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Sample records for acid oxidation mitochondrial

  1. Aspirin increases mitochondrial fatty acid oxidation.

    PubMed

    Uppala, Radha; Dudiak, Brianne; Beck, Megan E; Bharathi, Sivakama S; Zhang, Yuxun; Stolz, Donna B; Goetzman, Eric S

    2017-01-08

    The metabolic effects of salicylates are poorly understood. This study investigated the effects of aspirin on fatty acid oxidation. Aspirin increased mitochondrial long-chain fatty acid oxidation, but inhibited peroxisomal fatty acid oxidation, in two different cell lines. Aspirin increased mitochondrial protein acetylation and was found to be a stronger acetylating agent in vitro than acetyl-CoA. However, aspirin-induced acetylation did not alter the activity of fatty acid oxidation proteins, and knocking out the mitochondrial deacetylase SIRT3 did not affect the induction of long-chain fatty acid oxidation by aspirin. Aspirin did not change oxidation of medium-chain fatty acids, which can freely traverse the mitochondrial membrane. Together, these data indicate that aspirin does not directly alter mitochondrial matrix fatty acid oxidation enzymes, but most likely exerts its effects at the level of long-chain fatty acid transport into mitochondria. The drive on mitochondrial fatty acid oxidation may be a compensatory response to altered mitochondrial morphology and inhibited electron transport chain function, both of which were observed after 24 h incubation of cells with aspirin. These studies provide insight into the pathophysiology of Reye Syndrome, which is known to be triggered by aspirin ingestion in patients with fatty acid oxidation disorders.

  2. Nickel inhibits mitochondrial fatty acid oxidation.

    PubMed

    Uppala, Radha; McKinney, Richard W; Brant, Kelly A; Fabisiak, James P; Goetzman, Eric S

    2015-08-07

    Nickel exposure is associated with changes in cellular energy metabolism which may contribute to its carcinogenic properties. Here, we demonstrate that nickel strongly represses mitochondrial fatty acid oxidation-the pathway by which fatty acids are catabolized for energy-in both primary human lung fibroblasts and mouse embryonic fibroblasts. At the concentrations used, nickel suppresses fatty acid oxidation without globally suppressing mitochondrial function as evidenced by increased glucose oxidation to CO2. Pre-treatment with l-carnitine, previously shown to prevent nickel-induced mitochondrial dysfunction in neuroblastoma cells, did not prevent the inhibition of fatty acid oxidation. The effect of nickel on fatty acid oxidation occurred only with prolonged exposure (>5 h), suggesting that direct inhibition of the active sites of metabolic enzymes is not the mechanism of action. Nickel is a known hypoxia-mimetic that activates hypoxia inducible factor-1α (HIF1α). Nickel-induced inhibition of fatty acid oxidation was blunted in HIF1α knockout fibroblasts, implicating HIF1α as one contributor to the mechanism. Additionally, nickel down-regulated the protein levels of the key fatty acid oxidation enzyme very long-chain acyl-CoA dehydrogenase (VLCAD) in a dose-dependent fashion. In conclusion, inhibition of fatty acid oxidation by nickel, concurrent with increased glucose metabolism, represents a form of metabolic reprogramming that may contribute to nickel-induced carcinogenesis.

  3. Nickel Inhibits Mitochondrial Fatty Acid Oxidation

    PubMed Central

    Uppala, Radha; McKinney, Richard W.; Brant, Kelly A.; Fabisiak, James P.; Goetzman, Eric S.

    2015-01-01

    Nickel exposure is associated with changes in cellular energy metabolism which may contribute to its carcinogenic properties. Here, we demonstrate that nickel strongly represses mitochondrial fatty acid oxidation—the pathway by which fatty acids are catabolized for energy—in both primary human lung fibroblasts and mouse embryonic fibroblasts. At the concentrations used, nickel suppresses fatty acid oxidation without globally suppressing mitochondrial function as evidenced by increased glucose oxidation to CO2. Pre-treatment with L-carnitine, previously shown to prevent nickel-induced mitochondrial dysfunction in neuroblastoma cells, did not prevent the inhibition of fatty acid oxidation. The effect of nickel on fatty acid oxidation occurred only with prolonged exposure (>5 hr), suggesting that direct inhibition of the active sites of metabolic enzymes is not the mechanism of action. Nickel is a known hypoxia-mimetic that activates hypoxia inducible factor-1α (HIF1α). Nickel-induced inhibition of fatty acid oxidation was blunted in HIF1α knockout fibroblasts, implicating HIF1α as one contributor to the mechanism. Additionally, nickel down-regulated the protein levels of the key fatty acid oxidation enzyme very long-chain acyl-CoA dehydrogenase (VLCAD) in a dose-dependent fashion. In conclusion, inhibition of fatty acid oxidation by nickel, concurrent with increased glucose metabolism, represents a form of metabolic reprogramming that may contribute to nickel-induced carcinogenesis. PMID:26051273

  4. Ceramides and mitochondrial fatty acid oxidation in obesity.

    PubMed

    Fucho, Raquel; Casals, Núria; Serra, Dolors; Herrero, Laura

    2017-04-01

    Obesity is an epidemic, complex disease that is characterized by increased glucose, lipids, and low-grade inflammation in the circulation, among other factors. It creates the perfect scenario for the production of ceramide, the building block of the sphingolipid family of lipids, which is involved in metabolic disorders such as obesity, diabetes, and cardiovascular disease. In addition, obesity causes a decrease in fatty acid oxidation (FAO), which contributes to lipid accumulation within the cells, conferring more susceptibility to cell dysfunction. C16:0 ceramide, a specific ceramide species, has been identified recently as the principal mediator of obesity-derived insulin resistance, impaired fatty acid oxidation, and hepatic steatosis. In this review, we have sought to cover the importance of the ceramide species and their metabolism, the main ceramide signaling pathways in obesity, and the link between C16:0 ceramide, FAO, and obesity.-Fucho, R., Casals, N., Serra, D., Herrero, L. Ceramides and mitochondrial fatty acid oxidation in obesity.

  5. The inborn errors of mitochondrial fatty acid oxidation.

    PubMed

    Vianey-Liaud, C; Divry, P; Gregersen, N; Mathieu, M

    1987-01-01

    To date, seven inborn errors of mitochondrial fatty acid oxidation have been identified. A total of about 100 patients in the world have been reported. Clinically the beta-oxidation defects are more often characterized by episodic hypoglycaemia leading to a coma mimicking Reye's syndrome. The hypoglycaemia is non-ketotic since the synthesis of ketone bodies is deficient. Periods of decompensation occur when carbohydrate supply is poor, e.g. prolonged fasting, vomiting, or increased caloric requirements, as and when lipid stores are used. Defects in beta-oxidation have also been reported to be one cause of sudden infant death syndrome. The diagnosis of these inborn errors is by biochemical investigation since where symptoms suggest such a defect, the precise aetiology cannot be assessed. The biochemical diagnosis is based firstly on identification of abnormal plasma and of urinary metabolites during acute attacks. Derivatives of the omega-oxidation and omega-1-oxidation of medium chain fatty acids have been identified, as well as acylglycine and acylcarnitine conjugates. These metabolites are nearly always absent when patients are in good clinical condition. Secondly, the diagnosis must be based on the identification of the enzymatic defects: this involves global assays which allow a localization of the 'level' of the defect (i.e. the oxidation of long, medium or short chain fatty acids) and specific measurement of enzyme activities (acyl-CoA dehydrogenases and electron carriers: ETF and ETF-DH). The diagnosis of these disorders is of prime importance because of the severity of the clinical symptoms. These can be prevented, in some cases, by an appropriate diet (a high carbohydrate, low fat diet, sometimes supplemented with L-carnitine). In other cases, genetic counselling can be offered.

  6. Targeting oxidative stress attenuates malonic acid induced Huntington like behavioral and mitochondrial alterations in rats.

    PubMed

    Kalonia, Harikesh; Kumar, Puneet; Kumar, Anil

    2010-05-25

    Objective of the present study was to explore the possible role of oxidative stress in the malonic acid induced behavioral, biochemical and mitochondrial alterations in rats. In the present study, unilateral single injections of malonic acid at different doses (1.5, 3 and 6 micromol) were made into the ipsilateral striatum in rats. Behavioral parameters were accessed on 1st, 7th and 14th day post malonic acid administration. Oxidative stress parameters and mitochondrial enzyme functions were assessed on day 14 after behavioral observations. Ipsilateral striatal malonic acid (3 and 6 micromol) administration significantly reduced body weight, locomotor activity, motor coordination and caused oxidative damage (lipid peroxidation, nitrite, superoxide dismutase, catalase and glutathione) in the striatum as compared to sham treated animal. Mitochondrial enzyme complexes and MTT (3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolinium bromide) activity were significantly inhibited by malonic acid. Vitamin E treatment (50 and 100 mg/kg, p.o.) significantly reversed the various behavioral, biochemical and mitochondrial alterations in malonic acid treated animals. Our findings show that targeting oxidative stress by vitamin E in malonic acid model, results in amelioration of behavioral and mitochondrial alterations are linked to inhibition of oxidative damage. Based upon these finding present study hypothesize that protection exerted by vitamin E on behavioral, mitochondrial markers indicates the possible preservation of the functional status of the striatal neurons by targeting the deleterious actions of oxidative stress.

  7. Combined defects in oxidative phosphorylation and fatty acid β-oxidation in mitochondrial disease

    PubMed Central

    Nsiah-Sefaa, Abena; McKenzie, Matthew

    2016-01-01

    Mitochondria provide the main source of energy to eukaryotic cells, oxidizing fats and sugars to generate ATP. Mitochondrial fatty acid β-oxidation (FAO) and oxidative phosphorylation (OXPHOS) are two metabolic pathways which are central to this process. Defects in these pathways can result in diseases of the brain, skeletal muscle, heart and liver, affecting approximately 1 in 5000 live births. There are no effective therapies for these disorders, with quality of life severely reduced for most patients. The pathology underlying many aspects of these diseases is not well understood; for example, it is not clear why some patients with primary FAO deficiencies exhibit secondary OXPHOS defects. However, recent findings suggest that physical interactions exist between FAO and OXPHOS proteins, and that these interactions are critical for both FAO and OXPHOS function. Here, we review our current understanding of the interactions between FAO and OXPHOS proteins and how defects in these two metabolic pathways contribute to mitochondrial disease pathogenesis. PMID:26839416

  8. Mitochondrial free fatty acid β-oxidation supports oxidative phosphorylation and proliferation in cancer cells.

    PubMed

    Rodríguez-Enríquez, Sara; Hernández-Esquivel, Luz; Marín-Hernández, Alvaro; El Hafidi, Mohammed; Gallardo-Pérez, Juan Carlos; Hernández-Reséndiz, Ileana; Rodríguez-Zavala, José S; Pacheco-Velázquez, Silvia C; Moreno-Sánchez, Rafael

    2015-08-01

    Oxidative phosphorylation (OxPhos) is functional and sustains tumor proliferation in several cancer cell types. To establish whether mitochondrial β-oxidation of free fatty acids (FFAs) contributes to cancer OxPhos functioning, its protein contents and enzyme activities, as well as respiratory rates and electrical membrane potential (ΔΨm) driven by FFA oxidation were assessed in rat AS-30D hepatoma and liver (RLM) mitochondria. Higher protein contents (1.4-3 times) of β-oxidation (CPT1, SCAD) as well as proteins and enzyme activities (1.7-13-times) of Krebs cycle (KC: ICD, 2OGDH, PDH, ME, GA), and respiratory chain (RC: COX) were determined in hepatoma mitochondria vs. RLM. Although increased cholesterol content (9-times vs. RLM) was determined in the hepatoma mitochondrial membranes, FFAs and other NAD-linked substrates were oxidized faster (1.6-6.6 times) by hepatoma mitochondria than RLM, maintaining similar ΔΨm values. The contents of β-oxidation, KC and RC enzymes were also assessed in cells. The mitochondrial enzyme levels in human cervix cancer HeLa and AS-30D cells were higher than those observed in rat hepatocytes whereas in human breast cancer biopsies, CPT1 and SCAD contents were lower than in human breast normal tissue. The presence of CPT1 and SCAD in AS-30D mitochondria and HeLa cells correlated with an active FFA utilization in HeLa cells. Furthermore, the β-oxidation inhibitor perhexiline blocked FFA utilization, OxPhos and proliferation in HeLa and other cancer cells. In conclusion, functional mitochondria supported by FFA β-oxidation are essential for the accelerated cancer cell proliferation and hence anti-β-oxidation therapeutics appears as an alternative promising approach to deter malignant tumor growth.

  9. FABP4 reversed the regulation of leptin on mitochondrial fatty acid oxidation in mice adipocytes

    PubMed Central

    Gan, Lu; Liu, Zhenjiang; Cao, Weina; Zhang, Zhenzhen; Sun, Chao

    2015-01-01

    Fatty acid binding protein 4 (FABP4), plays key role in fatty acid transportation and oxidation, and increases with leptin synergistically during adipose inflammation process. However, the regulation mechanism between FABP4 and leptin on mitochondrial fatty acid oxidation remains unclear. In this study, we found that FABP4 reduced the expression of leptin, CPT-1 and AOX1 in mice adipocytes. Conversely, FABP4 was down-regulated in a time-dependent manner by leptin treatment. Additionally, forced expression of FABP4 attenuated the expression of PGC1-α, UCP2, CPT-1, AOX1 and COX2 compared with leptin incubation. Moreover, mitochondrial membrane potential, fatty acid oxidation enzyme medium-chain acyl-CoA dehydrogenase (MCAD), long-chain acyl-CoA dehydrogenase (LCAD) and Cyt C levels were reduced in response to the overexpression of FABP4. These reductions correspond well with the reduced release of free fatty acid and the inactivation of mitochondrial complexes I and III by FABP4 overexpression. Furthermore, addition of the Akt/mTOR pathway-specific inhibitor (MK2206) blocked the mitochondrial fatty acid oxidation and respiration factors, whereas interference of FABP4 overcame these effects. Taken together, FABP4 could reverse the activation of the leptin-induced mitochondrial fatty acid oxidation, and the inhibition of Akt/mTOR signal pathway played a key role in this process. PMID:26310911

  10. Overexpression of PGC-1α Increases Peroxisomal and Mitochondrial Fatty Acid Oxidation in Human Primary Myotubes.

    PubMed

    Huang, Tai-Yu; Zheng, Donghai; Houmard, Joseph A; Brault, Jeffrey J; Hickner, Robert C; Cortright, Ronald N

    2017-01-10

    Peroxisomes are indispensable organelles for lipid metabolism in humans and their biogenesis has been assumed to be under regulation by peroxisome proliferator-activated receptors (PPARs). However, recent studies in hepatocytes suggest that the mitochondrial proliferator PGC-1α (peroxisome proliferator-activated receptor gamma coactivator-1 alpha) also acts as an upstream transcriptional regulator for enhancing peroxisomal abundance and associated activity. It is unknown whether the regulatory mechanism(s) for enhancing peroxisomal function is through the same node as mitochondrial biogenesis in human skeletal muscle (HSkM) and whether fatty acid oxidation (FAO) is affected. Primary myotubes from vastus lateralis biopsies from lean donors (BMI =24.0 ± 0.6 kg/m(2), N = 6) were exposed to adenovirus encoding human PGC-1α or GFP control. Peroxisomal biogenesis proteins (Peroxins) and genes (PEXs) responsible for proliferation and functions were assessed by western blotting and real-time qRT-PCR respectively. 1-(14)C palmitic acid and 1-(14)C lignoceric acid (exclusive peroxisomal specific substrate) were used to assess mitochondrial oxidation of peroxisomal derived metabolites. Following overexpression of PGC-1α, 1) Peroxisomal membrane protein 70kD (PMP70), PEX19, and mitochondrial citrate synthetase protein content were significantly elevated (P<0.05) 2) PGC-1α, PMP70, key PEXs, and peroxisomal β-oxidation mRNA expression levels were significantly upregulated (P<0.05) and 3) A concomitant increase in lignoceric acid oxidation by both peroxisomal and mitochondrial activity was observed (P<0.05). These novel findings demonstrate that, in addition to the proliferative effect on mitochondria, PGC-1α can induce peroxisomes and accompanying elevations in long-chain and very-long-chain fatty acid oxidation by a peroxisomal-mitochondrial functional cooperation as observed in HSkM cells.

  11. Increasing mitochondrial muscle fatty acid oxidation induces skeletal muscle remodeling toward an oxidative phenotype.

    PubMed

    Hénique, Carole; Mansouri, Abdelhak; Vavrova, Eliska; Lenoir, Véronique; Ferry, Arnaud; Esnous, Catherine; Ramond, Elodie; Girard, Jean; Bouillaud, Frédéric; Prip-Buus, Carina; Cohen, Isabelle

    2015-06-01

    Adult skeletal muscle is a dynamic, remarkably plastic tissue, which allows myofibers to switch from fast/glycolytic to slow/oxidative types and to increase mitochondrial fatty acid oxidation (mFAO) capacity and vascularization in response to exercise training. mFAO is the main muscle energy source during endurance exercise, with carnitine palmitoyltransferase 1 (CPT1) being the key regulatory enzyme. Whether increasing muscle mFAO affects skeletal muscle physiology in adulthood actually remains unknown. To investigate this, we used in vivo electrotransfer technology to express in mouse tibialis anterior (TA), a fast/glycolytic muscle, a mutated CPT1 form (CPT1mt) that is active but insensitive to malonyl-CoA, its physiologic inhibitor. In young (2-mo-old) adult mice, muscle CPT1mt expression enhanced mFAO (+40%), but also increased the percentage of oxidative fibers (+28%), glycogen content, and capillary-to-fiber density (+45%). This CPT1mt-induced muscle remodeling, which mimicked exercise-induced oxidative phenotype, led to a greater resistance to muscle fatigue. In the context of aging, characterized by sarcopenia and reduced oxidative capacity, CPT1mt expression in TAs from aged (20-mo-old) mice partially reversed aging-associated sarcopenia and fiber-type transition, and increased muscle capillarity. These findings provide evidence that mFAO regulates muscle phenotype and may be a potential target to combat age-related decline in muscle function.

  12. Mitochondrial Dysfunction in Cancer and Neurodegenerative Diseases: Spotlight on Fatty Acid Oxidation and Lipoperoxidation Products

    PubMed Central

    Barrera, Giuseppina; Gentile, Fabrizio; Pizzimenti, Stefania; Canuto, Rosa Angela; Daga, Martina; Arcaro, Alessia; Cetrangolo, Giovanni Paolo; Lepore, Alessio; Ferretti, Carlo; Dianzani, Chiara; Muzio, Giuliana

    2016-01-01

    In several human diseases, such as cancer and neurodegenerative diseases, the levels of reactive oxygen species (ROS), produced mainly by mitochondrial oxidative phosphorylation, is increased. In cancer cells, the increase of ROS production has been associated with mtDNA mutations that, in turn, seem to be functional in the alterations of the bioenergetics and the biosynthetic state of cancer cells. Moreover, ROS overproduction can enhance the peroxidation of fatty acids in mitochondrial membranes. In particular, the peroxidation of mitochondrial phospholipid cardiolipin leads to the formation of reactive aldehydes, such as 4-hydroxynonenal (HNE) and malondialdehyde (MDA), which are able to react with proteins and DNA. Covalent modifications of mitochondrial proteins by the products of lipid peroxidation (LPO) in the course of oxidative cell stress are involved in the mitochondrial dysfunctions observed in cancer and neurodegenerative diseases. Such modifications appear to affect negatively mitochondrial integrity and function, in particular energy metabolism, adenosine triphosphate (ATP) production, antioxidant defenses and stress responses. In neurodegenerative diseases, indirect confirmation for the pathogenetic relevance of LPO-dependent modifications of mitochondrial proteins comes from the disease phenotypes associated with their genetic alterations. PMID:26907355

  13. Novel role of FATP1 in mitochondrial fatty acid oxidation in skeletal muscle cells

    PubMed Central

    Sebastián, David; Guitart, Maria; García-Martínez, Celia; Mauvezin, Caroline; Orellana-Gavaldà, Josep M.; Serra, Dolors; Gómez-Foix, Anna M.; Hegardt, Fausto G.; Asins, Guillermina

    2009-01-01

    Carnitine palmitoyltransferase 1 (CPT1) catalyzes the first step in long-chain fatty acid import into mitochondria, and it is believed to be rate limiting for β-oxidation of fatty acids. However, in muscle, other proteins may collaborate with CPT1. Fatty acid translocase/CD36 (FAT/CD36) may interact with CPT1 and contribute to fatty acid import into mitochondria in muscle. Here, we demonstrate that another membrane-bound fatty acid binding protein, fatty acid transport protein 1 (FATP1), collaborates with CPT1 for fatty acid import into mitochondria. Overexpression of FATP1 using adenovirus in L6E9 myotubes increased both fatty acid oxidation and palmitate esterification into triacylglycerides. Moreover, immunocytochemistry assays in transfected L6E9 myotubes showed that FATP1 was present in mitochondria and coimmunoprecipitated with CPT1 in L6E9 myotubes and rat skeletal muscle in vivo. The cooverexpression of FATP1 and CPT1 also enhanced mitochondrial fatty acid oxidation, similar to the cooverexpression of FAT/CD36 and CPT1. However, etomoxir, an irreversible inhibitor of CPT1, blocked all these effects. These data reveal that FATP1, like FAT/CD36, is associated with mitochondria and has a role in mitochondrial oxidation of fatty acids. PMID:19429947

  14. Mitochondrial and peroxisomal fatty acid oxidation in liver homogenates and isolated hepatocytes from control and clofibrate-treated rats.

    PubMed

    Mannaerts, G P; Debeer, L J; Thomas, J; De Schepper, P J

    1979-06-10

    Mitochondrial and peroxisomal fatty acid oxidation were compared in whole liver homogenates. Oxidation of 0.2 mM palmitoyl-CoA or oleate by mitochondria increased rapidly with increasing molar substrate:albumin ratios and became saturated at ratios below 3, while peroxisomal oxidation increased more slowly and continued to rise to reach maximal activity in the absence of albumin. Under the latter condition mitochondrial oxidation was severely depressed. In homogenates from normal liver peroxisomal oxidation was lower than mitochondrial oxidation at all ratios tested except when albumin was absent. In contrast with mitochondrial oxidation, peroxisomal oxidation did not produce ketones, was cyanide-insensitive, was not dependent on carnitine, and was not inhibited by (+)-octanoylcarnitine, malonyl-CoA and 4-pentenoate. Mitochondrial oxidation was inhibited by CoASH concentrations that were optimal for peroxisomal oxidation. In the presence of albumin, peroxisomal oxidation was stimulated by Triton X-100 but unaffected by freeze-thawing; both treatments suppressed mitochondrial oxidation. Clofibrate treatment increased mitochondrial and peroxisomal oxidation 2- and 6- to 8-fold, respectively. Peroxisomal oxidation remained unchanged in starvation and diabetes. Fatty acid oxidation was severely depressed by cyanide and (+)-octanoylcarnitine in hepatocytes from normal rats. Hepatocytes from clofibrate-treated rats, which displayed a 3- to 4-fold increase in fatty acid oxidation, were less inhibited by (+)-octanoylcarnitine. Hydrogen peroxide production was severalfold higher in hepatocytes from treated animals oxidizing fatty acids than in control hepatocytes. Assuming that all H2O2 produced during fatty acid oxidation was due to peroxisomal oxidation, it was calculated that the contribution of the peroxisomes to fatty acid oxidation was less than 10% both in cells from control and clofibrate-treated animals.

  15. Fatty acid binding protein facilitates sarcolemmal fatty acid transport but not mitochondrial oxidation in rat and human skeletal muscle

    PubMed Central

    Holloway, Graham P; Lally, Jamie; Nickerson, James G; Alkhateeb, Hakam; Snook, Laelie A; Heigenhauser, George J F; Calles-Escandon, Jorge; Glatz, Jan F C; Luiken, Joost J F P; Spriet, Lawrence L; Bonen, Arend

    2007-01-01

    The transport of long-chain fatty acids (LCFAs) across mitochondrial membranes is regulated by carnitine palmitoyltransferase I (CPTI) activity. However, it appears that additional fatty acid transport proteins, such as fatty acid translocase (FAT)/CD36, influence not only LCFA transport across the plasma membrane, but also LCFA transport into mitochondria. Plasma membrane-associated fatty acid binding protein (FABPpm) is also known to be involved in sacrolemmal LCFA transport, and it is also present on the mitochondria. At this location, it has been identified as mitochondrial aspartate amino transferase (mAspAT), despite being structurally identical to FABPpm. Whether this protein is also involved in mitochondrial LCFA transport and oxidation remains unknown. Therefore, we have examined the ability of FABPpm/mAspAT to alter mitochondrial fatty acid oxidation. Muscle contraction increased (P < 0.05) the mitochondrial FAT/CD36 content in rat (+22%) and human skeletal muscle (+33%). By contrast, muscle contraction did not alter the content of mitochondrial FABPpm/mAspAT protein in either rat or human muscles. Electrotransfecting rat soleus muscles, in vivo, with FABPpm cDNA increased FABPpm protein in whole muscle (+150%; P < 0.05), at the plasma membrane (+117%; P < 0.05) and in mitochondria (+80%; P < 0.05). In these FABPpm-transfected muscles, palmitate transport into giant vesicles was increased by +73% (P < 0.05), and fatty acid oxidation in intact muscle was increased by +18% (P < 0.05). By contrast, despite the marked increase in mitochondrial FABPpm/mAspAT protein content (+80%), the rate of mitochondrial palmitate oxidation was not altered (P > 0.05). However, electrotransfection increased mAspAT activity by +70% (P < 0.05), and the mitochondrial FABPpm/mAspAT protein content was significantly correlated with mAspAT activity (r= 0.75). It is concluded that FABPpm has two distinct functions depending on its subcellular location: (a) it contributes to

  16. Stereoselective and nonstereoselective effects of ibuprofen enantiomers on mitochondrial beta-oxidation of fatty acids

    SciTech Connect

    Freneaux, E.; Fromenty, B.; Berson, A.; Labbe, G.; Degott, C.; Letteron, P.; Larrey, D.; Pessayre, D. , Hopital Beaujon, Clichy )

    1990-11-01

    The effects of the R-(-) and S-(+)ibuprofen enantiomers were first studied in vitro with mouse liver mitochondria incubated in the presence of various concentrations of exogenous coenzyme A. In the presence of a low concentration of coenzyme A (2.5 microM), the R-(-)enantiomer (which forms an acylcoenzyme A) inhibited stereoselectively the beta oxidation of (1-{sup 14}C)palmitic acid but not that of (1-{sup 14}C)palmitoyl-L-carnitine (which can directly enter the mitochondria). In the presence, however, of a concentration of coenzyme A (50 microM) reproducing that present in liver cell cytosol, both enantiomers (2 mM) slightly inhibited the beta oxidation of (1-{sup 14}C)palmitic acid and markedly inhibited the beta oxidation of (1-{sup 14}C)octanoic acid and (1-{sup 14}C)butyric acid. In vivo, both enantiomers (1 mmol.kg-1) similarly inhibited the formation of ({sup 14}C)CO{sub 2} from (1-{sup 14}C)fatty acids. Both enantiomers similarly decreased plasma ketone bodies. Both similarly increased hepatic triglycerides, and both produced mild microvesicular steatosis of the liver. We conclude that both ibuprofen enantiomers inhibit beta oxidation of fatty acids in vitro and in vivo. In addition, the R-(-)enantiomer may stereoselectively sequester coenzyme A; at low concentrations of coenzyme A in vitro, this may stereoselectively inhibit the mitochondrial uptake and beta oxidation of long chain fatty acids.

  17. High concentrations of stavudine impair fatty acid oxidation without depleting mitochondrial DNA in cultured rat hepatocytes.

