Maintenance Of Redox Balance Research Articles

Excessive Thiol Nitrosylation Predisposes to Western Diet‐Induced Redox Imbalance and Erectile Dysfunction

ObjectivesBinding of nitric oxide (NO) to a cysteine thiol is termed S‐nitrosylation, which is emerging as an important means of NO‐mediated signal transduction independent of stimulation of soluble guanylate cyclase. However, the impact of S‐nitrosylation on erectile function remains unclear. S‐nitrosylation is a redox sensitive post‐translational modification, which is thought to be promoted by reactive oxygen species (ROS). Protein S‐nitrosylation levels are regulated in part by S‐nitrosoglutatione reductase (GSNOR), which acts to de‐nitrosylate protein thiols. The objectives of this study were to determine the role of GSNOR on maintenance of redox balance and erectile function in response to a Western diet (WD) high in fat and sugar.Materials and MethodsYoung male wild type (WT; n = 110) and GSNOR−/− (n = 108) mice were fed either a control diet or a WD ad libitum for 3, 6, 9, or 12 weeks. Following the dietary intervention, erectile function was assessed by measuring intracavernosal pressure (ICP) and mean arterial pressure (MAP) during cavernous nerve stimulation. In separate mice, in vivo ROS production was measured in the penis utilizing a novel microdialysis approach. Microdialysis probes were inserted into the penis of anesthetized mice and perfused with saline containing 100 uM Amplex Ultrared, 1 U/ml horseradish peroxidase, and 10 U/ml superoxide dismutase. ROS convert the reagents to a fluorescent byproduct resorufin, which was measured in the outflowing dialysate. Penile tissue was harvested from mice and processed for enrichment of nitrosylated thiols. Nitrosylated thiols were labeled with iodoTMT, and nitrosylated proteins were detected by mass spectrometry.ResultsNeither ROS generation (WT: 0.66 ± 0.09 vs. GSNOR−/− 0.96 ± 0.12 μM H2O2; p = 0.41) nor erectile function (WT:8.44 ± 1.4 vs. GSNOR−/− 9.6 ± 1.6 ICP area/MAP; p = 0.97) were different between genotypes fed the control diet. However, ROS were elevated following just 3 weeks of WD consumption in GSNOR−/− mice (1.44 ± 0.19; p = 0.01), whereas ROS increased following 6 weeks of WD in WT mice (1.29 ± 0.19; p = 0.02). Subsequent to increases in ROS, ED was evident after 6 weeks of WD in GSNOR−/− mice (5.0 ± 1.1, p = 0.04) and after 12 weeks of WD in WT mice (3.1 ± 0.6, p = 0.02). 249 unique peptides corresponding to 136 different proteins with nitrosylated thiol groups were identified by mass spectrometry.ConclusionsMice lacking GSNOR demonstrated an impaired ability to maintain redox balance in the face of the pro‐oxidant stimulus of the WD, prompting accelerated development of erectile dysfunction. These data implicate a protective role for GSNOR in the penis and suggest that excessive S‐nitrosylation may negatively regulate erectile function.Support or Funding InformationSupported by a mini grant from the National Kidney Foundation of Maryland and National Institutes of Health grant 1F32DK100082.

The transport activity of P-glycoprotein upon a change of the redox balance in lymphocytes of patients with chronic B-lymphocytic leukemia

The effect of drugs used in the treatment of chronic B-lymphocytic leukemia on the functional activity of P-glycoprotein, which is a membrane transporter associated with the phenomenon of multiple drug resistance, in B-lymphocytes of leukemia patients has been investigated. The level of reactive oxygen species and the content of low molecular weight antioxidants in the leukemic cells during the metabolic processing of chemotherapy drugs has been assessed. The degree of involvement of low molecular weight antioxidants in the maintenance of redox balance in the lymphocytes of drug-treated leukemia patients has been characterized. Variation of the levels of reactive oxygen species in leukemic B-lymphocytes within a certain range, as well as the subsequent return of these values to the levels of intact cells, was shown to enhance the P-glycoprotein transport activity. The viability of leukemic B-cells decreased upon exposure to antitumor compounds and depended on the redox balance. No statistically significant correlation between the P-glycoprotein transport activity and the viability of lymphocytes during the detoxification from drugs in B-cell chronic lymphocytic leukemia patients has been detected; however, the transport activity of the protein was directly correlated to the viability of leukemia cells within 15 h after exposure of the cells to H2O2.

IL-1β maintains the redox balance by regulating glutaredoxin 1 expression during oral carcinogenesis.

