Catalase deficiency induces reactive oxygen species mediated pexophagy and cell death in the liver during prolonged fasting.

Peroxisomes are dynamic organelles that participate in a diverse array of cellular processes, including ß-oxidation, which produces a considerable amount of reactive oxygen species (ROS). Although we showed that catalase depletion induces ROS-mediated pexophagy in cells, the effect of catalase deficiency during conditions that favor ROS generation remains elusive in mice. In this study, we reported that prolonged fasting in catalase-knockout (KO) mice drastically increased ROS production, which induced liver-specific pexophagy, an autophagic degradation of peroxisomes. In addition, increased ROS generation induced the production of pro-inflammatory cytokines in the liver tissues of catalase-KO mice. Furthermore, there was a significant increase in the levels of aspartate transaminase and alanine transaminase as well as apparent cell death in the liver of catalase-KO mice during prolonged fasting. However, an intra-peritoneal injection of the antioxidant N-acetyl-l-cysteine (NAC) and autophagy inhibitor chloroquine inhibited the inflammatory response, liver damage, and pexophagy in the liver of catalase-KO mice during prolonged fasting. Consistently, genetic ablation of autophagy, Atg5 led to suppression of pexophagy during catalase inhibition by 3-aminotriazole (3AT). Moreover, treatment with chloroquine also ameliorated the inflammatory response and cell death in embryonic fibroblast cells from catalase-KO mice. Taken together, our data suggest that ROS-mediated liver-specific pexophagy observed during prolonged fasting in catalase-KO mice may be responsible for the process associated with hepatic cell death.

Sod2 and catalase improve pathological conditions of intervertebral disc degeneration by modifying human adipose-derived mesenchymal stem cells.

OBJECTIVE: To investigate if the modification of human adipose-derived mesenchymal stem cells (hADSCs) by the antioxidants superoxide dismutase 2 (Sod2) and catalase (Cat) can attenuate the pathological conditions of intervertebral disc degeneration (IVD). METHODS: In vitro, MTT assay and qRT-PCR was used to detect cell proliferation and gene expressions in hADSCs transduced with Ad-null (an adenovirus vector containing no transgene expression cassette), Ad-Sod2 (recombinant adenovirus Sod2) and Ad-Cat. IVD mouse models were generated by needle puncture and treated with hADSCs with/without Ad-null/Ad-Sod2/Ad-Cat. X-ray evaluation, magnetic resonance imaging (MRI) analysis, histological analysis, immunohistochemistry, Western blots, ELISAs and qRT-PCR were performed. RESULTS: hADSCs transduced with Ad-Sod2 and Ad-Cat showed enhanced cell proliferation with the upregulation of SOX9, ACAN, and COL2. In vivo, IVD mice injected with hADSCs showed increased disc height index, MRI index and mean T2 intensities, as well as the attenuated histologic grading of the annulus fibrosus (AF) and NP accompanied by the upregulation of GAG and COL2, which were further improved in the Ad-Sod2 hADSC + IVD and Ad-Cat hADSC + IVD groups. Furthermore, the increased expression of IL-1ß, IL-6 and TNF-α was reduced in IVD mice injected with hADSCs. Compared with the hADSC + IVD group, the Ad-Sod2 hADSC/Ad-Cat hADSC + IVD groups had lower expression of pro-inflammatory factors. CONCLUSION: Modification of hADSCs by the antioxidants Sod2 and Cat improved the pathological condition of intervertebral disc tissues with increased GAG and COL2 expression, as well as reduced inflammation, thereby demonstrating a therapeutic effect in IVD.

Physiological and Biochemical Responses of four cassava cultivars to drought stress.