    PubMed

    Igoudjil, Anissa; Massart, Julie; Begriche, Karima; Descatoire, Véronique; Robin, Marie-Anne; Fromenty, Bernard

    2008-06-01

    The antiretroviral nucleoside reverse-transcriptase inhibitor (NRTI) stavudine (d4T) can induce mild to severe liver injuries such as steatosis (i.e. triglyceride accumulation), steatohepatitis and liver failure. NRTI-induced toxicity has been ascribed to the inhibition of mitochondrial DNA (mtDNA) replication causing mtDNA depletion and respiratory chain dysfunction. This can secondarily impair the tricarboxylic acid cycle and fatty acid oxidation (FAO), thus leading to lactic acidosis and hepatic steatosis. However, NRTIs could also impair mitochondrial function and induce hepatic steatosis through other mechanisms. In this study, we sought to determine whether d4T could inhibit mitochondrial FAO and induce triglyceride accumulation through a mtDNA-independent mechanism. Since human tumoral and non-tumoral hepatic cell lines were unable to efficiently oxidize palmitic acid, the effects of d4T on mitochondrial FAO were assessed on cultured rat hepatocytes. Our results showed that 750 microM of d4T significantly inhibited palmitic acid oxidation after 48 or 72 h of culture, without inducing cell death. Importantly, high concentrations of zidovudine and zalcitabine (two other NRTIs that can induce hepatic steatosis), or beta-aminoisobutyric acid (a d4T metabolite), did not impair FAO in rat hepatocytes. D4T-induced FAO inhibition was observed without mtDNA depletion and lactate production, and was fully prevented with l-carnitine or clofibrate coincubation. l-carnitine also prevented the accretion of neutral lipids within rat hepatocytes. High concentrations of d4T were unable to inhibit FAO on freshly isolated liver mitochondria. Moreover, a microarray analysis was performed to clarify the mechanism whereby d4T can inhibit mitochondrial FAO and induce triglyceride accumulation in rat hepatocytes. The microarray data, confirmed by quantitative real-time PCR analysis, showed that d4T increased the expression of sterol regulatory element-binding protein-1c (SREBP1c

  18. Phylogenomic Evidence for a Myxococcal Contribution to the Mitochondrial Fatty Acid Beta-Oxidation

    PubMed Central

    Schlüter, Agatha; Ruiz-Trillo, Iñaki; Pujol, Aurora

    2011-01-01

    Background The origin of eukaryotes remains a fundamental question in evolutionary biology. Although it is clear that eukaryotic genomes are a chimeric combination of genes of eubacterial and archaebacterial ancestry, the specific ancestry of most eubacterial genes is still unknown. The growing availability of microbial genomes offers the possibility of analyzing the ancestry of eukaryotic genomes and testing previous hypotheses on their origins. Methodology/Principal Findings Here, we have applied a phylogenomic analysis to investigate a possible contribution of the Myxococcales to the first eukaryotes. We conducted a conservative pipeline with homologous sequence searches against a genomic sampling of 40 eukaryotic and 357 prokaryotic genomes. The phylogenetic reconstruction showed that several eukaryotic proteins traced to Myxococcales. Most of these proteins were associated with mitochondrial lipid intermediate pathways, particularly enzymes generating reducing equivalents with pivotal roles in fatty acid β-oxidation metabolism. Our data suggest that myxococcal species with the ability to oxidize fatty acids transferred several genes to eubacteria that eventually gave rise to the mitochondrial ancestor. Later, the eukaryotic nucleocytoplasmic lineage acquired those metabolic genes through endosymbiotic gene transfer. Conclusions/Significance Our results support a prokaryotic origin, different from α-proteobacteria, for several mitochondrial genes. Our data reinforce a fluid prokaryotic chromosome model in which the mitochondrion appears to be an important entry point for myxococcal genes to enter eukaryotes. PMID:21760940

  19. A general introduction to the biochemistry of mitochondrial fatty acid β-oxidation

    PubMed Central

    Wanders, Ronald J. A.

    2010-01-01

    Over the years, the mitochondrial fatty acid β-oxidation (FAO) pathway has been characterised at the biochemical level as well as the molecular biological level. FAO plays a pivotal role in energy homoeostasis, but it competes with glucose as the primary oxidative substrate. The mechanisms behind this so-called glucose–fatty acid cycle operate at the hormonal, transcriptional and biochemical levels. Inherited defects for most of the FAO enzymes have been identified and characterised and are currently included in neonatal screening programmes. Symptoms range from hypoketotic hypoglycaemia to skeletal and cardiac myopathies. The pathophysiology of these diseases is still not completely understood, hampering optimal treatment. Studies of patients and mouse models will contribute to our understanding of the pathogenesis and will ultimately lead to better treatment. PMID:20195903

  20. High-intensity interval training increases intrinsic rates of mitochondrial fatty acid oxidation in rat red and white skeletal muscle.

    PubMed

    Hoshino, Daisuke; Yoshida, Yuko; Kitaoka, Yu; Hatta, Hideo; Bonen, Arend

    2013-03-01

    High-intensity interval training (HIIT) can increase mitochondrial volume in skeletal muscle. However, it is unclear whether HIIT alters the intrinsic capacity of mitochondrial fatty acid oxidation, or whether such changes are associated with changes in mitochondrial FAT/CD36, a regulator of fatty acid oxidation, or with reciprocal changes in the nuclear receptor coactivator (peroxisome proliferator-activated receptor gamma coactivator-1alpha (PGC-1α)) and the corepressor (receptor-interacting protein 140 (RIP140)). We examined whether HIIT alters fatty acid oxidation rates in the isolated subsarcolemmal (SS) and intermyofibrillar (IMF) mitochondria of red and white skeletal muscle and (or) induces changes in muscle PGC-1α and RIP140 proteins and mitochondrial FAT/CD36 protein content. Rats were divided into untrained or HIIT-trained groups. HIIT animals performed 10 bouts of 1-min high-intensity treadmill running (30-55 m·min(-1)), separated by 2 min of rest, for 5 days a week for 4 weeks. As expected, after the training period, HIIT increased mitochondrial enzymes (citrate synthase, COXIV, and β-hydroxyacyl CoA dehydrogenase) in red and white muscle, indicating that muscle mitochondrial volume had increased. HIIT also increased the rates of palmitate oxidation in mitochondria of red (37% for SS and 19% for IMF) and white (36% for SS and 12% for IMF) muscle. No changes occurred in SS and IMF mitochondrial FAT/CD36 proteins, despite increasing FAT/CD36 at the whole-muscle level (27% for red and 22% for white). Concurrently, muscle PGC-1α protein was increased in red (22%) and white (16%) muscle, but RIP140 was not altered. These results indicate that increases in SS and IMF mitochondrial fatty acid oxidation induced by HIIT are accompanied by an increase in PGC-1α, but not RIP140 or FAT/CD36.

  1. AMPK activation promotes lipid droplet dispersion on detyrosinated microtubules to increase mitochondrial fatty acid oxidation

    PubMed Central

    Herms, Albert; Bosch, Marta; Reddy, Babu J.N.; Schieber, Nicole L.; Fajardo, Alba; Rupérez, Celia; Fernández-Vidal, Andrea; Ferguson, Charles; Rentero, Carles; Tebar, Francesc; Enrich, Carlos; Parton, Robert G.; Gross, Steven P.; Pol, Albert

    2015-01-01

    Lipid droplets (LDs) are intracellular organelles that provide fatty acids (FAs) to cellular processes including synthesis of membranes and production of metabolic energy. While known to move bidirectionally along microtubules (MTs), the role of LD motion and whether it facilitates interaction with other organelles are unclear. Here we show that during nutrient starvation, LDs and mitochondria relocate on detyrosinated MT from the cell centre to adopt a dispersed distribution. In the cell periphery, LD–mitochondria interactions increase and LDs efficiently supply FAs for mitochondrial beta-oxidation. This cellular adaptation requires the activation of the energy sensor AMPK, which in response to starvation simultaneously increases LD motion, reorganizes the network of detyrosinated MTs and activates mitochondria. In conclusion, we describe the existence of a specialized cellular network connecting the cellular energetic status and MT dynamics to coordinate the functioning of LDs and mitochondria during nutrient scarcity. PMID:26013497

  2. Stilbenes and resveratrol metabolites improve mitochondrial fatty acid oxidation defects in human fibroblasts

    PubMed Central

    2014-01-01

    Background Inborn enzyme defects of mitochondrial fatty acid beta-oxidation (FAO) form a large group of genetic disorders associated to variable clinical presentations ranging from life-threatening pediatric manifestations up to milder late onset phenotypes, including myopathy. Very few candidate drugs have been identified in this group of disorders. Resveratrol (RSV) is a natural polyphenol with anti-oxidant and anti-inflammatory effects, recently shown to have beneficial metabolic properties in mice models. Our study explores its possible effects on FAO and mitochondrial energy metabolism in human cells, which are still very little documented. Methods Using cells from controls and from patients with Carnitine Palmitoyl Transferase 2 (CPT2) or Very Long Chain AcylCoA Dehydrogenase (VLCAD) deficiency we characterized the metabolic effects of RSV, RSV metabolites, and other stilbenes. We also focused on analysis of RSV uptake, and on the effects of low RSV concentrations, considering the limited bioavailability of RSV in vivo. Results Time course of RSV accumulation in fibroblasts over 48 h of treatment were consistent with the resulting stimulation or correction of FAO capacities. At 48 h, half maximal and maximal FAO stimulations were respectively achieved for 37,5 microM (EC50) and 75 microM RSV, but we found that serum content of culture medium negatively modulated RSV uptake and FAO induction. Indeed, decreasing serum from 12% to 3% led to shift EC50 from 37,5 to 13 microM, and a 2.6-3.6-fold FAO stimulation was reached with 20 microM RSV at 3% serum, that was absent at 12% serum. Two other stilbenes often found associated with RSV, i.e. cis- RSV and piceid, also triggered significant FAO up-regulation. Resveratrol glucuro- or sulfo- conjugates had modest or no effects. In contrast, dihydro-RSV, one of the most abundant circulating RSV metabolites in human significantly stimulated FAO (1.3-2.3-fold). Conclusions This study provides the first compared data on

  3. Impaired fatty acid oxidation in a Drosophila model of mitochondrial trifunctional protein (MTP) deficiency.

    PubMed

    Kishita, Yoshihito; Tsuda, Manabu; Aigaki, Toshiro

    2012-03-09

    Mitochondrial trifunctional protein (MTP), which consists of the MTPα and MTPβ subunits, catalyzes long-chain fatty acid β-oxidation. MTP deficiency in humans results in Reye-like syndrome. Here, we generated Drosophila models of MTP deficiency by targeting two genes encoding Drosophila homologs of human MTPα and MTPβ, respectively. Both Mtpα(KO) and Mtpβ(KO) flies were viable, but demonstrated reduced lifespan, defective locomotor activity, and reduced fecundity represented by the number of eggs laid by the females. The phenotypes of Mtpα(KO) flies were generally more striking than those of Mtpβ(KO) flies. Mtpα(KO) flies were hypersensitive to fasting, and retained lipid droplets in their fat body cells as in non-fasting conditions. The amount of triglyceride was also unchanged upon fasting in Mtpα(KO) flies, suggesting that lipid mobilization was disrupted. Finally, we showed that both Mtpα(KO) and Mtpβ(KO) flies accumulated acylcarnitine and hydroxyacylcarnitine, diagnostic markers of MTP deficiencies in humans. Our results indicated that both Mtpα(KO) and Mtpβ(KO) flies were impaired in long-chain fatty acid β-oxidation. These flies should be useful as a model system to investigate the molecular pathogenesis of MTP deficiency.

  4. Analysis of organic acids after incubation with (16-2H3)palmitic acid in fibroblasts from patients with mitochondrial beta-oxidation defects.

    PubMed

    Osorio, J H; Lluch, M; Ribes, A

    2003-01-01

    The analysis of acylcarnitines as products of incubation of intact fibroblasts with isotope-labelled precursors, usually (16-(2)H(3))hexadecanoic acid, is an advanced in vitro method for the study of mitochondrial beta-oxidation defects. We propose a technique for the measurement of the organic acid intermediates after hydrolysis of the acylcarnitines using electron-impact gas chromatography-mass spectrometry. For some mitochondrial beta-oxidation deficiencies, the characteristic profile enables us to approach the diagnosis with clear differentiation.

  5. Acyl-CoA thioesterase-2 facilitates mitochondrial fatty acid oxidation in the liver[S

    PubMed Central

    Moffat, Cynthia; Bhatia, Lavesh; Nguyen, Teresa; Lynch, Peter; Wang, Miao; Wang, Dongning; Ilkayeva, Olga R.; Han, Xianlin; Hirschey, Matthew D.; Claypool, Steven M.; Seifert, Erin L.

    2014-01-01

    Acyl-CoA thioesterase (Acot)2 localizes to the mitochondrial matrix and hydrolyses long-chain fatty acyl-CoA into free FA and CoASH. Acot2 is expressed in highly oxi­dative tissues and is poised to modulate mitochondrial FA oxidation (FAO), yet its biological role is unknown. Using a model of adenoviral Acot2 overexpression in mouse liver (Ad-Acot2), we show that Acot2 increases the utilization of FA substrate during the daytime in ad libitum-fed mice, but the nighttime switch to carbohydrate oxidation is similar to control mice. In further support of elevated FAO in Acot2 liver, daytime serum ketones were higher in Ad-Acot2 mice, and overnight fasting led to minimal hepatic steatosis as compared with control mice. In liver mitochondria from Ad-Acot2 mice, phosphorylating O2 consumption was higher with lipid substrate, but not with nonlipid substrate. This increase depended on whether FA could be activated on the outer mitochondrial membrane, suggesting that the FA released by Acot2 could be effluxed from mitochondria then taken back up again for oxidation. This circuit would prevent the build-up of inhibitory long-chain fatty acyl-CoA esters. Altogether, our findings indicate that Acot2 can enhance FAO, possibly by mitigating the accumulation of FAO intermediates within the mitochondrial matrix. PMID:25114170

  6. Fatty Acid Oxidation-Driven Src Links Mitochondrial Energy Reprogramming and Regulation of Oncogenic Properties in Triple Negative Breast Cancer

    PubMed Central

    Park, Jun Hyoung; Vithayathil, Sajna; Kumar, Santosh; Sung, Pi-Lin; Dobrolecki, Lacey Elizabeth; Putluri, Vasanta; Bhat, Vadiraja B.; Bhowmik, Salil Kumar; Gupta, Vineet; Arora, Kavisha; Wu, Danli; Tsouko, Efrosini; Zhang, Yiqun; Maity, Suman; Donti, Taraka R.; Graham, Brett H.; Frigo, Daniel E.; Coarfa, Cristian; Yotnda, Patricia; Putluri, Nagireddy; Sreekumar, Arun; Lewis, Michael T.; Creighton, Chad J.; Wong, Lee-Jun C.; Kaipparettu, Benny Abraham

    2016-01-01

    Summary Transmitochondrial cybrids and multiple OMICs approaches were used to understand mitochondrial reprogramming and mitochondria-regulated cancer pathways in triple negative breast cancer (TNBC). Analysis of cybrids and established breast cancer (BC) cell lines showed that metastatic TNBC maintains high levels of ATP through fatty acid β-oxidation (FAO) and activates Src oncoprotein through autophosphorylation at Y419. Manipulation of FAO including the knocking down of carnitine palmitoyltransferase-1 (CPT1) and 2 (CPT2), the rate-limiting proteins of FAO, and analysis of patient-derived xenograft models, confirmed the role of mitochondrial FAO in Src activation and metastasis. Analysis of TCGA and other independent BC clinical data further reaffirmed the role of mitochondrial FAO and CPT genes in Src regulation and their significance in BC metastasis. PMID:26923594

  7. Fatty Acid Oxidation-Driven Src Links Mitochondrial Energy Reprogramming and Oncogenic Properties in Triple-Negative Breast Cancer.

    PubMed

    Park, Jun Hyoung; Vithayathil, Sajna; Kumar, Santosh; Sung, Pi-Lin; Dobrolecki, Lacey Elizabeth; Putluri, Vasanta; Bhat, Vadiraja B; Bhowmik, Salil Kumar; Gupta, Vineet; Arora, Kavisha; Wu, Danli; Tsouko, Efrosini; Zhang, Yiqun; Maity, Suman; Donti, Taraka R; Graham, Brett H; Frigo, Daniel E; Coarfa, Cristian; Yotnda, Patricia; Putluri, Nagireddy; Sreekumar, Arun; Lewis, Michael T; Creighton, Chad J; Wong, Lee-Jun C; Kaipparettu, Benny Abraham

    2016-03-08

    Transmitochondrial cybrids and multiple OMICs approaches were used to understand mitochondrial reprogramming and mitochondria-regulated cancer pathways in triple-negative breast cancer (TNBC). Analysis of cybrids and established breast cancer (BC) cell lines showed that metastatic TNBC maintains high levels of ATP through fatty acid β oxidation (FAO) and activates Src oncoprotein through autophosphorylation at Y419. Manipulation of FAO including the knocking down of carnitine palmitoyltransferase-1A (CPT1) and 2 (CPT2), the rate-limiting proteins of FAO, and analysis of patient-derived xenograft models confirmed the role of mitochondrial FAO in Src activation and metastasis. Analysis of TCGA and other independent BC clinical data further reaffirmed the role of mitochondrial FAO and CPT genes in Src regulation and their significance in BC metastasis.

  8. Experimental Evidence that 3-Methylglutaric Acid Disturbs Mitochondrial Function and Induced Oxidative Stress in Rat Brain Synaptosomes: New Converging Mechanisms.

    PubMed

    Colín-González, Ana Laura; Paz-Loyola, Ariana Lizbeth; de Lima, María Eduarda; Galván-Arzate, Sonia; Seminotti, Bianca; Ribeiro, César Augusto João; Leipnitz, Guilhian; Souza, Diogo Onofre; Wajner, Moacir; Santamaría, Abel

    2016-10-01

    3-Methylglutaric acid (3MGA) is an organic acid that accumulates in various organic acidemias whose patients present neurodegeneration events in children coursing with metabolic acidurias. Limited evidence describes the toxic mechanisms elicited by 3MGA in the brain. Herein, we explored the effects of 3MGA on different toxic endpoints in synaptosomal and mitochondrial-enriched fractions of adult rat brains to provide novel information on early mechanisms evoked by this metabolite. At 1 and 5 mM concentration, 3MGA increased lipid peroxidation, but decreased mitochondrial function only at 5 mM concentration. Despite less intense effects were obtained at 1 mM concentration, its co-administration with the kynurenine pathway (KP) metabolite and N-methyl-D-aspartate receptor (NMDAr) agonist, quinolinic acid (QUIN, 50 and 100 µM), produced toxic synergism on markers of oxidative stress and mitochondrial function. The toxicity of 3MGA per se (5 mM) was prevented by the cannabinoid receptor agonist WIN55,212-2 and the NMDAr antagonist kynurenic acid (KYNA), suggesting cannabinoid and glutamatergic components in the 3MGA pattern of toxicity. The synergic model (3MGA + QUIN) was also sensitive to KYNA and the antioxidant S-allylcysteine, but not to the nitric oxide synthase inhibitor L-nitroarginine methyl ester. These findings suggest various underlying mechanisms involved in the neurotoxicity of 3MGA that may possibly contribute to the neurodegeneration observed in acidemias.

  9. Oleic acid increases mitochondrial reactive oxygen species production and decreases endothelial nitric oxide synthase activity in cultured endothelial cells.

    PubMed

    Gremmels, Hendrik; Bevers, Lonneke M; Fledderus, Joost O; Braam, Branko; van Zonneveld, Anton Jan; Verhaar, Marianne C; Joles, Jaap A

    2015-03-15

    Elevated plasma levels of free fatty acids (FFA) are associated with increased cardiovascular risk. This may be related to FFA-induced elevation of oxidative stress in endothelial cells. We hypothesized that, in addition to mitochondrial production of reactive oxygen species, endothelial nitric oxide synthase (eNOS)-mediated reactive oxygen species production contributes to oleic acid (OA)-induced oxidative stress in endothelial cells, due to eNOS uncoupling. We measured reactive oxygen species production and eNOS activity in cultured endothelial cells (bEnd.3) in the presence of OA bound to bovine serum albumin, using the CM-H2DCFDA assay and the L-arginine/citrulline conversion assay, respectively. OA induced a concentration-dependent increase in reactive oxygen species production, which was inhibited by the mitochondrial complex II inhibitor thenoyltrifluoroacetone (TTFA). OA had little effect on eNOS activity when stimulated by a calcium-ionophore, but decreased both basal and insulin-induced eNOS activity, which was restored by TTFA. Pretreatment of bEnd.3 cells with tetrahydrobiopterin (BH4) prevented OA-induced reactive oxygen species production and restored inhibition of eNOS activity by OA. Elevation of OA levels leads to both impairment in receptor-mediated stimulation of eNOS and to production of mitochondrial-derived reactive oxygen species and hence endothelial dysfunction.

  10. Effect of omega-3 fatty acid oxidation products on the cellular and mitochondrial toxicity of BDE 47.

    PubMed

    Yeh, Andrew; Kruse, Shane E; Marcinek, David J; Gallagher, Evan P

    2015-06-01

    High levels of the flame retardant 2,2',4,4'-tetrabromodiphenyl ether (BDE 47) have been detected in Pacific salmon sampled near urban areas, raising concern over the safety of salmon consumption. However, salmon fillets also contain the antioxidants eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), whose oxidation products induce cellular antioxidant responses. Because oxidative stress is a mechanism of BDE 47 toxicity, we hypothesized that oxidized EPA and DHA can ameliorate the cellular and mitochondrial toxicity of BDE 47. HepG2 cells were treated with a mixture of oxidized EPA and DHA (oxEPA/oxDHA) at a ratio relevant to salmon consumption (1.5/1 oxEPA/oxDHA) followed by exposure to 100 μM BDE 47. Pretreatment with oxEPA/oxDHA for 12 h prior to BDE 47 exposure prevented BDE 47-mediated depletion of glutathione, and increased expression of antioxidant response genes. oxEPA/oxDHA also reduced the level of reactive oxygen species production by BDE 47. The oxEPA/oxDHA antioxidant responses were associated with partial protection against BDE 47-induced loss of viability and also mitochondrial membrane potential. Mitochondrial electron transport system functional analysis revealed extensive inhibition of State 3 respiration and maximum respiratory capacity by BDE 47 were partially reversed by oxEPA/oxDHA. Our findings indicate that the antioxidant effects of oxEPA/oxDHA protect against short exposures to BDE 47, including a protective role of these compounds on maintaining cellular and mitochondrial function.

  11. In Vivo Determination of Mitochondrial Function Using Luciferase-Expressing Caenorhabditis elegans: Contribution of Oxidative Phosphorylation, Glycolysis, and Fatty Acid Oxidation to Toxicant-Induced Dysfunction.

    PubMed

    Luz, Anthony L; Lagido, Cristina; Hirschey, Matthew D; Meyer, Joel N

    2016-08-01

    Mitochondria are a target of many drugs and environmental toxicants; however, how toxicant-induced mitochondrial dysfunction contributes to the progression of human disease remains poorly understood. To address this issue, in vivo assays capable of rapidly assessing mitochondrial function need to be developed. Here, using the model organism Caenorhabditis elegans, we describe how to rapidly assess the in vivo role of the electron transport chain, glycolysis, or fatty acid oxidation in energy metabolism following toxicant exposure, using a luciferase-expressing ATP reporter strain. Alterations in mitochondrial function subsequent to toxicant exposure are detected by depleting steady-state ATP levels with inhibitors of the mitochondrial electron transport chain, glycolysis, or fatty acid oxidation. Differential changes in ATP following short-term inhibitor exposure indicate toxicant-induced alterations at the site of inhibition. Because a microplate reader is the only major piece of equipment required, this is a highly accessible method for studying toxicant-induced mitochondrial dysfunction in vivo. © 2016 by John Wiley & Sons, Inc.

  12. The enzymology of mitochondrial fatty acid beta-oxidation and its application to follow-up analysis of positive neonatal screening results.

    PubMed

    Wanders, Ronald J A; Ruiter, Jos P N; IJLst, Lodewijk; Waterham, Hans R; Houten, Sander M

    2010-10-01

    Oxidation of fatty acids in mitochondria is a key physiological process in higher eukaryotes including humans. The importance of the mitochondrial beta-oxidation system in humans is exemplified by the existence of a group of genetic diseases in man caused by an impairment in the mitochondrial oxidation of fatty acids. Identification of patients with a defect in mitochondrial beta-oxidation has long remained notoriously difficult, but the introduction of tandem-mass spectrometry in laboratories for genetic metabolic diseases has revolutionalized the field by allowing the rapid and sensitive analysis of acylcarnitines. Equally important is that much progress has been made with respect to the development of specific enzyme assays to identify the enzyme defect in patients subsequently followed by genetic analysis. In this review, we will describe the current state of knowledge in the field of fatty acid oxidation enzymology and its application to the follow-up analysis of positive neonatal screening results.

  13. Role of calcium signaling in the activation of mitochondrial nitric oxide synthase and citric acid cycle.

    PubMed

    Traaseth, Nathaniel; Elfering, Sarah; Solien, Joseph; Haynes, Virginia; Giulivi, Cecilia

    2004-07-23

    An apparent discrepancy arises about the role of calcium on the rates of oxygen consumption by mitochondria: mitochondrial calcium increases the rate of oxygen consumption because of the activation of calcium-activated dehydrogenases, and by activating mitochondrial nitric oxide synthase (mtNOS), decreases the rates of oxygen consumption because nitric oxide is a competitive inhibitor of cytochrome oxidase. To this end, the rates of oxygen consumption and nitric oxide production were followed in isolated rat liver mitochondria in the presence of either L-Arg (to sustain a mtNOS activity) or N(G)-monomethyl-L-Arg (NMMA, a competitive inhibitor of mtNOS) under State 3 conditions. In the presence of NMMA, the rates of State 3 oxygen consumption exhibited a K(0.5) of 0.16 microM intramitochondrial free calcium, agreeing with those required for the activation of the Krebs cycle. By plotting the difference between the rates of oxygen consumption in State 3 with L-Arg and with NMMA at various calcium concentrations, a K(0.5) of 1.2 microM intramitochondrial free calcium was obtained, similar to the K(0.5) (0.9 microM) of the dependence of the rate of nitric oxide production on calcium concentrations. The activation of dehydrogenases, followed by the activation of mtNOS, would lead to the modulation of the Krebs cycle activity by the modulation of nitric oxide on the respiratory rates. This would ensue in changes in the NADH/NAD and ATP/ADP ratios, which would influence the rate of the cycle and the oxygen diffusion.

  14. Characterization of reduced and oxidized dopamine and 3,4-dihydrophenylacetic acid, on brain mitochondrial electron transport chain activities.

