Influence of glutathione-related genetic variants on the oxidative stress profile of Mexican patients with psychotic disorders

Objective: The clinical trajectories of patients with psychotic disorders have divergent outcomes, which may result in part from glutathione (GSH)-related high-risk genotypes. We aimed to determine pharmacokinetics of clozapine, GSH levels, GSH peroxidase (GPx) activity, gene variants involved in the synthesis and metabolism of GSH, and their association with psychotic disorders in Mexican patients on clozapine monotherapy and controls. Methods: The sample included 75 patients with psychotic disorders on clozapine therapy and 40 paired healthy controls. Plasma clozapine/N-desmethylclozapine, GSH concentrations, and GPx activity were determined, along with genotyping of GCLC and GSTP1 variants and copy number variations of GSTP1, GSTT1, and GSTM1. Clinical, molecular and biochemical data were analyzed with a logistic regression model. Results: GSH levels were significantly reduced and, conversely, GPx activity was higher among patients than controls. GCLC_GAG-7/9 genotype (OR = 4.3, 95%CI = 1.40-14.31, p = 0.019) and hetero-/homozygous genotypes of GCLC_rs761142 (OR = 6.09, 95%CI = 1.93-22.59, p = 0.003) were found to be risk factors for psychosis. The genetic variants were not related to clozapine/N-desmethylclozapine levels or metabolic ratio. Conclusions: GCLC variants were associated with the oxidative stress profile of patients with psychotic disorders, raising opportunities for intervention to improve their antioxidant defenses. Further studies with larger samples should explore this proposal.

Influence of glutathione-related genetic variants on the oxidative stress profile of Mexican patients with psychotic disorders.

OBJECTIVE: The clinical trajectories of patients with psychotic disorders have divergent outcomes, which may result in part from glutathione (GSH)-related high-risk genotypes. We aimed to determine pharmacokinetics of clozapine, GSH levels, GSH peroxidase (GPx) activity, gene variants involved in the synthesis and metabolism of GSH, and their association with psychotic disorders in Mexican patients on clozapine monotherapy and controls. METHODS: The sample included 75 patients with psychotic disorders on clozapine therapy and 40 paired healthy controls. Plasma clozapine/N-desmethylclozapine, GSH concentrations, and GPx activity were determined, along with genotyping of GCLC and GSTP1 variants and copy number variations of GSTP1, GSTT1, and GSTM1. Clinical, molecular and biochemical data were analyzed with a logistic regression model. RESULTS: GSH levels were significantly reduced and, conversely, GPx activity was higher in PD patients compared to controls. GCLC_GAG-7/9 genotype (OR=4.3, CI95=1.40-14.31, p=0.019) and hetero-/homozygous genotypes of GCLC_rs761142 (OR=6.09, CI95=1.93-22.59, p=0.003) were found as risk factors for psychosis. The genetic variants were not related to clozapine/N-desmethylclozapine levels or to metabolic ratio. CONCLUSIONS: GCLC variants were associated with the oxidative stress profile of PD patients raising opportunities for intervention to improve their antioxidant defenses. Further studies with larger samples should explore this proposal.

Salicylic Acid Improves Antioxidant Defense System and Photosynthetic Performance in Aristotelia chilensis Plants Subjected to Moderate Drought Stress.

Salicylic acid (SA) has been shown to ameliorate drought stress. However, physiological and biochemical mechanisms involved in drought stress tolerance induced by SA in plants have not been well understood. Thus, this study aimed to study the role of SA application on enzymatic and non-enzymatic antioxidants, photosynthetic performance, and plant growth in A. chilensis plants subjected to moderate drought stress. One-year-old A. chilensis plants were subjected to 100% and 60% of field capacity. When plants reached moderate drought stress (average of stem water potential of -1.0 MPa, considered as moderate drought stress), a single SA application was performed on plants. Then, physiological and biochemical features were determined at different times during 14 days. Our study showed that SA application increased 13.5% plant growth and recovered 41.9% AN and 40.7% gs in drought-stressed plants on day 3 compared to drought-stressed plants without SA application. Interestingly, SOD and APX activities were increased 85% and 60%, respectively, in drought-stressed SA-treated plants on day 3. Likewise, SA improved 30% total phenolic content and 60% antioxidant capacity in drought-stressed A. chilensis plants. Our study provides insight into the SA mechanism to tolerate moderate drought stress in A. chilensis plants.