Interleukin-1 beta (IL-1β) is a pleiotropic cancer-inflammation-linked cytokine which has been reported upregulated in many cancers. In our previous study, IL-1β was found to be one of the key node genes during oral malignant transformation, and glutaredoxin 1 (Grx1) was identified as one of the downstream genes of IL-1β in tumor microenvironment. Grx1 is ubiquitous oxidoreductase which is necessary for scavenging reactive oxygen species (ROS) and the intracellular redox balance maintenance. Tissues from different stages of mucosal malignant transformation were obtained from 4NQO-induced rat oral carcinogenesis model and human mucosa for Grx1 expression detection by immunohistochemical staining. The intracellular ROS levels and Grx1 mRNA level of oral squamous carcinoma cell CAL27 were detected after IL-1β treatment with or without pretreatment of IL-1Ra or NAC, respectively. The ROS levels were detected in Leti-si-IL-1β and Leti-si-NC CAL27 cells after IL-1β stimulation. The invasion and migration abilities of CAL27 cells were tested by transwell assay after IL-1β stimulation with or without pretreatment of IL-1Ra. Grx1 expression was associated with the malignant transformation process in vivo. Exogenous IL-1β upregulated the intracellular ROS level and the expression of Grx1 in CAL27 cells, which could be counteracted by IL-1Ra. The intracellular ROS accumulation induced by exogenous IL-1β was responsible for the Grx1 upregulation. Endogenous IL-1β acted as a switch in regulating the ROS level by modulating Grx1 expression, which was involved in the invasion and migration of OSCC cells. IL-1β finely orchestrated the redox balance during carcinogenesis by modulating Grx1 expression.

Lactobacillus oligofermentans glucose, ribose and xylose transcriptomes show higher similarity between glucose and xylose catabolism-induced responses in the early exponential growth phase.

BackgroundLactobacillus oligofermentans has been mostly isolated from cold-stored packaged meat products in connection with their spoilage, but its precise role in meat spoilage is unknown. It belongs to the L. vaccinostercus group of obligate heterofermentative lactobacilli that generally ferment pentoses (e.g. xylose and ribose) more efficiently than hexoses (e.g. glucose). However, more efficient hexose utilization can be induced. The regulation mechanisms of the carbohydrate catabolism in such bacteria have been scarcely studied. To address this question, we provided the complete genome sequence of L. oligofermentans LMG 22743T and generated time course transcriptomes during its growth on glucose, ribose and xylose.ResultsThe genome was manually annotated and its main functional features were examined. L. oligofermentans was confirmed to be able to efficiently utilize several hexoses and maltose, which is, presumably, induced by its repeated cultivation with glucose in vitro. Unexpectedly, in the beginning of the exponential growth phase, glucose- and xylose-induced transcriptome responses were more similar, whereas toward the end of the growth phase xylose and ribose transcriptomes became more alike. The promoter regions of genes simultaneously upregulated both on glucose and xylose in comparison with ribose (particularly, hexose and xylose utilization genes) were found to be enriched in the CcpA- binding site. Transcriptionally, no glucose-induced carbon catabolite repression was detected. The catabolism of glucose, which requires initial oxidation, led to significant overexpression of the NAD(P)H re-oxidation genes, the upstream regions of which were found to contain a motif, which was highly similar to a Rex repressor binding site.ConclusionsThis paper presents the second complete genome and the first study of carbohydrate catabolism-dependent transcriptome response for a member of the L. vaccinostercus group. The transcriptomic changes detected in L. oligofermentans for growth with different carbohydrates differ significantly from those of facultative heterofermentative lactobacilli. The mechanism of CcpA regulation, putatively contributing to the observed similarities between glucose- and xylose-induced transcriptome responses and the absence of stringent carbon catabolite control, requires further studies. Finally, the cell redox balance maintenance, in terms of the NAD(P)+/NAD(P)H ratio, was predicted to be regulated by the Rex transcriptional regulator, supporting the previously made inference of Rex-regulons for members of the Lactobacillaceae family.Electronic supplementary materialThe online version of this article (doi:10.1186/s12864-016-2840-x) contains supplementary material, which is available to authorized users.