    PubMed

    Gautam, Alpa H; Zeevalk, Gail D

    2011-07-01

    Loss of dopamine (DA) homeostasis may be a contributing factor to cell damage in Parkinson's disease (PD). Past studies showing deleterious effects of DA on mitochondrial function, however, have been inconsistent raising questions about mitochondria as a downstream target for DA. Issues such as the dopamine species i.e., reduced or oxidized, time of exposure and the effect of major metabolites such as 3,4-dihydrophenylacetic acid (DOPAC) may contribute to the disparate findings. The present study used isolated, lysed rat brain mitochondria to characterize the effects of oxidized or reduced DA and DOPAC on complex activities of the electron transport chain (ETC). Time of exposure and quantitation of reduced or oxidized catachols for DA and DOPAC were monitored for all experiments. Reduced DA and DOPAC with or without a 30min preincubation had no affect on NADH oxidase activity which monitors the activities of complexes I, III and IV. Complex II activity was inhibited by reduced DA (≥500μM), but not by reduced DOPAC and was significantly attenuated by SOD suggesting reactive oxygen species involvement. In contrast, fully oxidized DA and DOPAC dose dependently inhibited NADH oxidase, complex I and complex III activities with IC(50s) in the 50-200μM range. No preincubation was required for inhibition with the catechols when they were fully oxidized. Oxidized DA inhibited complex I only when exposure occurred during stimulated electron flow, suggesting covalent binding of quinones to proteins within active sites of the complex. In intact, well coupled mitochondria, extramitochondrial DA was shown to access the mitochondrial matrix in a dose, time and energy-dependent fashion. The findings suggest that many of the reported inconsistencies with regards to the effects of DA and DOPAC on ETC function can be attributed to the oxidized state of the catechol at the time of exposure. In addition, the findings provide possible downstream targets for DA that could contribute

  15. Statins Increase Mitochondrial and Peroxisomal Fatty Acid Oxidation in the Liver and Prevent Non-Alcoholic Steatohepatitis in Mice

    PubMed Central

    Park, Han-Sol; Jang, Jung Eun; Ko, Myoung Seok; Woo, Sung Hoon; Kim, Bum Joong; Kim, Hyun Sik; Park, Hye Sun; Park, In-Sun; Koh, Eun Hee

    2016-01-01

    Background Non-alcoholic fatty liver disease is the most common form of chronic liver disease in industrialized countries. Recent studies have highlighted the association between peroxisomal dysfunction and hepatic steatosis. Peroxisomes are intracellular organelles that contribute to several crucial metabolic processes, such as facilitation of mitochondrial fatty acid oxidation (FAO) and removal of reactive oxygen species through catalase or plasmalogen synthesis. Statins are known to prevent hepatic steatosis and non-alcoholic steatohepatitis (NASH), but underlying mechanisms of this prevention are largely unknown. Methods Seven-week-old C57BL/6J mice were given normal chow or a methionine- and choline-deficient diet (MCDD) with or without various statins, fluvastatin, pravastatin, simvastatin, atorvastatin, and rosuvastatin (15 mg/kg/day), for 6 weeks. Histological lesions were analyzed by grading and staging systems of NASH. We also measured mitochondrial and peroxisomal FAO in the liver. Results Statin treatment prevented the development of MCDD-induced NASH. Both steatosis and inflammation or fibrosis grades were significantly improved by statins compared with MCDD-fed mice. Gene expression levels of peroxisomal proliferator-activated receptor α (PPARα) were decreased by MCDD and recovered by statin treatment. MCDD-induced suppression of mitochondrial and peroxisomal FAO was restored by statins. Each statin's effect on increasing FAO and improving NASH was independent on its effect of decreasing cholesterol levels. Conclusion Statins prevented NASH and increased mitochondrial and peroxisomal FAO via induction of PPARα. The ability to increase hepatic FAO is likely the major determinant of NASH prevention by statins. Improvement of peroxisomal function by statins may contribute to the prevention of NASH. PMID:27098507

  16. IDH1 mutations alter citric acid cycle metabolism and increase dependence on oxidative mitochondrial metabolism.

    PubMed

    Grassian, Alexandra R; Parker, Seth J; Davidson, Shawn M; Divakaruni, Ajit S; Green, Courtney R; Zhang, Xiamei; Slocum, Kelly L; Pu, Minying; Lin, Fallon; Vickers, Chad; Joud-Caldwell, Carol; Chung, Franklin; Yin, Hong; Handly, Erika D; Straub, Christopher; Growney, Joseph D; Vander Heiden, Matthew G; Murphy, Anne N; Pagliarini, Raymond; Metallo, Christian M

    2014-06-15

    Oncogenic mutations in isocitrate dehydrogenase 1 and 2 (IDH1/2) occur in several types of cancer, but the metabolic consequences of these genetic changes are not fully understood. In this study, we performed (13)C metabolic flux analysis on a panel of isogenic cell lines containing heterozygous IDH1/2 mutations. We observed that under hypoxic conditions, IDH1-mutant cells exhibited increased oxidative tricarboxylic acid metabolism along with decreased reductive glutamine metabolism, but not IDH2-mutant cells. However, selective inhibition of mutant IDH1 enzyme function could not reverse the defect in reductive carboxylation activity. Furthermore, this metabolic reprogramming increased the sensitivity of IDH1-mutant cells to hypoxia or electron transport chain inhibition in vitro. Lastly, IDH1-mutant cells also grew poorly as subcutaneous xenografts within a hypoxic in vivo microenvironment. Together, our results suggest therapeutic opportunities to exploit the metabolic vulnerabilities specific to IDH1 mutation.

  17. IDH1 Mutations Alter Citric Acid Cycle Metabolism and Increase Dependence on Oxidative Mitochondrial Metabolism

    PubMed Central

    Grassian, Alexandra R.; Parker, Seth J.; Davidson, Shawn M.; Divakarun, Ajit S.; Green, Courtney R.; Zhang, Xiamei; Slocum, Kelly L.; Pu, Minying; Lin, Fallon; Vickers, Chad; Joud-Caldwell, Carol; Chung, Franklin; Yin, Hong; Handly, Erika D.; Straub, Christopher; Growney, Joseph D.; Vander Heiden, Matthew G.; Murphy, Anne N.; Pagliarini, Raymond; Metallo, Christian M.

    2016-01-01

    Oncogenic mutations in isocitrate dehydrogenase 1 and 2 (IDH1/2) occur in several types of cancer, but the metabolic consequences of these genetic changes are not fully understood. In this study, we performed 13C metabolic flux analysis on a panel of isogenic cell lines containing heterozygous IDH1/2 mutations. We observed that under hypoxic conditions, IDH1-mutant cells exhibited increased oxidative tricarboxylic acid metabolism along with decreased reductive glutamine metabolism, but not IDH2-mutant cells. However, selective inhibition of mutant IDH1 enzyme function could not reverse the defect in reductive carboxylation activity. Furthermore, this metabolic reprogramming increased the sensitivity of IDH1-mutant cells to hypoxia or electron transport chain inhibition in vitro. Lastly, IDH1-mutant cells also grew poorly as subcutaneous xenografts within a hypoxic in vivo microenvironment. Together, our results suggest therapeutic opportunities to exploit the metabolic vulnerabilities specific to IDH1 mutation. PMID:24755473

  18. Human acyl-CoA dehydrogenase-9 plays a novel role in the mitochondrial beta-oxidation of unsaturated fatty acids.

    PubMed

    Ensenauer, Regina; He, Miao; Willard, Jan-Marie; Goetzman, Eric S; Corydon, Thomas J; Vandahl, Brian B; Mohsen, Al-Walid; Isaya, Grazia; Vockley, Jerry

    2005-09-16

    Unsaturated fatty acids play an important role in the prevention of human diseases such as diabetes, obesity, cancer, and neurodegeneration. However, their oxidation in vivo by acyl-CoA dehydrogenases (ACADs) that catalyze the first step of each cycle of mitochondrial fatty acid beta-oxidation is not entirely understood. Recently, a novel ACAD (ACAD-9) of unknown function that is highly homologous to human very-long-chain acyl-CoA dehydrogenase was identified by large-scale random sequencing. To characterize its enzymatic role, we have expressed ACAD-9 in Escherichia coli, purified it, and determined its pattern of substrate utilization. The N terminus of the mature form of the enzyme was identified by in vitro mitochondrial import studies of precursor protein. A 37-amino acid leader peptide was cleaved sequentially by two mitochondrial peptidases to yield a predicted molecular mass of 65 kDa for the mature subunit. Submitochondrial fractionation studies found native ACAD-9 to be associated with the mitochondrial membrane. Gel filtration analysis indicated that, like very-long-chain acyl-CoA dehydrogenase, ACAD-9 is a dimer, in contrast to the other known ACADs, which are tetramers. Purified mature ACAD-9 had maximal activity with long-chain unsaturated acyl-CoAs as substrates (C16:1-, C18:1-, C18:2-, C22:6-CoA). These results suggest a previously unrecognized role for ACAD-9 in the mitochondrial beta-oxidation of long-chain unsaturated fatty acids. Because of the substrate specificity and abundance of ACAD-9 in brain, we speculate that it may play a role in the turnover of lipid membrane unsaturated fatty acids that are essential for membrane integrity and structure.

  19. Pretreatment with Bacopa monnieri extract offsets 3-nitropropionic acid induced mitochondrial oxidative stress and dysfunctions in the striatum of prepubertal mouse brain.

    PubMed

    Shinomol, George K; Bharath, M M Srinivas; Muralidhara

    2012-05-01

    The present investigation was designed to determine the efficacy of Bacopa monnieri (Brahmi; BM) to offset 3-nitropropionic acid (3-NPA) induced oxidative stress and mitochondrial dysfunction in dopaminergic (N27) cells and prepubertal mouse brain. Pretreatment of N27 cells with BM ethanolic extract (BME) significantly attenuated 3-NPA-induced cytotoxicity. Further, we determined the degree of oxidative stress induction, redox status, enzymic antioxidants, and protein oxidation in the striatal mitochondria of mice given BME prophylaxis followed by 3-NPA challenge. While 3-NPA-induced marked oxidative stress in the mitochondria of the striatum, BME prophylaxis markedly prevented 3-NPA-induced oxidative dysfunctions and depletion of reduced glutathione and thiol levels. The activities of antioxidant enzymes (superoxide dismutase, glutathione peroxidase, glutathione reductase, thioredoxin reductase), Na(+),K(+)-ATPase, and citric acid cycle enzymes in the striatum discernible among 3-NPA mice were significantly restored with BME prophylaxis. Interestingly, BME offered protection against 3-NPA-induced mitochondrial dysfunctions as evidenced by the restoration of the activities of ETC enzymes (NADH:ubiquinone oxidoreductase, NADH:cytochrome c reductase, succinate-ubiquinone oxidoreductase, and cytochrome c oxidase) and mitochondrial viability. We hypothesize that the neuroprotective effects of BME may be wholly or in part related to its propensity to scavenge free radicals, maintain redox status, and upregulate antioxidant machinery in striatal mitochondria.

  20. Heterologous expression of human carnitine palmitoyltransferase (CPT) II in yeast: A model for the molecular analysis of mitochondrial fatty acid oxidation defects

    SciTech Connect

    Cavadini, P.; Invernizzi, F.; Baratta, S.

    1994-09-01

    The CPT enzyme system, which is composed of two distinct mitochondrial membrane-bound proteins (CPT I and CPT II), provides the mechanism whereby long-chain fatty acids are transferred from the cytosol to the mitochondrial matrix to undergo {beta}-oxidation. Here, we report the development of an expression system for investigating genotype/phenotype correlations in CPT II deficiency and, potentially, other mitochondrial fatty acid oxidation defects. To explore yeast as an expression system, we introduced a cDNA encoding the entire human CPT II precursor into Saccharomyces cerevisiae. Expression was programmed by using an inducible galactose operon promoter (GAL1). Following induction, human CPT II was expressed at high levels, with activity 4- to 16-fold greater than in human fibroblasts. Levels of expression paralleled those of respiration, being higher in cells grown on a nonfermentable carbon source than in those grown on glucose. Immunoprecipitation of pulse-labeled transformed cells demonstrated that human CPT II expressed in yeast was targeted to mitochondria with correct proteolytic processing of its 25-residue mitochondrial leader sequence. Preliminary results on the expression of a number of mutant CPT II alleles associated with different clinical phenotypes demonstrated the value of this system for examining the functional consequences of disease-causing mutations and investigating genotype/phenotype correlations in patients with CPT II deficiency.

  1. Long-chain 3-hydroxy fatty acids accumulating in long-chain 3-hydroxyacyl-CoA dehydrogenase and mitochondrial trifunctional protein deficiencies uncouple oxidative phosphorylation in heart mitochondria.

    PubMed

    Tonin, Anelise M; Amaral, Alexandre U; Busanello, Estela N B; Grings, Mateus; Castilho, Roger F; Wajner, Moacir

    2013-02-01

    Cardiomyopathy is a common clinical feature of some inherited disorders of mitochondrial fatty acid β-oxidation including mitochondrial trifunctional protein (MTP) and isolated long-chain 3-hydroxyacyl-CoA dehydrogenase (LCHAD) deficiencies. Since individuals affected by these disorders present tissue accumulation of various fatty acids, including long-chain 3-hydroxy fatty acids, in the present study we investigated the effect of 3-hydroxydecanoic (3 HDCA), 3-hydroxydodecanoic (3 HDDA), 3-hydroxytetradecanoic (3 HTA) and 3-hydroxypalmitic (3 HPA) acids on mitochondrial oxidative metabolism, estimated by oximetry, NAD(P)H content, hydrogen peroxide production, membrane potential (ΔΨ) and swelling in rat heart mitochondrial preparations. We observed that 3 HTA and 3 HPA increased resting respiration and diminished the respiratory control and ADP/O ratios using glutamate/malate or succinate as substrates. Furthermore, 3 HDDA, 3 HTA and 3 HPA decreased ΔΨ, the matrix NAD(P)H pool and hydrogen peroxide production. These data indicate that these fatty acids behave as uncouplers of oxidative phosphorylation. We also verified that 3 HTA-induced uncoupling-effect was not mediated by the adenine nucleotide translocator and that this fatty acid induced the mitochondrial permeability transition pore opening in calcium-loaded organelles since cyclosporin A prevented the reduction of mitochondrial ΔΨ and swelling provoked by 3 HTA. The present data indicate that major 3-hydroxylated fatty acids accumulating in MTP and LCHAD deficiencies behave as strong uncouplers of oxidative phosphorylation potentially impairing heart energy homeostasis.

  2. Neuroprotective effect of asiatic acid on rotenone-induced mitochondrial dysfunction and oxidative stress-mediated apoptosis in differentiated SH-SYS5Y cells.

    PubMed

    Nataraj, Jagatheesan; Manivasagam, Thamilarasan; Justin Thenmozhi, Arokiasamy; Essa, Musthafa Mohamed

    2016-02-08

    Parkinson's disease (PD) is a chronic neurodegenerative disease, manifested due to the loss of dopaminergic neurons, which ultimately leads to impaired movement in elderly populations. The pathogenesis of PD is associated with numerous factors including oxidative stress, mitochondrial dysfunction and apoptosis. There is no effective therapy available to cure or halt the progression of this disease still now. Asiatic acid (AA) is a triterpene extracted from Centella asiatica has been reported as an antioxidant and anti-inflammatory agent, that offers neuroprotection against glutamate toxicity. Therefore, in this study, we have investigated the effect of AA in a rotenone (an inhibitor of mitochondrial complex I) induced in vitro model of PD. Following the exposure of SH-SY5Y cells to rotenone, there was a marked overproduction of ROS, mitochondrial dysfunction (as indexed by the decrease in mitochondrial membrane potential) and apoptosis (Hoechst and dual staining, comet assay; expressions of pro-apoptotic and anti-apoptotic indices). Pre-treatment with AA reversed these changes might be due to its antioxidant, mitoprotective and anti-apoptotic properties. However further extensive studies on in vivo models of PD are warranted to prove AA neuroprotective effect before entering into the clinical trial.

  3. Oxidative Stress in Inherited Mitochondrial Diseases

    PubMed Central

    Hayashi, Genki; Cortopassi, Gino

    2015-01-01

    Mitochondria are a source of reactive oxygen species (ROS). Mitochondrial diseases are the result of inherited defects in mitochondrially-expressed genes. One potential pathomechanism for mitochondrial disease is oxidative stress. Oxidative stress can occur as the result of increased ROS production, or decreased ROS protection. The role of oxidative stresses in the five most common inherited mitochondrial diseases; Friedreich's ataxia (FA), LHON, MELAS, MERRF and Leigh Syndrome (LS) is discussed. Published reports for oxidative stress involvement in pathomechanism in these five mitochondrial diseases are reviewed. The strongest for oxidative stress pathomechanism among the five diseases was in Friedreich's ataxia. In addition, a meta-analysis was carried out to provide an unbiased evaluation of the role of oxidative stress in the five diseases, by searching for oxidative stress citation count frequency within each disease. Of the five most common mitochondrial diseases, the strongest support for oxidative stress is in Friedreich's ataxia (6.42%), followed by LHON (2.45%), MELAS (2.18%), MERRF (1.71%), and LS (1.03%). The increased frequency of oxidative stress citations was significant relative to the mean of the total pool of five diseases (p<0.01) and the mean of the four non-Friedreich's diseases (p<0.0001). Thus there is support for oxidative stress in all five most common mitochondrial diseases, but the strongest, significant support is for Friedreich's ataxia. PMID:26073122

  4. Mitochondrial nitric oxide synthase regulates mitochondrial matrix pH.

    PubMed

    Ghafourifar, P; Richter, C

    1999-01-01

    Nitric oxide (nitrogen monoxide, NO) exerts a wide profile of its biological activities via regulation of respiration and respiration-dependent functions. The presence of nitric oxide synthase (NOS) in mitochondria (mtNOS) was recently reported by us (Ghafourifar and Richter, FEBS Lett. 418, 291-296, 1997) and others (Giulivi et al., J. Biol. Chem. 273, 11038-11043, 1998). Here we report that NO, provided by an NO donor as well as by mtNOS stimulation, regulates mitochondrial matrix pH, transmembrane potential and Ca2+ buffering capacity. Exogenously-added NO causes a dose-dependent matrix acidification. Also mtNOS stimulation, induced by loading mitochondria with Ca2+, causes mitochondrial matrix acidification and a drop in mitochondrial transmembrane potential. Inhibition of mtNOS's basal activity causes mitochondrial matrix alkalinization and provides a resistance to the sudden drop of mitochondrial transmembrane potential induced by mitochondrial Ca2+ uptake. We conclude that mtNOS plays a critical role in regulating mitochondrial delta(pH).

  5. Analysis of carnitine esters by radio-high performance liquid chromatography in cultured skin fibroblasts from patients with mitochondrial fatty acid oxidation disorders.

    PubMed

    Schmidt-Sommerfeld, E; Bobrowski, P J; Penn, D; Rhead, W J; Wanders, R J; Bennett, M J

    1998-08-01

    Acylcarnitines are important diagnostic markers for inborn errors of fatty acid oxidation, but their analysis in body fluids may not always be reliable. Recently, disease-specific acylcarnitine profiles generated by cultured skin fibroblasts were reported to facilitate the diagnosis by localizing a specific enzymatic defect in the mitochondrial beta-oxidation pathway. Using a novel methodologic approach, fibroblasts from 16 patients with inborn errors of fatty acid oxidation and 13 control subjects were preincubated with L-[3H]carnitine to label the intracellular carnitine pool. Cells were subsequently incubated with unlabeled palmitic acid and, after methanol extraction of cells and media, labeled free carnitine and acylcarnitines were analyzed by radio-HPLC. Quantitation was based on the integrated radioactivity of individual peaks relative to the total radioactivity recovered. In control cell lines, all saturated acylcarnitines were detected, and reference values were established. With the exception of one cell line deficient in electron transfer flavoprotein, all mutant cell lines showed abnormal and disease-specific relative concentrations of acylcarnitines. Advantages of the method include use of a small number of cells, no need for trypsinization and permeabilization of cells before incubation, simple extraction without purification of the specimen before HPLC, and relatively inexpensive equipment. The method allows a focused approach to the subsequent, more laborious confirmation of a particular disease by direct enzymatic and/or molecular analysis. It remains to be established whether the method can replace widely used global measurements of fatty acid oxidation rates in vitro that do not provide specific information about the enzyme deficiency involved.

  6. Contribution of mitochondrial oxidative stress to hypertension

    PubMed Central

    Dikalov, Sergey I.; Dikalova, Anna E.

    2016-01-01

    Purpose of review In 1954 Harman proposed the free radical theory of aging, and in 1972 he suggested that mitochondria are both the source and the victim of toxic free radicals. Interestingly, hypertension is age-associated disease and clinical data show that by age 70, 70% of the population has hypertension and this is accompanied by oxidative stress. Antioxidant therapy however is not currently available and common antioxidants like ascorbate and vitamin E are ineffective in preventing hypertension. The present review focuses on molecular mechanisms of mitochondrial oxidative stress and therapeutic potential of targeting mitochondria in hypertension. Recent findings In the past several years, we have shown that the mitochondria become dysfunctional in hypertension and have defined novel role of mitochondrial superoxide radicals in this disease. We have shown that genetic manipulation of mitochondrial antioxidant enzyme superoxide dismutase (SOD2) affects blood pressure and have developed mitochondria-targeted therapies such as SOD2 mimetics that effectively lower blood pressure. The specific mechanism of mitochondrial oxidative stress in hypertension, however, remains unclear. Recent animal and clinical studies have demonstrated several hormonal, metabolic, inflammatory, and environmental pathways contributing to mitochondrial dysfunction and oxidative stress. Summary Nutritional supplements, calorie restriction, and life style change are the most effective preventive strategies to improve mitochondrial function and reduce mitochondrial oxidative stress. Aging associated mitochondrial dysfunction, however, reduces efficacy of these strategies. Therefore, we propose that new classes of mitochondria-targeted antioxidants can provide high therapeutic potential to improve endothelial function and reduce hypertension. PMID:26717313

  7. Acetylation of Mitochondrial Trifunctional Protein α-Subunit Enhances Its Stability To Promote Fatty Acid Oxidation and Is Decreased in Nonalcoholic Fatty Liver Disease

    PubMed Central

    Guo, Liang; Zhou, Shui-Rong; Wei, Xiang-Bo; Liu, Yuan; Chang, Xin-Xia; Liu, Yang; Ge, Xin; Dou, Xin; Huang, Hai-Yan; Qian, Shu-Wen; Li, Xi; Lei, Qun-Ying

    2016-01-01

    Nonalcoholic fatty liver disease (NAFLD) has become the most common liver disease, and decreased fatty acid oxidation is one of the important contributors to NAFLD. Mitochondrial trifunctional protein α-subunit (MTPα) functions as a critical enzyme for fatty acid β-oxidation, but whether dysregulation of MTPα is pathogenically connected to NAFLD is poorly understood. We show that MTPα is acetylated at lysine residues 350, 383, and 406 (MTPα-3K), which promotes its protein stability by antagonizing its ubiquitylation on the same three lysines (MTPα-3K) and blocking its subsequent degradation. Sirtuin 4 (SIRT4) has been identified as the deacetylase, deacetylating and destabilizing MTPα. Replacement of MTPα-3K with either MTPα-3KR or MTPα-3KQ inhibits cellular lipid accumulation both in free fatty acid (FFA)-treated alpha mouse liver 12 (AML12) cells and primary hepatocytes and in the livers of high-fat/high-sucrose (HF/HS) diet-fed mice. Moreover, knockdown of SIRT4 could phenocopy the effects of MTPα-3K mutant expression in mouse livers, and MTPα-3K mutants more efficiently attenuate SIRT4-mediated hepatic steatosis in HF/HS diet-fed mice. Importantly, acetylation of both MTPα and MTPα-3K is decreased while SIRT4 is increased in the livers of mice and humans with NAFLD. Our study reveals a novel mechanism of MTPα regulation by acetylation and ubiquitylation and a direct functional link of this regulation to NAFLD. PMID:27457618

  8. Impaired mitochondrial fat oxidation induces adaptive remodeling of muscle metabolism

    PubMed Central

    Wicks, Shawna E.; Vandanmagsar, Bolormaa; Haynie, Kimberly R.; Fuller, Scott E.; Warfel, Jaycob D.; Stephens, Jacqueline M.; Wang, Miao; Han, Xianlin; Zhang, Jingying; Noland, Robert C.; Mynatt, Randall L.

    2015-01-01

    The correlations between intramyocellular lipid (IMCL), decreased fatty acid oxidation (FAO), and insulin resistance have led to the hypothesis that impaired FAO causes accumulation of lipotoxic intermediates that inhibit muscle insulin signaling. Using a skeletal muscle-specific carnitine palmitoyltransferase-1 KO model, we show that prolonged and severe mitochondrial FAO inhibition results in increased carbohydrate utilization, along with reduced physical activity; increased circulating nonesterified fatty acids; and increased IMCLs, diacylglycerols, and ceramides. Perhaps more importantly, inhibition of mitochondrial FAO also initiates a local, adaptive response in muscle that invokes mitochondrial biogenesis, compensatory peroxisomal fat oxidation, and amino acid catabolism. Loss of its major fuel source (lipid) induces an energy deprivation response in muscle coordinated by signaling through AMP-activated protein kinase (AMPK) and peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC1α) to maintain energy supply for locomotion and survival. At the whole-body level, these adaptations result in resistance to obesity. PMID:26056297

  9. Impaired mitochondrial fat oxidation induces adaptive remodeling of muscle metabolism.

    PubMed

    Wicks, Shawna E; Vandanmagsar, Bolormaa; Haynie, Kimberly R; Fuller, Scott E; Warfel, Jaycob D; Stephens, Jacqueline M; Wang, Miao; Han, Xianlin; Zhang, Jingying; Noland, Robert C; Mynatt, Randall L

    2015-06-23

    The correlations between intramyocellular lipid (IMCL), decreased fatty acid oxidation (FAO), and insulin resistance have led to the hypothesis that impaired FAO causes accumulation of lipotoxic intermediates that inhibit muscle insulin signaling. Using a skeletal muscle-specific carnitine palmitoyltransferase-1 KO model, we show that prolonged and severe mitochondrial FAO inhibition results in increased carbohydrate utilization, along with reduced physical activity; increased circulating nonesterified fatty acids; and increased IMCLs, diacylglycerols, and ceramides. Perhaps more importantly, inhibition of mitochondrial FAO also initiates a local, adaptive response in muscle that invokes mitochondrial biogenesis, compensatory peroxisomal fat oxidation, and amino acid catabolism. Loss of its major fuel source (lipid) induces an energy deprivation response in muscle coordinated by signaling through AMP-activated protein kinase (AMPK) and peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC1α) to maintain energy supply for locomotion and survival. At the whole-body level, these adaptations result in resistance to obesity.

  10. In vivo stable isotope studies in three patients affected with mitochondrial fatty acid oxidation disorders: limited diagnostic use of 1-13C fatty acid breath test using bolus technique.

    PubMed

    Jakobs, C; Kneer, J; Martin, D; Boulloche, J; Brivet, M; Poll-The, B T; Saudubray, J M

    1997-08-01

    The in vivo oxidation of fatty acids (FA) of different chain length was investigated in three patients with documented mitochondrial FA oxidation disorders: one patient with mild multiple acyl-CoA dehydrogenase deficiency (MADM), one with medium chain acyl-CoA dehydrogenase deficiency (MCAD), and one with carnitine palmitoyltransferase I deficiency (CPT I). Breath tests were performed after oral administration of 1-13C butyric. 1-13C octanoic, and 1-13C palmitic acids. 13C/12C ratio in the expired oxidative end product CO2 was measured. The cumulative 13C elimination was calculated and expressed as a percentage of the administered dose. In the MADM patient the influence of carnitine therapy (or deprivation) on the utilization of 1-13C palmitic acid was also examined. In the MCAD and CPT I patients, the 1-13C butyric, 1-13C octanoic and 1-13C palmitic acids in vivo oxidation were similar to five healthy controls. In the MADM patient, the oxidation of 1-13C butyric and 1-13C octanoic acids were normal, whereas the metabolism of 1-13C palmitic acid ranged from 33% of 66% of controls. In this patient the serum carnitine level decreased from 60 to 27 mumol/l without carnitine supplementation. Clinically there was mild hypotonia. 1-13C palmitic acid oxidation compared to controls was 50%. After 2 further weeks of carnitine deprivation the serum carnitine was 10-15 mumol/l. Clinically he was very hypotonic and had a large liver. 1-13C Palmitic acid oxidation was 33%. After 6 weeks of readministration of carnitine (L-carnitine 100 mg/kg/day p.o.) the serum carnitine was 60 mumol/l and the patient was in good clinical condition. 1-13C palmitic acid oxidation was 66% compared to controls. Our study implies that this simple fatty acid breath test is not of diagnostic use for detection of enzymatic defects in FA oxidation disorders. The carnitine dependent 1-13C palmitic acid oxidation indicates that this test might be of some value in cases with primary or secondary carnitine

  11. Fatty Acid Oxidation Disorders

    MedlinePlus

    ... other health conditions > Fatty acid oxidation disorders Fatty acid oxidation disorders E-mail to a friend Please ... these disorders, go to genetests.org . What fatty acid oxidation disorders are tested for in newborn screening? ...

  12. Piracetam improves mitochondrial dysfunction following oxidative stress.

    PubMed

    Keil, Uta; Scherping, Isabel; Hauptmann, Susanne; Schuessel, Katin; Eckert, Anne; Müller, Walter E

    2006-01-01

    1.--Mitochondrial dysfunction including decrease of mitochondrial membrane potential and reduced ATP production represents a common final pathway of many conditions associated with oxidative stress, for example, hypoxia, hypoglycemia, and aging. 2.--Since the cognition-improving effects of the standard nootropic piracetam are usually more pronounced under such pathological conditions and young healthy animals usually benefit little by piracetam, the effect of piracetam on mitochondrial dysfunction following oxidative stress was investigated using PC12 cells and dissociated brain cells of animals treated with piracetam. 3.--Piracetam treatment at concentrations between 100 and 1000 microM improved mitochondrial membrane potential and ATP production of PC12 cells following oxidative stress induced by sodium nitroprusside (SNP) and serum deprivation. Under conditions of mild serum deprivation, piracetam (500 microM) induced a nearly complete recovery of mitochondrial membrane potential and ATP levels. Piracetam also reduced caspase 9 activity after SNP treatment. 4.--Piracetam treatment (100-500 mg kg(-1) daily) of mice was also associated with improved mitochondrial function in dissociated brain cells. Significant improvement was mainly seen in aged animals and only less in young animals. Moreover, the same treatment reduced antioxidant enzyme activities (superoxide dismutase, glutathione peroxidase, and glutathione reductase) in aged mouse brain only, which are elevated as an adaptive response to the increased oxidative stress with aging. 5.--In conclusion, therapeutically relevant in vitro and in vivo concentrations of piracetam are able to improve mitochondrial dysfunction associated with oxidative stress and/or aging. Mitochondrial stabilization and protection might be an important mechanism to explain many of piracetam's beneficial effects in elderly patients.

