Antioxidant intervention attenuates oxidative stress in children and teenagers with Down syndrome.

We previously demonstrated that systemic oxidative stress is present in Down syndrome (DS) patients. In the present study we investigated the antioxidant status in the peripheral blood of DS children and teenagers comparing such status before and after an antioxidant supplementation. Oxidative stress biomarkers were evaluated in the blood of DS patients (n=21) before and after a daily antioxidant intervention (vitamin E 400mg, C 500 mg) during 6 months. Healthy children (n=18) without DS were recruited as control group. The activity of superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx), glutathione reductase (GR), glutathione S-transferase (GST), gamma-glutamyltransferase (GGT), glucose-6-phosphate dehydrogenase (G6PD) and myeloperoxidase (MPO), as well as the contents of reduced glutathione (GSH), uric acid, vitamin E, thiobarbituric acid reactive substances (TBARS), and protein carbonyls (PC) were measured. Before the antioxidant therapy, DS patients presented decreased GST activity and GSH depletion; elevated SOD, CAT, GR, GGT and MPO activities; increased uric acid levels; while GPx and G6PD activities as well as vitamin E and TBARS levels were unaltered. After the antioxidant supplementation, SOD, CAT, GPx, GR, GGT and MPO activities were downregulated, while TBARS contents were strongly decreased in DS. Also, the antioxidant therapy did not change G6PD and GST activities as well as uric acid and PC levels, while it significantly increased GSH and vitamin E levels in DS patients. Our results clearly demonstrate that the antioxidant intervention with vitamins E and C attenuated the systemic oxidative damage present in DS patients.

Intracerebroventricular administration of N-acetylaspartic acid impairs antioxidant defenses and promotes protein oxidation in cerebral cortex of rats.

N-acetylaspartic acid (NAA) is the biochemical hallmark of Canavan Disease, an inherited metabolic disease caused by deficiency of aspartoacylase activity. NAA is an immediate precursor for the enzyme-mediated biosynthesis of N-acetylaspartylglutamic acid (NAAG), whose concentration is also increased in urine and cerebrospinal fluid of patients affected by CD. This neurodegenerative disorder is clinically characterized by severe mental retardation, hypotonia and macrocephaly, and generalized tonic and clonic type seizures. Considering that the mechanisms of brain damage in this disease remain not fully understood, in the present study we investigated whether intracerebroventricular administration of NAA or NAAG elicits oxidative stress in cerebral cortex of 30-day-old rats. NAA significantly reduced total radical-trapping antioxidant potential, catalase and glucose 6-phosphate dehydrogenase activities, whereas protein carbonyl content and superoxide dismutase activity were significantly enhanced. Lipid peroxidation indices and glutathione peroxidase activity were not affected by NAA. In contrast, NAAG did not alter any of the oxidative stress parameters tested. Our results indicate that intracerebroventricular administration of NAA impairs antioxidant defenses and induces oxidative damage to proteins, which could be involved in the neurotoxicity of NAA accumulation in CD patients.

Inhibition of yeast glutathione reductase by trehalose: possible implications in yeast survival and recovery from stress.

Accumulation of trehalose has been implicated in the tolerance of yeast cells to several forms of stress, including heat-shock and high ethanol levels. However, yeast lacking trehalase, the enzyme that degrades trehalose, exhibit poor survival after exposure to stress conditions. This suggests that optimal cell viability also depends on the capacity to rapidly degrade the high levels of trehalose that build up under stress. Here, we initially examined the effects of trehalose on the activity of an important antioxidant enzyme, glutathione reductase (GR), from Saccharomyces cerevisiae. At 25 degrees C, GR was inhibited by trehalose in a dose-dependent manner, with 70% inhibition at 1.5M trehalose. The inhibition was practically abolished at 40 degrees C, a temperature that induces a physiological response of trehalose accumulation in yeast. The inhibition of GR by trehalose was additive to the inhibition caused by ethanol, indicating that enzyme function is drastically affected upon ethanol-induced stress. Moreover, two other yeast enzymes, cytosolic pyrophosphatase and glucose 6-phosphate dehydrogenase, showed temperature dependences on inhibition by trehalose that were similar to the temperature dependence of GR inhibition. These results are discussed in terms of the apparent paradox represented by the induction of enzymes involved in both synthesis and degradation of trehalose under stress, and suggest that the persistence of high levels of trehalose after recovery from stress could lead to the inactivation of important yeast enzymes.