NADPH production, a growth marker, is stimulated by maslinic acid in gilthead sea bream by increased NADP-IDH and ME expression

NADPH plays a central role in reductive biosynthesis of membrane lipids, maintenance of cell integrity, protein synthesis and redox balance maintenance. Hence, NADPH is involved in the growth and proliferation processes. In addition, it has been shown that changes in nutritional conditions produced changes in NADPH levels and growth rate. Maslinic acid (MA), a pentacyclic triterpene of natural origin, is able to stimulate NADPH production, through regulation of the two oxidative phase dehydrogenases of the pentose phosphate pathway. Our main objective was to study the effects of MA on the kinetic behaviour and on the molecular expression of two NADPH-generating systems, NADP-dependent isocitrate dehydrogenase (NADP-IDH) and malic enzyme (ME), in the liver and white muscle of gilthead sea bream (Sparus aurata). Four groups of 12g of a mean body mass were fed for 210days in a fish farm, with diets containing 0 (control), and 0.1g of MA per kg of diet. Two groups were fed ad libitum (C-AL and MA-AL) and another's two, with restricted diet of 1% of fish weight (C-R and MA-R). Results showed that MA significantly increased the main kinetic parameter of the NADPH-forming enzymes (NADP-IDH and ME). In this sense, specific activity, maximum velocity, catalytic efficiency and activity ratio values were higher in MA conditions than control groups. Moreover, these changes were observed in both feeding regimen, AL and R. Meanwhile, the Michaelis constant changed mainly in groups fed with the MA and restricted diet, these changes are related to the best substrate affinity by enzyme. Moreover, in the Western-blot result, we found that MA increased both protein levels studied, this behaviour being consistent with the regulation of the number of enzyme molecules. All results, indicate that MA, independently of the fed regimen, could potentially be a nutritional additive for fish as it improved the metabolic state of fish, as consequence of increased activity and expression of NADP-IDH and ME enzymes.

Abstract A41: Leveraging metabolic dependencies in cancer: Cysteine addiction in pancreatic cancer cells

Abstract Pancreatic cancer is a devastating and lethal malignancy, estimated to kill over 40,000 Americans per year. Little has changed in the treatment of this disease over the past half-century, highlighting the need for an improved understanding of the molecular mechanisms driving tumor development and maintenance. The malignant behavior of tumor cells is underwritten by alterations to their cellular metabolism, which is optimized to meet the catabolic needs of a rapidly dividing cell. One of the consequences of this oncogenic growth is increased generation of reactive oxygen species (ROS), a potentially toxic byproduct of many metabolic reactions. Interestingly, oncogenes such as Kras, the dominant driver of pancreatic cancer, can counteract ROS toxicity through activation of an antioxidant program, creating a highly reducing intracellular environment. Cysteine is the rate-limiting precursor for the synthesis of glutathione (GSH), the preeminent antioxidant of the cell, and as such may represent a key metabolite in the maintenance of redox balance in tumor cells. In this study we explore the importance of cysteine metabolism in the viability of pancreatic cancer cells and test the effects of cysteine deprivation and the pharmacological inhibition of cysteine uptake on a variety of pancreatic cancer cell lines. To explore the role of cysteine in pancreatic cancer growth, we cultured a panel of human and murine pancreatic cancer cell lines in the absence of cysteine. We found that cysteine withdrawal rapidly induced a peculiar form of cell death called ferroptosis, which can result from the catastrophic accumulation of certain lipid peroxides. Cystine, the oxidized form of cysteine, is the predominant extracellular source of cysteine in most cells, and it is normally imported into the cell by a specific amino acid transporter known as system xc-. We show that pharmacological inhibition of sytem xc- causes rapid cell death in most pancreatic cancer cells through ferroptosis. We find, however, that some cells do exhibit baseline resistance to cysteine deprivation or system xc- inhibition. We hypothesized that these cells utilize methionine to synthesize cysteine through transsulfuration. We show that these cells can be sensitized to system xc- inhibition when co-treated with propargyglycine (PPG), an irreversible inhibitor of gamma-cystathionase, an enzyme required for transsulfuration. These findings imply that cysteine is an important substrate for appropriate redox homeostasis in these cells. To further explore if GSH synthesis is the key mediator of cysteine-deprivation induced cell death, we treated pancreatic tumor cells with buthionine sulfoxamine (BSO), a potent and specific inhibitor of GSH synthesis. To our surprise, cells treated with BSO do not exhibit any of the ill-effects we see in cysteine-deprived pancreatic cancer cells. Therefore, it is unlikely that the cell death seen in cells deprived of cysteine is due solely to GSH depletion, but possibly due to the activity of other antioxidant pathways that require cysteine. In conclusion, we find that pancreatic cancer cells are highly sensitive to cysteine-deprivation and to treatment with system xc- inhibitors, and that the cell death caused by this stress is an iron-dependent, oxidative form of cell death called ferroptosis. Furthermore, it appears that GSH, the primary antioxidant molecule of the cell, is not solely responsible for mediating this cell death, arguing that cysteine may play other important roles in maintaining redox homeostasis in these cells. Given that previous studies show that global system xc- knockout mice do not exhibit any changes in cellular GSH content, these findings may represent a targetable metabolic dependency in pancreatic cancer cells. Citation Format: Michael A. Badgley, Carmine F. Palermo, Stephen A. Sastra, Brent R. Stockwell, Kenneth P. Olive. Leveraging metabolic dependencies in cancer: Cysteine addiction in pancreatic cancer cells. [abstract]. In: Proceedings of the AACR Special Conference: Metabolism and Cancer; Jun 7-10, 2015; Bellevue, WA. Philadelphia (PA): AACR; Mol Cancer Res 2016;14(1_Suppl):Abstract nr A41.