  13. Mitochondrial Oxidative Damage in Aging and Alzheimer's Disease: Implications for Mitochondrially Targeted Antioxidant Therapeutics

    PubMed Central

    Reddy, P. Hemachandra

    2006-01-01

    The overall aim of this article is to review current therapeutic strategies for treating AD, with a focus on mitochondrially targeted antioxidant treatments. Recent advances in molecular, cellular, and animal model studies of AD have revealed that amyloid precursor protein derivatives, including amyloid beta (Aβ) monomers and oligomers, are likely key factors in tau hyperphosphorylation, mitochondrial oxidative damage, inflammatory changes, and synaptic failure in the brain tissue of AD patients. Several therapeutic strategies have been developed to treat AD, including anti-inflammatory, antioxidant, and antiamyloid approaches. Among these, mitochondrial antioxidant therapy has been found to be the most efficacious in reducing pathological changes and in not producing adverse effects; thus, mitochondrial antioxidant therapy is promising as a treatment for AD patients. However, a major limitation in applying mitochondrial antioxidants to AD treatment has been the inability of researchers to enhance antioxidant levels in mitochondria. Recently, however, there has been a breakthrough. Researchers have recently been able to promote the entry of certain antioxidants—including MitoQ, MitoVitE, MitoPBN, MitoPeroxidase, and amino acid and peptide-based SS tetrapeptides—into mitochondria, several hundred-fold more than do natural antioxidants. Once in the mitochondria, they rapidly neutralize free radicals and decrease mitochondrial toxicity. Thus, mitochondrially targeted antioxidants are promising candidates for treating AD patients. PMID:17047303

  14. Mitochondrial accumulation under oxidative stress is due to defects in autophagy.

    PubMed

    Luo, Cheng; Li, Yan; Wang, Hui; Feng, Zhihui; Li, Yuan; Long, Jiangang; Liu, Jiankang

    2013-01-01

    Mitochondrial dynamics maintains normal mitochondrial function by degrading damaged mitochondria and generating newborn mitochondria. The accumulation of damaged mitochondria influences the intracellular environment by promoting mitochondrial dysfunction, and thus initiating a vicious cycle. Oxidative stress induces mitochondrial malfunction, which is involved in many cardiovascular diseases. However, the mechanism of mitochondrial accumulation in cardiac myoblasts remains unclear. We observed mitochondrial dysfunction and an increase in mitochondrial mass under the oxidative conditions produced by tert-butyl hydroperoxide (tBHP) in cardiac myoblast H9c2 cells. However, in contrast to the increase in mitochondrial mass, mitochondrial DNA (mtDNA) decreased, suggesting that enhanced mitochondrial biogenesis may be not the primary cause of the mitochondrial accumulation. Therefore, we investigated changes in a number of proteins involved in autophagy. Beclin1, Atg12-Atg5 conjugate, Atg7 contents decreased but LC3-II accumulated in tBHP-treated H9c2 cells. Moreover, the capacity for acid hydrolysis decreased in H9c2 cells. We also demonstrated a decrease in DJ-1 protein under the oxidative conditions that deregulate mitochondrial dynamics. These results reveal that autophagy became defective under oxidative stress. We therefore suggest that defects in autophagy mediate mitochondrial accumulation under these conditions.

  15. Ageing, oxidative stress, and mitochondrial uncoupling.

    PubMed

    Harper, M-E; Bevilacqua, L; Hagopian, K; Weindruch, R; Ramsey, J J

    2004-12-01

    Mitochondria are a cell's single greatest source of reactive oxygen species. Reactive oxygen species are important for many life sustaining processes of cells and tissues, but they can also induce cell damage and death. If their production and levels within cells is not effectively controlled, then the detrimental effects of oxidative stress can accumulate. Oxidative stress is widely thought to underpin many ageing processes, and the oxidative stress theory of ageing is one of the most widely acknowledged theories of ageing. As well as being the major source of reactive oxygen species, mitochondria are also a major site of oxidative damage. The purpose of this review is a concise and current review of the effects of oxidative stress and ageing on mitochondrial function. Emphasis is placed upon the roles of mitochondrial proton leak, the uncoupling proteins, and the anti-ageing effects of caloric restriction.

  16. Proteomic and Biochemical Studies of Lysine Malonylation Suggest Its Malonic Aciduria-associated Regulatory Role in Mitochondrial Function and Fatty Acid Oxidation*

    PubMed Central

    Colak, Gozde; Pougovkina, Olga; Dai, Lunzhi; Tan, Minjia; te Brinke, Heleen; Huang, He; Cheng, Zhongyi; Park, Jeongsoon; Wan, Xuelian; Liu, Xiaojing; Yue, Wyatt W.; Wanders, Ronald J. A.; Locasale, Jason W.; Lombard, David B.; de Boer, Vincent C. J.; Zhao, Yingming

    2015-01-01

    The protein substrates of sirtuin 5-regulated lysine malonylation (Kmal) remain unknown, hindering its functional analysis. In this study, we carried out proteomic screening, which identified 4042 Kmal sites on 1426 proteins in mouse liver and 4943 Kmal sites on 1822 proteins in human fibroblasts. Increased malonyl-CoA levels in malonyl-CoA decarboxylase (MCD)-deficient cells induces Kmal levels in substrate proteins. We identified 461 Kmal sites showing more than a 2-fold increase in response to MCD deficiency as well as 1452 Kmal sites detected only in MCD−/− fibroblast but not MCD+/+ cells, suggesting a pathogenic role of Kmal in MCD deficiency. Cells with increased lysine malonylation displayed impaired mitochondrial function and fatty acid oxidation, suggesting that lysine malonylation plays a role in pathophysiology of malonic aciduria. Our study establishes an association between Kmal and a genetic disease and offers a rich resource for elucidating the contribution of the Kmal pathway and malonyl-CoA to cellular physiology and human diseases. PMID:26320211

  17. Understanding and preventing mitochondrial oxidative damage

    PubMed Central

    Murphy, Michael P.

    2016-01-01

    Mitochondrial oxidative damage has long been known to contribute to damage in conditions such as ischaemia–reperfusion (IR) injury in heart attack. Over the past years, we have developed a series of mitochondria-targeted compounds designed to ameliorate or determine how this damage occurs. I will outline some of this work, from MitoQ to the mitochondria-targeted S-nitrosating agent, called MitoSNO, that we showed was effective in preventing reactive oxygen species (ROS) formation in IR injury with therapeutic implications. In addition, the protection by this compound suggested that ROS production in IR injury was mainly coming from complex I. This led us to investigate the mechanism of the ROS production and using a metabolomic approach, we found that the ROS production in IR injury came from the accumulation of succinate during ischaemia that then drove mitochondrial ROS production by reverse electron transport at complex I during reperfusion. This surprising mechanism led us to develop further new therapeutic approaches to have an impact on the damage that mitochondrial ROS do in pathology and also to explore how mitochondrial ROS can act as redox signals. I will discuss how these approaches have led to a better understanding of mitochondrial oxidative damage in pathology and also to the development of new therapeutic strategies. PMID:27911703

  18. Sulforaphane prevents quinolinic acid-induced mitochondrial dysfunction in rat striatum.

    PubMed

    Luis-García, Erika Rubí; Limón-Pacheco, Jorge Humberto; Serrano-García, Norma; Hernández-Pérez, Alma Delia; Pedraza-Chaverri, José; Orozco-Ibarra, Marisol

    2017-02-01

    Quinolinic acid (QA) triggers striatal neuronal death by an excitotoxic cascade that involves oxidative stress, which in turns is tightly linked to mitochondria. Mitochondrial dysfunction is a molecular feature described in several brain pathologies. In this work, we determined whether the sulforaphane-neuroprotective effect in the rodent experimental model of Huntington's disease induced by QA is associated with mitochondrial function preservation. We found that QA impaired mitochondrial function within 24 h post-lesion. Sulforaphane effectively disrupted the mitochondrial dysfunction by preventing the decrease in respiratory control ratio, transmembrane potential, ability to synthetize ATP, and the activity of mitochondrial complexes I, II, and IV.

  19. S-Allylcysteine prevents the rat from 3-nitropropionic acid-induced hyperactivity, early markers of oxidative stress and mitochondrial dysfunction.

    PubMed

    Herrera-Mundo, María N; Silva-Adaya, Daniela; Maldonado, Perla D; Galván-Arzate, Sonia; Andrés-Martínez, Leticia; Pérez-De La Cruz, Verónica; Pedraza-Chaverrí, José; Santamaría, Abel

    2006-09-01

    We investigated the effects of S-allylcysteine (SAC) on early behavioral alterations, striatal changes in superoxide dismutase (SOD) activity, lipid peroxidation (LP) and mitochondrial dysfunction induced by the systemic infusion of 3-nitropropionic acid (3-NPA) to rats. SAC (300 mg/kg, i.p.), given to animals 30 min before 3-NPA (30 mg/kg, i.p.), prevented the hyperkinetic pattern evoked by the toxin. In addition, 3-NPA alone produced decreased activities of manganese- (Mn-SOD) and copper/zinc-dependent superoxide dismutase (Cu,Zn-SOD), increased LP (evaluated as the formation of lipid fluorescent products) and produced mitochondrial dysfunction in the striatum (measured as decreased 3-(3,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide reduction). In contrast, pretreatment of 3-NPA-injected rats with SAC resulted in a significant prevention of all these markers. Our findings suggest that the protective actions of SAC are related with its antioxidant properties, which in turn may be accounting for the preservation of SOD activity and primary mitochondrial tasks.

  20. Oxidative stress and mitochondrial dysfunction in sepsis.

    PubMed

    Galley, H F

    2011-07-01

    Sepsis-related organ dysfunction remains the most common cause of death in the intensive care unit (ICU), despite advances in healthcare and science. Marked oxidative stress as a result of the inflammatory responses inherent with sepsis initiates changes in mitochondrial function which may result in organ damage. Normally, a complex system of interacting antioxidant defences is able to combat oxidative stress and prevents damage to mitochondria. Despite the accepted role that oxidative stress-mediated injury plays in the development of organ failure, there is still little conclusive evidence of any beneficial effect of systemic antioxidant supplementation in patients with sepsis and organ dysfunction. It has been suggested, however, that antioxidant therapy delivered specifically to mitochondria may be useful.

  1. Fatty Acid Beta-Oxidation Disorders: A Brief Review

    PubMed Central

    Vishwanath, Vijay A.

    2016-01-01

    Background Mitochondrial fatty acid β-oxidation disorders (FAODs) are a heterogeneous group of defects in fatty acid transport and mitochondrial β-oxidation. They are inherited as autosomal recessive disorders and have a wide range of clinical presentations. Summary The background information and case report provide important insight into mitochondrial FAODs. The article provides a wealth of information describing the scope of these disorders. Key Messages This article presents a typical case of medium chain acyl-CoA dehydrogenase deficiency and summarizes the pathophysiology, clinical presentation, diagnosis and treatment of mitochondrial FAODs. PMID:27536022

  2. Age-associated mitochondrial oxidative decay: Improvement of carnitine acetyltransferase substrate-binding affinity and activity in brain by feeding old rats acetyl-l- carnitine and/or R-α-lipoic acid

    PubMed Central

    Liu, Jiankang; Killilea, David W.; Ames, Bruce N.

    2002-01-01

    We test whether the dysfunction with age of carnitine acetyltransferase (CAT), a key mitochondrial enzyme for fuel utilization, is due to decreased binding affinity for substrate and whether this substrate, fed to old rats, restores CAT activity. The kinetics of CAT were analyzed by using the brains of young and old rats and of old rats supplemented for 7 weeks with the CAT substrate acetyl-l-carnitine (ALCAR) and/or the mitochondrial antioxidant precursor R-α-lipoic acid (LA). Old rats, compared with young rats, showed a decrease in CAT activity and in CAT-binding affinity for both substrates, ALCAR and CoA. Feeding ALCAR or ALCAR plus LA to old rats significantly restored CAT-binding affinity for ALCAR and CoA, and CAT activity. To explore the underlying mechanism, lipid peroxidation and total iron and copper levels were assayed; all increased in old rats. Feeding old rats LA or LA plus ALCAR inhibited lipid peroxidation but did not decrease iron and copper levels. Ex vivo oxidation of young-rat brain with Fe(II) caused loss of CAT activity and binding affinity. In vitro oxidation of purified CAT with Fe(II) inactivated the enzyme but did not alter binding affinity. However, in vitro treatment of CAT with the lipid peroxidation products malondialdehyde or 4-hydroxy-nonenal caused a decrease in CAT-binding affinity and activity, thus mimicking age-related change. Preincubation of CAT with ALCAR or CoA prevented malondialdehyde-induced dysfunction. Thus, feeding old rats high levels of key mitochondrial metabolites can ameliorate oxidative damage, enzyme activity, substrate-binding affinity, and mitochondrial dysfunction. PMID:11854488

  3. Intervention of mitochondrial dysfunction-oxidative stress-dependent apoptosis as a possible neuroprotective mechanism of α-lipoic acid against rotenone-induced parkinsonism and L-dopa toxicity.

    PubMed

    Abdin, Amany A; Sarhan, Naglaa I

    2011-12-01

    The current study evidenced hypothesis that mitochondrial dysfunction-oxidative stress-dependent apoptotic pathways play a critical role in degeneration of dopaminergic neurons in Parkinson's disease. Model of rotenone-induced parkinsonism in rats produced decrease in striatal complex I activity and reduced glutathione with increase in nitrites concentration and caspase-3 activity. This was confirmed by significant correlation of catalepsy score with neurochemical parameters. Moreover, electron microscopic examination of striatal neurons displayed ultrastructure affection as hyperchromatic nuclei and disrupted mitochondria that are typical features of undergoing apoptosis. Administration of L-dopa as replacement therapy, although caused symptomatic improvement in catalepsy score, but further worsening in neurochemical parameters. Therefore, efforts are not only to improve effect of L-dopa, but also to introduce drugs provide antiparkinsonian and neuroprotective effects. In this study, α-lipoic acid exhibited noticeable neuroprotective effects by a mechanism via intervention of mitochondrial dysfunction-oxidative stress-dependent apoptotic pathways. Combination of α-lipoic acid efficiently halting deleterious toxic effects of L-dopa, revealed normalization of catalepsy score in addition to amelioration of neurochemical parameters and apparent preservation of striatal ultrastructure integrity, indicating benefit of both symptomatic and neuroprotective therapy. In conclusion, α-lipoic acid could be recommended as a disease-modifying therapy when given with L-dopa early in course of Parkinson's disease.

  4. CONTROL OF GLUTAMATE OXIDATION IN BRAIN AND LIVER MITOCHONDRIAL SYSTEMS.

    PubMed

    BALAZS, R

    1965-05-01

    1. Glutamate oxidation in brain and liver mitochondrial systems proceeds mainly through transamination with oxaloacetate followed by oxidation of the alpha-oxoglutarate formed. Both in the presence and absence of dinitrophenol in liver mitochondria this pathway accounted for almost 80% of the uptake of glutamate. In brain preparations the transamination pathway accounted for about 90% of the glutamate uptake. 2. The oxidation of [1-(14)C]- and [5-(14)C]-glutamate in brain preparations is compatible with utilization through the tricarboxylic acid cycle, either after the formation of alpha-oxoglutarate or after decarboxylation to form gamma-aminobutyrate. There is no indication of gamma-decarboxylation of glutamate. 3. The high respiratory control ratio obtained with glutamate as substrate in brain mitochondrial preparations is due to the low respiration rate in the absence of ADP: this results from the low rate of formation of oxaloacetate under these conditions. When oxaloacetate is made available by the addition of malate or of NAD(+), the respiration rate is increased to the level obtained with other substrates. 4. When the transamination pathway of glutamate oxidation was blocked with malonate, the uptake of glutamate was inhibited in the presence of ADP or ADP plus dinitrophenol by about 70 and 80% respectively in brain mitochondrial systems, whereas the inhibition was only about 50% in dinitrophenol-stimulated liver preparations. In unstimulated liver mitochondria in the presence of malonate there was a sixfold increase in the oxidation of glutamate by the glutamate-dehydrogenase pathway. Thus the operating activity of glutamate dehydrogenase is much less than the ;free' (non-latent) activity. 5. The following explanation is put forward for the control of glutamate metabolism in liver and brain mitochondrial preparations. The oxidation of glutamate by either pathway yields alpha-oxoglutarate, which is further metabolized. Since aspartate aminotransferase is

  5. Control of bovine hepatic fatty acid oxidation

    SciTech Connect

    Jesse, B.W.; Emery, R.S.; Thomas, J.W.

    1986-09-01

    Fatty acid oxidation by bovine liver slices and mitochondria was examined to determine potential regulatory sites of fatty acid oxidation. Conversion of 1-(/sup 14/C)palmitate to /sup 14/CO/sub 2/ and total (/sup 14/C)acid-soluble metabolites was used to measure fatty acid oxidation. Oxidation of palmitate (1 mM) was linear in both liver slice weight and incubation time. Carnitine stimulated palmitate oxidation; 2 mM dl-carnitine produced maximal stimulation of palmitate oxidation to both CO/sup 2/ and acid-soluble metabolites. Propionate (10 mM) inhibited palmitate oxidation by bovine liver slices. Propionate (.5 to 10 mM) had no effect on palmitate oxidation by mitochondria, but malonyl Coenzyme A, the first committed intermediate of fatty acid synthesis, inhibited mitochondrial palmitate oxidation (inhibition constant = .3 ..mu..M). Liver mitochonndrial carnitine palmitoyltransferase exhibited Michaelis constants for palmitoyl Coenzyme A and l-carnitine of 11.5 ..mu..M and .59 mM, respectively. Long-chain fatty acid oxidation in bovine liver is regulated by mechanisms similar to those in rats but adapted to the unique digestive physiology of the bovine.

  6. Ursolic acid mediates photosensitization by initiating mitochondrial-dependent apoptosis

    NASA Astrophysics Data System (ADS)

    Lee, Yuan-Hao; Wang, Exing; Kumar, Neeru; Glickman, Randolph D.

    2013-02-01

    The signaling pathways PI3K/Akt and MAPK play key roles in transcription, translation and carcinogenesis, and may be activated by light exposure. These pathways may be modulated or inhibited by naturally-occurring compounds, such as the triterpenoid, ursolic acid (UA). Previously, the transcription factors p53 and NF-kB, which transactivate mitochondrial apoptosis-related genes, were shown to be differentially modulated by UA. Our current work indicates that UA causes these effects via the mTOR and insulin-mediated pathways. UA-modulated apoptosis, following exposure to UV radiation, is observed to correspond to differential levels of oxidative stress in retinal pigment epithelial (RPE) and skin melanoma (SM) cells. Flow cytometry analysis, DHE (dihydroethidium) staining and membrane permeability assay showed that UA pretreatment potentiated cell cycle arrest and radiation-induced apoptosis selectively on SM cells while DNA photo-oxidative damage (i.e. strand breakage) was reduced, presumably by some antioxidant activity of UA in RPE cells. The UA-mediated NF-κB activation in SM cells was reduced by rapamycin pretreatment, which indicates that these agents exert inter-antagonistic effects in the PI3K/Akt/mTOR pathway. In contrast, the antagonistic effect of UA on the PI3K/Akt pathway was reversed by insulin leading to greater NF-κB and p53 activation in RPE cells. MitoTracker, a mitochondrial functional assay, indicated that mitochondria in RPE cells experienced reduced oxidative stress while those in SM cells exhibited increased oxidative stress upon UA pretreatment. When rapamycin administration was followed by UA, mitochondrial oxidative stress was increased in RPE cells but decreased in SM cells. These results indicate that UA modulates p53 and NF-κB, initiating a mitogenic response to radiation that triggers mitochondria-dependent apoptosis.

  7. Alpha-lipoic acid prevents mitochondrial damage and neurotoxicity in experimental chemotherapy neuropathy.

    PubMed

    Melli, Giorgia; Taiana, Michela; Camozzi, Francesca; Triolo, Daniela; Podini, Paola; Quattrini, Angelo; Taroni, Franco; Lauria, Giuseppe

    2008-12-01

    The study investigates if alpha-lipoic acid is neuroprotective against chemotherapy induced neurotoxicity, if mitochondrial damage plays a critical role in toxic neurodegenerative cascade, and if neuroprotective effects of alpha-lipoic acid depend on mitochondria protection. We used an in vitro model of chemotherapy induced peripheral neuropathy that closely mimic the in vivo condition by exposing primary cultures of dorsal root ganglion (DRG) sensory neurons to paclitaxel and cisplatin, two widely used and highly effective chemotherapeutic drugs. This approach allowed investigating the efficacy of alpha-lipoic acid in preventing axonal damage and apoptosis and the function and ultrastructural morphology of mitochondria after exposure to toxic agents and alpha-lipoic acid. Our results demonstrate that both cisplatin and paclitaxel cause early mitochondrial impairment with loss of membrane potential and induction of autophagic vacuoles in neurons. Alpha-lipoic acid exerts neuroprotective effects against chemotherapy induced neurotoxicity in sensory neurons: it rescues the mitochondrial toxicity and induces the expression of frataxin, an essential mitochondrial protein with anti-oxidant and chaperone properties. In conclusion mitochondrial toxicity is an early common event both in paclitaxel and cisplatin induced neurotoxicity. Alpha-lipoic acid protects sensory neurons through its anti-oxidant and mitochondrial regulatory functions, possibly inducing the expression of frataxin. These findings suggest that alpha-lipoic acid might reduce the risk of developing peripheral nerve toxicity in patients undergoing chemotherapy and encourage further confirmatory clinical trials.

  8. A coulombic hypothesis of mitochondrial oxidative phosphorylation.

    PubMed

    Malpress, F H

    1984-08-21

    A coulombic hypothesis of mitochondrial oxidative phosphorylation is presented, founded upon the evidence for negative fixed charge formation during electron transport chain activity. The intermediary force is electrostatic (psi H) and not electrochemical (delta mu H). The electrochemical potential of the chemiosmotic hypothesis is identified as a "phantom" parameter which owes its delusive existence to the procedures by which it is measured. The connection between psi H and the conditional delta mu H values is examined; it entails the use of a variable conversion factor, f, where delta mu H (mV) = f psi H, and the concept of the "protonic status" of the diffuse double layer. A number of problems which beset the chemiosmotic view are reappraised in the light of the new interpretation, and find authentic solutions.

  9. Expression of Mitochondrial Cytochrome C Oxidase Chaperone Gene (COX20) Improves Tolerance to Weak Acid and Oxidative Stress during Yeast Fermentation

    PubMed Central

    Kumar, Vinod; Hart, Andrew J.; Keerthiraju, Ethiraju R.; Waldron, Paul R.; Tucker, Gregory A.; Greetham, Darren

    2015-01-01

    Introduction Saccharomyces cerevisiae is the micro-organism of choice for the conversion of fermentable sugars released by the pre-treatment of lignocellulosic material into bioethanol. Pre-treatment of lignocellulosic material releases acetic acid and previous work identified a cytochrome oxidase chaperone gene (COX20) which was significantly up-regulated in yeast cells in the presence of acetic acid. Results A Δcox20 strain was sensitive to the presence of acetic acid compared with the background strain. Overexpressing COX20 using a tetracycline-regulatable expression vector system in a Δcox20 strain, resulted in tolerance to the presence of acetic acid and tolerance could be ablated with addition of tetracycline. Assays also revealed that overexpression improved tolerance to the presence of hydrogen peroxide-induced oxidative stress. Conclusion This is a study which has utilised tetracycline-regulated protein expression in a fermentation system, which was characterised by improved (or enhanced) tolerance to acetic acid and oxidative stress. PMID:26427054

  10. Salicylic acid binding of mitochondrial alpha-ketoglutarate dehydrogenase E2 affects mitochondrial oxidative phosphorylation and electron transport chain components and plays a role in basal defense against tobacco mosaic virus in tomato.

    PubMed

    Liao, Yangwenke; Tian, Miaoying; Zhang, Huan; Li, Xin; Wang, Yu; Xia, Xiaojian; Zhou, Jie; Zhou, Yanhong; Yu, Jingquan; Shi, Kai; Klessig, Daniel F

    2015-02-01

    Salicylic acid (SA) plays a critical role in plant defense against pathogen invasion. SA-induced viral defense in plants is distinct from the pathways mediating bacterial and fungal defense and involves a specific pathway mediated by mitochondria; however, the underlying mechanisms remain largely unknown. The SA-binding activity of the recombinant tomato (Solanum lycopersicum) alpha-ketoglutarate dehydrogenase (Slα-kGDH) E2 subunit of the tricarboxylic acid (TCA) cycle was characterized. The biological role of this binding in plant defenses against tobacco mosaic virus (TMV) was further investigated via Slα-kGDH E2 silencing and transient overexpression in plants. Slα-kGDH E2 was found to bind SA in two independent assays. SA treatment, as well as Slα-kGDH E2 silencing, increased resistance to TMV. SA did not further enhance TMV defense in Slα-kGDH E2-silenced tomato plants but did reduce TMV susceptibility in Nicotiana benthamiana plants transiently overexpressing Slα-kGDH E2. Furthermore, Slα-kGDH E2-silencing-induced TMV resistance was fully blocked by bongkrekic acid application and alternative oxidase 1a silencing. These results indicated that binding by Slα-kGDH E2 of SA acts upstream of and affects the mitochondrial electron transport chain, which plays an important role in basal defense against TMV. The findings of this study help to elucidate the mechanisms of SA-induced viral defense.

  11. Mechanism study on mitochondrial fragmentation under oxidative stress caused by high-fluence low-power laser irradiation

    NASA Astrophysics Data System (ADS)

    Wu, Shengnan; Zhou, Feifan; Xing, Da

    2012-03-01

    Mitochondria are dynamic organelles that undergo continual fusion and fission to maintain their morphology and functions, but the mechanism involved is still not clear. Here, we investigated the effect of mitochondrial oxidative stress triggered by high-fluence low-power laser irradiation (HF-LPLI) on mitochondrial dynamics in human lung adenocarcinoma cells (ASTC-a-1). Upon HF-LPLI-triggered oxidative stress, mitochondria displayed a fragmented structure, which was abolished by exposure to dehydroascorbic acid (DHA), a reactive oxygen species scavenger, indicating that oxidative stress can induce mitochondrial fragmentation. Mitochondrial translocation of the profission protein dynamin-related protein 1 (Drp1) was observed following HF-LPLI, demonstrating apoptosis-related activation of Drp1. Notably, DHA pre-treatment prevented HF-LPLI-induced Drp1 activation. We conclude that mitochondrial oxidative stress through activation of Drp1 causes mitochondrial fragmentation.

  12. Melatonin protects against common deletion of mitochondrial DNA-augmented mitochondrial oxidative stress and apoptosis.

