Changes in Noradrenergic Synthesis and Dopamine Beta-Hydroxylase Activity in Response to Oxidative Stress after Iron-induced Brain Injury.

Noradrenaline (NA) levels are altered during the first hours and several days after cortical injury. NA modulates motor functional recovery. The present study investigated whether iron-induced cortical injury modulated noradrenergic synthesis and dopamine beta-hydroxylase (DBH) activity in response to oxidative stress in the brain cortex, pons and cerebellum of the rat. Seventy-eight rats were divided into two groups: (a) the sham group, which received an intracortical injection of a vehicle solution; and (b) the injured group, which received an intracortical injection of ferrous chloride. Motor deficits were evaluated for 20 days post-injury. On the 3rd and 20th days, the rats were euthanized to measure oxidative stress indicators (reactive oxygen species (ROS), reduced glutathione (GSH) and oxidized glutathione (GSSG)) and catecholamines (NA, dopamine (DA)), plus DBH mRNA and protein levels. Our results showed that iron-induced brain cortex injury increased noradrenergic synthesis and DBH activity in the brain cortex, pons and cerebellum at 3 days post-injury, predominantly on the ipsilateral side to the injury, in response to oxidative stress. A compensatory increase in contralateral noradrenergic activity was observed, but without changes in the DBH mRNA and protein levels in the cerebellum and pons. In conclusion, iron-induced cortical injury increased the noradrenergic response in the brain cortex, pons and cerebellum, particularly on the ipsilateral side, accompanied by a compensatory response on the contralateral side. The oxidative stress was countered by antioxidant activity, which favored functional recovery following motor deficits.

UHPLC-HRMS-Based Analysis of S-Hydroxymethyl-Glutathione, GSH, and GSSG in Human Cells.

Glutathione (GSH) is one of the main antioxidant molecules present in cells. It harbors a thiol group responsible for sustaining cellular redox homeostasis. This moiety can react with cellular electrophiles such as formaldehyde yielding the compound S-hydroxymethyl-GSH (HSMGSH). HSMGSH is the substrate of the enzyme alcohol dehydrogenase 5 (ADH5) and thus a key intermediate in formaldehyde metabolism. In this work, we describe a method for the chemical synthesis of HSMGSH and a pipeline to identify this compound in complex cell extracts by means of ultra-high-performance liquid chromatography coupled to high-resolution spectrometry (UHPLC-HRMS). This method also allows determining GSH and oxidized disulfide (GSSG) in the same samples, thus providing broad information about formaldehyde-GSH metabolism.

Increased glutathione levels contribute to the beneficial effects of hydrogen sulfide and inducible nitric oxide inhibition in allergic lung inflammation.

OBJECTIVE: The interaction between nitric oxide (NO) and hydrogen sulfide (H2S) in the airways could have significant implications for the pathogenesis and therapeutic effects of both on lung diseases. In this study we investigated whether the beneficial effects of H2S on asthma could be comparable to that inhibition of inducible NO synthase (iNOS). METHODS: Female BALB/C mice sensitized with ovalbumin (OVA) received either the H2S donor sodium hydrosulfide (NaHS, 14µmol/kg) or the iNOS inhibitor 1400W (1mg/kg), 30min before each OVA challenge during six days. On the first, second and sixth days, the leucocyte infiltration in lung parenchyma and bronchoalveolar lavage was evaluated. The aconitase activity (a sensor of O2 formation) and lipid peroxidation, as well as levels of reduced glutathione (GSH) and oxidized glutathione (GSSG) were determined in the lung tissues. RESULTS: OVA-challenge caused a significant and time-dependent increase in the eosinophil number in the airways, which was accompanied by a significant decrease of aconitase activity and GSH/GSSG ratio along with enhanced lipid peroxidation in the lungs. Treatment with NaHS or 1400W significantly attenuated the airways eosinophilia that was paralleled by an increase in aconitase activity and decrease of lipid peroxidation. NaHS or 1400W treatments also reversed the decreased GSH/GSSG ratio seen after OVA-challenge. CONCLUSIONS: The present study shows for the first time that the increased GSH/GSSG ratio caused by either H2S supplementation or iNOS-inhibition is a potential mechanism protecting airways against oxidative stress and inflammatory lung diseases.