The antioxidant mechanism is crucial for resisting oxidative damage induced by drought stress in plants. Different antioxidant mechanisms may contribute to the tolerance of cassava to drought stress, but for a specific genotype, the response is still unknown. The objective of this study was to investigate antioxidant response and physiological changes of four cassava genotypes under water stress conditions, by keeping the soil moisture content as 80% (control), 50% (medium), 20% (severe) of field capacity for a week. Genotypes RS01 and SC124 were keeping higher relative water content (RWC) and relative chlorophyll content (SPAD value) and less affected by oxidative stress than SC205 and GR4 under drought stress. RS01 just showed slight membrane damage and oxidative stress even under severe drought conditions. A principal component analysis showed that cassava plant water status was closely related to the antioxidant mechanism. Antioxidant response in genotypes RS01 and SC124 under drought stress might attribute to the increased accumulation of ascorbate (AsA) and glutathione (GSH) content and higher superoxide dismutase (SOD) and catalase (CAT) activities, which explained by the up-regulation of Mn-SOD and CAT genes. However, Genotypes SC205 and GR4 mainly depended on the accumulation of total phenolics (TP) and increased glutathione reductase (GR) activity, which attribute to the up-regulation of the GR gene. Our findings could provide vital knowledge for refining the tactics of cultivation and molecular breeding with drought avoidance in cassava.

Neutrophil activation by Escherichia coli isolates from human intestine: effects of bacterial hydroperoxidase activity and surface hydrophobicity.

Successful colonization of the intestine requires that bacteria interact with the innate immune system and, in particular, neutrophils. Progression of inflammatory bowel diseases (IBD) is associated with alterations in gut microbiota, and dysbiosis in Crohn's disease (CD) patients is often associated with an expansion of Escherichia coli. Here, we investigated the ability of such E. coli isolates to avoid neutrophil activation and to utilize reactive oxygen species. Neutrophil activation was detected in vitro in normal human blood via luminol chemiluminescence (CL) induced by reactive oxygen and halogen species generated by neutrophils. No significant difference in neutrophil activation in vitro was detected between isolates from inflamed (23 isolates) vs healthy intestines (5 isolates), with 10-fold variation within both groups (2.9-61.2 mV). CL activity of isolates from the same patient differed by 1.5-5 times. Twenty-four isolates from ileal aspirate, biopsy, and feces of seven patients with CD and one patient with no intestine inflammation were tested for extracellular peroxidase and catalase activity and cell surface hydrophobicity. Average values between patients varied from 26 ± 3 to 73 ± 18 µmol·g-1 of air dry weight for peroxidase activity, from 15 ± 2 to 189 ± 56 mmol·g-1 of air dry weight for catalase activity, and from 5 ± 3 to 105 ± 9 a.u. for the hydrophobic probe fluorescence. Extracellular peroxidase activity and hydrophobicity of bacterial cell surface correlated negatively with stimulated neutrophil CL. The ability of some isolates to avoid neutrophil activation and to utilize reactive oxygen species may provide a strategy to survive assault by the innate immune system.

Oxidative stress and DNA damage in peripheral blood mononuclear cells from normal, obese, prediabetic and diabetic persons exposed to adrenaline in vitro.

Diabetes represents one of the major health concerns, especially in developed countries. Some hormones such as the stress hormone adrenaline can induce reactive oxygen species (ROS) and may worsen the diabetes. Therefore, the main aim of the investigation was to find out whether peripheral blood mononuclear cells (PBMCs) from normal persons have less DNA damage induced by adrenaline (0.1, 1 and 10 µM) in comparison to PBMCs from obese, prediabetic and diabetic patients. Also, the biochemical parameters of oxidative stress (TBARS, catalase) and lactate dehydrogenase were monitored. It was observed that higher concentrations of adrenaline (1 and 10 µM) induced DNA damage in the obese, prediabetic and diabetic groups. In healthy individuals only the highest concentration of adrenaline caused significant increase in the DNA damage. In summary, total comet score (TCS) comparison has shown significant differences between groups, and DNA damaging effects of adrenaline were most evident in diabetic patients. The results of the biochemical analysis also demonstrate that adrenaline exerts most obvious effects in diabetic individuals which is manifested as significant change of parameters of oxidative stress. In summary, the obtained results demonstrated that diabetics are more sensitive to genotoxic effects of adrenaline and this effect probably resulted from decreased antioxidative defence mechanisms in various stages of progression through diabetes. Therefore, these results could contribute to a better understanding of a role of endocrine factors to damage of cellular biomolecules which could be useful in finding novel therapeutic approaches and lifestyle changes with an aim to lower the possibility of diabetes complications.