Degradation of Acid Red 1 Catalyzed by Peroxidase Activity of Iron Oxide Nanoparticles and Detected by SERS.

Magnetic iron oxide nanoparticles (MIONPs) were synthesized using tannic acid and characterized by Raman, FTIR, UV, and DRX spectroscopy. In a heterogeneous Fenton-like reaction, the catalytic peroxidase-like activity of MIONPs in the degradation of Acid Red 1 (AR 1) dye was investigated. TEM/STEM was used to determine the quasi-spherical morphology and particle size (3.2 nm) of the synthesized MIONPs. The XRD powder patterns were indexed according to the reverse spinel structure of magnetite, and SEM-EDS analysis confirmed their chemical composition. At pH = 3.5, the decomposition of H2O2 in hydroxyl radicals by MIONPs results in high AR 1 degradation (99%). This behavior was attributed to the size and surface properties of the MIONPs. Finally, the Surface Enhanced Raman Spectroscopy (SERS) technique detected intermediary compounds in the degradation process.

Electron microscopy cytochemistry and three-dimensional reconstruction of labeled structures in Trypanosoma cruzi.

Significant advances have occurred in the area of high-resolution scanning electron microscopy (SEM), especially related to methodologies that allow the observation of intracellular structures that are exposed either by successive abrasion with a gallium ion beam or by sectioning in epoxy-embedded cells. Images of series of successively exposed surfaces can then be rendered into 3D models. Here, we report our observations by combining this approach with classical cytochemical methods to facilitate the 3D reconstruction of labeled structures and organelles. We used epimastigotes of Trypanosoma cruzi whose endocytic pathway was labeled with horseradish peroxidase, followed by fixation and detection of the peroxidase activity using the classical diaminobenzidine-osmium method followed by incubation with thiocarbohydrazide, which increases the concentration of osmium at the sites where the enzyme is located as well as the contrast of lipid-containing structures. This procedure allows not only a better visualization of membranous structures and lipid inclusions but can also easily identify the endocytic tracer (HRP) inside the cell. All structures involved in the endocytic activity could be traced and reconstructed.

Quantification of carbonate radical formation by the bicarbonate-dependent peroxidase activity of superoxide dismutase 1 using pyrogallol red bleaching.

Carbonate radicals (CO3-) are generated by the bicarbonate-dependent peroxidase activity of cytosolic superoxide dismutase (Cu,Zn-SOD, SOD-1). The present work explored the use of bleaching of pyrogallol red (PGR) dye to quantify the rate of CO3- formation from bovine and human SOD-1 (bSOD-1 and hSOD-1, respectively). This approach was compared to previously reported methods using electron paramagnetic resonance spin trapping with DMPO, and the oxidation of ABTS (2,2-azino-bis(3-ethylbenzothiazoline)-6-sulfonic acid). The kinetics of PGR consumption elicited by CO3- was followed by visible spectrophotometry. Solutions containing PGR (5-200 µM), SOD-1 (0.3-3 µM), H2O2 (2 mM) in bicarbonate buffer (200 mM, pH 7.4) showed a rapid loss of the PGR absorption band centered at 540 nm. The initial consumption rate (Ri) gave values independent of the initial PGR concentration allowing an estimate to be made of the rate of CO3- release of 24.6 ±â€¯4.3 µM min-1 for 3 µM bSOD-1. Both bSOD-1 and hSOD-1 showed a similar peroxidase activity, with enzymatic inactivation occurring over a period of 20 min. The single Trp residue (Trp32) present in hSOD-1 was rapidly consumed (initial consumption rate 1.2 ±â€¯0.1 µM min-1) with this occurring more rapidly than hSOD-1 inactivation, suggesting that these processes are not directly related. Added free Trp was rapidly oxidized in competition with PGR. These data indicate that PGR reacts rapidly and efficiently with CO3- resulting from the peroxidase activity of SOD-1, and that PGR-bleaching is a simple, fast and cheap method to quantify CO3- release from bSOD-1 and hSOD-1 peroxidase activity.