Water stress reveals differential antioxidant responses of tolerant and non-tolerant sugarcane genotypes.

The biochemical responses of the enzymatic antioxidant system of a drought-tolerant cultivar (IACSP 94-2094) and a commercial cultivar in Brazil (IACSP 95-5000) grown under two levels of soil water restriction (70% and 30% Soil Available Water Content) were investigated. IACSP 94-2094 exhibited one additional active superoxide dismutase (Cu/Zn-SOD VI) isoenzyme in comparison to IACSP 95-5000, possibly contributing to the heightened response of IACSP 94-2094 to the induced stress. The total glutathione reductase (GR) activity increased substantially in IACSP 94-2094 under conditions of severe water stress; however, the appearance of a new GR isoenzyme and the disappearance of another isoenzyme were found not to be related to the stress response because the cultivars from both treatment groups (control and water restrictions) exhibited identical changes. Catalase (CAT) activity seems to have a more direct role in H2O2 detoxification under water stress condition and the shift in isoenzymes in the tolerant cultivar might have contributed to this response, which may be dependent upon the location where the excessive H2O2 is being produced under stress. The improved performance of IACSP 94-2094 under drought stress was associated with a more efficient antioxidant system response, particularly under conditions of mild stress.

Cu(II)-disulfide complexes display simultaneous superoxide dismutase- and catalase-like activities.

Superoxide is a potentially toxic by-product of cellular metabolism. We have addressed here the in vitro ability of complexes formed between copper(II) ions and various biologically-occurring disulfides (RSSR: oxidized glutathione, cystine, homocystine and α-lipoic acid) to react with superoxide. The studied complexes were found to react with superoxide (generated by a xanthine/xanthine oxidase system) at rate constants (kCu(II)-RSSR) close to 10(6)M(-1)s(-1), which are three orders of magnitude lower than that reported for superoxide dismutase (SOD) but comparable to that of several other copper-containing complexes reported as SOD mimetics. The interaction between the tested Cu(II)-RSSR and superoxide, led to the generation and recovery of concentrations of hydrogen peroxide and oxygen that were, respectively, below and above those theoretically-expected from a sole SOD mimetic action. Interestingly, oxygen was generated when the Cu(II)-RSSR complexes were directly incubated with hydrogen peroxide. Taken together, these results reveal that the Cu(II)-RSSR complexes not only have the capacity to dismutate superoxide but also to simultaneously act like catalase mimetic molecules. When added to superoxide-overproducing mitochondria (condition attained by its exposure to diclofenac), three of the tested complexes were able (2-4µM), not only to totally restore, but also to lower below the basal level the mitochondrial production of superoxide. The present study is first in reporting on the potential of Cu(II)-disulfide complexes to act as SOD and catalase like molecules, suggesting a potential for these types of molecules to act as such under physiological and/or oxidative-stress conditions.

Homeostasis del glutatión / Glutathione homeostasis / Homeostase de glutationa

El glutatión (GSH) es una molécula única que participa en aspectos esenciales de la homeostasis celular, teniendo un rol central en la defensa contra el daño oxidativo. El GSH (L-g-glutamil-L-cisteinil-glicina) es un tripéptido hidrosoluble formado por los aminoácidos ácido glutámico, cisteína y glicina que se encuentra presente en el citoplasma de todas las células. La forma oxidada de la molécula, GSSG, se encuentra principalmente en forma extracelular. Las concentraciones de GSH y GSSG y su relación molar son indicadores de la funcionalidad celular y su alteración está relacionada con varios procesos patológicos en el hombre y en los animales de compañía. En esta revisión se abordan importantes aspectos de la homeostasis, las principales funciones biológicas y las metodologías analíticas disponibles para el análisis de GSH en sangre y plasma.(AU)