Novel molecular insights into the anti-oxidative stress response and structure-function of a salt-inducible cyanobacterial Mn-catalase.

KatB, a salt-inducible Mn-catalase, protects the cyanobacterium Anabaena from salinity/oxidative stress. In this report, we provide distinctive insights into the biological-biochemical function of KatB at the molecular level. Anabaena overexpressing the wild-type KatB protein (KatBWT) detoxified H2 O2 efficiently, showing reduced burden of reactive oxygen species compared with the strain overproducing KatBF2V (wherein F-2 is replaced by V). Correspondingly, the KatBWT protein also displayed several folds more activity than KatBF2V. Interestingly, the KatB variants with large hydrophobic amino acids (F/W/Y) were more compact, showed enhanced activity, and were resistant to thermal/chemical denaturation than variants with smaller residues (G/A/V) at the second position. X-ray crystallography-based analysis showed that F-2 was required for appropriate interactions between two subunits. These contacts provided stability to the hexamer, making it more compact. F-2, through its interaction with F-66 and W-43, formed the proper hydrophobic pocket that held the active site together. Consequently, only residues that supported activity (i.e., F/Y/W) were selected at the second position in Mn-catalases during evolution. This study (a) demonstrates that modification of nonactive site residues can alter the response of catalases to environmental stress and (b) has expanded the scope of amino acids that can be targeted for rational protein engineering in plants.

Bioaccumulation of decabromodiphenyl ether affects the antioxidant system in the clam Mactra veneriformis.

Antioxidant enzymes play vital roles against oxidative stress induced by decabromodiphenyl ether (BDE-209), being widespread in marine environment. However, the effect of BDE-209 on antioxidant enzymes remains poorly understood in marine bivalves. In this study, the clams Mactra veneriformis were exposed to 0.1, 1, and 10 µg/L BDE-209 for 7 days and then maintained in clean seawater for 3 days as the depuration. The bioaccumulation of BDE-209 and the effects on superoxide dismutase, catalase, and glutathione peroxidase were investigated. BDE-209 accumulation was concentration-dependent and decreased by 36%-52% after recovery. Malondialdehyde contents increased in a time- and dose-dependent manner. mRNA expression and activity of antioxidant enzymes changed with different patterns and recovered after depuration. These results suggested that antioxidant systems were triggered to protect the clams from oxidative damage caused by BDE-209. Thus, this research is helpful in elucidating the effect of BDE-209 on antioxidant system in marine bivalves.

Analysis of catalase mutants underscores the essential role of CATALASE2 for plant growth and day length-dependent oxidative signalling.

Three genes encode catalase in Arabidopsis. Although the role of CAT2 in photorespiration is well established, the importance of the different catalases in other processes is less clear. Analysis of cat1, cat2, cat3, cat1 cat2, and cat2 cat3 T-DNA mutants revealed that cat2 had the largest effect on activity in both roots and leaves. Root growth was inhibited in all cat2-containing lines, but this inhibition was prevented by growing plants at high CO2 , suggesting that it is mainly an indirect effect of stress at the leaf level. Analysis of double mutants suggested some overlap between CAT2 and CAT3 functions in leaves and CAT1 and CAT2 in seeds. When plants had been grown to a similar developmental stage in short days or long days, equal-time exposure to oxidative stress caused by genetic or pharmacological inhibition of catalase produced a much stronger induction of H2 O2 marker genes in short day plants. Together, our data (a) underline the importance of CAT2 in basal H2 O2 processing in Arabidopsis; (b) suggest that CAT1 and CAT3 are mainly "backup" or stress-specific enzymes; and (c) establish that day length-dependent responses to catalase deficiency are independent of the duration of oxidative stress.

