Altered Oxygen Metabolism Associated to Neurogenesis of Induced Pluripotent Stem Cells Derived From a Schizophrenic Patient

Schizophrenia has been defined as a neurodevelopmental disease that causes changes in the process of thoughts, perceptions. and emotions, usually leading to a mental deterioration and affective blunting. Studies have shown altered cell respiration and oxidative stress response in schizophrenia; however, most of the knowledge has been acquired from postmortem brain analyses or from nonneural cells. Here we describe that neural cells, derived from induced pluripotent stem cells generated from skin fibroblasts of a schizophrenic patient, presented a twofold increase in extramitochondrial oxygen consumption as well as elevated levels of reactive oxygen species (ROS), when compared to controls. This difference in ROS levels was reverted by the mood stabilizer valproic acid. Our model shows evidence that metabolic changes occurring during neurogenesis are associated with schizophrenia, contributing to a better understanding of the development of the disease and highlighting potential targets for treatment and drug screening.

Mitochondrial localization and structure-based phosphate activation mechanism of Glutaminase C with implications for cancer metabolism

Glutamine is an essential nutrient for cancer cell proliferation, especially in the context of citric acid cycle anaplerosis. In this manuscript we present results that collectively demonstrate that, of the three major mammalian glutaminases identified to date, the lesser studied splice variant of the gene gls, known as Glutaminase C (GAC), is important for tumor metabolism. We show that, although levels of both the kidney-type isoforms are elevated in tumor vs. normal tissues, GAC is distinctly mitochondrial. GAC is also most responsive to the activator inorganic phosphate, the content of which is supposedly higher in mitochondria subject to hypoxia. Analysis of X-ray crystal structures of GAC in different bound states suggests a mechanism that introduces the tetramerization-induced lifting of a "gating loop" as essential for the phosphate-dependent activation process. Surprisingly, phosphate binds inside the catalytic pocket rather than at the oligomerization interface. Phosphate also mediates substrate entry by competing with glutamate. A greater tendency to oligomerize differentiates GAC from its alternatively spliced isoform and the cycling of phosphate in and out of the active site distinguishes it from the liver-type isozyme, which is known to be less dependent on this ion.

EVALUATION OF GLUTATHIONE S-TRANSFERASE GSTM1 AND GSTT1 POLYMORPHISMS AND METHYLMERCURY METABOLISM IN AN EXPOSED AMAZON POPULATION

Over the last decades, the presence of methylmercury (MeHg) in the Amazon region of Brazil and its adverse human health effects have given rise to much concern. The biotransformation of MeHg occurs mainly through glutathione (GSH) in the bile mediated by conjugation with glutathione S-transferases (GST). Epidemiological evidence has shown that genetic polymorphisms may affect the metabolism of MeHg. The aim of this study was to evaluate the association between GST polymorphisms, GSH, and Hg levels in blood (B-Hg) and in hair (H-Hg) of an Amazon population chronically exposed to the metal through fish consumption. Blood and hair samples were collected from 144 volunteers (71 men, 73 women). B-Hg and H-Hg levels were determined by inductively coupled plasma-mass spectrometry, and GSH levels were evaluated by a spectrophotometric method. GSTM1 and T1 genotyping evaluation were carried out by multiplex polymerase chain reaction (PCR). Mean levels of B-Hg and H-Hg were 37.7 +/- 24.5 mu g/L and 10.4 +/- 7.4 mu g/g, respectively; GSH concentrations ranged from 0.52 to 2.89 mu M/ml of total blood. Distributions for GSTM1/T1, GSTM1/GSTT1*0, GSTM1*0/T1, and GSTM1*0/GSTT1*0 genotypes were 35.4, 22.2, 25.0, and 17.4%, respectively. GSTT1 genotype carriers presented lower levels of B-Hg and H-Hg when compared to other genotypes carriers. In addition, GSTM1*0/GSTT1*0 individuals presented higher Hg levels in blood and hair than subjects presenting any other genotypes. There appeared to be no evidence of an effect of polymorphisms on GSH levels. Therefore, our data suggest that GST polymorphisms may be associated with MeHg detoxification.

