Glutathione deficit in schizophrenia: strategies to increase glutathione levels in " in vitro " and clinical studies

Summary: Decrease in glutathione (GSH) levels was observed in cerebrospinal fluid, prefrontal cortex and post-mortem striatum of schizophrenia patients. Evidences suggest a defect in GSH synthesis at the levels of the rate-limiting synthesizing enzyme, glutamate cysteine ligase (GCL). Indeed, polymorphisms in the gene of the modifier subunit of GCL (GCLM) was shown to be associated with the disease in three different populations, GCLM gene expression is decreaséd in fibroblasts from patients and the increase in GCL activity induced by an oxidative stress is lower in patients' fibroblasts compared to controls. GSH being a major antioxydant and redox regulator, its presence is of high importance for protecting cells against oxidative stress. The aim of the present work was to use various substances to increase GSH levels by diverse strategies. Since the synthesizing enzyme GCL is defective, bypassing this enzyme was the first strategy we used. GSH ethyl ester (GSHEE), a membrane permeable analog of GSH, succeeded in replenishing GSH levels in cultured neurons and astrocytes previously depleted in GSH by L-buthionine-(S,R)-sulfoximine (BSO), an inhibitor of GCL. GSHEE also abolished dopamine-induced decrease of NMDA-mediated calcium response observed in BSO-treated neurons. y-Glutamylcysteine ethyl ester (GCSE), a membrane permeable analog of the product of GCL, increased GSH levels only in astrocytes. The second strategy was to boost the defective enzyme GCL. While quercetin (flavonoid) could increase GSH levels only in astrocytes, curcumin (polyphenol) and tertbutylhydroquinone (quinone) were successful in both neurons and astrocytes, via an increase in the gene expression of the two subunits of GCL and, consequently, an increase in the activity of the enzyme. However, FK506, an immunosupressant, was unefficient. Treating astrocytes from GCLM KO mice showed that the modulatory subunit is necessary for the action of the substances. Finally, since cysteine is the limiting precursor in the synthesis of GSH, we hypothesized that we could increase GSH levels by providing more of this precursor. N-acetyl-cysteine (NAC), a cysteine donor, was administered to schizophrenia patients, using adouble-blind and cross-over protocol. NAC significantly improved the mismatch negativity (MMN), a component of the auditory evoked potentials, thought to reflect selective current flowing through open, unblocked NMDA channels. Considering that NMDA function is reduced when GSH levels are low, increasing these levels with NAC could improve NMDA function as reflected by the improvement in the generation of the MMN. Résumé: Les taux de glutathion (GSH) dans le liquide céphalo-rachidien, le cortex préfrontal ainsi que le striatum post-mortem de patients schizophrènes, sont diminués. L'enzyme limitante dans la synthèse du GSH, la glutamyl-cysteine ligase (GCL), est défectueuse. En effet, des polymorphismes dans le gène de la sous-unité modulatrice de GCL (GCLM) sont associés à la maladie, l'expression du gène GCLM est diminuée dans les fibroblastes de patients et, lors d'un stress oxidative, l'augmentation de l'activité de GCL est plus faible chez les patients que chez les contrôles. Le GSH étant un important antioxydant et régulateur du status redox, sa présence est primordiale afin de protéger les cellules contre les stress oxydatifs. Au cours du présent travail, une variété de substances ont été utilisées dans le but d'augmenter les taux de GSH. Passer outre l'enzyme de synthèse GCL qui est défectueuse fut la première stratégie utilisée. L'éthylester de GSH (GSHEE), un analogue du GSH qui pénètre la membrane cellulaire, a augmenté les taux de GSH dans des neurones et des astrocytes déficitaires en GSH dû au L-buthionine-(S,R)-sulfoximine (BSO), un inhibiteur du GCL. Dans ces neurones, le GSHEE a aussi aboli la diminution de la réponse NMDA, induite parla dopamine. L'éthyl-ester de y-glutamylcysteine (GCEE), un analogue du produit de la GCL qui pénètre la membrane cellulaire, a augmenté les taux de GSH seulement dans les astrocytes. La seconde stratégie était d'augmenter l'activité de l'enzyme GCL. Tandis que la quercétine (flavonoïde) n'a pu augmenter les taux de GSH que dans les astrocytes, la curcumin (polyphénol) et le tert-butylhydroquinone (quinone) furent efficaces dans les deux types de cellules, via une augmentation de l'expression des gènes des deux sous-unités de GCL et de l'activité de l'enzyme. Le FK506 (immunosupresseur) n' a démontré aucune efficacité. Traiter des astrocytes provenant de souris GCLM KO a permis d'observer que la sous-unité modulatoire est nécessaire à l'action des substances. Enfin, puisque la cysteine est le substrat limitant dans la synthèse du GSH, fournir plus de ce présurseur pourrait augmenter les taux de GSH. Nacétyl-cystéine (NAC), un donneur de cystéine, a été administrée à des schizophrènes, lors d'une étude en double-aveugle et cross-over. NAC a amélioré le mismatch negativity (MMN), un composant des potentials évoqués auditifs, qui reflète le courant circulant via les canaux NMDA. Puisque la fonctionnalité des R-NMDA est diminuée lorsque les taux de GSH sont bas, augmenter ces taux avec NAC pourrait améliorer la fonction des R-NMDA, réflété par une augmentation de l'amplitude du MMN.