Silicon does not alleviate the adverse effects of drought stress in soybean plants / O silício não alivia os efeitos adversos do estresse hídrico em plantas de soja

Beneficial effects of silicon (Si) in the plants growth under conditions of drought stress have been associated with to uptake and accumulation ability of element by different species. However, the effects of Si on soybean under water stress are still incipient and inconclusive. This study investigated the effect of Si application as a way to confer greater soybean tolerance to drought stress. The experiment was carried out in 20-L pots under greenhouse conditions. Treatments were arranged in a randomized block design in a 2 × 4 factorial: two water regimes (no stress or water stress) and four Si rates (0, 50, 100 and 200 mg kg–1). Soybean plants were grown until beginning flowering (R1) growth stage with soil moisture content near at the field capacity, and then it started the differentiation of treatments under drought by the suspension of water supply. Changes in relative water content (RWC) in leaf, electrolyte leakage from cells, peroxidase activity, plant nutrition and growth were measured after 7 days of drought stress and 3 days recovery. The RWC in soybean leaves decreased with Si rates in the soil. Silicon supply in soil with average content of this element, reduced dry matter production of soybean under well-irrigated conditions and caused no effect on dry matter under drought stress. The nitrogen uptake by soybean plants is reduced with the Si application under drought stress. The results indicated that the Si application stimulated the defense mechanisms of soybean plants, but was not sufficient to mitigate the negative effects of drought stress on the RWC and dry matter production.

Os efeitos benéficos do silício (Si) no crescimento das plantas sob condições de déficit hídrico têm sido associados com a capacidade de absorção e acumulação deste elemento pelas diferentes espécies de plantas. No entanto, os efeitos de Si na cultura da soja em condições de déficit hídrico ainda são incipientes e inconclusivos. Este estudo investigou o efeito da aplicação de Si, como forma de conferir maior tolerância da soja ao estresse hídrico. O experimento foi conduzido em vasos de 20 L em condições de casa de vegetação. Os tratamentos foram dispostos em um delineamento em blocos casualizados, em esquema fatorial 2 × 4: dois regimes hídricos (sem ou com estresse hídrico) e quatro doses de Si (0, 50, 100 e 200 mg kg–1). As plantas de soja foram cultivadas até o início do florescimento (R1) com a teor de umidade do solo próximo à capacidade de campo, e então começou a diferenciação dos tratamentos sob déficit hídrico mediante a suspensão do fornecimento de água. Alterações no conteúdo relativo de água (CRA), extravasamento de eletrólitos das células, atividade de peroxidase, nutrição e crescimento das plantas foram mensurados após 7 dias de expossição do déficit hídrico e após 3 dias de recuperação. O CRA das folhas de soja diminuiu com a aplicação das doses de Si no solo. A aplicação de Si em solo com teor médio desse elemento resultou na redução da produção de matéria seca da soja no tratamento sem déficit hídrico e não causou efeito na produção de matéria seca em condições de déficit hídrico. A absorção de nitrogênio pelas plantas de soja foi reduzida com a aplicação de Si em condições de déficit hídrico. Os resultados deste estudo indicaram que a aplicação de Si estimulou os mecanismos de defesa das plantas de soja, mas não foi suficiente para minimizar os efeitos negativos do estresse hídrico no CRA e na produção de matéria seca das plantas.

Silicon does not alleviate the adverse effects of drought stress in soybean plants / O silício não alivia os efeitos adversos do estresse hídrico em plantas de soja

Beneficial effects of silicon (Si) in the plants growth under conditions of drought stress have been associated with to uptake and accumulation ability of element by different species. However, the effects of Si on soybean under water stress are still incipient and inconclusive. This study investigated the effect of Si application as a way to confer greater soybean tolerance to drought stress. The experiment was carried out in 20-L pots under greenhouse conditions. Treatments were arranged in a randomized block design in a 2 × 4 factorial: two water regimes (no stress or water stress) and four Si rates (0, 50, 100 and 200 mg kg–1). Soybean plants were grown until beginning flowering (R1) growth stage with soil moisture content near at the field capacity, and then it started the differentiation of treatments under drought by the suspension of water supply. Changes in relative water content (RWC) in leaf, electrolyte leakage from cells, peroxidase activity, plant nutrition and growth were measured after 7 days of drought stress and 3 days recovery. The RWC in soybean leaves decreased with Si rates in the soil. Silicon supply in soil with average content of this element, reduced dry matter production of soybean under well-irrigated conditions and caused no effect on dry matter under drought stress. The nitrogen uptake by soybean plants is reduced with the Si application under drought stress. The results indicated that the Si application stimulated the defense mechanisms of soybean plants, but was not sufficient to mitigate the negative effects of drought stress on the RWC and dry matter production.(AU)