[Effect of catalase on marginal microleakage of resin restoration after external bleaching].

PURPOSE: To investigate the effect of catalase on marginal microleakage of resin restoration after external tooth bleaching with 10% carbamide peroxide. METHODS: Forty extracted human premolars, both intact and health, were randomly divided into 4 groups: group 1, direct composite resin filling without bleaching; group 2, composite resin filling immediately after external bleaching; group 3, immersed in artificial saliva for 3 weeks after external bleaching ,then filling with composite resin; group 4 ,cavity treated with catalase after external bleaching and then filled with composite resin. After 2000 thermal cycles, the teeth were immersed in 2% methylene blue for 24 hours, then the microleakage of interface of resin restorations was observed under stereomicroscope. The data were analysed with SPSS17.0 software package. RESULTS: Group 1 displayed the least amount of microleakage, while group 2 showed the greatest amount of microleakage, group 3 behaved similarly as group 2, having great amount of microleakage, with no significant difference (P>0.05); the microleakage of group 4 decreased significantly (P<0.05) compared to group 2. CONCLUSIONS: The microleakage increases significantly after external bleaching with 10% carbamide peroxide, then decreases if cavities are treated with catalase, but delay filling can not improve microleakage effectively.

Reduction of Aspergillus niger Virulence in Apple Fruits by Deletion of the Catalase Gene cpeB.

Aspergillus niger, a common saprophytic fungus, causes rot in many fruits. We studied the role of a putative catalase-peroxidase-encoding gene, cpeB, in oxidative stress and virulence in fruit. The cpeB gene was deleted in A. niger by homologous recombination, and the Δ cpeB mutant showed decreased CAT activity compared with that of the wild type. The cpeB gene deletion caused increased sensitivity to H2O2 stress, and spore germination was significantly reduced; in addition, the reactive-oxygen-species (ROS) metabolites superoxide anions (·O2-), hydrogen peroxide (H2O2), and malondialdehyde (MDA) accumulated in the Δ cpeB mutant during H2O2 stress. Furthermore, ROS metabolism in A. niger infected apples was determined, and our results showed that the Δ cpeB mutant induced an attenuated response in apple fruit during the fruit-pathogen interaction; the cpeB gene deletion significantly reduced the development of lesions, suggesting that the cpeB gene in A. niger is essential for full virulence in apples.

Roles of the three Mycobacterium smegmatis katG genes for peroxide detoxification and isoniazid susceptibility.

Three different katG sequences (katGI, katGII and katGIII) were identified in the Mycobacterium smegmatis genome. The contributions of the three katG genes to survival of the bacterium were examined by constructing disruptants of these three genes. The katGIII sequence did not produce a functional catalase-peroxidase. Analyses of peroxidase activity and mRNA expression revealed that in wild type M. smegmatis, expression dominance between KatGI and KatGII was switched in the exponential and stationary growth phases. Susceptibility of the M. smegmatis gene disruptants to hydrogen peroxide (H2 O2 ) was tested in two growth phases. In the exponential phase, the katGI-null strain was more susceptible to H2 O2 than the katGII-null strain, indicating that KatGI plays a more important role in survival than KatGII in this growth phase. In contrast, in the stationary phase, growth of the katGII-null strain was inhibited at lower concentrations of H2 O2 . These results suggest that M. smegmatis has two types of catalase-peroxidases, expressions of which are controlled under different gene regulatory systems. Isoniazid (INH) susceptibilities of the katG-null strains were also examined and it was found that katGI is a major determinant of M. smegmatis susceptibility to INH.

Dysregulation of antioxidant responses in patients diagnosed with concomitant Primary Sclerosing Cholangitis/Inflammatory Bowel Disease.

