the Ría de Vigo (Galicia, NW Spain) were analysed for the presence and genotyping of HAV. A total of 160 shellfish samples were collected between March 2004 and December 2006, including 68 samples from cultured mussels (Mytilus galloprovincialis), 30 from wild clams (Rupitapes decussatus), 31 from wild cockles (Cerastoderma edule) and 31 from wild mussel. HAV detection, carried out by quantitative RT-PCR, was positive for 29 (42.6%) cultured and 40 (43.5%) wild samples, with levels ranging from 3.1 x 10(2) and 1.4 x 10(10) RNA copies/g of shellfish digestive tissue. The phylogenetic analysis of VP1-P2A and VP3-VP1 regions, separately or as concatenated sequences, revealed that all HAV strains analysed belong to subgenotype IB. These results indicate a high prevalence of this subgenotype in the area studied.

Oxidative inhibition of red blood cell ATPases by glyceraldehyde.

Glyceraldehyde and other simple monosaccharides autoxidize under physiological conditions, forming dicarbonyl compounds and hydrogen peroxide via intermediate free radicals. These products may have deleterious effects on cell components. In this paper we study the effect of glyceraldehyde autoxidation on red-cell ATPase activities. The autoxidation of glyceraldehyde in imidazole-glycylglycine buffer, measured by oxygen consumption, depends on the buffer concentration and decreases in the presence of superoxide dismutase and catalase. The addition of DETAPAC inhibits the autoxidation almost completely. When human red-blood-cell membranes are incubated with glyceraldehyde, the red-blood-cell ATPase activities decrease significantly. The addition of DETAPAC, GSH and DTE (dithioerythritol) protects the enzyme from inactivation, but superoxide dismutase and catalase have no effect. Methylglyoxal (a dicarbonyl which is analogous to hydroxypyruvaldehyde derived from glyceraldehyde autoxidation) proved to have a powerful inhibitory action on ATPase activities. The addition of DTE completely protects the enzyme from inactivation, suggesting that the sulphydryl groups of the active site of the enzyme are the critical targets for dicarbonyl compounds.

Different effects of thiol and nonthiol ace inhibitors on copper-induced lipid and protein oxidative modification.

Differences among angiotensin-converting enzyme inhibitors (ACEI) in scavenging reactive oxygen species were described and mainly attributed to the presence or absence of a thiol group. Plasma constituents and red cells are known targets for oxidative damage. Transition metals, like copper, are well known catalizers of free radical generation. In the present study we compared the abilities of captopril (a thiol ACEI), enalaprilat, and lisinopril (two nonthiol ACEI) for inhibiting copper-induced thiobarbituric acid reactive substances (TBARS) formation and fluorescence generation in whole human plasma and low-density lipoprotein. The effects of those ACEI on copper/hydrogen peroxide-induced fluorescence development and electrophoretic mobility modification in albumin and on copper-induced TBARS formation and hemolysis in human red cells were also compared. Captopril was more effective than the two nonthiol ACEI in inhibiting plasma and LDL lipid peroxidation, but it was ineffective in inhibiting the albumin oxidative modification that was moderately inhibited by enalaprilat and lisinopril. On the contrary, the inhibitory effects of the three ACEI on copper-induced lipid peroxidation and hemolysis in red cell suspensions were more uniform. This as yet unreported red cell protective effect may deserve pharmacological evaluation. Our results show that captopril is a more effective antioxidant than the nonthiol ACEI in some systems. However, the nonthiol ACEI also have the ability to partially protect some targets against oxidative damage. These observations suggest that the presence of a thiol group in the ACEI structure is not the only determinant for the antioxidant properties.

Scavenging of reactive oxygen species by silibinin dihemisuccinate.

Silibinin dihemisuccinate (SDH) is a flavonoid of plant origin with hepatoprotective effects which have been partially attributed to its ability to scavenge oxygen free radicals. In the present paper the antioxidant properties of SDH were evaluated by studying the ability of this drug to react with relevant biological oxidants such as superoxide anion radical (O2-), hydrogen peroxide (H2O2), hydroxyl radical (HO.) and hypochlorous acid (HOCl). In addition, its effect on lipid peroxidation was investigated. SDH is not a good scavenger of O2- and no reaction with H2O2 was detected within the sensitivity limit of our assay. However, it reacts rapidly with HO. radicals in free solution at approximately diffusion-controlled rate (K = (1.0-1.2) x 10(10)/M/sec) and appears to be a weak iron ion chelator. SDH at concentrations in the micromolar range protected alpha 1-antiproteinase against inactivation by HOCl, showing that it is a potent scavenger of this oxidizing species. Luminol-dependent chemiluminescence induced by HOCl was also inhibited by SDH. The reaction of SDH with HOCl was monitored by the modification of the UV-visible spectrum of SDH. The studies on rat liver microsome lipid peroxidation induced by Fe(III)/ascorbate showed that SDH has an inhibitory effect, which is dependent on its concentration and the magnitude of lipid peroxidation. This work supports the reactive oxygen species scavenger action ascribed to SDH.