Mecanismos de acción de xenobióticos en ovocitos de anfibios / Xenobiotic mechanisms of action in amphibian oocytes

Comprender los procesos que determinan la toxicidad de diversas sustancias a nivel celular constituye un desafío para el desarrollo tecnológico actual. Los ovocitos de diversas especies son utilizados con distintos propósitos en investigación. El tamaño de los ovocitos de anfibios y la facilidad con que puede obtenerse un gran número de ellos como células aisladas, hacen de los mismos el sistema de elección para múltiples propósitos, tales como: estudios de las etapas tempranas de la reproducción, técnicas de fertilización in vitro, expresión y caracterización de receptores de membranas plasmática, mecanismos de acción de agonistas, vías metabólicas, etc. El objetivo del presente trabajo con ovocitos de sapo Bufo arenarum es estudiar los mecanismos de acción de sustancias xenobióticas a nivel bioquímico y molecular, en relación con la capacidad de éstas gametas de ser fertilizadas y avanzar a través de los estadios tempranos del desarrollo embrionario. Se ha trabajado con diversos biomarcadores que permitieron reconocer los mecanismos de toxicidad de compuestos organoclorados y más recientemente de algunos metales pesados. Se demostró que la activación de una fosfolipasa C específica por el Dieldrin provoca depleción de mediadores lipídicos y como consecuencia inhibición de la fertilización (desarrollo hasta 4-8 células). Actualmente se estudian los efectos del Zn sobre la fertilización y su probable relación con alteraciones en la actividad de enzimas de la vía de las pentosas, directamente ligada a los mecanismos de detoxificación en presencia de stress oxidativo. El estudio de los mecanismos de citotoxicidad de diversas sustancias resulta de gran utilidad cuando se lleva a cabo en gametas, porque permite también evaluar los efectos de dichas sustancias sobre la reproducción de la especie. Los ovocitos de anfibios son además células de gran interés, por su utilidad en el estudio de la citotoxicidad de una amplia variedad de contaminantes hídricos (AU)

Liver microsomal parameters related to oxidative stress and antioxidant systems in hyperthyroid rats subjected to acute lindane treatment.

Liver microsomal functions related to xenobiotic biotransformation and free radical production were studied in control rats and in animals subjected to L-3,3',5-triiodothyronine (T3) and/or lindane administration as possible mechanisms contributing to oxidative stress, in relation to the activity of enzymes (superoxide dismutase (SOD), catalase, glutathione peroxidase (GPx), and glucose-6-phosphate dehydrogenase (G-6PDH)) and content of lipid-soluble vitamins (alpha-tocopherol, beta-carotene, and lycopene) affording antioxidant protection. Lindane treatment in euthyroid rats at a dosage of 20mg/kg did not modify the content of liver microsomal cytochromes P450 and b5, the activity of NADPH-cytochrome P450 reductase and NADH-cytochrome b5 reductase, and the production of superoxide radical (O2.-), as well as antioxidant systems, except for the reduction in lycopene levels. Hyperthyroidism elicited a calorigenic response and increased specific and molecular activities of NADPH-cytochrome P450 reductase, O2.- generation, and G-6PDH activity, concomitantly with diminution in liver SOD and catalase activities and in alpha-tocopherol, beta-carotene, and lycopene levels. The administration of lindane to hyperthyroid animals led to a further increase in the molecular activity of NADPH-cytochrome P450 reductase and in the O2.- production/SOD activity ratio, and decrease of hepatic alpha-tocopherol content, in a magnitude exceeding the sum of effects elicited by the separate treatments, as previously reported for reduced glutathione depletion. Collectively, these data support the contention that the increased susceptibility of the liver to the toxic effects of acute lindane treatment in hyperthyroid state is conditioned by potentiation of the hepatic oxidative stress status.

Carbohydrate protection of enzyme structure and function against guanidinium chloride treatment depends on the nature of carbohydrate and enzyme.