    PubMed

    Jou, Mei-Jie; Peng, Tsung-I; Yu, Pai-Zu; Jou, Shuo-Bin; Reiter, Russel J; Chen, Jin-Yi; Wu, Hong-Yueh; Chen, Chih-Chun; Hsu, Lee-Fen

    2007-11-01

    Defected mitochondrial respiratory chain (RC), in addition to causing a severe ATP deficiency, often augments reactive oxygen species (ROS) generation in mitochondria (mROS) which enhances pathological conditions and diseases. Previously, we demonstrated a potent endogenously RC defect-augmented mROS associated dose-dependently with a commonly seen large-scale deletion of 4977 base pairs of mitochondrial DNA (mtDNA), i.e. the common deletion (CD). As current treatments for CD-associated diseases are rather supplementary and ineffective, we investigated whether melatonin, a potential mitochondrial protector, provides beneficial protection for CD-augmented mitochondrial oxidative stress and apoptosis particularly upon the induction of a secondary oxidative stress. Detailed mechanistic investigations were performed by using laser scanning dual fluorescence imaging microscopy to provide precise spatial and temporal resolution of mitochondrial events at single cell level. We demonstrate, for the first time, that melatonin significantly prevents CD-augmented mROS formation under basal conditions as well as at early time-points upon secondary oxidative stress induced by H2O2 exposure. Thus, melatonin prevents mROS-mediated depolarization of mitochondrial membrane potential (DeltaPsim) and subsequent opening of the mitochondrial permeability transition pore (MPTP) and cytochrome c release. Moreover, melatonin prevents depletion of cardiolipin which appears to be crucial for postponing later MPTP opening, disruption of the mitochondrial membrane and apoptosis. Finally, the protection provided by melatonin is superior to those caused by the suppression of mitochondrial Ca2+ regulators including the mitochondrial Na+-Ca2) exchanger, the MPTP, and the mitochondrial Ca2+ uniporter and by antioxidants including vitamin E and mitochondria-targeted coenzyme Q, MitoQ. As RC defect-augmented endogenous mitochondrial oxidative stress is centrally involved in a variety of pathological

  13. Impaired Mitochondrial Fat Oxidation Induces FGF21 in Muscle.

    PubMed

    Vandanmagsar, Bolormaa; Warfel, Jaycob D; Wicks, Shawna E; Ghosh, Sujoy; Salbaum, J Michael; Burk, David; Dubuisson, Olga S; Mendoza, Tamra M; Zhang, Jingying; Noland, Robert C; Mynatt, Randall L

    2016-05-24

    Fatty acids are the primary fuel source for skeletal muscle during most of our daily activities, and impaired fatty acid oxidation (FAO) is associated with insulin resistance. We have developed a mouse model of impaired FAO by deleting carnitine palmitoyltransferase-1b specifically in skeletal muscle (Cpt1b(m-/-)). Cpt1b(m-/-) mice have increased glucose utilization and are resistant to diet-induced obesity. Here, we show that inhibition of mitochondrial FAO induces FGF21 expression specifically in skeletal muscle. The induction of FGF21 in Cpt1b-deficient muscle is dependent on AMPK and Akt1 signaling but independent of the stress signaling pathways. FGF21 appears to act in a paracrine manner to increase glucose uptake under low insulin conditions, but it does not contribute to the resistance to diet-induced obesity.

  14. Impaired mitochondrial fat oxidation induces FGF21 in muscle

    PubMed Central

    Vandanmagsar, Bolormaa; Warfel, Jaycob D.; Wicks, Shawna E.; Ghosh, Sujoy; Salbaum, J. Michael; Burk, David; Dubuisson, Olga S.; Mendoza, Tamra M.; Zhang, Jingying; Noland, Robert C.; Mynatt, Randall L.

    2016-01-01

    SUMMARY Fatty acids are the primary fuel source for skeletal muscle during most of our daily activities and impaired fatty acid oxidation (FAO) is associated with insulin resistance. We have developed a mouse model of impaired FAO by deleting carnitine palmitoyltransferase-1b specifically in skeletal muscle (Cpt1bm−/−). Cpt1bm−/− mice have increased glucose utilization and are resistant to diet induced obesity. Here we show that inhibition of mitochondrial FAO induces FGF21 expression specifically in skeletal muscle. The induction of FGF21 in Cpt1b-deficient muscle is dependent on AMPK and Akt1 signaling but independent on the stress signaling pathways. FGF21 appears to act in a paracrine manner to increase glucose uptake under low insulin conditions, but does not contribute to the resistance to diet induced obesity. PMID:27184848

  15. Salvianolic acid B inhibits mitochondrial dysfunction by up-regulating mortalin

    PubMed Central

    Liu, Yunxia; Hu, Yingying; E, Qiukai; Zuo, Ji; Yang, Ling; Liu, Wen

    2017-01-01

    Salvianolic acid B is an antioxidative ingredient derived from Radix Salviae miltiorrhizae that has been widely used to treat liver diseases. However, the therapeutic mechanism underlying Salvianolic acid B has remained largely unknown. Our studies verified that Salvianolic acid B efficiently blocked mitochondrial deformation and dysfunction induced by H2O2 in the human hepatocyte cell line HL7702. Mortalin, a mitochondrial molecular chaperone, maintains mitochondrial morphology stabilization and function integrity. Previous results showed that mortalin overexpression has been observed in hematoma carcinoma cells and that mortalin maintains mitochondrial homeostasis and antagonizes oxidative stress damage. We found that Salvianolic acid B significantly up-regulated mortalin protein expression levels. In addition, Salvianolic acid B lost the function of preventing mitochondrial deformation and dysfunction induced by oxidative stress under mortalin knockdown conditions. We further found that mortalin overexpression increases the mRNA expression of mitofusin-related factor Mfn1 and mitofission-related factor hFis1. In conclusion, Salvianolic acid B maintains the mitochondrial structure stabilization and functional integrity by up-regulating mortalin, which may be associated with increased mitofusin factor Mfn1 and reduced mitofission factor hFis1. PMID:28251987

  16. Salvianolic acid B inhibits mitochondrial dysfunction by up-regulating mortalin.

    PubMed

    Liu, Yunxia; Hu, Yingying; E, Qiukai; Zuo, Ji; Yang, Ling; Liu, Wen

    2017-03-02

    Salvianolic acid B is an antioxidative ingredient derived from Radix Salviae miltiorrhizae that has been widely used to treat liver diseases. However, the therapeutic mechanism underlying Salvianolic acid B has remained largely unknown. Our studies verified that Salvianolic acid B efficiently blocked mitochondrial deformation and dysfunction induced by H2O2 in the human hepatocyte cell line HL7702. Mortalin, a mitochondrial molecular chaperone, maintains mitochondrial morphology stabilization and function integrity. Previous results showed that mortalin overexpression has been observed in hematoma carcinoma cells and that mortalin maintains mitochondrial homeostasis and antagonizes oxidative stress damage. We found that Salvianolic acid B significantly up-regulated mortalin protein expression levels. In addition, Salvianolic acid B lost the function of preventing mitochondrial deformation and dysfunction induced by oxidative stress under mortalin knockdown conditions. We further found that mortalin overexpression increases the mRNA expression of mitofusin-related factor Mfn1 and mitofission-related factor hFis1. In conclusion, Salvianolic acid B maintains the mitochondrial structure stabilization and functional integrity by up-regulating mortalin, which may be associated with increased mitofusin factor Mfn1 and reduced mitofission factor hFis1.

  17. The Mitochondrial-Derived Peptide Humanin Protects RPE Cells From Oxidative Stress, Senescence, and Mitochondrial Dysfunction

    PubMed Central

    Sreekumar, Parameswaran G.; Ishikawa, Keijiro; Spee, Chris; Mehta, Hemal H.; Wan, Junxiang; Yen, Kelvin; Cohen, Pinchas; Kannan, Ram; Hinton, David R.

    2016-01-01

    Purpose To investigate the expression of humanin (HN) in human retinal pigment epithelial (hRPE) cells and its effect on oxidative stress–induced cell death, mitochondrial bioenergetics, and senescence. Methods Humanin localization in RPE cells and polarized RPE monolayers was assessed by confocal microscopy. Human RPE cells were treated with 150 μM tert-Butyl hydroperoxide (tBH) in the absence/presence of HN (0.5–10 μg/mL) for 24 hours. Mitochondrial respiration was measured by XF96 analyzer. Retinal pigment epithelial cell death and caspase-3 activation, mitochondrial biogenesis and senescence were analyzed by TUNEL, immunoblot analysis, mitochondrial DNA copy number, SA-β-Gal staining, and p16INK4a expression and HN levels by ELISA. Oxidative stress–induced changes in transepithelial resistance were studied in RPE monolayers with and without HN cotreatment. Results A prominent localization of HN was found in the cytoplasmic and mitochondrial compartments of hRPE. Humanin cotreatment inhibited tBH-induced reactive oxygen species formation and significantly restored mitochondrial bioenergetics in hRPE cells. Exogenous HN was taken up by RPE and colocalized with mitochondria. The oxidative stress–induced decrease in mitochondrial bioenergetics was prevented by HN cotreatment. Humanin treatment increased mitochondrial DNA copy number and upregulated mitochondrial transcription factor A, a key biogenesis regulator protein. Humanin protected RPE cells from oxidative stress–induced cell death by STAT3 phosphorylation and inhibiting caspase-3 activation. Humanin treatment inhibited oxidant-induced senescence. Polarized RPE demonstrated elevated cellular HN and increased resistance to cell death. Conclusions Humanin protected RPE cells against oxidative stress–induced cell death and restored mitochondrial function. Our data suggest a potential role for HN therapy in the prevention of retinal degeneration, including AMD. PMID:26990160

  18. Mitochondrial respiratory dysfunction-elicited oxidative stress and posttranslational protein modification in mitochondrial diseases.

    PubMed

    Wu, Yu-Ting; Wu, Shi-Bei; Lee, Wan-Yu; Wei, Yau-Huei

    2010-07-01

    Pathogenic mutation in mtDNA and mitochondrial dysfunction are associated with mitochondrial diseases. In this review, we discuss the oxidative stress-elicited mitochondrial protein modifications that may contribute to the pathophysiology of mitochondrial diseases. We demonstrated that excess ROS produced by defective mitochondria could increase the acetylation of microtubule proteins through the suppression of Sirt2, which results in perinuclear distribution of mitochondria in skin fibroblasts of patients with CPEO syndrome. Our recent work showed that mitochondrial dysfunction-induced oxidative stress can disrupt protein degradation system by inhibiting the ubiquitin-proteasome pathway and protease activity in human cells harboring mutant mtDNA. This in turn causes accumulation of aberrant proteins in mitochondria and renders the mutant cells more susceptible to apoptosis induced by oxidative stress. Furthermore, oxidative stress can modulate phosphorylation of mitochondrial proteins, which can affect metabolism in a number of diseases. Taken together, we suggest that oxidative stress-triggered protein modifications and defects in protein turnover play an important role in the pathogenesis and progression of mitochondrial diseases.

  19. Organochloride pesticides impaired mitochondrial function in hepatocytes and aggravated disorders of fatty acid metabolism

    PubMed Central

    Liu, Qian; Wang, Qihan; Xu, Cheng; Shao, Wentao; Zhang, Chunlan; Liu, Hui; Jiang, Zhaoyan; Gu, Aihua

    2017-01-01

    p,p’-dichlorodiphenyldichloroethylene (p, p’-DDE) and β-hexachlorocyclohexane (β-HCH) were two predominant organochlorine pesticides (OCPs) metabolites in human body associated with disorders of fatty acid metabolism. However, the underlying mechanisms have not been fully clarified. In this study, adult male C57BL/6 mice were exposed to low dose of p, p’-DDE and β-HCH for 8 wk. OCPs accumulation in organs, hepatic fatty acid composition, tricarboxylic acid cycle (TCA) metabolites and other metabolite profiles were analyzed. Expression levels of genes involved in hepatic lipogenesis and β-oxidation were measured. Mitochondrial function was evaluated in HepG2 cells exposed to OCPs. High accumulation of p, p’-DDE and β-HCH was found in liver and damaged mitochondria was observed under electron microscopy. Expression of genes in fatty acid synthesis increased and that in mitochondrial fatty acid β-oxidation decreased in OCPs treatment groups. OCPs changed metabolite profiles in liver tissues, varied hepatic fatty acid compositions and levels of several TCA cycle metabolites. Furthermore, MitoTracker Green fluorescence, ATP levels, mitochondrial membrane potential and OCR decreased in HepG2 cells exposed to OCPs. In conclusion, chronic exposure to OCPs at doses equivalent to internal exposures in humans impaired mitochondrial function, decreased fatty acid β-oxidation and aggravated disorders of fatty acid metabolism.

  20. Effect of mitochondrial ascorbic acid synthesis on photosynthesis.

    PubMed

    Senn, M E; Gergoff Grozeff, G E; Alegre, M L; Barrile, F; De Tullio, M C; Bartoli, C G

    2016-07-01

    Ascorbic acid (AA) is synthesized in plant mitochondria through the oxidation of l-galactono-1,4-lactone (l-GalL) and then distributed to different cell compartments. AA-deficient Arabidopsis thaliana mutants (vtc2) and exogenous applications of l-GalL were used to generate plants with different AA content in their leaves. This experimental approach allows determining specific AA-dependent effects on carbon metabolism. No differences in O2 uptake, malic and citric acid and NADH content suggest that AA synthesis or accumulation did not affect mitochondrial activity; however, l-GalL treatment increased CO2 assimilation and photosynthetic electron transport rate in vtc2 (but not wt) leaves demonstrating a stimulation of photosynthesis after l-GalL treatment. Increased CO2 assimilation correlated with increased leaf stomatal conductance observed in l-GalL-treated vtc2 plants.

  1. The Warburg effect in tumor progression: Mitochondrial oxidative metabolism as an anti-metastasis mechanism

    PubMed Central

    Lu, Jianrong; Tan, Ming; Cai, Qingsong

    2014-01-01

    Compared to normal cells, cancer cells strongly upregulate glucose uptake and glycolysis to give rise to increased yield of intermediate glycolytic metabolites and the end product pyruvate. Moreover, glycolysis is uncoupled from the mitochondrial tricarboxylic acid (TCA) cycle and oxidative phosphorylation (OXPHOS) in cancer cells. Consequently, the majority of glycolysis-derived pyruvate is diverted to lactate fermentation and kept away from mitochondrial oxidative metabolism. This metabolic phenotype is known as the Warburg effect. While it has become widely accepted that the glycolytic intermediates provide essential anabolic support for cell proliferation and tumor growth, it remains largely elusive whether and how the Warburg metabolic phenotype may play a role in tumor progression. We hereby review the cause and consequence of the restrained oxidative metabolism, in particular in tumor metastasis. Cells change or lose their extracellular matrix during the metastatic process. Inadequate/inappropriate matrix attachment generates reactive oxygen species (ROS) and causes a specific type of cell death, termed anoikis, in normal cells. Although anoikis is a barrier to metastasis, cancer cells have often acquired elevated threshold for anoikis and hence heightened metastatic potential. As ROS are inherent byproducts of oxidative metabolism, forced stimulation of glucose oxidation in cancer cells raises oxidative stress and restores cells’ sensitivity to anoikis. Therefore, by limiting the pyruvate flux into mitochondrial oxidative metabolism, the Warburg effect enables cancer cells to avoid excess ROS generation from mitochondrial respiration and thus gain increased anoikis resistance and survival advantage for metastasis. Consistent with this notion, pro-metastatic transcription factors HIF and Snail attenuate oxidative metabolism, whereas tumor suppressor p53 and metastasis suppressor KISS1 promote mitochondrial oxidation. Collectively, these findings reveal

  2. The Warburg effect in tumor progression: mitochondrial oxidative metabolism as an anti-metastasis mechanism.

    PubMed

    Lu, Jianrong; Tan, Ming; Cai, Qingsong

    2015-01-28

    Compared to normal cells, cancer cells strongly upregulate glucose uptake and glycolysis to give rise to increased yield of intermediate glycolytic metabolites and the end product pyruvate. Moreover, glycolysis is uncoupled from the mitochondrial tricarboxylic acid (TCA) cycle and oxidative phosphorylation (OXPHOS) in cancer cells. Consequently, the majority of glycolysis-derived pyruvate is diverted to lactate fermentation and kept away from mitochondrial oxidative metabolism. This metabolic phenotype is known as the Warburg effect. While it has become widely accepted that the glycolytic intermediates provide essential anabolic support for cell proliferation and tumor growth, it remains largely elusive whether and how the Warburg metabolic phenotype may play a role in tumor progression. We hereby review the cause and consequence of the restrained oxidative metabolism, in particular in the context of tumor metastasis. Cells change or lose their extracellular matrix during the metastatic process. Inadequate/inappropriate matrix attachment generates reactive oxygen species (ROS) and causes a specific type of cell death, termed anoikis, in normal cells. Although anoikis is a barrier to metastasis, cancer cells have often acquired elevated threshold for anoikis and hence heightened metastatic potential. As ROS are inherent byproducts of oxidative metabolism, forced stimulation of glucose oxidation in cancer cells raises oxidative stress and restores cells' sensitivity to anoikis. Therefore, by limiting the pyruvate flux into mitochondrial oxidative metabolism, the Warburg effect enables cancer cells to avoid excess ROS generation from mitochondrial respiration and thus gain increased anoikis resistance and survival advantage for metastasis. Consistent with this notion, pro-metastatic transcription factors HIF and Snail attenuate oxidative metabolism, whereas tumor suppressor p53 and metastasis suppressor KISS1 promote mitochondrial oxidation. Collectively, these

  3. Mitochondrial matrix P53 sensitizes cells to oxidative stress☆

    PubMed Central

    Koczor, Christopher A.; Torres, Rebecca A.; Fields, Earl J.; Boyd, Amy; Lewis, William

    2013-01-01

    A mitochondrial matrix-specific p53 construct (termed p53–290) in HepG2 cells was utilized to determine the impact of p53 in the mitochondrial matrix following oxidative stress. H2O2 exposure reduced cellular proliferation similarly in both p53–290 and vector cells, and p53–290 cells demonstrating decreased cell viability at 1 mM H2O2 (~85% viable). Mitochondrial DNA (mtDNA) abundance was decreased in a dose-dependent manner in p53–290 cells while no change was observed in vector cells. Oximetric analysis revealed reduced maximal respiration and reserve capacity in p53–290 cells. Our results demonstrate that mitochondrial matrix p53 sensitizes cells to oxidative stress by reducing mtDNA abundance and mitochondrial function. PMID:23499753

  4. Mitochondrial and peroxisomal beta-oxidation capacities of organs from a non-oilseed plant.

    PubMed

    Masterson, C; Wood, C

    2001-09-22

    Until recently, beta-oxidation was believed to be exclusively located in the peroxisomes of all higher plants. Whilst this is true for germinating oilseeds undergoing gluconeogenesis, evidence demonstrating mitochondrial beta-oxidation in other plant systems has refuted this central dogma of plant lipid metabolism. This report describes a comparative study of the dual mitochondrial and peroxisomal beta-oxidation capacities of plant organs. Oxidation of [1-(14)C] palmitate was measured in the cotyledons, plumules and radicles of Pisum sativum L., which is a starchy seed, over a 14 day period from the commencement of imbibition. Respiratory chain inhibitors were used for differentiating between mitochondrial and peroxisomal beta-oxidation. Peroxisomal beta-oxidation gave a steady, baseline rate and, in the early stages of seedling development, accounted for 70-100% of the beta-oxidation observed. Mitochondrial beta-oxidation gave peaks of activity at days 7 and 10-11, accounting for up to 82% of the total beta-oxidation activity at these times. These peaks coincide with key stages of seedling development and were not observed when normal development was disrupted by growth in the dark. Peroxisomal beta-oxidation was unaffected by etiolation. Since mitochondrial beta-oxidation was overt only during times of intense biosynthetic activity it might be switched on or off during seedling development. In contrast, peroxisomes maintained a continuous, low beta-oxidation activity that could be essential in removing harmful free fatty acids, e.g. those produced by protein and lipid turnover.

  5. Oxidative stress induces mitochondrial fragmentation in frataxin-deficient cells

    SciTech Connect

    Lefevre, Sophie; Sliwa, Dominika; Rustin, Pierre; Camadro, Jean-Michel; Santos, Renata

    2012-02-10

    Highlights: Black-Right-Pointing-Pointer Yeast frataxin-deficiency leads to increased proportion of fragmented mitochondria. Black-Right-Pointing-Pointer Oxidative stress induces complete mitochondrial fragmentation in {Delta}yfh1 cells. Black-Right-Pointing-Pointer Oxidative stress increases mitochondrial fragmentation in patient fibroblasts. Black-Right-Pointing-Pointer Inhibition of mitochondrial fission in {Delta}yfh1 induces oxidative stress resistance. -- Abstract: Friedreich ataxia (FA) is the most common recessive neurodegenerative disease. It is caused by deficiency in mitochondrial frataxin, which participates in iron-sulfur cluster assembly. Yeast cells lacking frataxin ({Delta}yfh1 mutant) showed an increased proportion of fragmented mitochondria compared to wild-type. In addition, oxidative stress induced complete fragmentation of mitochondria in {Delta}yfh1 cells. Genetically controlled inhibition of mitochondrial fission in these cells led to increased resistance to oxidative stress. Here we present evidence that in yeast frataxin-deficiency interferes with mitochondrial dynamics, which might therefore be relevant for the pathophysiology of FA.

  6. Mitochondrial targeting of bilirubin regulatory enzymes: An adaptive response to oxidative stress

    SciTech Connect

    Muhsain, Siti Nur Fadzilah; Lang, Matti A.; Abu-Bakar, A'edah

    2015-01-01

    The intracellular level of bilirubin (BR), an endogenous antioxidant that is cytotoxic at high concentrations, is tightly controlled within the optimal therapeutic range. We have recently described a concerted intracellular BR regulation by two microsomal enzymes: heme oxygenase 1 (HMOX1), essential for BR production and cytochrome P450 2A5 (CYP2A5), a BR oxidase. Herein, we describe targeting of these enzymes to hepatic mitochondria during oxidative stress. The kinetics of microsomal and mitochondrial BR oxidation were compared. Treatment of DBA/2J mice with 200 mg pyrazole/kg/day for 3 days increased hepatic intracellular protein carbonyl content and induced nucleo-translocation of Nrf2. HMOX1 and CYP2A5 proteins and activities were elevated in microsomes and mitoplasts but not the UGT1A1, a catalyst of BR glucuronidation. A CYP2A5 antibody inhibited 75% of microsomal BR oxidation. The inhibition was absent in control mitoplasts but elevated to 50% after treatment. An adrenodoxin reductase antibody did not inhibit microsomal BR oxidation but inhibited 50% of mitochondrial BR oxidation. Ascorbic acid inhibited 5% and 22% of the reaction in control and treated microsomes, respectively. In control mitoplasts the inhibition was 100%, which was reduced to 50% after treatment. Bilirubin affinity to mitochondrial and microsomal CYP2A5 enzyme is equally high. Lastly, the treatment neither released cytochrome c into cytoplasm nor dissipated membrane potential, indicating the absence of mitochondrial membrane damage. Collectively, the observations suggest that BR regulatory enzymes are recruited to mitochondria during oxidative stress and BR oxidation by mitochondrial CYP2A5 is supported by mitochondrial mono-oxygenase system. The induced recruitment potentially confers membrane protection. - Highlights: • Pyrazole induces oxidative stress in the mouse liver. • Pyrazole-induced oxidative stress induces mitochondrial targeting of key bilirubin regulatory enzymes, HMOX1

  7. Dietary fatty acids affect mitochondrial phospholipid compositions and mitochondrial gene expression of rainbow trout liver at different ages.

    PubMed

    Almaida-Pagán, P F; De Santis, C; Rubio-Mejía, O L; Tocher, D R

    2015-01-01

    Mitochondria are among the first responders to various stressors that challenge the homeostasis of cells and organisms. Mitochondrial decay is generally associated with impairment in the organelle bioenergetics function and increased oxidative stress, and it appears that deterioration of mitochondrial inner membrane phospholipids (PL), particularly cardiolipin (CL), and accumulation of mitochondrial DNA (mtDNA) mutations are among the main mechanisms involved in this process. In the present study, liver mitochondrial membrane PL compositions, lipid peroxidation, and mtDNA gene expression were analyzed in rainbow trout fed three diets with the same base formulation but with lipid supplied either by fish oil (FO), rapeseed oil (RO), or high DHA oil (DHA) during 6 weeks. Specifically, two feeding trials were performed using fish from the same population of two ages (1 and 3 years), and PL class compositions of liver mitochondria, fatty acid composition of individual PL classes, TBARS content, and mtDNA expression were determined. Dietary fatty acid composition strongly affected mitochondrial membrane composition from trout liver but observed changes did not fully reflect the diet, particularly when it contained high DHA. The changes were PL specific, CL being particularly resistant to changes in DHA. Some significant differences observed in expression of mtDNA with diet may suggest long-term dietary effects in mitochondrial gene expression which could affect electron transport chain function. All the changes were influenced by fish age, which could be related to the different growth rates observed between 1- and 3-year-old trout but that could also indicate age-related changes in the ability to maintain structural homeostasis of mitochondrial membranes.

  8. Oxidative DNA damage causes mitochondrial genomic instability in Saccharomyces cerevisiae.

    PubMed

    Doudican, Nicole A; Song, Binwei; Shadel, Gerald S; Doetsch, Paul W

    2005-06-01

    Mitochondria contain their own genome, the integrity of which is required for normal cellular energy metabolism. Reactive oxygen species (ROS) produced by normal mitochondrial respiration can damage cellular macromolecules, including mitochondrial DNA (mtDNA), and have been implicated in degenerative diseases, cancer, and aging. We developed strategies to elevate mitochondrial oxidative stress by exposure to antimycin and H(2)O(2) or utilizing mutants lacking mitochondrial superoxide dismutase (sod2Delta). Experiments were conducted with strains compromised in mitochondrial base excision repair (ntg1Delta) and oxidative damage resistance (pif1Delta) in order to delineate the relationship between these pathways. We observed enhanced ROS production, resulting in a direct increase in oxidative mtDNA damage and mutagenesis. Repair-deficient mutants exposed to oxidative stress conditions exhibited profound genomic instability. Elimination of Ntg1p and Pif1p resulted in a synergistic corruption of respiratory competency upon exposure to antimycin and H(2)O(2). Mitochondrial genomic integrity was substantially compromised in ntg1Delta pif1Delta sod2Delta strains, since these cells exhibit a total loss of mtDNA. A stable respiration-defective strain, possessing a normal complement of mtDNA damage resistance pathways, exhibited a complete loss of mtDNA upon exposure to antimycin and H(2)O(2). This loss was preventable by Sod2p overexpression. These results provide direct evidence that oxidative mtDNA damage can be a major contributor to mitochondrial genomic instability and demonstrate cooperation of Ntg1p and Pif1p to resist the introduction of lesions into the mitochondrial genome.

  9. NITRIC OXIDE, MITOCHONDRIAL HYPERPOLARIZATION AND T-CELL ACTIVATION

    PubMed Central

    Nagy, Gyorgy; Koncz, Agnes; Fernandez, David; Perl, Andras

    2007-01-01

    T lymphocyte activation is associated with nitric oxide (NO) production that plays an essential role in multiple T cell functions. NO acts as a messenger, activating soluble guanyl cyclase and participating in the transduction signaling pathways involving cyclic GMP. NO modulates mitochondrial events that are involved in apoptosis and regulates mitochondrial membrane potential and mitochondrial biogenesis in many cell types, including lymphocytes. Mitochondrial hyperpolarization (MHP), an early and reversible event during both T lymphocyte activation and apoptosis, is regulated by NO. Here, we discuss recent evidence that NO-induced MHP represents a molecular switch in multiple T cell signaling pathways. Overproduction of NO in systemic lupus erythematosus (SLE) induces mitochondrial biogenesis and alters Ca2+ signaling. Thus, while NO plays a physiological role in lymphocyte cell signaling, its overproduction may disturb normal T cell function, contributing to the pathogenesis of autoimmunity. PMID:17462531

  10. Cardiac Mitochondrial Proteome Dynamics with Heavy Water Reveals Stable Rate of Mitochondrial Protein Synthesis in Heart Failure Despite Decline in Mitochondrial Oxidative Capacity

    PubMed Central

    Shekar, Kadambari Chandra; Li, Ling; Dabkowski, Erinne R.; Xu, Wenhong; Ribeiro, Rogerio Faustino; Hecker, Peter A.; Recchia, Fabio A.; Sadygov, Rovshan G.; Willard, Belinda; Kasumov, Takhar; Stanley, William C.