Vitamin C in Health and Disease: Its Role in the Metabolism of Cells and Redox State in the Brain.

Ever since Linus Pauling published his studies, the effects of vitamin C have been surrounded by contradictory results. This may be because its effects depend on a number of factors such as the redox state of the body, the dose used, and also on the tissue metabolism. This review deals with vitamin C pharmacokinetics and its participation in neurophysiological processes, as well as its role in the maintenance of redox balance. The distribution and the concentration of vitamin C in the organs depend on the ascorbate requirements of each and on the tissue distribution of sodium-dependent vitamin C transporter 1 and 2 (SVCT1 and SVCT2). This determines the specific distribution pattern of vitamin C in the body. Vitamin C is involved in the physiology of the nervous system, including the support and the structure of the neurons, the processes of differentiation, maturation, and neuronal survival; the synthesis of catecholamine, and the modulation of neurotransmission. This antioxidant interacts with self-recycling mechanisms, including its participation in the endogenous antioxidant system. We conclude that the pharmacokinetic properties of ascorbate are related to the redox state and its functions and effects in tissues.

The MIA pathway: a key regulator of mitochondrial oxidative protein folding and biogenesis.

ConspectusMitochondria are fundamental intracellular organelles with key roles in important cellular processes like energy production, Fe/S cluster biogenesis, and homeostasis of lipids and inorganic ions. Mitochondrial dysfunction is consequently linked to many human pathologies (cancer, diabetes, neurodegeneration, stroke) and apoptosis. Mitochondrial biogenesis relies on protein import as most mitochondrial proteins (about 10–15% of the human proteome) are imported after their synthesis in the cytosol. Over the last several years many mitochondrial translocation pathways have been discovered. Among them, the import pathway that targets proteins to the intermembrane space (IMS) stands out as it is the only one that couples import to folding and oxidation and results in the covalent modification of the incoming precursor that adopt internal disulfide bonds in the process (the MIA pathway). The discovery of this pathway represented a significant paradigm shift as it challenged the prevailing dogma that the endoplasmic reticulum is the only compartment of eukaryotic cells where oxidative folding can occur.The concept of the oxidative folding pathway was first proposed on the basis of folding and import data for the small Tim proteins that have conserved cysteine motifs and must adopt intramolecular disulfides after import so that they are retained in the organelle. The introduction of disulfides in the IMS is catalyzed by Mia40 that functions as a chaperone inducing their folding. The sulfhydryl oxidase Erv1 generates the disulfide pairs de novo using either molecular oxygen or, cytochrome c and other proteins as terminal electron acceptors that eventually link this folding process to respiration. The solution NMR structure of Mia40 (and supporting biochemical experiments) showed that Mia40 is a novel type of disulfide donor whose recognition capacity for its substrates relies on a hydrophobic binding cleft found adjacent to a thiol active CPC motif. Targeting of the substrates to this pathway is guided by a novel type of IMS targeting signal called ITS or MISS. This consists of only 9 amino acids, found upstream or downstream of a unique Cys that is primed for docking to Mia40 when the substrate is accommodated in the Mia40 binding cleft. Different routes exist to complete the folding of the substrates and their final maturation in the IMS. Identification of new Mia40 substrates (some even without the requirement of their cysteines) reveals an expanded chaperone-like activity of this protein in the IMS. New evidence on the targeting of redox active proteins like thioredoxin, glutaredoxin, and peroxiredoxin into the IMS suggests the presence of redox-dependent regulatory mechanisms of the protein folding and import process in mitochondria. Maintenance of redox balance in mitochondria is crucial for normal cell physiology and depends on the cross-talk between the various redox signaling processes and the mitochondrial oxidative folding pathway.