Effect of inulin on the growth and metabolism of a probiotic strain of Lactobacillus rhamnosus in co-culture with Streptococcus thermophilus

Metabolic studies are very important to improve quality of functional dairy products. For this purpose, the behaviors of pure cultures of Streptococcus thermophilus (St) and Lactobacillus rhamnosus (Lr) as well a co-culture of them (St-Lr) were investigated during skim milk fermentation, and the inulin effect as prebiotic was assessed. Lr was able to metabolize 6 g/100 g more galactose than St and St-Lr. Final lactic acid production by Lr was higher (9.8 g/L) compared to St (9.1 g/L) and St-Lr (9.1 g/L). Acetic acid concentration varied from 0.8 g/L (St-Lr) to 1.5 g/L (Lr) and that of ethanol from only 0.2 g/L (St-Lr) to 0.4 g/L (Lr), which suggests the occurrence in Lr of a NADH oxidase activity and citrate co-metabolization via pyruvate, both dissipating a part of the reducing power. Diacetyl and acetoin accumulated at the highest levels (18.4 and 0.8 mg/L, respectively) with St-Lr, which suggests possible synergistic interactions between these microorganisms as well as the Lr capability of co-metabolizing citrate in the presence of lactose. Inulin stimulated both biomass growth and levels of all end-products, as the likely result of fructose release from its partial hydrolysis and subsequent metabolization as an additional carbon and energy source. Crown Copyright (C) 2012 Published by Elsevier Ltd. All rights reserved.

Effects of Natural Radiation, Photosynthetically Active Radiation and Artificial Ultraviolet Radiation-B on the Chloroplast Organization and Metabolism of Porphyra acanthophora var. brasiliensis (Rhodophyta, Bangiales)

We undertook a study of Porphyra acanthophora var. brasiliensis to determine its responses under ambient conditions, photosynthetically active radiation (PAR), and PAR+UVBR (ultraviolet radiation-B) treatment, focusing on changes in ultrastructure, and cytochemistry. Accordingly, control ambient samples were collected in the field, and two different treatments were performed in the laboratory. Plants were exposed to PAR at 60 mu mol photons m(-2) s(-1) and PAR+UVBR at 0.35 W m(-2) for 3 h per day during 21 days of in vitro cultivation. Confocal laser scanning microscopy analysis of the vegetative cells showed single stellate chloroplast in ambient and PAR samples, but in PAR+UVBR-exposed plants, the chloroplast showed alterations in the number and form of arms. Under PAR+UVBR treatment, the thylakoids of the chloroplasts were disrupted, and an increase in the number of plastoglobuli was observed, in addition to mitochondria, which appeared with irregular, disrupted morphology compared to ambient and PAR samples. After UVBR exposure, the formation of carpospores was also observed. Plants under ambient conditions, as well as those treated with PAR and PAR+UVBR, all showed different concentrations of enzymatic response, including glutathione peroxidase and reductase activity. In summary, the present study demonstrates that P. acanthophora var. brasiliensis shows the activation of distinct mechanisms against natural radiation, PAR and PAR+UVBR.

Genetic polymorphisms modulate the folate metabolism of Brazilian individuals with Down syndrome

Individuals with Down syndrome (DS) carry three copies of the Cystathionine beta-synthase (C beta S) gene. The increase in the dosage of this gene results in an altered profile of metabolites involved in the folate pathway, including reduced homocysteine (Hcy), methionine, S-adenosylhomocysteine (SAH) and S-adenosylmethionine (SAM). Furthermore, previous studies in individuals with DS have shown that genetic variants in genes involved in the folate pathway influence the concentrations of this metabolism's products. The purpose of this study is to investigate whether polymorphisms in genes involved in folate metabolism affect the plasma concentrations of Hcy and methylmalonic acid (MMA) along with the concentration of serum folate in individuals with DS. Twelve genetic polymorphisms were investigated in 90 individuals with DS (median age 1.29 years, range 0.07-30.35 years; 49 male and 41 female). Genotyping for the polymorphisms was performed either by polymerase chain reaction (PCR) based techniques or by direct sequencing. Plasma concentrations of Hcy and MMA were measured by liquid chromatography-tandem mass spectrometry as previously described, and serum folate was quantified using a competitive immunoassay. Our results indicate that the MTHFR C677T, MTR A2756G, TC2 C776G and BHMT G742A polymorphisms along with MMA concentration are predictors of Hcy concentration. They also show that age and Hcy concentration are predictors of MMA concentration. These findings could help to understand how genetic variation impacts folate metabolism and what metabolic consequences these variants have in individuals with trisomy 21.