The inhibition of lipid peroxidation by cinnarizine. Possible implications to its therapeutic and side-effects.

Cinnarizine has antivasoconstrictor properties and improves red-cell deformability. Its major side-effects are the induction of extrapyramidal reactions. It is a calcium antagonist, but it was suggested that its effects may depend on other mechanisms, namely on antiperoxidant properties. We have studied these properties in different biological systems, intact red-cells included. The occurrence of lipid peroxidation was determined by the formation of 2-thiobarbituric acid reactive products. Cinnarizine was found to inhibit spontaneous lipid peroxidation in rat liver homogenates, copper-induced lipid peroxidation in human plasma and copper-induced and hydrogen peroxide-induced lipid peroxidation in human red-cells. In red-cells, the inhibition of lipid peroxidation is accompanied by the inhibition of hemolysis. Copper-induced red-cell lipid peroxidation is 85% inhibited by as little as 5 microM cinnarizine. The antioxidant activity of cinnarizine may contribute to explain some of the effects of this drug.

Protective effects of a 21-aminosteroid against copper-induced erythrocyte and plasma lipid peroxidation.

The 21-aminosteroids, or lazaroids, are a novel class of antioxidant drugs designed to inhibit iron-dependent lipid peroxidation in biological lipid environments. They have been shown to be of therapeutic value in several animal models of traumatic, ischemic and hemorrhagic injury of the central nervous system. Our purpose was to evaluate the ability of 21-aminosteroids to protect human erythrocytes and plasma against oxidative damage in vitro. We found that the 21-aminosteroid U74500A inhibited erythrocyte and plasma lipid peroxidation. U74500A at 1 microM significantly reduced copper-induced and hydrogen peroxide-induced erythrocyte lipid peroxidation by 76.5 and 27.6%, respectively. The inhibition of erythrocyte lipid peroxidation was accompanied by an inhibition of hemolysis. Copper-induced plasma lipid peroxidation was also significantly reduced by as little as 1 microM U74500A. These results suggest that 21-aminosteroids may prove useful in preventive or therapeutic interventions in situations where erythrocyte or plasma components are subjected to oxidative stress and in situations related to copper-induced oxidative damage.

Superoxide anion radical generation during the oxidation of various amines by diamine oxidase.

Diamine oxidase (DAO) or histaminase is an enzyme which deaminates histamine and several aliphatic amines to their corresponding aldehydes. Hydrogen peroxide and ammonia are side products of this reaction. The purpose of the present work was to evaluate if determination of produced hydrogen peroxide reflects DAO activity or if intermediate formation of the superoxide radical could be a reason for lack of correspondence between oxygen uptake and hydrogen peroxide production at different pH. Superoxide radical formation was determined by cytochrome c reduction in the presence and absence of superoxide dismutase (SOD). Oxygen uptake was measured with an oxygen electrode and hydrogen peroxide production by a spectrophotometric method. At pH 6.6 there was no superoxide production, but at pH 7.4 there was some, and it increased markedly at pH 9.5. Oxygen uptake also increased with increasing pH, especially with histamine as substrate. These results lead us to suggest that the mechanism of action of DAO involves the intermediate generation of superoxide radicals.

Early increase in histamine concentration in the islets of Langerhans isolated from rats made diabetic with streptozotocin.

Sprague-Dawley rats were separated in 4 groups. G1 received streptozotocin (ST). G2 received nicotinamide (NC) followed by ST. G3 was a NC control and G4 was a citrate control. The rats were sacrificed after 28 h and the islets isolated. Histamine and histaminase were determined. In the islets there was an increase in histamine content in G1 and a smaller increase in G2. The first two groups differ significantly and also in relation to the control groups. A decrease in islet histaminase does not seem responsible for the increased histamine, since group 2 (NC + ST) which had no diabetes, had a lower activity than group 1 (ST). It is suggested that histamine liberation by ST may be related to the diabetogenic effect of this drug.

Measurement of relative antioxidant activity of compounds: a methodological note.