    2017-01-01

    We recently developed a method to measure mitochondrial proteome dynamics with heavy water (2H2O)-based metabolic labeling and high resolution mass spectrometry. We reported the half-lives and synthesis rates of several proteins in the two cardiac mitochondrial subpopulations, subsarcolemmal and interfibrillar (SSM and IFM), in Sprague Dawley rats. In the present study, we tested the hypothesis that the mitochondrial protein synthesis rate is reduced in heart failure, with possible differential changes in SSM versus IFM. Six to seven week old male Sprague Dawley rats underwent transverse aortic constriction (TAC) and developed moderate heart failure after 22 weeks. Heart failure and sham rats of the same age received heavy water (5% in drinking water) for up to 80 days. Cardiac SSM and IFM were isolated from both groups and the proteins were separated by 1D gel electrophoresis. Heart failure reduced protein content and increased the turnover rate of several proteins involved in fatty acid oxidation, electron transport chain and ATP synthesis, while it decreased the turnover of other proteins, including pyruvate dehydrogenase subunit in IFM, but not in SSM. Because of these bidirectional changes, the average overall half-life of proteins was not altered by heart failure in both SSM and IFM. The kinetic measurements of individual mitochondrial proteins presented in this study may contribute to a better understanding of the mechanisms responsible for mitochondrial alterations in the failing heart. PMID:24995939

  11. Oxidized mitochondrial DNA activates the NLRP3 inflammasome during apoptosis.

    PubMed

    Shimada, Kenichi; Crother, Timothy R; Karlin, Justin; Dagvadorj, Jargalsaikhan; Chiba, Norika; Chen, Shuang; Ramanujan, V Krishnan; Wolf, Andrea J; Vergnes, Laurent; Ojcius, David M; Rentsendorj, Altan; Vargas, Mario; Guerrero, Candace; Wang, Yinsheng; Fitzgerald, Katherine A; Underhill, David M; Town, Terrence; Arditi, Moshe

    2012-03-23

    We report that in the presence of signal 1 (NF-κB), the NLRP3 inflammasome was activated by mitochondrial apoptotic signaling that licensed production of interleukin-1β (IL-1β). NLRP3 secondary signal activators such as ATP induced mitochondrial dysfunction and apoptosis, resulting in release of oxidized mitochondrial DNA (mtDNA) into the cytosol, where it bound to and activated the NLRP3 inflammasome. The antiapoptotic protein Bcl-2 inversely regulated mitochondrial dysfunction and NLRP3 inflammasome activation. Mitochondrial DNA directly induced NLRP3 inflammasome activation, because macrophages lacking mtDNA had severely attenuated IL-1β production, yet still underwent apoptosis. Both binding of oxidized mtDNA to the NLRP3 inflammasome and IL-1β secretion could be competitively inhibited by the oxidized nucleoside 8-OH-dG. Thus, our data reveal that oxidized mtDNA released during programmed cell death causes activation of the NLRP3 inflammasome. These results provide a missing link between apoptosis and inflammasome activation, via binding of cytosolic oxidized mtDNA to the NLRP3 inflammasome.

  12. Oxidized Mitochondrial DNA Activates the NLRP3 Inflammasome During Apoptosis

    PubMed Central

    Shimada, Kenichi; Crother, Timothy R.; Karlin, Justin; Dagvadorj, Jargalsaikhan; Chiba, Norika; Chen, Shuang; Ramanujan, V. Krishnan; Wolf, Andrea J.; Vergnes, Laurent; Ojcius, David M.; Rentsendorj, Altan; Vargas, Mario; Guerrero, Candace; Wang, Yinsheng; Fitzgerald, Katherine A.; Underhill, David M.; Town, Terrence; Arditi, Moshe

    2012-01-01

    SUMMARY We report that in the presence of signal 1 (NF-κB), the NLRP3 inflammasome was activated by mitochondrial apoptotic signaling that licensed production of interleukin-1β (IL-1β). NLRP3 secondary signal activators such as ATP induced mitochondrial dysfunction and apoptosis, resulting in release of oxidized mitochondrial DNA (mtDNA) into the cytosol, where it bound to and activated the NLRP3 inflammasome. The anti-apoptotic protein Bcl-2 inversely regulated mitochondrial dysfunction and NLRP3 inflammasome activation. Mitochondrial DNA directly induced NLRP3 inflammasome activation, because macrophages lacking mtDNA had severely attenuated IL-1β production, yet still underwent apoptosis. Both binding of oxidized mtDNA to the NLRP3 inflammasome and IL-1β secretion could be competitively inhibited by the oxidized nucleoside, 8-OH-dG. Thus, our data reveal that oxidized mtDNA released during programmed cell death causes activation of the NLRP3 inflammasome. These results provide a missing link between apoptosis and inflammasome activation, via binding of cytosolic oxidized mtDNA to the NLRP3 inflammasome. PMID:22342844

  13. Elevated mitochondrial oxidative stress impairs metabolic adaptations to exercise in skeletal muscle.

    PubMed

    Crane, Justin D; Abadi, Arkan; Hettinga, Bart P; Ogborn, Daniel I; MacNeil, Lauren G; Steinberg, Gregory R; Tarnopolsky, Mark A

    2013-01-01

    Mitochondrial oxidative stress is a complex phenomenon that is inherently tied to energy provision and is implicated in many metabolic disorders. Exercise training increases mitochondrial oxidative capacity in skeletal muscle yet it remains unclear if oxidative stress plays a role in regulating these adaptations. We demonstrate that the chronic elevation in mitochondrial oxidative stress present in Sod2 (+/-) mice impairs the functional and biochemical mitochondrial adaptations to exercise. Following exercise training Sod2 (+/-) mice fail to increase maximal work capacity, mitochondrial enzyme activity and mtDNA copy number, despite a normal augmentation of mitochondrial proteins. Additionally, exercised Sod2 (+/-) mice cannot compensate for their higher amount of basal mitochondrial oxidative damage and exhibit poor electron transport chain complex assembly that accounts for their compromised adaptation. Overall, these results demonstrate that chronic skeletal muscle mitochondrial oxidative stress does not impact exercise induced mitochondrial biogenesis, but impairs the resulting mitochondrial protein function and can limit metabolic plasticity.

  14. Uncoupling of mitochondrial oxidative phosphorylation by DNA gyrase inhibitors

    SciTech Connect

    Gallagher, M.; Weinberg, R.; Simpson, M.V.

    1986-05-01

    Supercoiled mtDNA and the swivel requirement for its replication suggest the existence of a mtDNA gyrase. The authors published studies on isolated mitochondria showing that novobiocin, coumermycin, nalidixic acid, and oxolinic acid promote relaxed DNA formation at the expense of supercoiled DNA are in accord with this view. However, their inability to directly detect the enzyme led them to ask whether these drugs act elsewhere. Their results with isolated rat liver mitochondria show that novo, nal, but not oxo, stimulate O/sub 2/ uptake as much as does 2.4-dinitrophenol (DNP). This possible uncoupling effect was confirmed by a standard (/sup 32/P) assay showing the following inhibitions of ATP synthesis: 0.2 mM novo, 95% (0.4 mM, 100%) 0.4 mM nal, 37%; oxo to at least 1.9 mM, 0%; (0.5 mM 2,4-DNP, 100%). Thus, oxo remains a useful tool for intact mitochondrial studies. Because these three drugs, especially novo, are being used to study the role of DNA superhelicity on pro- and eucaryotic (and mitochondrial) gene expression, the authors studied their effect on oxidative phosphorylation in such cells. In these cases the drugs did not affect DNP-sensitive (/sup 14/C)glutamine transport into E. coli cells (an established measure of ATP level), nor, in an S. cerevisiae mutant permeable to novo, did novo affect the steady state ATP level. Studies on cultured mammalian cells are in progress.

  15. D-saccharic acid-1,4-lactone ameliorates alloxan-induced diabetes mellitus and oxidative stress in rats through inhibiting pancreatic beta-cells from apoptosis via mitochondrial dependent pathway

    SciTech Connect

    Bhattacharya, Semantee; Manna, Prasenjit; Sil, Parames C.

    2011-12-15

    Oxidative stress plays a vital role in diabetic complications. To suppress the oxidative stress mediated damage in diabetic pathophysiology, a special focus has been given on naturally occurring antioxidants present in normal diet. D-saccharic acid 1,4-lactone (DSL), a derivative of D-glucaric acid, is present in many dietary plants and is known for its detoxifying and antioxidant properties. The aim of the present study was to evaluate the beneficial role of DSL against alloxan (ALX) induced diabetes in the pancreas tissue of Swiss albino rats. A dose-dependent study for DSL (20-120 mg/kg body weight) was carried out to find the effective dose of the compound in ALX-induced diabetic rats. ALX exposure elevated the blood glucose, glycosylated Hb, decreased the plasma insulin and disturbed the intra-cellular antioxidant machineries whereas oral administration of DSL at a dose of 80 mg/kg body weight restored these alterations close to normal. Investigating the mechanism of the protective activity of DSL we observed that it prevented the pancreatic {beta}-cell apoptosis via mitochondria-dependent pathway. Results showed decreased mitochondrial membrane potential, enhanced cytochrome c release in the cytosol and reciprocal regulation of Bcl-2 family proteins in the diabetic rats. These events were also found to be associated with increased level of Apaf-1, caspase 9, and caspase 3 that ultimately led to pancreatic {beta}-cell apoptosis. DSL treatment, however, counteracted these changes. In conclusion, DSL possesses the capability of ameliorating the oxidative stress in ALX-induced diabetes and thus could be a promising approach in lessening diabetic complications. Highlights: Black-Right-Pointing-Pointer Oxidative stress is suggested as a key event in the pathogenesis of diabetes. Black-Right-Pointing-Pointer D-saccharic acid 1,4-lactone (DSL) reduces the alloxan-induced diabetes mellitus. Black-Right-Pointing-Pointer DSL normalizes cellular antioxidant machineries

  16. Tamoxifen inhibits mitochondrial oxidative stress damage induced by copper orthophenanthroline.

    PubMed

    Buelna-Chontal, Mabel; Hernández-Esquivel, Luz; Correa, Francisco; Díaz-Ruiz, Jorge Luis; Chávez, Edmundo

    2016-12-01

    In this work, we studied the effect of tamoxifen and cyclosporin A on mitochondrial permeability transition caused by addition of the thiol-oxidizing pair Cu(2+) -orthophenanthroline. The findings indicate that tamoxifen and cyclosporin A circumvent the oxidative membrane damage manifested by matrix Ca(2+) release, mitochondrial swelling, and transmembrane electrical gradient collapse. Furthermore, it was found that tamoxifen and cyclosporin A prevent the generation of TBARs promoted by Cu(2+) -orthophenanthroline, as well as the inactivation of the mitochondrial enzyme aconitase and disruption of mDNA. Electrophoretic analysis was unable to demonstrate a cross-linking reaction between membrane proteins. Yet, it was found that Cu(2+) -orthophenanthroline induced the generation of reactive oxygen species. It is thus plausible that membrane leakiness is due to an oxidative stress injury.

  17. Mitochondrial glycolate oxidation contributes to photorespiration in higher plants.

    PubMed

    Niessen, Markus; Thiruveedhi, Krishnaveni; Rosenkranz, Ruben; Kebeish, Rashad; Hirsch, Heinz-Josef; Kreuzaler, Fritz; Peterhänsel, Christoph

    2007-01-01

    The oxidation of glycolate to glyoxylate is an important reaction step in photorespiration. Land plants and charophycean green algae oxidize glycolate in the peroxisome using oxygen as a co-factor, whereas chlorophycean green algae use a mitochondrial glycolate dehydrogenase (GDH) with organic co-factors. Previous analyses revealed the existence of a GDH in the mitochondria of Arabidopsis thaliana (AtGDH). In this study, the contribution of AtGDH to photorespiration was characterized. Both RNA abundance and mitochondrial GDH activity were up-regulated under photorespiratory growth conditions. Labelling experiments indicated that glycolate oxidation in mitochondrial extracts is coupled to CO(2) release. This effect could be enhanced by adding co-factors for aminotransferases, but is inhibited by the addition of glycine. T-DNA insertion lines for AtGDH show a drastic reduction in mitochondrial GDH activity and CO(2) release from glycolate. Furthermore, photorespiration is reduced in these mutant lines compared with the wild type, as revealed by determination of the post-illumination CO(2) burst and the glycine/serine ratio under photorespiratory growth conditions. The data show that mitochondrial glycolate oxidation contributes to photorespiration in higher plants. This indicates the conservation of chlorophycean photorespiration in streptophytes despite the evolution of leaf-type peroxisomes.

  18. Nox2 as a potential target of mitochondrial superoxide and its role in endothelial oxidative stress

    PubMed Central

    Nazarewicz, Rafal R.; Bikineyeva, Alfiya; Dikalov, Sergey I.

    2013-01-01

    Superoxide (O2·−) production by the NADPH oxidases is implicated in the pathogenesis of many cardiovascular diseases, including hypertension. We have previously shown that activation of NADPH oxidases increases mitochondrial O2·− which is inhibited by the ATP-sensitive K+ channel (mitoKATP) inhibitor 5-hydroxydecanoic acid and that scavenging of mitochondrial or cytoplasmic O2·− inhibits hypertension. We hypothesized that mitoKATP-mediated mitochondrial O2·− potentiates cytoplasmic O2·− by stimulation of NADPH oxidases. In this work we studied Nox isoforms as a potential target of mitochondrial O2·−. We tested contribution of reverse electron transfer (RET) from complex II to complex I in mitochondrial O2·− production and NADPH oxidase activation in human aortic endothelial cells. Activation of mitoKATP with low dose of diazoxide (100 nM) decreased mitochondrial membrane potential (tetramethylrhodamine methyl ester probe) and increased production of mitochondrial and cytoplasmic O2·− measured by site-specific probes and mitoSOX. Inhibition of RET with complex II inhibitor (malonate) or complex I inhibitor (rotenone) attenuated the production of mitochondrial and cytoplasmic O2·−. Supplementation with a mitochondria-targeted SOD mimetic (mitoTEMPO) or a mitochondria-targeted glutathione peroxidase mimetic (mitoEbselen) inhibited production of mitochondrial and cytoplasmic O2·−. Inhibition of Nox2 (gp91ds) or Nox2 depletion with small interfering RNA but not Nox1, Nox4, or Nox5 abolished diazoxide-induced O2·− production in the cytoplasm. Treatment of angiotensin II-infused mice with RET inhibitor dihydroethidium (malate) significantly reduced blood pressure. Our study suggests that mitoKATP-mediated mitochondrial O2·− stimulates cytoplasmic Nox2, contributing to the development of endothelial oxidative stress and hypertension. PMID:23955717

  19. p-Bromophenacyl bromide prevents cumene hydroperoxide-induced mitochondrial permeability transition by inhibiting pyridine nucleotide oxidation.

    PubMed

    Zhukova, A; Gogvadze, G; Gogvadze, V

    2004-01-01

    Mitochondrial permeability transition is commonly characterized as a Ca2+ -dependent non-specific increase in inner membrane permeability that results in swelling of mitochondria and their de-energization. In the present study, the effect of different inhibitors of phospholipase A2--p-bromophenacyl bromide, dibucaine, and aristolochic acid--on hydroperoxide-induced permeability transitions in rat liver mitochondria was tested. p-Bromophenacyl bromide completely prevented the hydroperoxide-induced mitochondrial permeability transition while the effects of dibucaine or aristolochic acid were negligible. Organic hydroperoxides added to mitochondria undergo reduction to corresponding alcohols by mitochondrial glutathione peroxidase. This reduction occurs at the expense of GSH which, in turn, can be reduced by glutathione reductase via oxidation of mitochondrial pyridine nucleotides. The latter is considered a prerequisite step for mitochondrial permeability transition. Among all the inhibitors tested, only p-bromophenacyl bromide completely prevented hydroperoxide-induced oxidation of mitochondrial pyridine nucleotides. Interestingly, p-bromophenacyl bromide had no affect on mitochondrial glutathione peroxidase, but reacted with mitochondrial glutathione that prevented pyridine nucleotides from being oxidized. Our data suggest that p-bromophenacyl bromide prevents hydroperoxide-induced deterioration of mitochondria via interaction with glutathione rather than through inhibition of phospholipase A2.

  20. Mitochondrial fat oxidation is essential for lipid-induced inflammation in skeletal muscle in mice.

    PubMed

    Warfel, Jaycob D; Bermudez, Estrellita M; Mendoza, Tamra M; Ghosh, Sujoy; Zhang, Jingying; Elks, Carrie M; Mynatt, Randall; Vandanmagsar, Bolormaa

    2016-11-28

    Inflammation, lipotoxicity and mitochondrial dysfunction have been implicated in the pathogenesis of obesity-induced insulin resistance and type 2 diabetes. However, how these factors are intertwined in the development of obesity/insulin resistance remains unclear. Here, we examine the role of mitochondrial fat oxidation on lipid-induced inflammation in skeletal muscle. We used skeletal muscle-specific Cpt1b knockout mouse model where the inhibition of mitochondrial fatty acid oxidation results in accumulation of lipid metabolites in muscle and elevated circulating free fatty acids. Gene expression of pro-inflammatory cytokines, chemokines, and cytokine- and members of TLR-signalling pathways were decreased in Cpt1b(m-/-) muscle. Inflammatory signalling pathways were not activated when evaluated by multiplex and immunoblot analysis. In addition, the inflammatory response to fatty acids was reduced in primary muscle cells derived from Cpt1b(m-/-) mice. Gene expression of Cd11c, the M1 macrophage marker, was decreased; while Cd206, the M2 macrophage marker, was increased in skeletal muscle of Cpt1b(m-/-) mice. Finally, expression of pro-inflammatory markers was decreased in white adipose tissue of Cpt1b(m-/-) mice. We show that the inflammatory response elicited by elevated intracellular lipids in skeletal muscle is repressed in Cpt1b(m-/-) mice, strongly supporting the hypothesis that mitochondrial processing of fatty acids is essential for the lipid-induction of inflammation in muscle.

  1. Mitochondrial fat oxidation is essential for lipid-induced inflammation in skeletal muscle in mice

    PubMed Central

    Warfel, Jaycob D.; Bermudez, Estrellita M.; Mendoza, Tamra M.; Ghosh, Sujoy; Zhang, Jingying; Elks, Carrie M.; Mynatt, Randall; Vandanmagsar, Bolormaa

    2016-01-01

    Inflammation, lipotoxicity and mitochondrial dysfunction have been implicated in the pathogenesis of obesity-induced insulin resistance and type 2 diabetes. However, how these factors are intertwined in the development of obesity/insulin resistance remains unclear. Here, we examine the role of mitochondrial fat oxidation on lipid-induced inflammation in skeletal muscle. We used skeletal muscle-specific Cpt1b knockout mouse model where the inhibition of mitochondrial fatty acid oxidation results in accumulation of lipid metabolites in muscle and elevated circulating free fatty acids. Gene expression of pro-inflammatory cytokines, chemokines, and cytokine- and members of TLR-signalling pathways were decreased in Cpt1bm−/− muscle. Inflammatory signalling pathways were not activated when evaluated by multiplex and immunoblot analysis. In addition, the inflammatory response to fatty acids was reduced in primary muscle cells derived from Cpt1bm−/− mice. Gene expression of Cd11c, the M1 macrophage marker, was decreased; while Cd206, the M2 macrophage marker, was increased in skeletal muscle of Cpt1bm−/− mice. Finally, expression of pro-inflammatory markers was decreased in white adipose tissue of Cpt1bm−/− mice. We show that the inflammatory response elicited by elevated intracellular lipids in skeletal muscle is repressed in Cpt1bm−/− mice, strongly supporting the hypothesis that mitochondrial processing of fatty acids is essential for the lipid-induction of inflammation in muscle. PMID:27892502

  2. Oxidation of alpha-ketoglutarate is required for reductive carboxylation in cancer cells with mitochondrial defects

    PubMed Central

    Mullen, Andrew R.; Hu, Zeping; Shi, Xiaolei; Jiang, Lei; Boroughs, Lindsey K.; Kovacs, Zoltan; Boriack, Richard; Rakheja, Dinesh; Sullivan, Lucas B.; Linehan, W. Marston; Chandel, Navdeep S.; DeBerardinis, Ralph J.

    2014-01-01

    Summary Mammalian cells generate citrate by decarboxylating pyruvate in the mitochondria to supply the tricarboxylic acid (TCA) cycle. In contrast, hypoxia and other impairments of mitochondrial function induce an alternative pathway that produces citrate by reductively carboxylating α-ketoglutarate (AKG) via NADPH-dependent isocitrate dehydrogenase (IDH). It is unknown how cells generate reducing equivalents necessary to supply reductive carboxylation in the setting of mitochondrial impairment. Here we identified shared metabolic features in cells using reductive carboxylation. Paradoxically, reductive carboxylation was accompanied by concomitant AKG oxidation in the TCA cycle. Inhibiting AKG oxidation decreased reducing equivalent availability and suppressed reductive carboxylation. Interrupting transfer of reducing equivalents from NADH to NADPH by nicotinamide nucleotide transhydrogenase increased NADH abundance and decreased NADPH abundance while suppressing reductive carboxylation. The data demonstrate that reductive carboxylation requires bidirectional AKG metabolism along oxidative and reductive pathways, with the oxidative pathway producing reducing equivalents used to operate IDH in reverse. PMID:24857658

  3. Oxidative stress and mitochondrial damage in coronary artery bypass graft surgery: effects of antioxidant treatments.

    PubMed

    Milei, J; Ferreira, R; Grana, D R; Boveris, A

    2001-01-01

    We examined antioxidant actions in 73 patients undergoing coronary artery surgery by assessing mitochondrial damage and oxidative stress in ventricular biopsies obtained at preischemia and postreperfusion. Those patients who received antioxidant therapy benefited by less oxidative stress and mitochondrial damage.

  4. Sirt3 Protects Dopaminergic Neurons from Mitochondrial Oxidative Stress.

    PubMed

    Shi, Han; Deng, Han-Xiang; Gius, David; Schumacker, Paul T; James Surmeier, D; Ma, Yong-Chao

    2017-03-24

    Age-dependent elevation in mitochondrial oxidative stress is widely posited to be a major factor underlying the loss of substantia nigra pars compacta (SNc) dopaminergic neurons in Parkinson's disease (PD). However, mechanistic links between aging and oxidative stress are not well understood. Sirtuin-3 (Sirt3) is a mitochondrial deacetylase that could mediate this connection. Indeed, genetic deletion of Sirt3 increased oxidative stress and decreased the membrane potential of mitochondria in SNc dopaminergic neurons. This change was attributable to increased acetylation and decreased activity of manganese superoxide dismutase (MnSOD). Site directed mutagenesis of lysine 68 to glutamine (K68Q), mimicking acetylation, decreased MnSOD activity in SNc dopaminergic neurons, whereas mutagenesis of lysine 68 to arginine (K68R), mimicking deacetylation, increased activity. Introduction of K68R MnSOD rescued mitochondrial redox status and membrane potential of SNc dopaminergic neurons from Sirt3 knockouts. Moreover, deletion of DJ-1, which helps orchestrate nuclear oxidant defenses, and Sirt3 in mice led to a clear age-related loss of SNc dopaminergic neurons. Lastly, K68 acetylation of MnSOD was significantly increased in the SNc of PD patients. Taken together, our studies suggest that an age-related decline in Sirt3 protective function is a major factor underlying increasing mitochondrial oxidative stress and loss of SNc dopaminergic neurons in PD.

  5. Mitochondrial dysfunction, oxidative stress, and major depressive disorder

    PubMed Central

    Tobe, Edward H

    2013-01-01

    There is controversy about depression being a physical illness, in part because a reproducible, sensitive, and specific biologic marker is not available. However, there is evidence that mitochondrial dysfunction and oxidative stress may be associated with abnormal brain function and mood disorders, such as depression. This paper reviews selected human and animal studies providing evidence that intracellular mitochondrial metabolic dysfunction in specific brain regions is associated with major depressive disorder. This supports the hypothesis that chronic mitochondrial dysfunction in specific tissues may be associated with depression. Evaluation of mitochondrial dysfunction in specific tissues may broaden the perspective of depression beyond theories about neurotransmitters or receptor sites, and may explain the persistent signs and symptoms of depression. PMID:23650447

  6. In vitro treatment of HepG2 cells with saturated fatty acids reproduces mitochondrial dysfunction found in nonalcoholic steatohepatitis.

    PubMed

    García-Ruiz, Inmaculada; Solís-Muñoz, Pablo; Fernández-Moreira, Daniel; Muñoz-Yagüe, Teresa; Solís-Herruzo, José A

    2015-02-01

    Activity of the oxidative phosphorylation system (OXPHOS) is decreased in humans and mice with nonalcoholic steatohepatitis. Nitro-oxidative stress seems to be involved in its pathogenesis. The aim of this study was to determine whether fatty acids are implicated in the pathogenesis of this mitochondrial defect. In HepG2 cells, we analyzed the effect of saturated (palmitic and stearic acids) and monounsaturated (oleic acid) fatty acids on: OXPHOS activity; levels of protein expression of OXPHOS complexes and their subunits; gene expression and half-life of OXPHOS complexes; nitro-oxidative stress; and NADPH oxidase gene expression and activity. We also studied the effects of inhibiting or silencing NADPH oxidase on the palmitic-acid-induced nitro-oxidative stress and subsequent OXPHOS inhibition. Exposure of cultured HepG2 cells to saturated fatty acids resulted in a significant decrease in the OXPHOS activity. This effect was prevented in the presence of a mimic of manganese superoxide dismutase. Palmitic acid reduced the amount of both fully-assembled OXPHOS complexes and of complex subunits. This reduction was due mainly to an accelerated degradation of these subunits, which was associated with a 3-tyrosine nitration of mitochondrial proteins. Pretreatment of cells with uric acid, an antiperoxynitrite agent, prevented protein degradation induced by palmitic acid. A reduced gene expression also contributed to decrease mitochondrial DNA (mtDNA)-encoded subunits. Saturated fatty acids induced oxidative stress and caused mtDNA oxidative damage. This effect was prevented by inhibiting NADPH oxidase. These acids activated NADPH oxidase gene expression and increased NADPH oxidase activity. Silencing this oxidase abrogated totally the inhibitory effect of palmitic acid on OXPHOS complex activity. We conclude that saturated fatty acids caused nitro-oxidative stress, reduced OXPHOS complex half-life and activity, and decreased gene expression of mtDNA-encoded subunits

  7. Oxidative stress and mitochondrial dysfunction-linked neurodegenerative disorders.

    PubMed

    Islam, Md Torequl

    2017-01-01

    Reactive species play an important role in physiological functions. Overproduction of reactive species, notably reactive oxygen (ROS) and nitrogen (RNS) species along with the failure of balance by the body's antioxidant enzyme systems results in destruction of cellular structures, lipids, proteins, and genetic materials such as DNA and RNA. Moreover, the effects of reactive species on mitochondria and their metabolic processes eventually cause a rise in ROS/RNS levels, leading to oxidation of mitochondrial proteins, lipids, and DNA. Oxidative stress has been considered to be linked to the etiology of many diseases, including neurodegenerative diseases (NDDs) such as Alzheimer diseases, Amyotrophic lateral sclerosis, Friedreich's ataxia, Huntington's disease, Multiple sclerosis, and Parkinson's diseases. In addition, oxidative stress causing protein misfold may turn to other NDDs include Creutzfeldt-Jakob disease, Bovine Spongiform Encephalopathy, Kuru, Gerstmann-Straussler-Scheinker syndrome, and Fatal Familial Insomnia. An overview of the oxidative stress and mitochondrial dysfunction-linked NDDs has been summarized in this review.

  8. Hepatic mitochondrial dysfunction induced by fatty acids and ethanol.

    PubMed

    Gyamfi, Daniel; Everitt, Hannah E; Tewfik, Ihab; Clemens, Dahn L; Patel, Vinood B

    2012-12-01

    Understanding the key aspects of the pathogenesis of alcoholic fatty liver disease particularly alterations to mitochondrial function remains to be resolved. The role of fatty acids in this regard requires further investigation due to their involvement in fatty liver disease and obesity. This study aimed to characterize the early effects of saturated and unsaturated fatty acids alone on liver mitochondrial function and during concomitant ethanol exposure using isolated liver mitochondria and VA-13 cells (Hep G2 cells that efficiently express alcohol dehydrogenase). Liver mitochondria or VA-13 cells were treated with increasing concentrations of palmitic or arachidonic acid (1 to 160 μM) for 24 h with or without 100 mM ethanol. The results showed that in isolated liver mitochondria both palmitic and arachidonic acids significantly reduced state 3 respiration in a concentration-dependent manner (P<0.001), implicating their ionophoric activities. Increased ROS production occurred in a dose-dependent manner especially in the presence of rotenone (complex I inhibitor), which was significantly more prominent in arachidonic acid at 80 μM (+970%, P<0.001) than palmitic acid (+40%, P<0.01). In VA-13 cells, ethanol alone and both fatty acids (40 μM) were able to decrease the mitochondrial membrane potential and cellular ATP levels and increase lipid formation. ROS production was significantly increased with arachidonic acid (+110%, P<0.001) exhibiting a greater effect than palmitic acid (+39%, P<0.05). While in the presence of ethanol, the drop in the mitochondrial membrane potential, cellular ATP levels, and increased lipid formation were further enhanced by both fatty acids, but with greater effect in the case of arachidonic acid, which also correlated with significant cytotoxicity (P<0.001). This study confirms the ability of fatty acids to promote mitochondrial injury in the development of alcoholic fatty liver disease.