Os efeitos benéficos do silício (Si) no crescimento das plantas sob condições de déficit hídrico têm sido associados com a capacidade de absorção e acumulação deste elemento pelas diferentes espécies de plantas. No entanto, os efeitos de Si na cultura da soja em condições de déficit hídrico ainda são incipientes e inconclusivos. Este estudo investigou o efeito da aplicação de Si, como forma de conferir maior tolerância da soja ao estresse hídrico. O experimento foi conduzido em vasos de 20 L em condições de casa de vegetação. Os tratamentos foram dispostos em um delineamento em blocos casualizados, em esquema fatorial 2 × 4: dois regimes hídricos (sem ou com estresse hídrico) e quatro doses de Si (0, 50, 100 e 200 mg kg–1). As plantas de soja foram cultivadas até o início do florescimento (R1) com a teor de umidade do solo próximo à capacidade de campo, e então começou a diferenciação dos tratamentos sob déficit hídrico mediante a suspensão do fornecimento de água. Alterações no conteúdo relativo de água (CRA), extravasamento de eletrólitos das células, atividade de peroxidase, nutrição e crescimento das plantas foram mensurados após 7 dias de expossição do déficit hídrico e após 3 dias de recuperação. O CRA das folhas de soja diminuiu com a aplicação das doses de Si no solo. A aplicação de Si em solo com teor médio desse elemento resultou na redução da produção de matéria seca da soja no tratamento sem déficit hídrico e não causou efeito na produção de matéria seca em condições de déficit hídrico. A absorção de nitrogênio pelas plantas de soja foi reduzida com a aplicação de Si em condições de déficit hídrico. Os resultados deste estudo indicaram que a aplicação de Si estimulou os mecanismos de defesa das plantas de soja, mas não foi suficiente para minimizar os efeitos negativos do estresse hídrico no CRA e na produção de matéria seca das plantas.(AU)

Nanostructures for peroxidases.

Peroxidases are enzymes catalyzing redox reactions that cleave peroxides. Their active redox centers have heme, cysteine thiols, selenium, manganese, and other chemical moieties. Peroxidases and their mimetic systems have several technological and biomedical applications such as environment protection, energy production, bioremediation, sensors and immunoassays design, and drug delivery devices. The combination of peroxidases or systems with peroxidase-like activity with nanostructures such as nanoparticles, nanotubes, thin films, liposomes, micelles, nanoflowers, nanorods and others is often an efficient strategy to improve catalytic activity, targeting, and reusability.

Oxidation of the tryptophan 32 residue of human superoxide dismutase 1 caused by its bicarbonate-dependent peroxidase activity triggers the non-amyloid aggregation of the enzyme.

The role of oxidative post-translational modifications of human superoxide dismutase 1 (hSOD1) in the amyotrophic lateral sclerosis (ALS) pathology is an attractive hypothesis to explore based on several lines of evidence. Among them, the remarkable stability of hSOD1(WT) and several of its ALS-associated mutants suggests that hSOD1 oxidation may precede its conversion to the unfolded and aggregated forms found in ALS patients. The bicarbonate-dependent peroxidase activity of hSOD1 causes oxidation of its own solvent-exposed Trp(32) residue. The resulting products are apparently different from those produced in the absence of bicarbonate and are most likely specific for simian SOD1s, which contain the Trp(32) residue. The aims of this work were to examine whether the bicarbonate-dependent peroxidase activity of hSOD1 (hSOD1(WT) and hSOD1(G93A) mutant) triggers aggregation of the enzyme and to comprehend the role of the Trp(32) residue in the process. The results showed that Trp(32) residues of both enzymes are oxidized to a similar extent to hSOD1-derived tryptophanyl radicals. These radicals decayed to hSOD1-N-formylkynurenine and hSOD1-kynurenine or to a hSOD1 covalent dimer cross-linked by a ditryptophan bond, causing hSOD1 unfolding, oligomerization, and non-amyloid aggregation. The latter process was inhibited by tempol, which recombines with the hSOD1-derived tryptophanyl radical, and did not occur in the absence of bicarbonate or with enzymes that lack the Trp(32) residue (bovine SOD1 and hSOD1(W32F) mutant). The results support a role for the oxidation products of the hSOD1-Trp(32) residue, particularly the covalent dimer, in triggering the non-amyloid aggregation of hSOD1.