The relative activities of some hydrogen-donating antioxidants were assessed by comparing their activities with that of Trolox (Trolox equivalent antioxidant capacity, TEAC) for scavenging the ABTS radical cation (ABTS.+) generated in the aqueous phase. We have verified, however, that TEAC values may change with the concentration of compounds and with the measuring times used. Not withstanding, TEAC values do not differ significantly if the compounds have kinetic curves of ABTS.+ formation similar to that of Trolox. This is the case with ascorbic acid, whose TEAC values, determined by using five concentrations at three different measuring times, are very close. For the flavonoids studied (catechin, rutin, naringenin and silibinin) which have kinetic curves of ABTS.+ formation different from that of Trolox, the TEAC values decrease with increasing concentrations of the compounds for each measuring time, and increase with increasing measuring times for each concentration. In the present study, we conclude that, in order to evaluate relative antioxidant activities of compounds by the ABTS assay, it is essential to perform kinetic studies to assess scavenging of ABTS.+ by these compounds. Therefore, when the TEAC values of compounds are determined for more than one measuring time, we may be sure that all the antioxidant potential of compounds is being considered and whether or not it is possible to establish a hierarchy for their antioxidant activities.

pH dependence of lipid peroxidation and albumin oxidative modification: possible implications to the pH paradox.

The hydrogen ion concentration may have an important influence in biological free radical reactions. We studied the effect of an acidic pH on two models of free radical-mediated damage: copper-induced lipid peroxidation in plasma and copper/hydrogen peroxide-induced oxidative modification of albumin. A reduction of pH from 7·4 to 6·6 decreased diene conjugation by 32%, thiobarbituric acid-reactive substances formation by 25% and fluorescence generation by 53% in plasma exposed to cupric chloride. At pH values lower than 6·6 an even greater inhibition of lipid peroxidation in plasma was obtained. Visible fluorescence development in albumin by exposure to site-specific generation of free radicals was also increasingly reduced by decreasing pH values. From pH 7·4 to 6·6 there was a 50% fluorescence generation inhibition. The observed partial protection of lipids and proteins against oxidative damage by an acidic pH alerts to the need for rigorously controlling the pH values when assaying compounds for antioxidant properties in vitro. It may also contribute to the explanation for the protective effect of an acidic pH against anoxic cell injury and for cell death that is precipitated by a rapid return to a normal pH following reperfusion (the 'pH paradox').

Effects of zinc on copper-induced and spontaneous lipid peroxidation.

Zinc (Zn) is an essential nonredox metal that has been regarded as having antioxidant properties. Some epidemiological indications and therapeutic results point to a role of Zn in restricting the development and the progression of some diseases. Redox-active metals like iron and copper are involved in oxidative injury mechanisms, and a decrease in the Zn:Cu ratio may be associated with certain pathologies. We studied the effect of Zn on the copper-induced lipid peroxidation in diluted human plasma. Lipid peroxidation was evaluated by measuring the formation of conjugated dienes and of thiobarbituric acid reactive products. We found that 20 microM Zn reduced the 125-microM copper-dependent formation of conjugated dienes by 27% and of thiobarbituric acid reactive products by 49%, during a 3-h incubation period. The inhibition of lipid peroxidation by 125 microM Zn is almost total in the same conditions. The time-course study of the inhibitory effect of 125 microM Zn showed that it lasted for 7 h, which was the maximum incubation period tested. We also found that Zn had an inhibitory effect on the spontaneous lipid peroxidation in rat brain whole homogenates. Our results support the antioxidant properties of Zn, which may be potentially relevant to the protection of human plasma constituents, competing with the transition metals for redox reactions.

Kinetics of the inhibition of xanthine dehydrogenase and of the reversible and irreversible forms of xanthine oxidase by silibinin and bendazac.

Xanthine oxidase exists in vivo predominantly as a NAD(+)-dependent dehydrogenase form (xanthine dehydrogenase) which can be transformed into oxygen-dependent oxidase forms as a result of sulfhydryl oxidation (reversible xanthine oxidase) or proteolysis (irreversible xanthine oxidase). Xanthine oxidase has been hypothesized to be a potential source of oxygen-derived free radicals during reperfusion of ischemic tissues. Xanthine dehydrogenase was purified from rat liver and converted into reversible xanthine oxidase by heating at 37 °C and into irreversible xanthine oxidase by proteolysis with trypsin. Silibinin and bendazac are compounds used in therapeutics and to which free radical scavenging properties were ascribed. The effects of the compounds silibinin and bendazac on the different forms of the enzyme were studied. Silibinin inhibited all the forms of the enzyme but bendazac inhibited only reversible and irreversible xanthine oxidase. The inhibitions seem to be mixed non-competitive-competitive. The authors discuss the hypothesis that selective inhibitors of xanthine oxidase, preventing the interruption of uric acid formation, may have some advantage over the inhibitors of both xanthine dehydrogenase and xanthine oxidase in the treatment and prevention of situations such as ischemia and reperfusion syndromes.

Ischemia, reperfusion and oxygen free radicals.