  9. Pseudomonas aeruginosa Pyocyanin Induces Neutrophil Death via Mitochondrial Reactive Oxygen Species and Mitochondrial Acid Sphingomyelinase

    PubMed Central

    Managò, Antonella; Becker, Katrin Anne; Carpinteiro, Alexander; Wilker, Barbara; Soddemann, Matthias; Seitz, Aaron P.; Edwards, Michael J.; Grassmé, Heike

    2015-01-01

    Abstract Aims: Pulmonary infections with Pseudomonas aeruginosa are a serious clinical problem and are often lethal. Because many strains of P. aeruginosa are resistant to antibiotics, therapeutic options are limited. Neutrophils play an important role in the host's early acute defense against pulmonary P. aeruginosa. Therefore, it is important to define the mechanisms by which P. aeruginosa interacts with host cells, particularly neutrophils. Results: Here, we report that pyocyanin, a membrane-permeable pigment and toxin released by P. aeruginosa, induces the death of wild-type neutrophils; its interaction with the mitochondrial respiratory chain results in the release of reactive oxygen species (ROS), the activation of mitochondrial acid sphingomyelinase, the formation of mitochondrial ceramide, and the release of cytochrome c from mitochondria. A genetic deficiency in acid sphingomyelinase prevents both the activation of this pathway and pyocyanin-induced neutrophil death. This reduced death, on the other hand, is associated with an increase in the release of interleukin-8 from pyocyanin-activated acid sphingomyelinase-deficient neutrophils but not from wild-type cells. Innovation: These studies identified the mechanisms by which pyocyanin induces the release of mitochondrial ROS and by which ROS induce neutrophil death via mitochondrial acid sphingomyelinase. Conclusion: These findings demonstrate a novel mechanism of pyocyanin-induced death of neutrophils and show how this apoptosis balances innate immune reactions. Antioxid. Redox Signal. 22, 1097–1110. PMID:25686490

  10. Mitochondrial biology and oxidative stress in Parkinson disease pathogenesis.

    PubMed

    Henchcliffe, Claire; Beal, M Flint

    2008-11-01

    Parkinson disease (PD) is associated with progressive loss of dopaminergic neurons in the substantia nigra, as well as with more-widespread neuronal changes that cause complex and variable motor and nonmotor symptoms. Recent rapid advances in PD genetics have revealed a prominent role for mitochondrial dysfunction in the pathogenesis of the disease, and the products of several PD-associated genes, including SNCA, Parkin, PINK1, DJ-1, LRRK2 and HTR2A, show a degree of localization to the mitochondria under certain conditions. Impaired mitochondrial function is likely to increase oxidative stress and might render cells more vulnerable to this and other related processes, including excitotoxicity. The mitochondria, therefore, represent a highly promising target for the development of disease biomarkers by use of genetic, biochemical and bioimaging approaches. Novel therapeutic interventions that modify mitochondrial function are currently under development, and a large phase III clinical trial is underway to examine whether high-dose oral coenzyme Q10 will slow disease progression. In this Review, we examine evidence for the roles of mitochondrial dysfunction and increased oxidative stress in the neuronal loss that leads to PD and discuss how this knowledge might further improve patient management and aid in the development of 'mitochondrial therapy' for PD.

  11. Mitochondrial-Nuclear Communication by Prohibitin Shuttling Under Oxidative Stress

    PubMed Central

    Sripathi, Srinivas; He, Weilue; Atkinson, Cameron; Smith, Joey; Liu, Zhicong; Elledge, Beth; Jahng, Wan Jin

    2017-01-01

    Mitochondrial-nuclear communication is critical to maintain mitochondrial activity under stress conditions. Adaptation of the mitochondria-nucleus network to changes in the intracellular oxidation and reduction milieu is critical for the survival of retinal and retinal pigment epithelial (RPE) cells, in relation to their high oxygen demand and rapid metabolism. However, the generation and transmittal of mitochondrial signal to the nucleus remains elusive. Previously, our in vivo study revealed that prohibitin is up-regulated in the retina but is down-regulated in RPE in the aging and diabetic model. In this study, the functional role of prohibitin in the retina and the RPE was studied using biochemical methods, including lipid binding assay, 2D gel electrophoresis, immunocytochemistry, Western blot, and knockdown approach. Protein depletion by siRNA characterized prohibitin as an anti-apoptotic molecule in mitochondria, while lipid binding assay demonstrated subcellular communications between mitochondria and the nucleus under oxidative stress. The changes of the expressions and localization of mitochondrial prohibitin triggered by reactive oxygen species are crucial for mitochondrial integrity. We propose that prohibitin shuttles between mitochondria and the nucleus as an anti-apoptotic molecule and a transcriptional regulator under stress environment in the retina and RPE. PMID:21879722

  12. Mitochondrial dysfunction associated with nitric oxide pathways in glutamate neurotoxicity.

    PubMed

    Manucha, Walter

    Multiple mechanisms underlying glutamate-induced neurotoxicity have recently been discussed. Likewise, a clear deregulation of the mitochondrial respiratory mechanism has been described in patients with neurodegeneration, oxidative stress, and inflammation. This article highlights nitric oxide, an atypical neurotransmitter synthesized and released on demand by the post-synaptic neurons, and has many important implications for nerve cell survival and differentiation. Consequently, synaptogenesis, synapse elimination, and neurotransmitter release, are nitric oxide-modulated. Interesting, an emergent role of nitric oxide pathways has been discussed as regards neurotoxicity from glutamate-induced apoptosis. These findings suggest that nitric oxide pathways modulation could prevent oxidative damage to neurons through apoptosis inhibition. This review aims to highlight the emergent aspects of nitric oxide-mediated signaling in the brain, and how they can be related to neurotoxicity, as well as the development of neurodegenerative diseases development.

  13. Oxidized mitochondrial nucleoids released by neutrophils drive type I interferon production in human lupus

    PubMed Central

    Caielli, Simone; Athale, Shruti; Domic, Bojana; Murat, Elise; Chandra, Manjari; Banchereau, Romain; Baisch, Jeanine; Phelps, Kate; Clayton, Sandra; Gong, Mei; Wright, Tracey; Punaro, Marilynn; Palucka, Karolina; Guiducci, Cristiana; Banchereau, Jacques

    2016-01-01

    Autoantibodies against nucleic acids and excessive type I interferon (IFN) are hallmarks of human systemic lupus erythematosus (SLE). We previously reported that SLE neutrophils exposed to TLR7 agonist autoantibodies release interferogenic DNA, which we now demonstrate to be of mitochondrial origin. We further show that healthy human neutrophils do not complete mitophagy upon induction of mitochondrial damage. Rather, they extrude mitochondrial components, including DNA (mtDNA), devoid of oxidized (Ox) residues. When mtDNA undergoes oxidation, it is directly routed to lysosomes for degradation. This rerouting requires dissociation from the transcription factor A mitochondria (TFAM), a dual high-mobility group (HMG) protein involved in maintenance and compaction of the mitochondrial genome into nucleoids. Exposure of SLE neutrophils, or healthy IFN-primed neutrophils, to antiribonucleotide protein autoantibodies blocks TFAM phosphorylation, a necessary step for nucleoid dissociation. Consequently, Ox nucleoids accumulate within mitochondria and are eventually extruded as potent interferogenic complexes. In support of the in vivo relevance of this phenomenon, mitochondrial retention of Ox nucleoids is a feature of SLE blood neutrophils, and autoantibodies against Ox mtDNA are present in a fraction of patients. This pathway represents a novel therapeutic target in human SLE. PMID:27091841

  14. Mitochondrial dysfunction and oxidative damage in parkin-deficient mice.

    PubMed

    Palacino, James J; Sagi, Dijana; Goldberg, Matthew S; Krauss, Stefan; Motz, Claudia; Wacker, Maik; Klose, Joachim; Shen, Jie

    2004-04-30

    Loss-of-function mutations in parkin are the predominant cause of familial Parkinson's disease. We previously reported that parkin-/- mice exhibit nigrostriatal deficits in the absence of nigral degeneration. Parkin has been shown to function as an E3 ubiquitin ligase. Loss of parkin function, therefore, has been hypothesized to cause nigral degeneration via an aberrant accumulation of its substrates. Here we employed a proteomic approach to determine whether loss of parkin function results in alterations in abundance and/or modification of proteins in the ventral midbrain of parkin-/- mice. Two-dimensional gel electrophoresis followed by mass spectrometry revealed decreased abundance of a number of proteins involved in mitochondrial function or oxidative stress. Consistent with reductions in several subunits of complexes I and IV, functional assays showed reductions in respiratory capacity of striatal mitochondria isolated from parkin-/- mice. Electron microscopic analysis revealed no gross morphological abnormalities in striatal mitochondria of parkin-/- mice. In addition, parkin-/- mice showed a delayed rate of weight gain, suggesting broader metabolic abnormalities. Accompanying these deficits in mitochondrial function, parkin-/- mice also exhibited decreased levels of proteins involved in protection from oxidative stress. Consistent with these findings, parkin-/- mice showed decreased serum antioxidant capacity and increased protein and lipid peroxidation. The combination of proteomic, genetic, and physiological analyses reveal an essential role for parkin in the regulation of mitochondrial function and provide the first direct evidence of mitochondrial dysfunction and oxidative damage in the absence of nigral degeneration in a genetic mouse model of Parkinson's disease.

  15. Mitochondrial dysfunction and oxidative stress in aging and cancer

    PubMed Central

    Kudryavtseva, Anna V.; Krasnov, George S.; Dmitriev, Alexey A.; Alekseev, Boris Y.; Kardymon, Olga L.; Sadritdinova, Asiya F.; Fedorova, Maria S.; Pokrovsky, Anatoly V.; Melnikova, Nataliya V.; Kaprin, Andrey D.; Moskalev, Alexey A.; Snezhkina, Anastasiya V.

    2016-01-01

    Aging and cancer are the most important issues to research. The population in the world is growing older, and the incidence of cancer increases with age. There is no doubt about the linkage between aging and cancer. However, the molecular mechanisms underlying this association are still unknown. Several lines of evidence suggest that the oxidative stress as a cause and/or consequence of the mitochondrial dysfunction is one of the main drivers of these processes. Increasing ROS levels and products of the oxidative stress, which occur in aging and age-related disorders, were also found in cancer. This review focuses on the similarities between ageing-associated and cancer-associated oxidative stress and mitochondrial dysfunction as their common phenotype. PMID:27270647

  16. Mechanisms underlying skeletal muscle insulin resistance induced by fatty acids: importance of the mitochondrial function

    PubMed Central

    2012-01-01

    Insulin resistance condition is associated to the development of several syndromes, such as obesity, type 2 diabetes mellitus and metabolic syndrome. Although the factors linking insulin resistance to these syndromes are not precisely defined yet, evidence suggests that the elevated plasma free fatty acid (FFA) level plays an important role in the development of skeletal muscle insulin resistance. Accordantly, in vivo and in vitro exposure of skeletal muscle and myocytes to physiological concentrations of saturated fatty acids is associated with insulin resistance condition. Several mechanisms have been postulated to account for fatty acids-induced muscle insulin resistance, including Randle cycle, oxidative stress, inflammation and mitochondrial dysfunction. Here we reviewed experimental evidence supporting the involvement of each of these propositions in the development of skeletal muscle insulin resistance induced by saturated fatty acids and propose an integrative model placing mitochondrial dysfunction as an important and common factor to the other mechanisms. PMID:22360800

  17. The mitochondrial acyl carrier protein (ACP) coordinates mitochondrial fatty acid synthesis with iron sulfur cluster biogenesis

    PubMed Central

    Van Vranken, Jonathan G; Jeong, Mi-Young; Wei, Peng; Chen, Yu-Chan; Gygi, Steven P; Winge, Dennis R; Rutter, Jared

    2016-01-01

    Mitochondrial fatty acid synthesis (FASII) and iron sulfur cluster (FeS) biogenesis are both vital biosynthetic processes within mitochondria. In this study, we demonstrate that the mitochondrial acyl carrier protein (ACP), which has a well-known role in FASII, plays an unexpected and evolutionarily conserved role in FeS biogenesis. ACP is a stable and essential subunit of the eukaryotic FeS biogenesis complex. In the absence of ACP, the complex is destabilized resulting in a profound depletion of FeS throughout the cell. This role of ACP depends upon its covalently bound 4’-phosphopantetheine (4-PP)-conjugated acyl chain to support maximal cysteine desulfurase activity. Thus, it is likely that ACP is not simply an obligate subunit but also exploits the 4-PP-conjugated acyl chain to coordinate mitochondrial fatty acid and FeS biogenesis. DOI: http://dx.doi.org/10.7554/eLife.17828.001 PMID:27540631

  18. Mitochondrial oxidant stress in locus coeruleus is regulated by activity and nitric oxide synthase

    PubMed Central

    Sanchez–Padilla, J.; Guzman, J.N.; Ilijic, E.; Kondapalli, J.; Galtieri, D.J.; Yang, B.; Schieber, S.; Oertel, W.; Wokosin, D.; Schumacker, P. T.; Surmeier, D. J.

    2014-01-01

    Summary Loss of noradrenergic locus coeruleus (LC) neurons is a prominent feature of aging–related neurodegenerative diseases, like Parkinson’s disease (PD). The basis of this vulnerability is not understood. To explore possible physiological determinants, LC neurons were studied using electrophysiological and optical approaches in ex vivo mouse brain slices. These studies revealed that autonomous activity in LC neurons was accompanied by oscillations in dendritic Ca2+ concentration attributable to opening of L–type Ca2+ channels. This oscillation elevated mitochondrial oxidant stress and was attenuated by inhibition of nitric oxide synthase. The relationship between activity and stress was malleable, as arousal and carbon dioxide, each increased the spike rate, but differentially affected mitochondrial oxidant stress. Oxidant stress also was increased in an animal model of PD. Thus, our results point to activity–dependent Ca2+ entry and a resulting mitochondrial oxidant stress as factors contributing to the vulnerability of LC neurons. PMID:24816140

  19. Exportability of the mitochondrial oxidative phosphorylation machinery into myelin sheath.

    PubMed

    Morelli, Alessandro; Ravera, Silvia; Calzia, Daniela; Panfoli, Isabella

    2011-01-01

    White matter comprises over half of the brain, and its role in axonal survival is being reconsidered, consistently with the observation that axonal degeneration follows demyelination. The recent evidence of an extra-mitochondrial aerobic ATP production in isolated myelin vesicles, thanks to the expression therein of the mitochondrial Oxydative Phosphorylation (OXPHOS) machinery, stands in for myelin playing a functional bioenergetic role in ATP supply for the axon. The observation that subunits of the OXPHOS encoded by the mitochondrial genome are expressed in myelin, suggests that they can be the same as those of the inner mitochondrial membrane. This would mean that the OXPHOS is exportable. Here the hypothesis is exposed that the mitochondrion is the unique site of the assembly of the OXPHOS, so that this is exported to those sub cellular districts displaying high energy demand, such as myelin sheath. There the OXPHOS would display a higher efficiency in oxidative ATP production than inside the mitochondrion itself In this respect, the role of the glia in the nervous conduction is shed new light and the oligodendrocyte mitochondrial OXPHOS are hypothesized to be delivered to nascent myelin.

  20. Diseases of the human mitochondrial oxidative phosphorylation system.

    PubMed

    Montoya, Julio; López-Gallardo, Ester; Herrero-Martín, María Dolores; Martínez-Romero, Iñigo; Gómez-Durán, Aurora; Pacheu, David; Carreras, Magdalena; Díez-Sánchez, Carmen; López-Pérez, Manuel J; Ruiz-Pesini, Eduardo

    2009-01-01

    Mitochondrial diseases, or diseases of the oxidative phosphorylation system, consist of a group of disorders originated by a deficient synthesis of ATP. This system is composed of proteins codified in the two genetic systems of the cell, the nuclear and the mitochondrial genomes, and, therefore, the mode of inheritance could be either mendelian or maternal. The diseases can also appear sporadically. Due to the central role that mitochondria play in cellular physiology, these diseases are a social and health problem of great importance. They are considered rare diseases; however, together they constitute a large variety of genetic disorders. It is also believed that mitochondria are involved, directly or indirectly, in many other human diseases, mainly in age-related diseases. This review will focus mainly on describing the special characteristics of the mitochondrial genetic system and the diseases caused by mitochondrial DNA mutations. We will also note the difficulties in studying these pathologies, and the possible involvement of the genetic variability of the mitochondrial genome in the development of these diseases.

  1. Current Experience in Testing Mitochondrial Nutrients in Disorders Featuring Oxidative Stress and Mitochondrial Dysfunction: Rational Design of Chemoprevention Trials

    PubMed Central

    Pagano, Giovanni; Aiello Talamanca, Annarita; Castello, Giuseppe; Cordero, Mario D.; d’Ischia, Marco; Gadaleta, Maria Nicola; Pallardó, Federico V.; Petrović, Sandra; Tiano, Luca; Zatterale, Adriana

    2014-01-01

    An extensive number of pathologies are associated with mitochondrial dysfunction (MDF) and oxidative stress (OS). Thus, mitochondrial cofactors termed “mitochondrial nutrients” (MN), such as α-lipoic acid (ALA), Coenzyme Q10 (CoQ10), and l-carnitine (CARN) (or its derivatives) have been tested in a number of clinical trials, and this review is focused on the use of MN-based clinical trials. The papers reporting on MN-based clinical trials were retrieved in MedLine up to July 2014, and evaluated for the following endpoints: (a) treated diseases; (b) dosages, number of enrolled patients and duration of treatment; (c) trial success for each MN or MN combinations as reported by authors. The reports satisfying the above endpoints included total numbers of trials and frequencies of randomized, controlled studies, i.e., 81 trials testing ALA, 107 reports testing CoQ10, and 74 reports testing CARN, while only 7 reports were retrieved testing double MN associations, while no report was found testing a triple MN combination. A total of 28 reports tested MN associations with “classical” antioxidants, such as antioxidant nutrients or drugs. Combinations of MN showed better outcomes than individual MN, suggesting forthcoming clinical studies. The criteria in study design and monitoring MN-based clinical trials are discussed. PMID:25380523

  2. Nitro-Arachidonic Acid Prevents Angiotensin II-Induced Mitochondrial Dysfunction in a Cell Line of Kidney Proximal Tubular Cells.

    PubMed

    Sánchez-Calvo, Beatriz; Cassina, Adriana; Rios, Natalia; Peluffo, Gonzalo; Boggia, José; Radi, Rafael; Rubbo, Homero; Trostchansky, Andres

    2016-01-01

    Nitro-arachidonic acid (NO2-AA) is a cell signaling nitroalkene that exerts anti-inflammatory activities during macrophage activation. While angiotensin II (ANG II) produces an increase in reactive oxygen species (ROS) production and mitochondrial dysfunction in renal tubular cells, little is known regarding the potential protective effects of NO2-AA in ANG II-mediated kidney injury. As such, this study examines the impact of NO2-AA on ANG II-induced mitochondrial dysfunction in an immortalized renal proximal tubule cell line (HK-2 cells). Treatment of HK-2 cells with ANG II increases the production of superoxide (O2●-), nitric oxide (●NO), inducible nitric oxide synthase (NOS2) expression, peroxynitrite (ONOO-) and mitochondrial dysfunction. Using high-resolution respirometry, it was observed that the presence of NO2-AA prevented ANG II-mediated mitochondrial dysfunction. Attempting to address mechanism, we treated isolated rat kidney mitochondria with ONOO-, a key mediator of ANG II-induced mitochondrial damage, in the presence or absence of NO2-AA. Whereas the activity of succinate dehydrogenase (SDH) and ATP synthase (ATPase) were diminished upon exposure to ONOO-, they were restored by pre-incubating the mitochondria with NO2-AA. Moreover, NO2-AA prevents oxidation and nitration of mitochondrial proteins. Combined, these data demonstrate that ANG II-mediated oxidative damage and mitochondrial dysfunction is abrogated by NO2-AA, identifying this compound as a promising pharmacological tool to prevent ANG II-induced renal disease.

  3. Treatment of Fatty Acid Oxidation Disorders

    MedlinePlus

    ... of fatty acid oxidation disorders Treatment of fatty acid oxidation disorders E-mail to a friend Please ... page It's been added to your dashboard . Fatty acid oxidation disorders are rare health conditions that affect ...

  4. Exercise increases mitochondrial glutamate oxidation in the mouse cerebral cortex.

    PubMed

    Herbst, Eric A F; Holloway, Graham P

    2016-07-01

    The present study investigated the impact of acute exercise on stimulating mitochondrial respiratory function in mouse cerebral cortex. Where pyruvate-stimulated respiration was not affected by acute exercise, glutamate respiration was enhanced following the exercise bout. Additional assessment revealed that this affect was dependent on the presence of malate and did not occur when substituting glutamine for glutamate. As such, our results suggest that glutamate oxidation is enhanced with acute exercise through activation of the malate-aspartate shuttle.

  5. Apricot Melanoidins Prevent Oxidative Endothelial Cell Death by Counteracting Mitochondrial Oxidation and Membrane Depolarization

    PubMed Central

    Giordo, Roberta; Emanueli, Costanza; Sanguinetti, Anna Maria; Piscopo, Amalia; Poiana, Marco; Capobianco, Giampiero; Piga, Antonio; Pintus, Gianfranco

    2012-01-01

    The cardiovascular benefits associated with diets rich in fruit and vegetables are thought to be due to phytochemicals contained in fresh plant material. However, whether processed plant foods provide the same benefits as unprocessed ones is an open question. Melanoidins from heat-processed apricots were isolated and their presence confirmed by colorimetric analysis and browning index. Oxidative injury of endothelial cells (ECs) is the key step for the onset and progression of cardiovascular diseases (CVD), therefore the potential protective effect of apricot melanoidins on hydrogen peroxide-induced oxidative mitochondrial damage and cell death was explored in human ECs. The redox state of cytoplasmic and mitochondrial compartments was detected by using the redox-sensitive, fluorescent protein (roGFP), while the mitochondrial membrane potential (MMP) was assessed with the fluorescent dye, JC-1. ECs exposure to hydrogen peroxide, dose-dependently induced mitochondrial and cytoplasmic oxidation. Additionally detected hydrogen peroxide-induced phenomena were MMP dissipation and ECs death. Pretreatment of ECs with apricot melanoidins, significantly counteracted and ultimately abolished hydrogen peroxide-induced intracellular oxidation, mitochondrial depolarization and cell death. In this regard, our current results clearly indicate that melanoidins derived from heat-processed apricots, protect human ECs against oxidative stress. PMID:23144984

  6. Ascorbic acid extends replicative life span of human embryonic fibroblast by reducing DNA and mitochondrial damages.

    PubMed

    Hwang, Won-Sang; Park, Seong-Hoon; Kim, Hyun-Seok; Kang, Hong-Jun; Kim, Min-Ju; Oh, Soo-Jin; Park, Jae-Bong; Kim, Jaebong; Kim, Sung Chan; Lee, Jae-Yong

    2007-01-01

    Ascorbic acid has been reported to extend replicative life span of human embryonic fibroblast (HEF). Since the detailed molecular mechanism of this phenomenon has not been investigated, we attempted to elucidate. Continuous treatment of HEF cells with ascorbic acid (at 200 microM) from 40 population doubling (PD) increased maximum PD numbers by 18% and lowered SA-beta-gal positive staining, an aging marker, by 2.3 folds, indicating that ascorbic acid extends replicative life span of HEF cells. Ascorbic acid treatment lowered DCFH by about 7 folds and Rho123 by about 70%, suggesting that ascorbic acid dramatically decreased ROS formation. Ascorbic acid also increased aconitase activity, a marker of mitochondrial aging, by 41%, indicating that ascorbic acid treatment restores age-related decline of mitochondrial function. Cell cycle analysis by flow cytometry revealed that ascorbic acid treatment decreased G1 population up to 12%. Further western blot analysis showed that ascorbic acid treatment decreased levels of p53, phospho-p53 at ser 15, and p21, indicating that ascorbic acid relieved senescence-related G1 arrest. Analysis of AP (apurinic/apyrimidinic) sites showed that ascorbic acid treatment decreased AP site formation by 35%. We also tested the effect of hydrogen peroxide treatment, as an additional oxidative stress. Continuous treatment of 20 microM of hydrogen peroxide from PD 40 of HEF cells resulted in premature senescence due to increased ROS level, and increased AP sites. Taken together, the results suggest that ascorbic acid extends replicative life span of HEF cells by reducing mitochondrial and DNA damages through lowering cellular ROS.

  7. Mitochondrial Oxidative Stress, Mitochondrial DNA Damage and Their Role in Age-Related Vascular Dysfunction

    PubMed Central

    Mikhed, Yuliya; Daiber, Andreas; Steven, Sebastian

    2015-01-01

    The prevalence of cardiovascular diseases is significantly increased in the older population. Risk factors and predictors of future cardiovascular events such as hypertension, atherosclerosis, or diabetes are observed with higher frequency in elderly individuals. A major determinant of vascular aging is endothelial dysfunction, characterized by impaired endothelium-dependent signaling processes. Increased production of reactive oxygen species (ROS) leads to oxidative stress, loss of nitric oxide (•NO) signaling, loss of endothelial barrier function and infiltration of leukocytes to the vascular wall, explaining the low-grade inflammation characteristic for the aged vasculature. We here discuss the importance of different sources of ROS for vascular aging and their contribution to the increased cardiovascular risk in the elderly population with special emphasis on mitochondrial ROS formation and oxidative damage of mitochondrial DNA. Also the interaction (crosstalk) of mitochondria with nicotinamide adenosine dinucleotide phosphate (NADPH) oxidases is highlighted. Current concepts of vascular aging, consequences for the development of cardiovascular events and the particular role of ROS are evaluated on the basis of cell culture experiments, animal studies and clinical trials. Present data point to a more important role of oxidative stress for the maximal healthspan (healthy aging) than for the maximal lifespan. PMID:26184181

  8. Curcumin alleviates oxidative stress and mitochondrial dysfunction in astrocytes.

    PubMed

    Daverey, Amita; Agrawal, Sandeep K

    2016-10-01

    Oxidative stress plays a critical role in various neurodegenerative diseases, thus alleviating oxidative stress is a potential strategy for therapeutic intervention and/or prevention of neurodegenerative diseases. In the present study, alleviation of oxidative stress through curcumin is investigated in A172 (human glioblastoma cell line) and HA-sp (human astrocytes cell line derived from the spinal cord) astrocytes. H2O2 was used to induce oxidative stress in astrocytes (A172 and HA-sp). Data show that H2O2 induces activation of astrocytes in dose- and time-dependent manner as evident by increased expression of GFAP in A172 and HA-sp cells after 24 and 12h respectively. An upregulation of Prdx6 was also observed in A172 and HA-sp cells after 24h of H2O2 treatment as compared to untreated control. Our data also showed that curcumin inhibits oxidative stress-induced cytoskeleton disarrangement, and impedes the activation of astrocytes by inhibiting upregulation of GFAP, vimentin and Prdx6. In addition, we observed an inhibition of oxidative stress-induced inflammation, apoptosis and mitochondria fragmentation after curcumin treatment. Therefore, our results suggest that curcumin not only protects astrocytes from H2O2-induced oxidative stress but also reverses the mitochondrial damage and dysfunction induced by oxidative stress. This study also provides evidence for protective role of curcumin on astrocytes by showing its effects on attenuating reactive astrogliosis and inhibiting apoptosis.