Glutathione is a unique molecule that participates in key cellular homeostasis, having a central role in defense against oxidative damage. GSH (L-g-glutamyl-L-cysteinyl-glycine) is a water soluble tripeptide composed of amino acid glutamic acid, cysteine and glycine. GSH is present in every cell cytoplasm. The oxidized form of the molecule, GSSG, is found primarily in extracellular form. GSH and GSSG concentrations and their molar ratio are indicators of cell function and its alteration is associated with several disease processes in humans and in companion animals. This review focuses on important aspects of homeostasis, major biological functions and available analytical methodologies for the analysis of GSH in blood and plasma.(AU)

A glutationa (GSH) é urna molécula única envolvida em aspectos essenciais da homeostase celular, tendo um papel central na defesa contra o dano oxidativo. O GSH (L-g-glutamil-L-cisteinil-glicina) é um tripeptídeo hidrossolúvel formado pelos aminoácidos: ácido glutámico, cisteína e glicina que se encontra presente no citoplasma de todas as células. A forma oxidada da molécula, GSSG, acha-se principalmente em forma extracelular. As concentragoes de GSH e GSSG e a sua relagáo molar sao indicadores da funcionalidade celular e a sua alteragao está relacionada com vários processos patológicos no homem e nos animais de estimagáo. A presente revisáo aborda questoes importantes da homeostase, as principais fungoes biológicas e as metodologias analíticas disponíveis para a análise de GSH em sangue e plasma.(AU)

Homeostasis del glutatión / Glutathione homeostasis / Homeostase de glutationa

El glutatión (GSH) es una molécula única que participa en aspectos esenciales de la homeostasis celular, teniendo un rol central en la defensa contra el daño oxidativo. El GSH (L-g-glutamil-L-cisteinil-glicina) es un tripéptido hidrosoluble formado por los aminoácidos ácido glutámico, cisteína y glicina que se encuentra presente en el citoplasma de todas las células. La forma oxidada de la molécula, GSSG, se encuentra principalmente en forma extracelular. Las concentraciones de GSH y GSSG y su relación molar son indicadores de la funcionalidad celular y su alteración está relacionada con varios procesos patológicos en el hombre y en los animales de compañía. En esta revisión se abordan importantes aspectos de la homeostasis, las principales funciones biológicas y las metodologías analíticas disponibles para el análisis de GSH en sangre y plasma.

Glutathione is a unique molecule that participates in key cellular homeostasis, having a central role in defense against oxidative damage. GSH (L-g-glutamyl-L-cysteinyl-glycine) is a water soluble tripeptide composed of amino acid glutamic acid, cysteine and glycine. GSH is present in every cell cytoplasm. The oxidized form of the molecule, GSSG, is found primarily in extracellular form. GSH and GSSG concentrations and their molar ratio are indicators of cell function and its alteration is associated with several disease processes in humans and in companion animals. This review focuses on important aspects of homeostasis, major biological functions and available analytical methodologies for the analysis of GSH in blood and plasma.

A glutationa (GSH) é urna molécula única envolvida em aspectos essenciais da homeostase celular, tendo um papel central na defesa contra o dano oxidativo. O GSH (L-g-glutamil-L-cisteinil-glicina) é um tripeptídeo hidrossolúvel formado pelos aminoácidos: ácido glutámico, cisteína e glicina que se encontra presente no citoplasma de todas as células. A forma oxidada da molécula, GSSG, acha-se principalmente em forma extracelular. As concentragoes de GSH e GSSG e a sua relagáo molar sao indicadores da funcionalidade celular e a sua alteragao está relacionada com vários processos patológicos no homem e nos animais de estimagáo. A presente revisáo aborda questoes importantes da homeostase, as principais fungoes biológicas e as metodologias analíticas disponíveis para a análise de GSH em sangue e plasma.