Abstract B47: Modulating the NQO1-dependent ‘kiss of death’ mechanism of action of NQO1 bioactivatable drugs

Abstract Metabolic adaptations in tumors are necessary to balance the need for cellular energy supply, macromoleclar biosynthesis and maintenance of redox balance. A critical molecule produced as a result of this altered metabolism is nicotinamide adenine dinucleotide phosphate (NADPH). NADPH is a critical co-factor that provides reducing power to maintain cytoprotective concentrations of reduced glutathione against reactive oxygen species (ROS) that are produced during rapid growth and proliferation. In pancreatic cancers, NADPH can be formed by the NAD+ salvage pathway through the enzyme, Nicotinamide phosphoribosyltransferase (NamPT), as well as through the non-canonical metabolism of glutamine through glutaminase-1 (GLS1). While NamPT and GLS1 inhibitors have shown some anti-tumor activity in vitro and in vivo, these inhibitors lack tumor specificity and can have an extensive toxicity profile due to the requirement of NADPH synthesis in proliferating normal cells/tissue. We recently discovered that NAD(P)H:quinone oxidoreductase 1 (NQO1) levels were elevated 5- to 40-fold in >80% of pancreatic tumors vs associated normal tissue. As important, we noted that catalase levels were inversely expressed comparatively, elevated in normal and suppressed in tumor tissue. Thus, NQO1 represents a unique target to exploit for the therapeutic elimination of pancreatic cancers. While NQO1 is normally a detoxifying (Phase II) enzyme, we have discovered two unique classes of compounds that are ‘bioactivated’ in a futile redox cycle by the enzyme. NQO1 bioactivatable drugs, such as β-lapachone (β-lap) and deoxynyboquinone (DNQ), generate hydrogen peroxide as a mechanism to hyperactivate poly(ADP-ribose) polymerase 1 (PARP1), and to rapidly deplete NAD+ and ATP in a selective manner to kill tumors. Normal cells are protected due to low NQO1 and relatively high Catalase levels. We found that the tumor specificity and efficacy of NamPT or GLS1 inhibition can be greatly increased when small molecule NamPT inhibitors (e.g., FK866) or GLS1 inhibitors (e.g., BPTES) are used in combination with NQO1-bioactivatable drugs. With combination treatment, cells are primed for ROS-mediated damage due to reduced glutathione and NADPH levels; they are unable to recover ATP, NAD+, and NADPH synthesis even after only a 2 h exposure to NQO1 bioactivatable drugs, and they rapidly die through a caspase-independent, parthanatos (programmed necrosis) mechanism. Our findings indicate that NQO1-bioactivatable drugs may improve the sensitivities of clinical grade GLS1 and NamPT inhibitors in pancreatic cancer patients. This work was supported by an AACR/Pancreatic Cancer Action Network - George and June Block Foundation - Innovator Award and an NIH/NCI R01 CA102971 grant to DAB. Citation Format: Zachary Moore, Gaurab Chakrabarti, Costas Lyssiotis, Ralph Deberardinis, Boothman A. David. Modulating the NQO1-dependent ‘kiss of death’ mechanism of action of NQO1 bioactivatable drugs. [abstract]. In: Proceedings of the AACR Special Conference on Pancreatic Cancer: Innovations in Research and Treatment; May 18-21, 2014; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2015;75(13 Suppl):Abstract nr B47.

MNADK, a Long-Awaited Human Mitochondrion-Localized NAD Kinase.

Nicotinamide adenine dinucleotide (NAD) and its phosphorylated form, NADP, play essential roles in numerous cellular processes in all organisms. NADP maintains a pool of its reducing equivalent, NADPH, which regenerates cellular oxidative defense systems to counteract oxidative damages. Mitochondria represent a major source of oxidative stress, because the majority of superoxide, a reactive oxygen species, is generated from the mitochondrial respiratory chain. Therefore, as universal electron carriers in cellular electron transfer reactions, the pyridine nucleotides are required by mitochondria for both antioxidant protection and biosynthetic pathways. The NAD kinase (NADK) is the sole NADP biosynthetic enzyme. Because NADP is membrane-impermeable, eukaryotes need compartment-specific NADKs for different organelles. Consistently, in both yeast and plants, three compartment-specific NADKs have been identified. In contrast, even though the first human NADK, a cytosolic one, was identified in 2001, the identity of a hypothesized mitochondrial NADK remained elusive, until a recent discovery that the uncharacterized human gene C5ORF33 encodes a mitochondrion-localized NADK, referred to as MNADK. Three groups have characterized MNADK functions based on distinct systems involving yeast, mouse, and human studies, from aspects of both in vitro and in vivo evidence. MNADK is a mitochondrial NADK that is enriched and nutritionally-regulated in mouse liver, and a MNADK-deficient patient exhibits symptoms characteristic of mitochondrial disease. The identification of MNADK provides a key clue to the mechanism involved in mitochondrial NADPH production and the maintenance of redox balance in mammalian cells. The roles of MNADK in physiological and pathological processes have yet to be discovered.