Baker's yeast cells accumulate osmolytes as a response to several stress conditions such as high-temperature and low-temperature shifts, dehydration, or osmotic stress. One of the major osmolytes that accumulates is trehalose, which plays an essential role affecting the survival of yeast at the time of stress. In this report, we show that trehalose efficiently protects the function and the structure of two yeast cytosolic enzymes against chemical denaturation by guanidinium chloride. Other sugars tested also protected yeast pyrophosphatase and glucose-6-phosphate dehydrogenase structure against guanidinium chloride effects, but were not as efficient at protecting enzyme activity. The thermostable pyrophosphatase from Bacillus stearothermophilus was also protected by several sugars against the chaotropic properties of guanidinium chloride, but was only protected by trehalose against functional inactivation. The function of the membrane-embedded H+-ATPase from yeast could not be protected by any of the tested sugars, although all of the sugars protected its structure from guanidinium-chloride-induced unfolding. The results presented in this study suggest that several sugars are able to prevent protein unfolding induced by a chaotropic compound. However, prevention of functional inactivation depends on the nature of the sugar. Trehalose was the most efficient, being able to protect many cytosolic enzymes against guanidinium chloride. The efficiency of protection also depended on the nature of the protein tested. This might explain why trehalose is one of the osmolytes accumulated in yeast and also why it is not the only osmolyte to accumulate.

Effects of ivermectin on the activity of enzymes in mammalian cells in vitro.

The effects of ivermectin on the activities of lactate dehydrogenase, glucose-6-phosphate dehydrogenase, and glucose-6-phosphatase have been estimated in IB-RS-2 cells in vitro. A 72-hr time course following ivermectin exposure indicated a decrease in the activity of lactate dehydrogenase. The activities of glucose-6-phosphate dehydrogenase and glucose-6-phosphatase remained essentially unchanged.

Anemias hemoliticas por defectos enzimáticos: Consideraciones generales / Hemolytic anemia caused by enzymatic disorders: General aspects

Las anemias hemolíticas congénitas por defectos enzimáticos fueron reconocidas como entidades diferentes, en este trabajo se hizo una revisión sobre las consideraciones generales en cuanto a la deficienia de glucosafosfato deshidrogenasa, y como anormalidad heredada a través del cromosoma X

Actividad de la glucosa-6-fosfato deshidrogenasa en pacientes diabéticos tratados con pirofosfato de tiamina o cocarboxilasa / Activity of clucose-6-phosphate dehydrogenase on diabetic patients treated with thiramine pyrophosphate or cocarboxilase

Trabajos realizados con anterioridad tanto a nivel clínico como experimental en el Instituto de Investigaciones Cientificas Hans Selye, permiten proponer que la cocarboxilasa o pirofosfato de tiamina (PPT) estable en solución, al aplicarse a pacientes con hiperglucemia, mejora la incorporación de la glucosa a los tejidos. Con base en tales hechos, un posterior estudio pretende analizar si el PPT ejerce acción alguna sobre el ciclo de las pentosas. Para demostrar esto, se estudió la conducta de la glucosa-6-fosfato deshidrogenasa (G-6-PD) en el eritrocitos humanos, toda vez que su actividad se ha encontrado disminuida en los pacientes afectados por diabetes mellitus. Para comprobarlo se seleccionaron 10 pacientes de uno y otro sexo, de 45 a 70 años de edad y con diagnóstico de diabetes tipo II. De cada uno se tomó una muestra sanguínea antes de iniciar el traqtamiento con PPT y posteriormente a los 15, 30, 60, 90 y 120 días después de obtener la primera muestra, a fin de determinar la actividad de G-6-PD, así como las de glucosa, colesterol total, HDL, LDL, triglicériados y hemoglobina glucosilada. Los resultados obtenidos demuestran que la glucemia tiende a registrar niveles normales, alcanzando una recuperación física plena en 70 por ciento de los pacientes; en 100 por ciento de ellos también se normalizaron los niveles de colesterol y de triglicéridos. La hemoglobina glucosilada disminuye considerablemente en relación con los valores registrados antes de iniciar el tratamiento. Con respecto a la actividad de la G-6-PD, los autores observaron que ésta se incrementa, en promedio, de 3 a 5 U/g de hemoglobina en los pacientes hiperglucémicos: el máximo incremento de actividad se registra entre el segundo y tercer mes de tratamiento, lo cual indica que PPT participa en: 1) la incorporación de glucosa al interior de los eritrocitos, toda vez que dicha sustancia disminuye la glucosa sérica: 2) la utilización de la glucosa en el metabolismo de esta célilas, y 3) el ciclo de las pentosas, dado que la actividad de la G-6-PD aumenta en forma considerable.