L-Carnitine induces recovery of liver lipid metabolism in cancer cachexia

Cancer cachexia causes metabolic alterations with a marked effect on hepatic lipid metabolism. l-Carnitine modulates lipid metabolism and its supplementation has been proposed as a therapeutic strategy in many diseases. In the present study, the effects of l-carnitine supplementation on gene expression and on liver lipid metabolism-related proteins was investigated in cachectic tumour-bearing rats. Wistar rats were assigned to receive 1 g/kg of l-carnitine or saline. After 14 days, supplemented and control animals were assigned to a control (N), control supplemented with l-carnitine (CN), tumour-bearing Walker 256 carcinosarcoma (TB) and tumour-bearing supplemented with l-carnitine (CTB) group. The mRNA expression of carnitine palmitoyltransferase I and II (CPT I and II), microsomal triglyceride transfer protein (MTP), liver fatty acid-binding protein (L-FABP), fatty acid translocase (FAT/CD36), peroxisome proliferator-activated receptor-alpha (PPAR-alpha) and organic cation transporter 2 (OCTN2) was assessed, and the maximal activity of CPT I and II in the liver measured, along with plasma and liver triacylglycerol content. The gene expression of MTP, and CPT I catalytic activity were reduced in TB, who also showed increased liver (150%) and plasma (3.3-fold) triacylglycerol content. l-Carnitine supplementation was able to restore these parameters back to control values (p < 0.05). These data show that l-carnitine preserves hepatic lipid metabolism in tumour-bearing animals, suggesting its supplementation to be of potential interest in cachexia.

Biomimetic in vitro oxidation of lapachol: A model to predict and analyse the in vivo phase I metabolism of bioactive compounds

The bioactive naphtoquinone lapachol was studied in vitro by a biomimetic model with Jacobsen catalyst (manganese(III) salen) and iodosylbenzene as oxidizing agent. Eleven oxidation derivatives were thus identified and two competitive oxidation pathways postulated. Similar to Mn(III) porphyrins, Jacobsen catalyst mainly induced the formation of para-naphtoquinone derivatives of lapachol, but also of two ortho-derivatives. The oxidation products were used to develop a GC MS (SIM mode) method for the identification of potential phase I metabolites in vivo. Plasma analysis of Wistar rats orally administered with lapachol revealed two metabolites, alpha-lapachone and dehydro-alpha-lapachone. Hence, the biomimetic model with a manganese salen complex has evidenced its use as a valuable tool to predict and elucidate the in vivo phase I metabolism of lapachol and possibly also of other bioactive natural compounds. (C) 2012 Elsevier Masson SAS. All rights reserved.

Interstrain differences in the severity of liver injury induced by a choline- and folate-deficient diet in mice are associated with dysregulation of genes involved in lipid metabolism

Nonalcoholic fatty liver disease (NAFLD) is a major health problem and a leading cause of chronic liver disease in the United States and developed countries. In humans, genetic factors greatly influence individual susceptibility to NAFLD. The goals of this study were to compare the magnitude of interindividual differences in the severity of liver injury induced by methyl-donor deficiency among individual inbred strains of mice and to investigate the underlying mechanisms associated with the variability. Feeding mice a choline-and folate-deficient diet for 12 wk caused liver injury similar to NAFLD. The magnitude of liver injury varied among the strains, with the order of sensitivity being A/J approximate to C57BL/6J approximate to C3H/HeJ < 129S1/SvImJ approximate to CAST/EiJ < PWK/PhJ < WSB/EiJ. The interstrain variability in severity of NAFLD liver damage was associated with dysregulation of genes involved in lipid metabolism, primarily with a down-regulation of the peroxisome proliferator receptor alpha (PPAR alpha)-regulated lipid catabolic pathway genes. Markers of oxidative stress and oxidative stress-induced DNA damage were also elevated in the livers but were not correlated with severity of liver damage. These findings suggest that the PPAR alpha-regulated metabolism network is one of the key mechanisms determining interstrain susceptibility and severity of NAFLD in mice.-Tryndyak, V., de Conti, A., Kobets, T., Kutanzi, K., Koturbash, I., Han, T., Fuscoe, J. C., Latendresse, J. R., Melnyk, S., Shymonyak, S., Collins, L., Ross, S. A., Rusyn, I., Beland, F. A., Pogribny, I. P. Interstrain differences in the severity of liver injury induced by a choline-and folate-deficient diet in mice are associated with dysregulation of genes involved in lipid metabolism. FASEB J. 26, 4592-4602 (2012). www.fasebj.org