OBJECTIVE: Primary Sclerosing Cholangitis (PSC) is a chronic cholestatic liver disease that is characterized by severe peri-biliary tract inflammation and fibrosis, elevated oxidative stress and hepatocellular injury. A hallmark of PSC patients is the concurrent diagnosis of Inflammatory Bowel Disease occurring in approximately 70%-80% of PSC patients (PSC/IBD). The objective of this study was to determine the impact of end stage PSC/IBD on cellular antioxidant responses and the formation of protein carbonylation. METHODS: Using hepatic tissue and whole cell extracts isolated from age-matched healthy humans and patients diagnosed with end stage PSC/IBD, overall inflammation, oxidative stress, and protein carbonylation were assessed by Western blotting, and immunohistochemistry. RESULTS: Increased immunohistochemical staining for CD3+ (lymphocyte), CD68 (Kupffer cell) and myeloperoxidase (neutrophil) colocalized with the extensive Picrosirius red stained fibrosis confirming the inflammatory aspect of PSC. Importantly, the increased inflammation also colocalized with elevated periportal post-translational modification by the reactive aldehydes 4-HNE, MDA and acrolein. 4-HNE, MDA and acrolein IHC all displayed a significant component in hepatocytes adjacent to fibrotic regions. Furthermore, acrolein was also elevated within the nuclei of periportal inflammatory cells whereas MDA staining was increased in hepatocytes across the lobule. Prussian Blue staining, when compared to the positive controls (ALD, NASH), did not display any evidence of iron accumulation in PSC/IBD livers. Western analysis of PSC/IBD anti-oxidant responses revealed elevated expression of SOD2, GSTπ as well as upregulation of Akt Ser473 phosphorylation. In contrast, expression of GSTµ, GSTA4, catalase, Gpx1 and Hsp70 were suppressed. These data were further supported by a significant decrease in measured GST activity. Dysregulation of anti-oxidant responses in the periportal region of the liver was supported by elevated SOD2 and GSTπ IHC signals in periportal hepatocytes and cholangiocytes. Expression of the Nrf2-regulated proteins HO-1, NAD(P)H quinone reductase (NQO1) and Gpx1 was primarily localized to macrophages. In contrast, catalase staining decreased within periportal hepatocytes and was not evident within cholangiocytes. CONCLUSIONS: Results herein provide additional evidence that cholestasis induces significant increases in periportal oxidative stress and suggest that there are significant differences in the cellular and subcellular generation of reactive aldehydes formed during cholestatic liver injury. Furthermore, these data suggest that anti-oxidant responses are dysregulated during end-stage PSC/IBD supporting pathological data. This work was funded by NIH5R37AA009300-22 D.R.P.

HOCl and the control of oncogenesis.

Sustained generation of extracellular superoxide anions by membrane-associated NADPH oxidase-1 is a hallmark of malignant transformation. The resulting H2O2 drives the proliferation of malignant cells and is converted to HOCl by a Dual oxidase-related peroxidase domain that acts analogously to myeloperoxidase. Whereas H2O2 induces apoptosis nonselectively in nontransformed and transformed cells, HOCl selectively affects malignant cells, as the interaction between HOCl and extracellular superoxide anions allows for site-specific generation of apoptosis-inducing hydroxyl radicals. Transformed cells (early stages of tumor progression) and bona fide tumor cells (representing late stages of tumor progression) respond to exogenous HOCl or HOCl generated by professional phagocytes with induction of apoptosis. In contrast, only transformed cells have the potential to synthesize HOCl through interaction between their superoxide anions/H2O2 and Dual oxidase-related peroxidase released by themselves or neighbouring nontransformed or transformed effector cells. Tumor cells prevent HOCl synthesis through membrane-associated catalase that decomposes H2O2, the substrate for peroxidase, and thus prevents HOCl synthesis. Elimination of malignant cells through HOCl signaling is prevented by Helicobacter pylori-associated catalase and superoxide dismutase, whereas it is enhanced by low dose irradiation and by H2O2-producing lactobacilli in the presence of myeloperoxidase. Peroxidase and catalase that are involved in the control of HOCl signaling are also affecting apoptosis-inducing pathways based on reactive nitrogen species. Modification of tumor cell proteins by HOCl enhances the establishment of a T cell response and thus might be involved in immunogenic modulation. Therefore, targeting the control of HOCl signaling system should allow one to establish novel rational therapeutic approaches.