Redox homeostasis via gene families of ascorbate-glutathione pathway

The imposition of environmental stresses on plants brings about disturbance in their metabolism thereby negatively affecting their growth and development and leading to reduction in the productivity. One of the manifestations of abiotic and biotic stress conditions is the enhanced production of reactive oxygen species (ROS) which can be hazardous to cells. Therefore, in order to protect themselves against toxic ROS, plant cells employ the anti-oxidant defense system. The ascorbate-glutathione pathway (Halliwell-Asada cycle) is an indispensible component of the ROS homeostasis mechanism of plants. This pathway entails the antioxidant metabolites: ascorbate, glutathione and NADPH along with the enzymes linking them. The ascorbate-glutathione pathway is functional in different subcellular compartments and all the enzymes of this pathway exist as multiple isoforms. The expression of different isoforms of the enzymes of ascorbate-glutathione pathway is developmentally as well as spatially regulated. Moreover, various abiotic and biotic stress conditions modulate the expression of the enzyme- isoforms differently. It is the intricate regulation of expression of different isoforms of the ascorbate-glutathione pathway enzymes that helps in the maintenance of redox balance in plants under various abiotic and biotic stress conditions. The present review provides an insight into the gene families of the ascorbate-glutathione pathway, shedding light on their role in different abiotic and biotic stress conditions as well as in the growth and development of plants.

Thiamine increases the resistance of baker's yeast Saccharomyces cerevisiae against oxidative, osmotic and thermal stress, through mechanisms partly independent of thiamine diphosphate-bound enzymes.

Numerous recent studies have established a hypothesis that thiamine (vitamin B1 ) is involved in the responses of different organisms against stress, also suggesting that underlying mechanisms are not limited to the universal role of thiamine diphosphate (TDP) in the central cellular metabolism. The current work aimed at characterising the effect of exogenously added thiamine on the response of baker's yeast Saccharomyces cerevisiae to the oxidative (1mM H2 O2 ), osmotic (1M sorbitol) and thermal (42°C) stress. As compared to the yeast culture in thiamine-free medium, in the presence of 1.4μM external thiamine, (1) the relative mRNA levels of major TDP-dependent enzymes under stress conditions vs. unstressed control (the 'stress/control ratio') were moderately lower, (2) the stress/control ratio was strongly decreased for the transcript levels of several stress markers localised to the cytoplasm, peroxisomes, the cell wall and (with the strongest effect observed) the mitochondria (e.g. Mn-superoxide dismutase), (3) the production of reactive oxygen and nitrogen species under stress conditions was markedly decreased, with the significant alleviation of concomitant protein oxidation. The results obtained suggest the involvement of thiamine in the maintenance of redox balance in yeast cells under oxidative stress conditions, partly independent of the functions of TDP-dependent enzymes.

Magmas functions as a ROS regulator and provides cytoprotection against oxidative stress-mediated damages.

Redox imbalance generates multiple cellular damages leading to oxidative stress-mediated pathological conditions such as neurodegenerative diseases and cancer progression. Therefore, maintenance of reactive oxygen species (ROS) homeostasis is most important that involves well-defined antioxidant machinery. In the present study, we have identified for the first time a component of mammalian protein translocation machinery Magmas to perform a critical ROS regulatory function. Magmas overexpression has been reported in highly metabolically active tissues and cancer cells that are prone to oxidative damage. We found that Magmas regulates cellular ROS levels by controlling its production as well as scavenging. Magmas promotes cellular tolerance toward oxidative stress by enhancing antioxidant enzyme activity, thus preventing induction of apoptosis and damage to cellular components. Magmas enhances the activity of electron transport chain (ETC) complexes, causing reduced ROS production. Our results suggest that J-like domain of Magmas is essential for maintenance of redox balance. The function of Magmas as a ROS sensor was found to be independent of its role in protein import. The unique ROS modulatory role of Magmas is highlighted by its ability to increase cell tolerance to oxidative stress even in yeast model organism. The cytoprotective capability of Magmas against oxidative damage makes it an important candidate for future investigation in therapeutics of oxidative stress-related diseases.

Oxidative stress and adrenocortical insufficiency.

Maintenance of redox balance is essential for normal cellular functions. Any perturbation in this balance due to increased reactive oxygen species (ROS) leads to oxidative stress and may lead to cell dysfunction/damage/death. Mitochondria are responsible for the majority of cellular ROS production secondary to electron leakage as a consequence of respiration. Furthermore, electron leakage by the cytochrome P450 enzymes may render steroidogenic tissues acutely vulnerable to redox imbalance. The adrenal cortex, in particular, is well supplied with both enzymatic (glutathione peroxidases and peroxiredoxins) and non-enzymatic (vitamins A, C and E) antioxidants to cope with this increased production of ROS due to steroidogenesis. Nonetheless oxidative stress is implicated in several potentially lethal adrenal disorders including X-linked adrenoleukodystrophy, triple A syndrome and most recently familial glucocorticoid deficiency. The finding of mutations in antioxidant defence genes in the latter two conditions highlights how disturbances in redox homeostasis may have an effect on adrenal steroidogenesis.