In silico and in vivo analysis reveal a novel gene in Saccharomyces cerevisiae trehalose metabolism

Abstract Background The ability to respond rapidly to fluctuations in environmental changes is decisive for cell survival. Under these conditions trehalose has an essential protective function and its concentration increases in response to enhanced expression of trehalose synthase genes, TPS1, TPS2, TPS3 and TSL1. Intriguingly, the NTH1 gene, which encodes neutral trehalase, is highly expressed at the same time. We have previously shown that trehalase remains in its inactive non-phosphorylated form by the action of an endogenous inhibitor. Recently, a comprehensive two-hybrid analysis revealed a 41-kDa protein encoded by the YLR270w ORF, which interacts with NTH1p. Results In this work we investigate the correlation of this Trehalase Associated Protein, in trehalase activity regulation. The neutral trehalase activity in the ylr270w mutant strain was about 4-fold higher than in the control strain. After in vitro activation by PKA the ylr270w mutant total trehalase activity increased 3-fold when compared to a control strain. The expression of the NTH1 gene promoter fused to the heterologous reporter lacZ gene was evaluated. The mutant strain lacking YLR270w exhibited a 2-fold increase in the NTH1-lacZ basal expression when compared to the wild type strain. Conclusions These results strongly indicate a central role for Ylr270p in inhibiting trehalase activity, as well as in the regulation of its expression preventing a wasteful futile cycle of synthesis-degradation of trehalose.

Cell organisation, sulphur metabolism and ion transport-related genes are differentially expressed in Paracoccidioides brasiliensis mycelium and yeast cells

Abstract Background Mycelium-to-yeast transition in the human host is essential for pathogenicity by the fungus Paracoccidioides brasiliensis and both cell types are therefore critical to the establishment of paracoccidioidomycosis (PCM), a systemic mycosis endemic to Latin America. The infected population is of about 10 million individuals, 2% of whom will eventually develop the disease. Previously, transcriptome analysis of mycelium and yeast cells resulted in the assembly of 6,022 sequence groups. Gene expression analysis, using both in silico EST subtraction and cDNA microarray, revealed genes that were differential to yeast or mycelium, and we discussed those involved in sugar metabolism. To advance our understanding of molecular mechanisms of dimorphic transition, we performed an extended analysis of gene expression profiles using the methods mentioned above. Results In this work, continuous data mining revealed 66 new differentially expressed sequences that were MIPS(Munich Information Center for Protein Sequences)-categorised according to the cellular process in which they are presumably involved. Two well represented classes were chosen for further analysis: (i) control of cell organisation – cell wall, membrane and cytoskeleton, whose representatives were hex (encoding for a hexagonal peroxisome protein), bgl (encoding for a 1,3-β-glucosidase) in mycelium cells; and ags (an α-1,3-glucan synthase), cda (a chitin deacetylase) and vrp (a verprolin) in yeast cells; (ii) ion metabolism and transport – two genes putatively implicated in ion transport were confirmed to be highly expressed in mycelium cells – isc and ktp, respectively an iron-sulphur cluster-like protein and a cation transporter; and a putative P-type cation pump (pct) in yeast. Also, several enzymes from the cysteine de novo biosynthesis pathway were shown to be up regulated in the yeast form, including ATP sulphurylase, APS kinase and also PAPS reductase. Conclusion Taken together, these data show that several genes involved in cell organisation and ion metabolism/transport are expressed differentially along dimorphic transition. Hyper expression in yeast of the enzymes of sulphur metabolism reinforced that this metabolic pathway could be important for this process. Understanding these changes by functional analysis of such genes may lead to a better understanding of the infective process, thus providing new targets and strategies to control PCM.