Examination of the superoxide/hydrogen peroxide forming and quenching potential of mouse liver mitochondria.

Pyruvate dehydrogenase (PDHC) and α-ketoglutarate dehydrogenase complex (KGDHC) are important sources of reactive oxygen species (ROS). In addition, it has been found that mitochondria can also serve as sinks for cellular hydrogen peroxide (H2O2). However, the ROS forming and quenching capacity of liver mitochondria has never been thoroughly examined. Here, we show that mouse liver mitochondria use catalase, glutathione (GSH), and peroxiredoxin (PRX) systems to quench ROS. Incubation of mitochondria with catalase inhibitor 3-amino-1,2,4-triazole (triazole) induced a significant increase in pyruvate or α-ketoglutarate driven O2-/H2O2 formation. 1-Choro-2,4-dinitrobenzene (CDNB), which depletes glutathione (GSH), elicited a similar effect. Auranofin (AF), a thioredoxin reductase-2 (TR2) inhibitor which disables the PRX system, did not significantly change O2-/H2O2 formation. By contrast catalase, GSH, and PRX were all required to scavenging extramitochondrial H2O2. In this study, the ROS forming potential of PDHC, KGDHC, Complex I, and Complex III was also profiled. Titration of mitochondria with 3-methyl-2-oxovaleric acid (KMV), a specific inhibitor for O2-/H2O2 production by KGDHC, induced a ~86% and ~84% decrease in ROS production during α-ketoglutarate and pyruvate oxidation. Titration of myxothiazol, a Complex III inhibitor, decreased O2-/H2O2 formation by ~45%. Rotenone also lowered ROS production in mitochondria metabolizing pyruvate or α-ketoglutarate indicating that Complex I does not contribute to ROS production during forward electron transfer from NADH. Taken together, our results indicate that KGDHC and Complex III are high capacity sites for O2-/H2O2 production in mouse liver mitochondria. We also confirm that catalase plays a role in quenching either exogenous or intramitochondrial H2O2.

Hydrogen Peroxide-Resistant CotA and YjqC of Bacillus altitudinis Spores Are a Promising Biocatalyst for Catalyzing Reduction of Sinapic Acid and Sinapine in Rapeseed Meal.

For the more efficient detoxification of phenolic compounds, a promising avenue would be to develop a multi-enzyme biocatalyst comprising peroxidase, laccase and other oxidases. However, the development of this multi-enzyme biocatalyst is limited by the vulnerability of fungal laccases and peroxidases to hydrogen peroxide (H2O2)-induced inactivation. Therefore, H2O2-resistant peroxidase and laccase should be exploited. In this study, H2O2-stable CotA and YjqC were isolated from the outer coat of Bacillus altitudinis SYBC hb4 spores. In addition to the thermal and alkali stability of catalytic activity, CotA also exhibited a much higher H2O2 tolerance than fungal laccases from Trametes versicolor and Trametes trogii. YjqC is a sporulation-related manganese (Mn) catalase with striking peroxidase activity for sinapic acid (SA) and sinapine (SNP). In contrast to the typical heme-containing peroxidases, the peroxidase activity of YjqC was also highly resistant to inhibition by H2O2 and heat. CotA could also catalyze the oxidation of SA and SNP. CotA had a much higher affinity for SA than B. subtilis CotA. CotA and YjqC rendered from B. altitudinis spores had promising laccase and peroxidase activities for SA and SNP. Specifically, the B. altitudinis spores could be regarded as a multi-enzyme biocatalyst composed of CotA and YjqC. The B. altitudinis spores were efficient for catalyzing the degradation of SA and SNP in rapeseed meal. Moreover, efficiency of the spore-catalyzed degradation of SA and SNP was greatly improved by the presence of 15 mM H2O2. This effect was largely attributed to synergistic biocatalysis of the H2O2-resistant CotA and YjqC toward SA and SNP.