Indol-butyric acid levels on cashew cloning by air-layering process

A study was conducted to determine the possibility of cashew (Anacardium occidentale) cloning by air-layering and influence of IBA (indol-butyric acid) on this process. It was adopted a completely randomized design with 4 treatments, 10 air layers each and 4 replications, reaching 160 air layers. The IBA levels on the treatments were, as follow: 0, 1000, 3000 and 5000 mg.kg-1. It was evaluated: survival, callus and rooting percentage, average number and length of roots. The highest survival rate (67.5%) was registered with no growth regulator and IBA at 1000 mg.kg-1, while the best rooting percentage (82%) referred to 1000 mg.kg-1. In spite of average number and length of roots, the highest results were observed with IBA at 5000 mg.kg-1. IBA concentrations had no influence on cashew air-layering formation.

Predicting neurological outcome after cardiac arrest.

Purpose of reviewTherapeutic hypothermia and aggressive management of postresuscitation disease considerably improved outcome after adult cardiac arrest over the past decade. However, therapeutic hypothermia alters prognostic accuracy. Parameters for outcome prediction, validated by the American Academy of Neurology before the introduction of therapeutic hypothermia, need further update.Recent findingsTherapeutic hypothermia delays the recovery of motor responses and may render clinical evaluation unreliable. Additional modalities are required to predict prognosis after cardiac arrest and therapeutic hypothermia. Electroencephalography (EEG) can be performed during therapeutic hypothermia or shortly thereafter; continuous/reactive EEG background strongly predicts good recovery from cardiac arrest. On the contrary, unreactive/spontaneous burst-suppression EEG pattern, together with absent N20 on somatosensory evoked potentials (SSEP), is almost 100% predictive of irreversible coma. Therapeutic hypothermia alters the predictive value of serum markers of brain injury [neuron-specific enolase (NSE), S-100B]. Good recovery can occur despite NSE levels >33 mu g/l, thus this cut-off value should not be used to guide therapy. Diffusion MRI may help predicting long-term neurological sequelae of hypoxic-ischemic encephalopathy.SummaryAwakening from postanoxic coma is increasingly observed, despite early absence of motor signs and frank elevation of serum markers of brain injury. A new multimodal approach to prognostication is therefore required, which may particularly improve early prediction of favorable clinical evolution after cardiac arrest.

Modulation of genetic associations with serum urate levels by body-mass-index in humans.

We tested for interactions between body mass index (BMI) and common genetic variants affecting serum urate levels, genome-wide, in up to 42569 participants. Both stratified genome-wide association (GWAS) analyses, in lean, overweight and obese individuals, and regression-type analyses in a non BMI-stratified overall sample were performed. The former did not uncover any novel locus with a major main effect, but supported modulation of effects for some known and potentially new urate loci. The latter highlighted a SNP at RBFOX3 reaching genome-wide significant level (effect size 0.014, 95% CI 0.008-0.02, Pinter= 2.6 x 10-8). Two top loci in interaction term analyses, RBFOX3 and ERO1LB-EDARADD, also displayed suggestive differences in main effect size between the lean and obese strata. All top ranking loci for urate effect differences between BMI categories were novel and most had small magnitude but opposite direction effects between strata. They include the locus RBMS1-TANK (men, Pdifflean-overweight= 4.7 x 10-8), a region that has been associated with several obesity related traits, and TSPYL5 (men, Pdifflean-overweight= 9.1 x 10-8), regulating adipocytes-produced estradiol. The top-ranking known urate loci was ABCG2, the strongest known gout risk locus, with an effect halved in obese compared to lean men (Pdifflean-obese= 2 x 10-4). Finally, pathway analysis suggested a role for N-glycan biosynthesis as a prominent urate-associated pathway in the lean stratum. These results illustrate a potentially powerful way to monitor changes occurring in obesogenic environment.