Osmotic adjustment and maintenance of the redox balance in root tissue may be key points to overcome a mild water deficit during the early growth of wheat

In this investigation we analyzed in detail the consequences of water deficit during the first 4 days of wheat development, focusing on root growth as affected by eventual changes in cell cycle regulation and oxidative processes. Root elongation decreased under water restriction in correlation with the intensity of this limitation, but the total number of cells between the quiescent center and the start of the rapid elongation zone in the root apex did not vary. Neither lipid peroxidation nor protein carbonylation increased in the roots of water-starved seedlings (ψw: −0.6 MPa); accordingly, catalase activity increased, and transcript levels of cat2 gene were enhanced. Superoxide dismutase activity rose at day 2 and 3 and, unlike catalase, displayed quite similar levels on comparing roots and coleoptiles. Proline and total soluble carbohydrates increased in the roots of water-starved seedling. Total conductivity and osmolality were also augmented. No changes in the transcript levels of the markers related to G1-S transition phase of cell cycle could be detected. However, two expansin genes (TaEXPB8 and TaEXPA5) were up-regulated in roots under water deficit. We conclude that wheat root elongation in water-deprived seedlings was simply hampered by lack of water income to cells. The enhanced expression of two root expansin genes is probably related to the eventual need of a quick cell wall expansion to allow the existing root cells to recover normal turgor, in case of sudden rewatering. Maintenance of redox balance, osmotic adjustment and modification of cell wall plasticity may be key points for wheat seedlings to overcome a water restriction during early growth.

Glucose-6-phosphate Dehydrogenase: a Biomarker and Potential Therapeutic Target for Cancer

Re-programming of metabolic pathways is a hallmark of pathological changes in cancer cells. The expression of certain genes that directly control the rate of key metabolic pathways including glycolysis, lipogenesis and nucleotide synthesis is dysregulated for the adaptation and progression of tumor cells to become more aggressive phenotypes. The pentose phosphate pathway controlled by glucose- 6-phosphate dehydrogenase (G6PD) has been appreciated largely to its role as a provider of reducing power and ribose phosphate to the cell for maintenance of redox balance and biosynthesis of nucleotides and lipids. Recently, G6PD has been revealed to be involved in apoptosis, angiogenesis, and the efficacy to anti-cancer therapy, making it as a promising target in cancer therapy. This review summarizes the information about the latest progress relating the activity of the G6PD to cell proliferation, angiogenesis, and resistance to therapy in cancer cells, and discusses the possibility of G6PD as a diagnostic biomarker of cancer and the therapeutic potentials of G6PD inhibitors in cancer treatment. The available data show that G6PD plays a critical role in survival, proliferation, and metastasis of cancer cells. Development of potent and selective G6PD inhibitors would provide novel opportunity for cancer therapy.

Antioxidant defense system responses and role of nitrate reductase in the redox balance maintenance in Bradyrhizobium japonicum strains exposed to cadmium

In this work, we evaluated the effects of cadmium (Cd) on the antioxidant defense system responses and the role of nitrate reductase (NR) in the redox balance maintenance in Bradyrhizobium japonicum strains. For that, B. japonicum USDA110 and its NR defective mutant strain (GRPA1) were used. Results showed that the addition of 10μM Cd did not modify the aerobic growth of the wild type strain while the mutant strain was strongly affected. Anaerobic growth revealed that only the parental strain was able to grow under this condition. Cd reduced drastically the NR activity in B. japonicum USDA110 and increased lipid peroxide content in both strains. Cd decreased reduced glutathione (GSH)/oxidized glutathione (GSSG) ratio in B. japonicum USDA110 although, a significant increased was observed in the mutant GRPA1. GSH-related enzymes were induced by Cd, being more evident the increase in the mutant strain. This different behavior observed between strains suggests that NR enzyme plays an important role in the redox balance maintenance in B. japonicum USDA 110 exposed to Cd.