Activity of Redox Enzymes in the Thallus of Anthoceros natalensis.

Anthocerotophyta (hornworts) belong to a group of ancient nonvascular plants and originate from a common ancestor with contemporary vascular plants. Hornworts represent a unique model for investigating mechanisms of formation of stress resistance in higher plants due to their high tolerance to the action of adverse environmental factors. In this work, we demonstrate that the thallus of Anthoceros natalensis exhibits high redox activity changing under stress. Dehydration of the thallus is accompanied by the decrease in activities of intracellular peroxidases, DOPA-peroxidases, and tyrosinases, while catalase activity increases. Subsequent rehydration results in the increase in peroxidase and catalase activities. Kinetic features of peroxidases and tyrosinases were characterized as well as the peroxidase isoenzyme composition of different fractions of the hornwort cell wall proteins. It was shown that the hornwort peroxidases are functionally similar to peroxidases of higher vascular plants including their ability to form superoxide anion-radical. The biochemical mechanism was elucidated, supporting the possible participation of peroxidases in the formation of reactive oxygen species (ROS) via substrate-substrate interactions in the hornwort thallus. It has been suggested that the ROS formation by peroxidases is an evolutionarily ancient process that emerged as a protective mechanism for enhancing adaptive responses of higher land plants and their adaptation to changing environmental conditions and successful colonization of various ecological niches.

Compensatory role of the Nrf2-ARE pathway against paraquat toxicity: Relevance of 26S proteasome activity.

Oxidative stress and the ubiquitin-proteasome system play a key role in the pathogenesis of Parkinson disease. Although the herbicide paraquat is an environmental factor that is involved in the etiology of Parkinson disease, the role of 26S proteasome in paraquat toxicity remains to be determined. Using PC12 cells overexpressing a fluorescent protein fused to the proteasome degradation signal, we report here that paraquat yielded an inhibitory effect on 26S proteasome activity without an obvious decline in 20S proteasome activity. Relative low concentrations of proteasome inhibitors caused the accumulation of nuclear factor erythroid 2-related factor 2 (Nrf2), which is targeted to the ubiquitin-proteasome system, and activated the antioxidant response element (ARE)-dependent transcription. Paraquat also upregulated the protein level of Nrf2 without increased expression of Nrf2 mRNA, and activated the Nrf2-ARE pathway. Consequently, paraquat induced expression of Nrf2-dependent ARE-driven genes, such as γ-glutamylcysteine synthetase, catalase, and hemeoxygenase-1. Knockdown of Nrf2 or inhibition of γ-glutamylcysteine synthetase and catalase exacerbated paraquat-induced toxicity, whereas suppression of hemeoxygenase-1 did not. These data indicate that the compensatory activation of the Nrf2-ARE pathway via inhibition of 26S proteasome serves as part of a cellular defense mechanism to protect against paraquat toxicity.

Reducción de la esteatosis hepática, insulino resistencia y pérdida de la defensa antioxidante en ratones alimentados con dieta alta en grasa suplementados con AGPICL n-3 más aceite de oliva extra virgen / Reduction of hepatic steatosis, insulin resistance and loss of antioxidant defense in high fat diet fed mice suplemented with n-3 LCPUFA more extra virgin olive oil