Metabolic responses of guava trees irrigated withdifferent N and K levels in São Francisco Valley

The guava (Psidium guajava L.) cv. Paluma has been cultivated in São Francisco Valley, Northeastern of Brazil, for in natura consumption and processing purposes. In spite of its importance, there are few scientific knowledge regarding guava physiology, nutrition, irrigation and fertigation. The objective of this work was to evaluate the effect of weather conditions and different concentrations of N and K applied by fertigation in foliar contents of reducing sugars, total soluble sugars, starch, sucrose, amino acids, and proteins. The field experiment was carried out at Bebedouro Experimental Field and the biochemical evaluations at the Laboratory of Seed and Plant Physiology, both located at Embrapa Semi-Árido, Petrolina-PE. The doses of 200 g N and 100 g K2O; 400 g N and 200 g K2O; 600 g N and 300 g K2O; and 800 g N and 400 g K2O per plant were applied in an experiment field. The experimental design was totally randomized blocks, with four treatments and five blocks. The weather conditions influenced the plant photosynthesis, which affects the plants metabolism. Guava presented specific responses to N and K fertigation for each parameter evaluated. The weather conditions during the evaluation period influenced guava responses to N and K fertigation.

Assessment on nutrient levels in the aerial biomass of irrigated guava in São Francisco Valley, Brazil

In perennial crops, fertilizers are applied to restore the amount of nutrient removed from the plant by harvesting. Nevertheless, the immobilized nutrient inside the whole plant should be considered. Based on it, the nutrient concentrations in the guava leaf and fruit and the nutrient contents removed by fruits and post-harvest pruning were evaluated in an irrigated orchard, in Petrolina, State of Pernambuco, Brazil. The greatest contents removed by fruits were N and K among the macronutrients, while Fe, Zn and Mn presented higher levels among the micronutrients. More than 60% of the total content of N, P, K, Mg, Fe, and B removed by pruning were immobilized in the leaf and non-marketable fruit.

Effects of levels of nitrogen and potassium on yield and fruit maturation of irrigated guava trees in the São Francisco Valley

Yield and physical and chemical characteristics of 'Paluma' guava fruit were evaluated as a function of the harvest at different maturity stages, under influence of nitrogen and potassium fertilization, in Petrolina, State of Pernambuco, Brazil. Fertilizer rates were 67 kg N + 33 kg K2O, 133 kg N + 67 kg K2O, 200 kg N + 100 kg K2O and 267 kg N + 133 kg K2O per hectare. Fruits were evaluated at maturity stages 2, 3, 4 and 5, established according to peel color. Higher doses of N and K induced higher yields. Nevertheless, fertilization with 200 kg of N + 100 kg of K per hectare improved fruit quality, delaying ascorbic acid breakdown and conserving pulp firmness. Main changes took place at maturity stages 4 and 5, when the fruit should present ideal conditions for consumption, namely the increase on soluble solids and soluble sugars content.

Primary root protophloem differentiation requires balanced phosphatidylinositol-4,5-biphosphate levels and systemically affects root branching.

Protophloem is a specialized vascular tissue in growing plant organs, such as root meristems. In Arabidopsis mutants with impaired primary root protophloem differentiation, brevis radix (brx) and octopus (ops), meristematic activity and consequently overall root growth are strongly reduced. Second site mutation in the protophloem-specific presumed phosphoinositide 5-phosphatase COTYLEDON VASCULAR PATTERN 2 (CVP2), but not in its homolog CVP2-LIKE 1 (CVL1), partially rescues brx defects. Consistent with this finding, CVP2 hyperactivity in a wild-type background recreates a brx phenotype. Paradoxically, however, while cvp2 or cvl1 single mutants display no apparent root defects, the root phenotype of cvp2 cvl1 double mutants is similar to brx or ops, although, as expected, cvp2 cvl1 seedlings contain more phosphatidylinositol-4,5-biphosphate. Thus, tightly balanced phosphatidylinositol-4,5-biphosphate levels appear essential for proper protophloem differentiation. Genetically, OPS acts downstream of phosphatidylinositol-4,5-biphosphate levels, as cvp2 mutation cannot rescue ops defects, whereas increased OPS dose rescues cvp2 cvl1 defects. Finally, all three mutants display higher density and accelerated emergence of lateral roots, which correlates with increased auxin response in the root differentiation zone. This phenotype is also created by application of peptides that suppress protophloem differentiation, CLAVATA3/EMBRYO SURROUNDING REGION 26 (CLE26) and CLE45. Thus, local changes in the primary root protophloem systemically shape overall root system architecture.

Female major histocompatibility complex type affects male testosterone levels and sperm number in the horse (Equus caballus).