Serum Oxidant and Antioxidant Status Following an All-Out 21-km Run in Adolescent Runners Undergoing Professional Training—A One-Year Prospective Trial

This study investigated the 1-year longitudinal effect of professional training in adolescent runners on redox balance during intense endurance exercise. Changes in selected serum oxidant and antioxidant status in response to a 21-km running time trial in 10 runners (15.5 ± 1.3 years) undergoing professional training were evaluated twice in 12 months (pre- and post-evaluation). Venous blood samples were collected immediately before and 4-h following the 21-km run for analysis of serum concentrations of thiobarbituric acid-reactive substances (TBARS), xanthine oxidase (XO), catalase (CAT), reduced glutathione (GSH), superoxide dismutase (SOD), and total antioxidant capacity (T-AOC). In pre-evaluation trial, serum TBARS and SOD decreased after the 21-km run (p < 0.05) while XO, GSH, CAT and TAOC were unchanged. In post-evaluation trial, serum TBARS and SOD decreased, whereas XO and CAT increased post-exercise (p < 0.05). Furthermore, pre-exercise serum T-AOC, post-exercise serum XO, CAT, T-AOC (p < 0.05), and GSH (p = 0.057) appeared to be higher than the corresponding pre-evaluation values. The current findings suggest that a professional training regime in adolescent runners is not likely to jeopardize the development of their antioxidant defense. However, uncertainties in the maintenance of redox balance in runners facing increased exercise-induced oxidative stress as a consequence of training-induced enhancement of exercise capacity await further elucidation.

The cardiac acetyl-lysine proteome.

In the heart, lysine acetylation has been implicated in processes ranging from transcriptional control of pathological remodeling, to cardioprotection arising from caloric restriction. Given the emerging importance of this post-translational modification, we used a proteomic approach to investigate the broader role of lysine acetylation in the heart using a guinea pig model. Briefly, hearts were fractionated into myofilament-, mitochondrial- and cytosol-enriched fractions prior to proteolysis and affinity-enrichment of acetylated peptides. LC-MS/MS analysis identified 1075 acetylated peptides, harboring 994 acetylation sites that map to 240 proteins with a global protein false discovery rate <0.8%. Mitochondrial targets account for 59% of identified proteins and 64% of sites. The majority of the acetyl-proteins are enzymes involved in fatty acid metabolism, oxidative phosphorylation or the TCA cycle. Within the cytosolic fraction, the enzymes of glycolysis, fatty acid synthesis and lipid binding are prominent. Nuclear targets included histones and the transcriptional regulators E1A(p300) and CREB binding protein. Comparison of our dataset with three previous global acetylomic studies uniquely revealed 53 lysine-acetylated proteins. Specifically, newly-identified acetyl-proteins include Ca2+-handling proteins, RyR2 and SERCA2, and the myofilament proteins, myosin heavy chain, myosin light chains and subunits of the Troponin complex, among others. These observations were confirmed by anti-acetyl-lysine immunoblotting. In summary, cardiac lysine acetylation may play a role in cardiac substrate selection, bioenergetic performance, and maintenance of redox balance. New sites suggest a host of potential mechanisms by which excitation-contraction coupling may also be modulated.

Quantitative 1H-NMR-Metabolomics Reveals Extensive Metabolic Reprogramming and the Effect of the Aquaglyceroporin FPS1 in Ethanol-Stressed Yeast Cells

A metabolomic analysis using high resolution 1H NMR spectroscopy coupled with multivariate statistical analysis was used to characterize the alterations in the endo- and exo-metabolome of S. cerevisiae BY4741 during the exponential phase of growth in minimal medium supplemented with different ethanol concentrations (0, 2, 4 and 6% v/v). This study provides evidence that supports the notion that ethanol stress induces reductive stress in yeast cells, which, in turn, appears to be counteracted by the increase in the rate of NAD+ regenerating bioreactions. Metabolomics data also shows increased intra- and extra-cellular accumulation of most amino acids and TCA cycle intermediates in yeast cells growing under ethanol stress suggesting a state of overflow metabolism in turn of the pyruvate branch-point. Given its previous implication in ethanol stress resistance in yeast, this study also focused on the effect of the expression of the aquaglyceroporin encoded by FPS1 in the yeast metabolome, in the absence or presence of ethanol stress. The metabolomics data collected herein shows that the deletion of the FPS1 gene in the absence of ethanol stress partially mimics the effect of ethanol stress in the parental strain. Moreover, the results obtained suggest that the reported action of Fps1 in mediating the passive diffusion of glycerol is a key factor in the maintenance of redox balance, an important feature for ethanol stress resistance, and may interfere with the ability of the yeast cell to accumulate trehalose. Overall, the obtained results corroborate the idea that metabolomic approaches may be crucial tools to understand the function and/or the effect of membrane transporters/porins, such as Fps1, and may be an important tool for the clear-cut design of improved process conditions and more robust yeast strains aiming to optimize industrial fermentation performance.