Non-alcoholic fatty liver disease (NAFLD) is directly associated with insulin resistance and oxidative stress. In NAFLD is established a reduction in n-3 LCPUFA (EPA + DHA) levels and hepatic activity of transcription factor PPAR-alpha. EPA and DHA inhibit lipogenesis and stimulate fatty acid oxidation in the liver. Extra virgin olive oil (EVOO) has important antioxidant properties. This study evaluated the prevention of insulin resistance and prevention of depletion of hepatic antioxidant defense inC57BL/6J mice fed high-fat diet (HFD), supplemented with n-3 LCPUFA plus EVOO. HFD generated insulin resistance and hepatic steatosis, together with significant reduction in i) n-3 LCPUFA hepatic levels, ii) DNA binding activity of PPAR-alpha, iii) activity of antioxidant enzymes (catalase and superoxide dismutase), respect to control group (fed with control diet). Supplementation with n-3 LCPUFA plus EVOO prevent development insulin resistance and attenuate increased of fat in liver (p < 0.05), together with a normalization of i) DNA binding activity of PPAR-á, ii) activity of antioxidant enzymes (catalase and superoxide dismutase) and iii) reducing depletion of n-3 LCPUFA levels in liver tissue, compared to the control group (p < 0.05). Supplementation with n-3 LCPUFA plus EVOO reduced hepatic steatosis and prevent development of insulin resistance, along with preserving the antioxidant defense in liver. Projecting the use of this mixture of AGPICL n-3 plus EVOO as a potential treatment of NAFLD.

Potentiation of Hypoxic Pulmonary Vasoconstriction by Hydrogen Sulfide Precursors 3-Mercaptopyruvate and D-Cysteine Is Blocked by the Cystathionine γ Lyase Inhibitor Propargylglycine.

Although the gasotransmitter hydrogen sulfide (H(2)S) generally dilates systemic arteries in mammals, it causes constriction of pulmonary arteries. In isolated rat pulmonary arteries, we have shown that the H(2)S precursor cysteine enhances both hypoxic pulmonary vasoconstriction and tension development caused by the agonist prostaglandin F(2α) under normoxic conditions. These effects were blocked by propargylglycine (PAG), a blocker of the enzyme cystathionine γ lyase which metabolises cysteine to sulfide. In the present study, we evaluated whether 3-mercaptopyruvate (3-MP), a sulfide precursor which is thought to give rise to sulfide when it is metabolised by the enzyme mercaptopyruvate sulfurtransferase, also enhanced contraction. Application of 3-MP prior to hypoxic challenge caused a marked enhancement of HPV which was completely blocked by both L- and D,L-PAG (both 1 mM). Cumulative application of 3-1,000 µM 3-MP during an ongoing contraction to PGF(2α) under normoxic conditions also caused a marked increase in tension. Application of D-cysteine (1 mM) also enhanced HPV, and this effect was prevented by both the D-amino acid oxidase inhibitor sodium benzoate (500 µM) and 1 mM L-PAG.

A Single Nucleotide Polymorphism in Catalase Is Strongly Associated with Ovarian Cancer Survival.

Ovarian cancer is the deadliest of all gynecologic cancers. Recent evidence demonstrates an association between enzymatic activity altering single nucleotide polymorphisms (SNP) with human cancer susceptibility. We sought to evaluate the association of SNPs in key oxidant and antioxidant enzymes with increased risk and survival in epithelial ovarian cancer. Individuals (n = 143) recruited were divided into controls, (n = 94): healthy volunteers, (n = 18), high-risk BRCA1/2 negative (n = 53), high-risk BRCA1/2 positive (n = 23) and ovarian cancer cases (n = 49). DNA was subjected to TaqMan SNP genotype analysis for selected oxidant and antioxidant enzymes. Of the seven selected SNP studied, no association with ovarian cancer risk (Pearson Chi-square) was found. However, a catalase SNP was identified as a predictor of ovarian cancer survival by the Cox regression model. The presence of this SNP was associated with a higher likelihood of death (hazard ratio (HR) of 3.68 (95% confidence interval (CI): 1.149-11.836)) for ovarian cancer patients. Kaplan-Meier survival analysis demonstrated a significant median overall survival difference (108 versus 60 months, p<0.05) for those without the catalase SNP as compared to those with the SNP. Additionally, age at diagnosis greater than the median was found to be a significant predictor of death (HR of 2.78 (95% CI: 1.022-7.578)). This study indicates a strong association with the catalase SNP and survival of ovarian cancer patients, and thus may serve as a prognosticator.