Metabolic flux analysis for directing metabolism of Streptomyces olindensis from growth to anti-tumor drug (cosmomycin) production

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior, CAPES, Brazil

Body weight, metabolism and clock genes

Biological rhythms are present in the lives of almost all organisms ranging from plants to more evolved creatures. These oscillations allow the anticipation of many physiological and behavioral mechanisms thus enabling coordination of rhythms in a timely manner, adaption to environmental changes and more efficient organization of the cellular processes responsible for survival of both the individual and the species. Many components of energy homeostasis exhibit circadian rhythms, which are regulated by central (suprachiasmatic nucleus) and peripheral (located in other tissues) circadian clocks. Adipocyte plays an important role in the regulation of energy homeostasis, the signaling of satiety and cellular differentiation and proliferation. Also, the adipocyte circadian clock is probably involved in the control of many of these functions. Thus, circadian clocks are implicated in the control of energy balance, feeding behavior and consequently in the regulation of body weight. In this regard, alterations in clock genes and rhythms can interfere with the complex mechanism of metabolic and hormonal anticipation, contributing to multifactorial diseases such as obesity and diabetes. The aim of this review was to define circadian clocks by describing their functioning and role in the whole body and in adipocyte metabolism, as well as their influence on body weight control and the development of obesity.

Lipoprotein metabolism in patients with type 1 diabetes under intensive insulin treatment

Background Type 1 diabetes (T1DM) is frequently accompanied by dyslipidemia related with insulin-dependent steps of the intravascular lipoprotein metabolism. T1DM dyslipidemia may predispose to precocious cardiovascular disease and the lipid status in T1DM under intensive insulin treatment has not been sufficiently explored. The aim was to investigate the plasma lipids and the metabolism of LDL and HDL in insulin-treated T1DM patients with high glycemic levels. Methods Sixteen male patients with T1DM (26 ± 7 yrs) with glycated hemoglobin >7%, and 15 control subjects (28 ± 6 yrs) were injected with a lipid nanoemulsion (LDE) resembling LDL and labeled with 14C-cholesteryl ester and 3H-free-cholesterol for determination of fractional clearance rates (FCR, in h-1) and cholesterol esterification kinetics. Transfer of labeled lipids from LDE to HDL was assayed in vitro. Results LDL-cholesterol (83 ± 15 vs 100 ± 29 mg/dl, p=0.08) tended to be lower in T1DM than in controls; HDL-cholesterol and triglycerides were equal. LDE marker 14C-cholesteryl ester was removed faster from plasma in T1DM patients than in controls (FCR=0.059 ± 0.022 vs 0.039 ± 0.022h-1, p=0.019), which may account for their lower LDL-cholesterol levels. Cholesterol esterification kinetics and transfer of non-esterified and esterified cholesterol, phospholipids and triglycerides from LDE to HDL were also equal. Conclusion T1DM patients under intensive insulin treatment but with poor glycemic control had lower LDL-cholesterol with higher LDE plasma clearance, indicating that LDL plasma removal was even more efficient than in controls. Furthermore, HDL-cholesterol and triglycerides, cholesterol esterification and transfer of lipids to HDL, an important step in reverse cholesterol transport, were all normal. Coexistence of high glycemia levels with normal intravascular lipid metabolism may be related to differences in exogenous insulin bioavailabity and different insulin mechanisms of action on glucose and lipids. Those findings may have important implications for prevention of macrovascular disease by intensive insulin treatment.

Absorption and metabolism of volatile fatty acids by rumen and omasum

Volatile fatty acids (VFA) absorption and metabolic capacity of rumen and omasum were compared, in vitro. Fragments of rumen wall and omasum laminae were taken from eight adult crossbred bovines. An isolated fragment of the mucosa was fitted in a tissue diffusion chamber. Valeric acid and CrEDTA were added to ruminal fluid and placed on the mucosal side and buffer solution was placed on the serosal side. Fractional absorption rates were measured by exponential VFA:Cr ratio decay over time. Metabolism rate was determined as the difference between VFA absorbed and VFA which appeared on the serosal side over time. Mitotic index was higher in omasum (0.52%) than in rumen epithelium (0.28%). VFA fractional absorption rate was higher in omasum (4.6%/h.cm²) than in rumen (0.4%/h.cm²). Acetate, propionate, butyrate, and valerate showed similar fractional absorption rates in both fragments. Percentage of metabolized acetate and propionate was lower than butyrate and valerate in both stomach compartments. In the rumen, individual VFA metabolism rates were similar (mean of 7.7 , but in the omasum, valerate (90.0 was more metabolized than butyrate (59.6 propionate (69.8 and acetate (51.7 . Correlation between VFA metabolism and mitotic index was positive in the rumen and in the omasum. In conclusion, VFA metabolism and absorption potential per surface of the omasum is higher than that of the rumen. Variations on rumen and omasum absorption capacities occur in the same way, and there are indications that factors capable of stimulating rumen wall proliferation are similarly capable of stimulating omasum walls.