Protective effect of ((4-tert-butylcyclohexylidene) methyl) (4-methoxystyryl) sulfide, a novel unsymmetrical divinyl sulfide, on an oxidative stress model induced by sodium nitroprusside in mouse brain: involvement of glutathione peroxidase activity.

OBJECTIVES: In this study, the antioxidant action of ((4-tert-butylcyclohexylidene) methyl) (4-methoxystyryl) sulfide, a novel unsymmetrical divinyl sulfide, against oxidative damage induced by sodium nitroprusside (SNP) in brains of mice was investigated. METHODS: Mice received SNP (0.335 µmol/site, intracerebroventricular) 30 min after administration of sulfide (10 mg/kg, intragastrically). After 1 h, animals were sacrificed and the brains were removed to biochemistry analysis. Thiobarbituric acid reactive species (TBARS), protein carbonyl (PC) and non-protein thiol (NPSH) levels, as well as catalase (CAT), superoxide dismutase (SOD), glutathione peroxidase (GPx), glutathione reductase (GR) and glutathione S-transferase (GST) activities were determined. KEY FINDINGS: SNP increased TBARS and PC levels, CAT, GPx, GR and GST activities and reduced NPSH levels. Administration of the sulfide attenuated the changes produced by SNP and increased per se GPx activity in brains of mice. Toxicological parameters revealed that this compound did not cause acute renal or hepatic damage. CONCLUSIONS: In conclusion, ((4-tert-butylcyclohexylidene) methyl) (4-methoxystyryl) sulfide protected against oxidative damage caused by SNP in mouse brain. GPx activity is involved, at least in part, in the cerebral antioxidant action of this compound.

In vitro antibacterial activity of meclofenamate metal complexes with Cd(II), Pb(II), Co(II), and Cu(II). Crystal structures of [Cd(C14H10NO2Cl2)2∙(CH3OH)]n and [Cu(C14H10NO2Cl2)2(C5H5N)2].

The synthesis and characterization of five metal complexes derived from sodium meclofenamate (1) are reported: [Cd(C14H10NO2Cl2)2∙(CH3OH)]n∙nCH3OH (6), [Pb(C14H10NO2Cl2)2]n (7), [Co(C14H10NO2Cl2)]n (8), [Cu(C14H10NO2Cl2)]n (9), and [Cu(C14H10NO2Cl2)2(C5H5N)2] (10) (C14H10NO2Cl2=meclofenamate; C5H5N=pyridine). The characterization of the compounds was based on FTIR and UV-visible spectroscopy, mass spectrometry and, in the case of complexes 6 and 10, single crystal X-ray diffraction analysis. For compound 6, the structural analysis revealed a 1-D polymeric chain structure, in which pentagonal planar [Cd(RCOO)2(CH3OH)] units were linked through bridging carboxylate functions of the meclofenamate ligands. The overall coordination environment of the Cd(II) ions was seven-coordinate, since each carboxylate group exhibited a µ3-bridging coordination mode. On the other hand, for complex 10 a discrete mononuclear structure was observed, in which the six-coordinate copper(II) metal atoms were coordinated by two pyridine molecules and the carboxylate functions of two meclofenamate entities, in an anisobidentate coordination mode. The antibacterial activity of compounds 6-9 against four strains of Gram positive (Staphylococcus aureus and Bacillus subtilis) and Gram negative (Escherichia coli and Pseudomonas aeruginosa) bacteria was examined, finding that only complex 6 was active. Additionally, it was found that the Co(II) and Cu(II) complexes 8 and 9 showed peroxidase activity.