Odours of vertebrates often contain information about the major histocompatibility complex (MHC), and are used in kin recognition, mate choice or female investment in pregnancy. It is, however, still unclear whether MHC-linked signals can also affect male reproductive strategies. We used horses (Equus caballus) to study this question under experimental conditions. Twelve stallions were individually exposed either to an unfamiliar MHC-similar mare and then to an unfamiliar MHC-dissimilar mare, or vice versa. Each exposure lasted over a period of four weeks. Peripheral blood testosterone levels were determined weekly. Three ejaculates each were collected in the week after exposure to both mares (i.e. in the ninth week) to determine mean sperm number and sperm velocity. We found high testosterone levels when stallions were kept close to MHC-dissimilar mares and significantly lower ones when kept close to MHC-similar mares. Mean sperm number per ejaculate (but not sperm velocity) was positively correlated to mean testosterone levels and also affected by the order of presentation of mares: sperm numbers were higher if MHC-dissimilar mares were presented last than if MHC-similar mares were presented last. We conclude that MHC-linked signals influence testosterone secretion and semen characteristics, two indicators of male reproductive strategies.

Prevalence of iodine inadequacy in Switzerland assessed by the estimated average requirement cut-point method in relation to the impact of iodized salt.

OBJECTIVE: To assess the iodine status of Swiss population groups and to evaluate the influence of iodized salt as a vector for iodine fortification. DESIGN: The relationship between 24 h urinary iodine and Na excretions was assessed in the general population after correcting for confounders. Single-day intakes were estimated assuming that 92 % of dietary iodine was excreted in 24 h urine. Usual intake distributions were derived for male and female population groups after adjustment for within-subject variability. The estimated average requirement (EAR) cut-point method was applied as guidance to assess the inadequacy of the iodine supply. SETTING: Public health strategies to reduce the dietary salt intake in the general population may affect its iodine supply. SUBJECTS: The study population (1481 volunteers, aged ≥15 years) was randomly selected from three different linguistic regions of Switzerland. RESULTS: The 24 h urine samples from 1420 participants were determined to be properly collected. Mean iodine intakes obtained for men (n 705) and women (n 715) were 179 (sd 68.1) µg/d and 138 (sd 57.8) µg/d, respectively. Urinary Na and Ca, and BMI were significantly and positively associated with higher iodine intake, as were men and non-smokers. Fifty-four per cent of the total iodine intake originated from iodized salt. The prevalence of inadequate iodine intake as estimated by the EAR cut-point method was 2 % for men and 14 % for women. CONCLUSIONS: The estimated prevalence of inadequate iodine intake was within the optimal target range of 2-3 % for men, but not for women.

A need for determination of arsenic species at low levels in cereal-based food and infant cereals. Validation of a method by LC-ICPMS.

The present study arose from the need to determine inorganic arsenic (iAs) at low levels in cereal-based food. Validated methods with a low limit of detection (LOD) are required to analyse these kinds of food. An analytical method for the determination of iAs, methylarsonic acid (MA) and dimethylarsinic acid (DMA) in cereal-based food and infant cereals is reported. The method was optimised and validated to achieve low LODs. Ion chromatography-inductively coupled plasma mass spectrometry (LC-ICPMS) was used for arsenic speciation. The main quality parameters were established. To expand the applicability of the method, different cereal products were analysed: bread, biscuits, breakfast cereals, wheat flour, corn snacks, pasta and infant cereals. The total and inorganic arsenic content of 29 cereal-based food samples ranged between 3.7-35.6 and 3.1-26.0 microg As kg-1, respectively. The present method could be considered a valuable tool for assessing inorganic arsenic contents in cereal-based foods.

Leaf nutritional levels in peach and nectarine grown in subtropical climate

The study evaluated the leaf nutritional levels of peach and nectarine trees under subtropical climate in order to improve the fertilization practices. The experiment was carried out in São Paulo state University, Botucatu, São Paulo State, Brazil. The experimental design consisted of subdivided plots, in which plots corresponded to cultivars and subplots to the leaf sample periods. The evaluated peach cultivars were: Marli, Turmalina, Precocinho, Jubileu, Cascata 968, Cascata 848, CP 951C, CP 9553CYN, and Tropic Beauty, and that of nectarine was 'Sun Blaze'. The sample periods were: after harvest, plants in vegetative period; dormancy; beginning of flowering and fruiting (standard sample). Results indicated significant variations in the levels of N, P, K, Ca, Mg, S, B, Cu, Fe, Mn and Zn for the sampling period and in N, Ca, Mg, S, B, Fe and Mn levels for the cultivars.