Impact of exercise training on redox signaling in cardiovascular diseases.

Reactive oxygen and nitrogen species regulate a wide array of signaling pathways that governs cardiovascular physiology. However, oxidant stress resulting from disrupted redox signaling has an adverse impact on the pathogenesis and progression of cardiovascular diseases. In this review, we address how redox signaling and oxidant stress affect the pathophysiology of cardiovascular diseases such as ischemia-reperfusion injury, hypertension and heart failure. We also summarize the benefits of exercise training in tackling the hyperactivation of cellular oxidases and mitochondrial dysfunction seen in cardiovascular diseases.

Neuronal damage and cognitive impairment associated with hypoglycemia: An integrated view.

The aim of the present review is to offer a current perspective about the consequences of hypoglycemia and its impact on the diabetic disorder due to the increasing incidence of diabetes around the world. The main consequence of insulin treatment in type 1 diabetic patients is the occurrence of repetitive periods of hypoglycemia and even episodes of severe hypoglycemia leading to coma. In the latter, selective neuronal death is observed in brain vulnerable regions both in humans and animal models, such as the cortex and the hippocampus. Cognitive damage subsequent to hypoglycemic coma has been associated with neuronal death in the hippocampus. The mechanisms implicated in selective damage are not completely understood but many factors have been identified including excitotoxicity, oxidative stress, zinc release, PARP-1 activation and mitochondrial dysfunction. Importantly, the diabetic condition aggravates neuronal damage and cognitive failure induced by hypoglycemia. In the absence of coma prolonged and severe hypoglycemia leads to increased oxidative stress and discrete neuronal death mainly in the cerebral cortex. The mechanisms responsible for cell damage in this condition are still unknown. Recurrent moderate hypoglycemia is far more common in diabetic patients than severe hypoglycemia and currently important efforts are being done in order to elucidate the relationship between cognitive deficits and recurrent hypoglycemia in diabetics. Human studies suggest impaired performance mainly in memory and attention tasks in healthy and diabetic individuals under the hypoglycemic condition. Only scarce neuronal death has been observed under moderate repetitive hypoglycemia but studies suggest that impaired hippocampal synaptic function might be one of the causes of cognitive failure. Recent studies have also implicated altered mitochondrial function and mitochondrial oxidative stress.

Redox regulation of the proteasome via S-glutathionylation.

The proteasome is a multimeric and multicatalytic intracellular protease responsible for the degradation of proteins involved in cell cycle control, various signaling processes, antigen presentation, and control of protein synthesis. The central catalytic complex of the proteasome is called the 20S core particle. The majority of these are flanked on one or both sides by regulatory units. Most common among these units is the 19S regulatory unit. When coupled to the 19S unit, the complex is termed the asymmetric or symmetric 26S proteasome depending on whether one or both sides are coupled to the 19S unit, respectively. The 26S proteasome recognizes poly-ubiquitinylated substrates targeted for proteolysis. Targeted proteins interact with the 19S unit where they are deubiquitinylated, unfolded, and translocated to the 20S catalytic chamber for degradation. The 26S proteasome is responsible for the degradation of major proteins involved in the regulation of the cellular cycle, antigen presentation and control of protein synthesis. Alternatively, the proteasome is also active when dissociated from regulatory units. This free pool of 20S proteasome is described in yeast to mammalian cells. The free 20S proteasome degrades proteins by a process independent of poly-ubiquitinylation and ATP consumption. Oxidatively modified proteins and other substrates are degraded in this manner. The 20S proteasome comprises two central heptamers (ß-rings) where the catalytic sites are located and two external heptamers (α-rings) that are responsible for proteasomal gating. Because the 20S proteasome lacks regulatory units, it is unclear what mechanisms regulate the gating of α-rings between open and closed forms. In the present review, we discuss 20S proteasomal gating modulation through a redox mechanism, namely, S-glutathionylation of cysteine residues located in the α-rings, and the consequence of this post-translational modification on 20S proteasomal function.