Efeito do gênero (masculino e feminino) sobre a cárie dentária e parâmetros salivares de crianças / Effect of gender on dental caries and salivary parameters of children

Objetivo: avaliar o índice de cárie dentária e parâmetros salivares comparando meninos e meninas.Métodos: a saliva total estimulada mecanicamente com parafilm foi coletada de 46 crianças saudáveis, com quatro a seis anos de idade, sendo 24 meninos e 22 meninas. As crianças foram divididas em subgrupos de acordo com o gênero (meninos e meninas) e presença de cárie dentária (cavidades de cárie, CC; livres de cárie, CF). A cárie dentária foi avaliada usando os critérios da OMS e Kappa=0,87. O grupo CC foi definido pela presença de no mínimo três superfícies com necessidade de restauração e o grupo CF pela ausência de dentes com lesões de cárie clinicamente detectáveis (ceo-s=o). Os parâmetros salivares avaliados foram o fluxo salivar, a concentração de proteína total e a atividade enzimática da peroxidase. Os resultados foram comparados por teste t de Student e Análise de Variância e teste de Tukey para contraste de média (p≤0,05). Resultados: crianças com cárie dentária apresentaram redução de 33% do fluxo salivar comparadas com crianças sem cárie dentária (p≤0,05). Entre as crianças com cárie dentária, as meninas apresentaram maior índice de cárie dentária comparadas com os meninos (98%, p≤0.05). As meninas apresentaram diferenças salivares mais pronunciadas comparados aos meninos, com maior concentração de proteína total e menor fluxo salivar e atividade da peroxidase (p≤0,05). Conclusão: o estudo sugere que o gênero pode influenciar o desenvolvimento de cárie dentária e parâmetros salivares de crianças...

Objective: To evaluate the dental caries index and salivary parametersin boys and girls. Method: Whole stimulated saliva by chewing Parafilm® was collected from 46 healthy children (24 boys and 22 girls) aged 4 to 6 years. The children were assigned to subgroups according to gender (boys andgirls) and dental caries (CC û caries cavities; CF û caries free). Dentalcaries was evaluated using the WHO criteria and kappa=0.87. The CC group was defined by the presence of at least three surfaces requiring restoration while the CF group was characterized by the absence of clinically detectable caries (ceo-s=0). The following salivary parameters were evaluated: salivary flow, total protein concentration andperoxidase enzymatic activity. The results were compared by theStudentÆs t-test, ANOVA and TukeyÆs multiple-comparison test (p≤0.05).Results: Children with dental caries presented a 33% reduction of salivary flow compared to the caries-free children (p≤0.05). Among the children with dental caries, the girls had a greater caries index than the boys (98%, p≤0.05). The girls presented more accentuated salivary parameters than boys, with greater total protein concentration, lower salivary flow and lower peroxidase activity (p≤0.05).Conclusion: This study suggests that gender might influence the development of dental caries and the salivary parameters in children...

Atividade peroxidásica da enzima superóxido dismutase 1 humana: produção do radical carbonato, dimerização covalente da enzima e implicações para a esclerose lateral amiotrófica / Peroxidase activity of human superoxide dismutase 1: production of the carbonate radical, covalent dimerization of the enzyme, and implications to amyotrophic lateral sclerosis

A esclerose lateral amiotrófica (ELA) é uma doença neurodegenerativa que afeta os neurônios motores levando a atrofia muscular e morte por insuficiência respiratória. Esta patologia se manifesta de forma esporádica ou familiar, que são indistinguíveis clinicamente. Mutações na enzima antioxidante superóxido dismutase 1 (hSod1) respondem por aproximadamente 20% dos casos familiares de ELA. Além disso, o caráter autossômico dominante destas mutações revela que a hSod1 adquire propriedades tóxicas aos neurônios motores. Atualmente, duas hipóteses não mutuamente excludentes existem para explicar o caráter tóxico das mutantes da hSod1 relacionadas à ELA. A primeira refere-se à produção de oxidantes pela atividade peroxidásica exacerbada das mutantes contribuindo para o estresse oxidativo observado em ELA. A segunda refere-se à agregação de proteínas como ocorre em outras doenças neurodegenerativas. Digno de nota, o radical carbonato produzido na atividade peroxidásica da hSod1 causa a formação de um dímero covalente da proteína análogo a uma espécie de hSod1 frequentemente detectada em modelos experimentais e pacientes da doença e associada à propriedade tóxica das mutantes. Desta forma, o presente trabalho buscou esclarecer o mecanismo de produção do radical carbonato pela hSod1, bem como caracterizar o dímero covalente da proteína para posterior estudo de sua formação em um modelo de ELA em ratos que superexpressam a mutante G93A da hSod1. Os estudos cinéticos da variação do pH sobre os efeitos de bicarbonato/CO2, nitrito e formato na atividade peroxidásica da hSod1, medidos pelo consumo de peróxido de hidrogênio e produção de radical, permitiram excluir o mecanismo de Fenton para explicar o ciclo peroxidativo da enzima em tampão bicarbonato em favor de outros intermediários reativos. Já, os experimentos de 13C RMN, modelagem molecular e cinética de fluxo interrompido com mistura assimétrica demonstraram que o ânion peroxomonocarbonato constitui o precursor...

Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disease of motors neurons that causes muscle atrophy, weakness, and death by respiratory failure. This pathology occurs in both sporadic and familiar forms that are clinically indistinguishable. Mutations in the antioxidant enzyme superoxide dismutase 1 (hSod1) respond to about 20% of the familiar cases of ALS. Besides, the autosomal dominant nature of these hSod1-associated ALS suggests that the mutants gain toxic properties to motor neurons. Currently, two hypotheses exist to explain the toxicity of hSod1 mutants but they do not exclude each other. The first one is related to the production of oxidants by the increased peroxidase activity of the ALS-linked mutants that could contribute to the oxidative stress reported in ALS. The second refers to protein aggregation as proposed in other neurodegenerative diseases. Noteworthy, the carbonate radical produced during hSod1 peroxidase activity leads to the formation of a covalent dimer of the protein similar to a hSod1 species often detected in experimental models and patients of the disease and implicated in the toxic properties of hSod1 mutants. Thus, the present work aimed to determine the mechanism of carbonate radical production by hSod1 and to characterize the covalent dimer of the protein in vitro followed by the study of covalent aggregates of hSod1 in a rat model of ALS that overexpresses the G93A mutant of the protein. The kinetic studies of the effect of bicarbonate/CO2, nitrite and formate in the peroxidase activity of hSod1 at various pH, measured by hydrogen peroxide consumption and radical production, permitted to exclude the Fenton mechanism to explain the enzyme peroxidative cycle in bicarbonate buffer in favor of other reactive intermediates. Furthermore, 13C NMR, molecular docking and stopped-flow experiments with asymmetric mixing demonstrated that the anion peroxomonocarbonate is the precursor of the carbonate radical produced by...

Influencia de la época del año en la respuesta in vitro del cafeto Coffea canephora P Var Robusta / Influence of the time of year on the in vitro response of Coffea canephora P Var Robusta coffee trees

Generalmente, las plantas responden de manera diferenciada a las técnicas de cultivo in vitro, dependiendode la respuesta inicial del cultivo y su capacidad embriogénica del genotipo, el estado fisiológico de la planta yel explante, la edad de la planta donante, los factores físicos, los reguladores del crecimiento y las condiciones nutricionales, así como el cambio estacional. De aquí que el presente estudio se realizó con el objetivo de evaluar la influencia de la época del año en que fue tomada la fuente de los explantes en la respuesta in vitro de genotipos de Robusta. Se realizaron muestreos durante todos los meses del año, los explantes foliares fueron cultivados en un medio contentivo de las sales minerales de Murashige y Skoog (1962) y 0,5 y 2,0 mg.L-1 de 2,4-D y kinetina, respectivamente. Se observó que la época del año en que se tomó la muestra ejerció un marcado efecto en la respuesta de los explantes de los genotipos evaluados. Para las condiciones estudiadas resultó favorable la toma de las muestras foliares en los períodos mayo-junio, enero-febrero y noviembre-diciembre, dados los bajos índices de actividad enzimática peroxidasa, oxidación fenólica y contaminación fúngica, así como un elevado porcentaje de formación de callos y mayor contenido de proteínas totales presentes en los mismos. Se determinó que la actividad enzimática peroxidasa pudiera constituir un importante marcador en la etapa inicial de selección de las muestras.