Alterations which occur during ischemia are reviewed. They modify the metabolic status in such a way they prepare the cell to an anomalous response to reoxygenation. The consequence of this disturbance is the generation of oxygen free radicals through several mechanisms, including the mitochondrial oxidative phosphorylation, the arachidonic acid cascade, the activation of xanthine oxidase, activation of phagocytes, iron mobilization, etc. Reduced glutathione is exhausted, proteins are inactivated. Lipid peroxidation induces membrane breakdown and cellular death.

[The beta-adrenergic receptor]. / O receptor beta adrenérgico.

The Authors review the constitution and mechanism of action of the beta adrenergic receptor. It is part of a large family which includes visual pigments, muscarinic, serotonergic, olfactive and substance K receptors. Catecholamines given an electron to the receptor. It goes then successively to the alpha submit of Gs protein ant to adenylyl cyclase. The process of activation consists in a successive transfer of one electron.

[Antioxidant effect of drugs used in cardiovascular therapy]. / Efeito antioxidante de Fármacos usados em terapêutica cardiovascular.

Oxidative stress is implicated in the pathogenesis of various cardiovascular disorders. The knowledge, characterisation and comparison of the different antioxidant properties of some cardiovascular drugs might lead to new therapeutic approaches. Blood constituents are biological products easier to obtain. Their oxidative damage is accepted to be involved in several pathogenic pathways such as atherogenesis. Blood products can be utilised as in vitro models of macromolecular and cellular oxidative damage. It is well known that transition metals catalyse the generation of more damaging reactive oxygen species. We studied the antioxidant effect of four beta-adrenergic blocking agents (pindolol, propranolol, atenolol and metoprolol) and of one calcium channel antagonist (nifedipine), on the plasma oxidative damage induced by copper. Lipid peroxidation was evaluated by measuring fluorescent substances. We added the drugs to the assay system, before and after the induction of peroxidation by copper, to elucidate their ability to prevent and/or block lipid peroxidation. We observed that pindolol, propranolol and nifedipine have antioxidant properties, in the assay system, when added 15 min before the addition of copper. This effect is demonstrated by the delay in initiation and decrease in formation of lipid peroxidation products. Nifedipine also has a remarkable chain breaking effect. We compared the drugs with ascorbic acid in their relative antioxidant effect: nifedipine > ascorbate > pindolol > propranolol.

[Ketanserin as antioxidant]. / A cetanserina como antioxidante.

Free radicals have been related to the pathogenesis of some cardiovascular diseases. Several drugs used to treat these diseases were shown to have antioxidant properties. Our purpose was to evaluate if ketanserin, a selective S2 receptor antagonist with proven antihypertensive efficacy and which beneficially affects hemorheology, also is able to inhibit lipid peroxidation. Lipid peroxidation was induced in different biological systems in vitro and evaluated by the formation of thiobarbituric acid-reactive products. Ketanserin 50 microM inhibited copper-dependent lipid peroxidation in human red-cell suspensions by 40.8% and the subsequent hemolysis by 57,3%. It was less efficient in inhibiting hydrogen peroxide-dependent lipid peroxidation and hemolysis in the same system. Ketanserin 100 microM inhibited lipid peroxidation induced by a mixture of copper(II) and hydrogen peroxide in hepatic microsomal suspensions and in brain total homogenates by 86.2% and 56.7%, respectively. These results proved an antioxidant effect for ketanserin which was unknown, although its therapeutic relevance remains undetermined.

[Stress]. / Stress.

The general adaptation syndrome is discussed on the light of recent discoveries on hypothalamic peptides and of their possible influence in survival and in induction of diseases. The problem of stress in alcoholism is reviewed. The author ends with a short souvenir of Hans Selye.

[Alcohol and free radicals. Various consequences: protein synthesis, endocrine disorders, immunity. Role of stress]. / Alcool e radicais livres. Algumas consequências: síntese proteica, disendocrinias, imunidade. Importância do stress.

Ethanol is a powerful generator of oxygen free radicals, when metabolized in the liver or in other organs. Isoenzyme 2E1 of cytochrome P450 and aldehyde oxidase are the main mechanisms for the generation of these radicals. A consequence of free radical generation is a decrease in protein synthesis. As a result we have endocrine and immunity alterations. The paper ords with a brief discussion of stress associated to alcoholism.

[Pathologic states related to alcoholism]. / Patologias relacionadas com o alcoolismo.

The Author reviews the pathological consequences of chronic alcoholism with particular emphasis to the consequences of malnutrition, especially vitamin B-1 deficiency, immune deficiencies, adaptation of cytochrome 2E1, testicular atrophy, alterations of the skin, bones, liver, muscles, cardiac and neuropsychiatric.