Plants generally respond in different ways to in vitro cultivation techniques, depending on the crop’s initial response and the embryogenic capacity of its genotype, the physiological state of the plant and the explant, the age of the donor plant, physical factors, growth regulators, nutritional conditions and seasonal change. The present study was thus aimed at evaluating the influence of the time of year during which the source of explants was taken on the in vitro response of Robusta genotypes. Samples were taken during each month ofthe year, foliar explants were cultured in a medium containing Murashige and Skoog mineral salts (1962) and 0,5 and 2,0 mg.L-1 2,4-D and kinetine, respectively. It was observed that the time of the year when a sample was taken exercised as marked effect on the response of the explants from the genotypes evaluated. Taking foliar samples during May-June, January-February and November-December proved favourable in the conditions studied here given the low indices of peroxidase enzymatic activity, phenol oxidation and fungal contamination, as well as the high percentage of callus formation and their greater total protein content. It was determined that peroxidase enzymatic activity could constitute an important marker during the initial stage of selecting samples.

Tolerância à salinidade em feijão (Phaseolus vulgaris L)

One of the applications of the tissue culture technique in plant improvement is the identification of cell lines which show salinity tolerance. Several authors were able to obtain saline stress-tolerant cell lines and show that mechanisms of tolerance to salts have a strong correlation between this phenomenon and a high macromolecule concentration in plant tissues. Callus obtained from embrionic axis of Phaseolus vulgarís cv. IAC carioca in solid medium, supplemented with 0 to 60 mM NaCl, as the salt treatment, were used. Callus harvesting was done on the 13th day, when they were processed for relative growth, protein, proline content and peroxidase acivity. The results show both, a decrease of the relative growth and of protein content in response to the NaCl treatment, as compared to controls. However, there was a significant increase on the proline content and on the peroxidase activity.

Uma das aplicações das técnicas da cultura de tecidos no melhoramento é a identificação de linhas de células que apresentam tolerância à salinidade. Vários autores obtiveram linhas de células tolerantes ao estresse salino; e estudo de mecanismos bioquímicos da tolerância a sais em plantas tem demonstrado altas correlações entre estes e o acúmulo de macromoléculas em tecido de plantas superiores. Para verificar essas correlações em feijão (Phaseolus vulgaris cv IAC carioca), calos oriundos de eixos embrionários foram cultivados em meio sólido, suplementado com NaCl nas concentrações de 0 a 60 mM. Após 13 dias de incubação, os calos foram coletados e analisados quanto ao crescimento relativo, teor de proteínas, teor de prolina e atividade da peroxidase. Os parâmetros analisados mostraram decréscimo no crescimento relativo e no de proteínas em resposta ao NaCl. Paralelamente, observou-se aumento significativo no conteúdo de prolina e atividade da enzima peroxidase.

Tolerância à salinidade em feijão (Phaseolus vulgaris L)

One of the applications of the tissue culture technique in plant improvement is the identification of cell lines which show salinity tolerance. Several authors were able to obtain saline stress-tolerant cell lines and show that mechanisms of tolerance to salts have a strong correlation between this phenomenon and a high macromolecule concentration in plant tissues. Callus obtained from embrionic axis of Phaseolus vulgarís cv. IAC carioca in solid medium, supplemented with 0 to 60 mM NaCl, as the salt treatment, were used. Callus harvesting was done on the 13th day, when they were processed for relative growth, protein, proline content and peroxidase acivity. The results show both, a decrease of the relative growth and of protein content in response to the NaCl treatment, as compared to controls. However, there was a significant increase on the proline content and on the peroxidase activity.

Uma das aplicações das técnicas da cultura de tecidos no melhoramento é a identificação de linhas de células que apresentam tolerância à salinidade. Vários autores obtiveram linhas de células tolerantes ao estresse salino; e estudo de mecanismos bioquímicos da tolerância a sais em plantas tem demonstrado altas correlações entre estes e o acúmulo de macromoléculas em tecido de plantas superiores. Para verificar essas correlações em feijão (Phaseolus vulgaris cv IAC carioca), calos oriundos de eixos embrionários foram cultivados em meio sólido, suplementado com NaCl nas concentrações de 0 a 60 mM. Após 13 dias de incubação, os calos foram coletados e analisados quanto ao crescimento relativo, teor de proteínas, teor de prolina e atividade da peroxidase. Os parâmetros analisados mostraram decréscimo no crescimento relativo e no de proteínas em resposta ao NaCl. Paralelamente, observou-se aumento significativo no conteúdo de prolina e atividade da enzima peroxidase.