  9. Metabolic rewiring in cancer cells overexpressing the glucocorticoid-induced leucine zipper protein (GILZ): Activation of mitochondrial oxidative phosphorylation and sensitization to oxidative cell death induced by mitochondrial targeted drugs.

    PubMed

    André, Fanny; Trinh, Anne; Balayssac, Stéphane; Maboudou, Patrice; Dekiouk, Salim; Malet-Martino, Myriam; Quesnel, Bruno; Idziorek, Thierry; Kluza, Jérome; Marchetti, Philippe

    2017-04-01

    Cancer cell metabolism is largely controlled by oncogenic signals and nutrient availability. Here, we highlighted that the glucocorticoid-induced leucine zipper (GILZ), an intracellular protein influencing many signaling pathways, reprograms cancer cell metabolism to promote proliferation. We provided evidence that GILZ overexpression induced a significant increase of mitochondrial oxidative phosphorylation as evidenced by the augmentation in basal respiration, ATP-linked respiration as well as respiratory capacity. Pharmacological inhibition of glucose, glutamine and fatty acid oxidation reduced the activation of GILZ-induced mitochondrial oxidative phosphorylation. At glycolysis level, GILZ-overexpressing cells enhanced the expression of glucose transporters in their plasmatic membrane and showed higher glycolytic reserve. (1)H NMR metabolites quantification showed an up-regulation of amino acid biosynthesis. The GILZ-induced metabolic reprograming is present in various cancer cell lines regardless of their driver mutations status and is associated with higher proliferation rates persisting under metabolic stress conditions. Interestingly, high levels of OXPHOS made GILZ-overexpressing cells vulnerable to cell death induced by mitochondrial pro-oxidants. Altogether, these data indicate that GILZ reprograms cancer metabolism towards mitochondrial OXPHOS and sensitizes cancer cells to mitochondria-targeted drugs with pro-oxidant activities.

  10. Neuropsychological Outcomes in Fatty Acid Oxidation Disorders: 85 Cases Detected by Newborn Screening

    ERIC Educational Resources Information Center

    Waisbren, Susan E.; Landau, Yuval; Wilson, Jenna; Vockley, Jerry

    2013-01-01

    Mitochondrial fatty acid oxidation disorders include conditions in which the transport of activated acyl-Coenzyme A (CoA) into the mitochondria or utilization of these substrates is disrupted or blocked. This results in a deficit in the conversion of fat into energy. Most patients with fatty acid oxidation defects are now identified through…

  11. Aging-induced alterations in gene transcripts and functional activity of mitochondrial oxidative phosphorylation complexes in the heart.

    PubMed

    Preston, Claudia C; Oberlin, Andrew S; Holmuhamedov, Ekhson L; Gupta, Anu; Sagar, Sandeep; Syed, Rashad H Khazi; Siddiqui, Sabeeh A; Raghavakaimal, Sreekumar; Terzic, Andre; Jahangir, Arshad

    2008-06-01

    Aging is associated with progressive decline in energetic reserves compromising cardiac performance and tolerance to injury. Although deviations in mitochondrial functions have been documented in senescent heart, the molecular bases for the decline in energy metabolism are only partially understood. Here, high-throughput transcription profiles of genes coding for mitochondrial proteins in ventricles from adult (6-months) and aged (24-months) rats were compared using microarrays. Out of 614 genes encoding for mitochondrial proteins, 94 were differentially expressed with 95% downregulated in the aged. The majority of changes affected genes coding for proteins involved in oxidative phosphorylation (39), substrate metabolism (14) and tricarboxylic acid cycle (6). Compared to adult, gene expression changes in aged hearts translated into a reduced mitochondrial functional capacity, with decreased NADH-dehydrogenase and F(0)F(1) ATPase complex activities and capacity for oxygen-utilization and ATP synthesis. Expression of genes coding for transcription co-activator factors involved in the regulation of mitochondrial metabolism and biogenesis were downregulated in aged ventricles without reduction in mitochondrial density. Thus, aging induces a selective decline in activities of oxidative phosphorylation complexes I and V within a broader transcriptional downregulation of mitochondrial genes, providing a substrate for reduced energetic efficiency associated with senescence.

  12. Aging-Induced Alterations in Gene Transcripts and Functional Activity of Mitochondrial Oxidative Phosphorylation Complexes in the Heart

    PubMed Central

    Preston, Claudia C.; Oberlin, Andrew S.; Holmuhamedov, Ekhson L.; Gupta, Anu; Sagar, Sandeep; Khazi Syed, Rashad H.; Siddiqui, Sabeeh; Raghavakaimal, Sreekumar; Terzic, Andre; Jahangir, Arshad

    2008-01-01

    Aging is associated with progressive decline in energetic reserves compromising cardiac performance and tolerance to injury. Although deviations in mitochondrial functions have been documented in senescent heart, the molecular bases for the decline in energy metabolism are only partially understood. Here, high-throughput transcription profiles of genes coding for mitochondrial proteins in ventricles from adult (6-months) and aged (24-months) rats were compared using microarrays. Out of 614 genes encoding for mitochondrial proteins, 94 were differentially expressed with 95% downregulated in the aged. The majority of changes affected genes coding for proteins involved in oxidative phosphorylation (39), substrate metabolism (14) and tricarboxylic acid cycle (6). Compared to adult, gene expression changes in aged hearts translated into a reduced mitochondrial functional capacity, with decreased NADH-dehydrogenase and F0F1-ATPase complex activities and capacity for oxygen-utilization and ATP synthesis. Expression of genes coding for transcription co-activator factors involved in the regulation of mitochondrial metabolism and biogenesis were downregulated in aged ventricles without reduction in mitochondrial density. Thus, aging induces a selective decline in activities of oxidative phosphorylation complexes I and V within a broader transcriptional downregulation of mitochondrial genes, providing a substrate for reduced energetic efficiency associated with senescence. PMID:18400259

  13. Diabetes and mitochondrial function: Role of hyperglycemia and oxidative stress

    SciTech Connect

    Rolo, Anabela P.; Palmeira, Carlos M. . E-mail: palmeira@ci.uc.pt

    2006-04-15

    Hyperglycemia resulting from uncontrolled glucose regulation is widely recognized as the causal link between diabetes and diabetic complications. Four major molecular mechanisms have been implicated in hyperglycemia-induced tissue damage: activation of protein kinase C (PKC) isoforms via de novo synthesis of the lipid second messenger diacylglycerol (DAG), increased hexosamine pathway flux, increased advanced glycation end product (AGE) formation, and increased polyol pathway flux. Hyperglycemia-induced overproduction of superoxide is the causal link between high glucose and the pathways responsible for hyperglycemic damage. In fact, diabetes is typically accompanied by increased production of free radicals and/or impaired antioxidant defense capabilities, indicating a central contribution for reactive oxygen species (ROS) in the onset, progression, and pathological consequences of diabetes. Besides oxidative stress, a growing body of evidence has demonstrated a link between various disturbances in mitochondrial functioning and type 2 diabetes. Mutations in mitochondrial DNA (mtDNA) and decreases in mtDNA copy number have been linked to the pathogenesis of type 2 diabetes. The study of the relationship of mtDNA to type 2 diabetes has revealed the influence of the mitochondria on nuclear-encoded glucose transporters, glucose-stimulated insulin secretion, and nuclear-encoded uncoupling proteins (UCPs) in {beta}-cell glucose toxicity. This review focuses on a range of mitochondrial factors important in the pathogenesis of diabetes. We review the published literature regarding the direct effects of hyperglycemia on mitochondrial function and suggest the possibility of regulation of mitochondrial function at a transcriptional level in response to hyperglycemia. The main goal of this review is to include a fresh consideration of pathways involved in hyperglycemia-induced diabetic complications.

  14. Some aspects of the mitochondrial oxidative metabolism in human atrial tissue during cardiopulmonary by-pass.

    PubMed

    Corbucci, G G; Gasparetto, A; Antonelli, M; Bufi, M; De Blasi, R A

    1987-01-01

    Following previous research on the hypoxic cell in human circulatory shock, the present work has investigated some mitochondrial oxidative aspects in atrial biopsies taken during cardiopulmonary by-pass. Cardioplegic solution and hypothermia were administered to 10 patients and the atrial samples were collected before and after aortic clamping. The results show a cellular protective effect of cardioplegia and hypothermia on the electron-transport chain, even if the enzymes with high KmO2 appear to be more sensitive to ischaemia. The results suggest a metabolic injury rather than an oxidative damage due to the induced ischaemia, alterations to fatty-acid beta-oxidation being especially notable. Because of the unchanged oxidative capacities, the oxyradical generation and the peroxidative damage appear to be irrelevant in the ischaemic period and during the course of reperfusion. Further studies are needed to elucidate the metabolic damage and the therapeutic implications due to the induced ischaemia in the myocardial cell during the aortic clamping.

  15. Antioxidant properties of Neu2000 on mitochondrial free radicals and oxidative damage.

    PubMed

    Visavadiya, Nishant P; McEwen, Melanie L; Pandya, Jignesh D; Sullivan, Patrick G; Gwag, Byoung Joo; Springer, Joe E

    2013-03-01

    Neu2000 [2-hydroxy-5-(2,3,5,6-tetrafluoro-4 trifluoromethylbenzylamino) benzoic acid] is a dual-acting neuroprotective agent that functions both as a noncompetitive N-methyl-d-aspartate (NMDA) receptor antagonist and a free radical scavenger. In the present study, we investigated the scavenging activity of Neu2000 on various classes of reactive oxygen species and reactive nitrogen species (ROS/RNS) as well as its efficacy for reducing free radicals and oxidative stress/damage induced in spinal cord mitochondrial preparations. Neu2000 exerted scavenging activity against superoxide, nitric oxide, and hydroxyl radicals, and efficiently scavenged peroxynitrite. In the mitochondrial studies, Neu2000 markedly inhibited ROS/RNS and hydrogen peroxide levels following antimycin treatment. In addition, Neu2000 effectively scavenged hydroxyl radicals generated by iron(III)-ascorbate, reduced protein carbonyl formation mediated by hydroxyl radicals and peroxynitrite, and prevented glutathione oxidation caused by tert-butyl hydroperoxide in isolated mitochondria. Interestingly, incubation of isolated mitochondria with Neu2000 followed by centrifugation and removal of the supernatant also resulted in a concentration-dependent decrease in lipid peroxidation. This observation suggests that Neu2000 enters mitochondria to target free radicals or indirectly affects mitochondrial function in a manner that promotes antioxidant activity. The results of the present study demonstrate that Neu2000 possesses potent in vitro antioxidant activity due, most likely, to its active phenoxy group.

  16. Impairment of mitochondrial β-oxidation in rats under cold-hypoxic environment

    NASA Astrophysics Data System (ADS)

    Dutta, Arkadeb; Vats, Praveen; Singh, Vijay K.; Sharma, Yogendra K.; Singh, Som N.; Singh, Shashi B.

    2009-09-01

    Mitochondrial ß-oxidation of fatty acid provides a major source of energy in mammals. High altitude (HA), characterized by hypobaric hypoxia and low ambient temperatures, causes alteration in metabolic homeostasis. Several studies have depicted that hypoxic exposure in small mammals causes hypothermia due to hypometabolic state. Moreover, cold exposure along with hypoxia reduces hypoxia tolerance in animals. The present study investigated the rate of β-oxidation and key enzymes, carnitine palmitoyl transferase-I (CPT-I) and hydroxyacyl CoA dehydrogenase (HAD), in rats exposed to cold-hypobaric hypoxic environment. Male Sprague Dawley rats (190-220 g) were randomly divided into eight groups ( n = 6 rats in each group): 1 day hypoxia (H1); 7 days hypoxia (H7); 1 day cold (C1); 7 days cold (C7); 1 day cold-hypoxia (CH1); 7 days cold-hypoxia (CH7) exposed; and unexposed control for 1 and 7 days (UC1 and UC7). After exposure, animals were anaesthetized with ketamine (50 mg/kg body weight) and xylazine (10 mg/kg body weight) intraperitonialy and sacrificed. Mitochondrial CPT-I, HAD, 14C-palmitate oxidation in gastrocnemius muscle and liver, and plasma leptin were measured. Mitochondrial CPT-I was significantly reduced in muscle and liver in CH1 and CH7 as compared to respective controls. HAD activity was significantly reduced in H1 and CH7, and in H1, H7, CH1, and CH7 as compared to unexposed controls in muscle and liver, respectively. A concomitant decrease in 14C-palmitate oxidation was found. Significant reduction in plasma leptin in hypoxia and cold-hypoxia suggested hypometabolic state. It can be concluded that ß-oxidation of fatty acids is reduced in rats exposed to cold-hypoxic environment due to the persisting hypometabolic state in cold-hypoxia exposure.

  17. Inhibitors of mitochondrial fission as a therapeutic strategy for diseases with oxidative stress and mitochondrial dysfunction.

    PubMed

    Reddy, P Hemachandra

    2014-01-01

    Mitochondria are essential cytoplasmic organelles, critical for cell survival and death. Recent mitochondrial research revealed that mitochondrial dynamics-the balance of fission and fusion in normal mitochondrial dynamics--is an important cellular mechanism in eukaryotic cell and is involved in the maintenance of mitochondrial morphology, structure, number, distribution, and function. Research into mitochondria and cell function has revealed that mitochondrial dynamics is impaired in a large number of aging and neurodegenerative diseases, and in several inherited mitochondrial diseases, and that this impairment involves excessive mitochondrial fission, resulting in mitochondrial structural changes and dysfunction, and cell damage. Attempts have been made to develop molecules to reduce mitochondrial fission while maintaining normal mitochondrial fusion and function in those diseases that involve excessive mitochondrial fission. This review article discusses mechanisms of mitochondrial fission in normal and diseased states of mammalian cells and discusses research aimed at developing therapies, such as Mdivi, Dynasore and P110, to prevent or to inhibit excessive mitochondrial fission.

  18. Status Epilepticus in Immature Rats Is Associated with Oxidative Stress and Mitochondrial Dysfunction

    PubMed Central

    Folbergrová, Jaroslava; Ješina, Pavel; Kubová, Hana; Druga, Rastislav; Otáhal, Jakub

    2016-01-01

    Epilepsy is a neurologic disorder, particularly frequent in infants and children where it can lead to serious consequences later in life. Oxidative stress and mitochondrial dysfunction are implicated in the pathogenesis of many neurological disorders including epilepsy in adults. However, their role in immature epileptic brain is unclear since there have been two contrary opinions: oxidative stress is age-dependent and does not occur in immature brain during status epilepticus (SE) and, on the other hand, evidence of oxidative stress in immature brain during a specific model of SE. To solve this dilemma, we have decided to investigate oxidative stress following SE induced in immature 12-day-old rats by three substances with a different mechanism of action, namely 4-aminopyridine, LiCl-pilocarpine or kainic acid. Fluoro-Jade-B staining revealed mild brain damage especially in hippocampus and thalamus in each of the tested models. Decrease of glucose and glycogen with parallel rises of lactate clearly indicate high rate of glycolysis, which was apparently not sufficient in 4-AP and Li-Pilo status, as evident from the decreases of PCr levels. Hydroethidium method revealed significantly higher levels of superoxide anion (by ∼60%) in the hippocampus, cerebral cortex and thalamus of immature rats during status. SE lead to mitochondrial dysfunction with a specific pronounced decrease of complex I activity that persisted for a long period of survival. Complexes II and IV activities remained in the control range. Antioxidant treatment with SOD mimetic MnTMPYP or peroxynitrite scavenger FeTPPS significantly attenuated oxidative stress and inhibition of complex I activity. These findings bring evidence that oxidative stress and mitochondrial dysfunction are age and model independent, and may thus be considered a general phenomenon. They can have a clinical relevance for a novel approach to the treatment of epilepsy, allowing to target the mechanisms which play a crucial or

  19. Mitochondrial dysfunction and death in motor neurons exposed to the glutathione-depleting agent ethacrynic acid.

    PubMed

    Rizzardini, M; Lupi, M; Bernasconi, S; Mangolini, A; Cantoni, L

    2003-03-15

    This study investigated the mechanisms of toxicity of glutathione (GSH) depletion in one cell type, the motor neuron. Ethacrynic acid (EA) (100 microM) was added to immortalized mouse motor neurons (NSC-34) to deplete both cytosolic and mitochondrial glutathione rapidly. This caused a drop in GSH to 25% of the initial level in 1 h and complete loss in 4 h. This effect was accompanied by enhanced generation of reactive oxygen species (ROS) with a peak after 2 h of exposure, and by signs of mitochondrial dysfunction such as a decrease in 3-(4,5-dimethyl-2-thiazoyl)-2,5-diphenyltetrazolium bromide (MTT) (30% less after 4 h). The increase in ROS and the MTT reduction were both EA concentration-dependent. Expression of heme oxygenase-1 (HO-1), a marker of oxidative stress, also increased. The mitochondrial damage was monitored by measuring the mitochondrial membrane potential (MMP) from the uptake of rhodamine 123 into mitochondria. MMP dropped (20%) after only 1 h exposure to EA, and slowly continued to decline until 3 h, with a steep drop at 5 h (50% decrease), i.e. after the complete GSH loss. Quantification of DNA fragmentation by the TUNEL technique showed that the proportion of cells with fragmented nuclei rose from 10% after 5 h EA exposure to about 65% at 18 h. These results indicate that EA-induced GSH depletion rapidly impairs the mitochondrial function of motor neurons, and this precedes cell death. This experimental model of oxidative toxicity could be useful to study mechanisms of diseases like spinal cord injury (SCI) and amyotrophic lateral sclerosis (ALS), where motor neurons are the vulnerable population and oxidative stress has a pathogenic role.

  20. Avocado Oil Improves Mitochondrial Function and Decreases Oxidative Stress in Brain of Diabetic Rats.

    PubMed

    Ortiz-Avila, Omar; Esquivel-Martínez, Mauricio; Olmos-Orizaba, Berenice Eridani; Saavedra-Molina, Alfredo; Rodriguez-Orozco, Alain R; Cortés-Rojo, Christian

    2015-01-01

    Diabetic encephalopathy is a diabetic complication related to the metabolic alterations featuring diabetes. Diabetes is characterized by increased lipid peroxidation, altered glutathione redox status, exacerbated levels of ROS, and mitochondrial dysfunction. Although the pathophysiology of diabetic encephalopathy remains to be clarified, oxidative stress and mitochondrial dysfunction play a crucial role in the pathogenesis of chronic diabetic complications. Taking this into consideration, the aim of this work was to evaluate the effects of 90-day avocado oil intake in brain mitochondrial function and oxidative status in streptozotocin-induced diabetic rats (STZ rats). Avocado oil improves brain mitochondrial function in diabetic rats preventing impairment of mitochondrial respiration and mitochondrial membrane potential (ΔΨ m ), besides increasing complex III activity. Avocado oil also decreased ROS levels and lipid peroxidation and improved the GSH/GSSG ratio as well. These results demonstrate that avocado oil supplementation prevents brain mitochondrial dysfunction induced by diabetes in association with decreased oxidative stress.

  1. Avocado Oil Improves Mitochondrial Function and Decreases Oxidative Stress in Brain of Diabetic Rats

    PubMed Central

    Ortiz-Avila, Omar; Esquivel-Martínez, Mauricio; Olmos-Orizaba, Berenice Eridani; Saavedra-Molina, Alfredo; Rodriguez-Orozco, Alain R.; Cortés-Rojo, Christian

    2015-01-01

    Diabetic encephalopathy is a diabetic complication related to the metabolic alterations featuring diabetes. Diabetes is characterized by increased lipid peroxidation, altered glutathione redox status, exacerbated levels of ROS, and mitochondrial dysfunction. Although the pathophysiology of diabetic encephalopathy remains to be clarified, oxidative stress and mitochondrial dysfunction play a crucial role in the pathogenesis of chronic diabetic complications. Taking this into consideration, the aim of this work was to evaluate the effects of 90-day avocado oil intake in brain mitochondrial function and oxidative status in streptozotocin-induced diabetic rats (STZ rats). Avocado oil improves brain mitochondrial function in diabetic rats preventing impairment of mitochondrial respiration and mitochondrial membrane potential (ΔΨm), besides increasing complex III activity. Avocado oil also decreased ROS levels and lipid peroxidation and improved the GSH/GSSG ratio as well. These results demonstrate that avocado oil supplementation prevents brain mitochondrial dysfunction induced by diabetes in association with decreased oxidative stress. PMID:26180820

  2. Oxidative stress, mitochondrial and proteostasis malfunction in adrenoleukodystrophy: A paradigm for axonal degeneration.

    PubMed

    Fourcade, Stéphane; Ferrer, Isidre; Pujol, Aurora

    2015-11-01

    Peroxisomal and mitochondrial malfunction, which are highly intertwined through redox regulation, in combination with defective proteostasis, are hallmarks of the most prevalent multifactorial neurodegenerative diseases-including Alzheimer's (AD) and Parkinson's disease (PD)-and of the aging process, and are also found in inherited conditions. Here we review the interplay between oxidative stress and axonal degeneration, taking as groundwork recent findings on pathomechanisms of the peroxisomal neurometabolic disease adrenoleukodystrophy (X-ALD). We explore the impact of chronic redox imbalance caused by the excess of very long-chain fatty acids (VLCFA) on mitochondrial respiration and biogenesis, and discuss how this impairs protein quality control mechanisms essential for neural cell survival, such as the proteasome and autophagy systems. As consequence, prime molecular targets in the pathogenetic cascade emerge, such as the SIRT1/PGC-1α axis of mitochondrial biogenesis, and the inhibitor of autophagy mTOR. Thus, we propose that mitochondria-targeted antioxidants; mitochondrial biogenesis boosters such as the antidiabetic pioglitazone and the SIRT1 ligand resveratrol; and the autophagy activator temsirolimus, a derivative of the mTOR inhibitor rapamycin, hold promise as disease-modifying therapies for X-ALD.

  3. Nitric oxide-mediated mitochondrial damage in the brain: mechanisms and implications for neurodegenerative diseases.

    PubMed

    Bolaños, J P; Almeida, A; Stewart, V; Peuchen, S; Land, J M; Clark, J B; Heales, S J

    1997-06-01

    Within the CNS and under normal conditions, nitric oxide (.NO) appears to be an important physiological signalling molecule. Its ability to increase cyclic GMP concentration suggests that .NO is implicated in the regulation of important metabolic pathways in the brain. Under certain circumstances .NO synthesis may be excessive and .NO may become neurotoxic. Excessive glutamate-receptor stimulation may lead to neuronal death through a mechanism implicating synthesis of both .NO and superoxide (O2.-) and hence peroxynitrite (ONOO-) formation. In response to lipopolysaccharide and cytokines, glial cells may also be induced to synthesize large amounts of .NO, which may be deleterious to the neighbouring neurones and oligodendrocytes. The precise mechanism of .NO neurotoxicity is not fully understood. One possibility is that it may involve neuronal energy deficiency. This may occur by ONOO- interfering with key enzymes of the tricarboxylic acid cycle, the mitochondrial respiratory chain, mitochondrial calcium metabolism, or DNA damage with subsequent activation of the energy-consuming pathway involving poly(ADP-ribose) synthetase. Possible mechanisms whereby ONOO- impairs the mitochondrial respiratory chain and the relevance for neurotoxicity are discussed. The intracellular content of reduced glutathione also appears important in determining the sensitivity of cells to ONOO- production. It is concluded that neurotoxicity elicited by excessive .NO production may be mediated by mitochondrial dysfunction leading to an energy deficiency state.

  4. NMR Metabolomics Show Evidence for Mitochondrial Oxidative Stress in a Mouse Model of Polycystic Ovary Syndrome.

    PubMed

    Selen, Ebru Selin; Bolandnazar, Zeinab; Tonelli, Marco; Bütz, Daniel E; Haviland, Julia A; Porter, Warren P; Assadi-Porter, Fariba M

    2015-08-07

    Polycystic ovary syndrome (PCOS) is associated with metabolic and endocrine disorders in women of reproductive age. The etiology of PCOS is still unknown. Mice prenatally treated with glucocorticoids exhibit metabolic disturbances that are similar to those seen in women with PCOS. We used an untargeted nuclear magnetic resonance (NMR)-based metabolomics approach to understand the metabolic changes occurring in the plasma and kidney over time in female glucocorticoid-treated (GC-treated) mice. There are significant changes in plasma amino acid levels (valine, tyrosine, and proline) and their intermediates (2-hydroxybutyrate, 4-aminobutyrate, and taurine), whereas in kidneys, the TCA cycle metabolism (citrate, fumarate, and succinate) and the pentose phosphate (PP) pathway products (inosine and uracil) are significantly altered (p < 0.05) from 8 to 16 weeks of age. Levels of NADH, NAD(+), NAD(+)/NADH, and NADH redox in kidneys indicate increased mitochondrial oxidative stress from 8 to 16 weeks in GC-treated mice. These results indicate that altered metabolic substrates in the plasma and kidneys of treated mice are associated with altered amino acid metabolism, increased cytoplasmic PP, and increased mitochondrial activity, leading to a more oxidized state. This study identifies biomarkers associated with metabolic dysfunction in kidney mitochondria of a prenatal gluococorticoid-treated mouse model of PCOS that may be used as early predictive biomarkers of oxidative stress in the PCOS metabolic disorder in women.

  5. Nutritional and Hormonal Regulation of Citrate and Carnitine/Acylcarnitine Transporters: Two Mitochondrial Carriers Involved in Fatty Acid Metabolism

    PubMed Central

    Giudetti, Anna M.; Stanca, Eleonora; Siculella, Luisa; Gnoni, Gabriele V.; Damiano, Fabrizio

    2016-01-01

    The transport of solutes across the inner mitochondrial membrane is catalyzed by a family of nuclear-encoded membrane-embedded proteins called mitochondrial carriers (MCs). The citrate carrier (CiC) and the carnitine/acylcarnitine transporter (CACT) are two members of the MCs family involved in fatty acid metabolism. By conveying acetyl-coenzyme A, in the form of citrate, from the mitochondria to the cytosol, CiC contributes to fatty acid and cholesterol synthesis; CACT allows fatty acid oxidation, transporting cytosolic fatty acids, in the form of acylcarnitines, into the mitochondrial matrix. Fatty acid synthesis and oxidation are inversely regulated so that when fatty acid synthesis is activated, the catabolism of fatty acids is turned-off. Malonyl-CoA, produced by acetyl-coenzyme A carboxylase, a key enzyme of cytosolic fatty acid synthesis, represents a regulator of both metabolic pathways. CiC and CACT activity and expression are regulated by different nutritional and hormonal conditions. Defects in the corresponding genes have been directly linked to various human diseases. This review will assess the current understanding of CiC and CACT regulation; underlining their roles in physio-pathological conditions. Emphasis will be placed on the molecular basis of the regulation of CiC and CACT associated with fatty acid metabolism. PMID:27231907

  6. Induction of mitochondrial biogenesis and respiration is associated with mTOR regulation in hepatocytes of rats treated with the pan-PPAR activator tetradecylthioacetic acid (TTA)

    SciTech Connect

    Hagland, Hanne R.; Nilsson, Linn I.H.; Burri, Lena; Nikolaisen, Julie; Berge, Rolf K.; Tronstad, Karl J.

    2013-01-11

    Highlights: Black-Right-Pointing-Pointer We investigated mechanisms of mitochondrial regulation in rat hepatocytes. Black-Right-Pointing-Pointer Tetradecylthioacetic acid (TTA) was employed to activate mitochondrial oxidation. Black-Right-Pointing-Pointer Mitochondrial biogenesis and respiration were induced. Black-Right-Pointing-Pointer It was confirmed that PPAR target genes were induced. Black-Right-Pointing-Pointer The mechanism involved activation mTOR. -- Abstract: The hypolipidemic effect of peroxisome proliferator-activated receptor (PPAR) activators has been explained by increasing mitochondrial fatty acid oxidation, as observed in livers of rats treated with the pan-PPAR activator tetradecylthioacetic acid (TTA). PPAR-activation does, however, not fully explain the metabolic adaptations observed in hepatocytes after treatment with TTA. We therefore characterized the mitochondrial effects, and linked this to signalling by the metabolic sensor, the mammalian target of rapamycin (mTOR). In hepatocytes isolated from TTA-treated rats, the changes in cellular content and morphology were consistent with hypertrophy. This was associated with induction of multiple mitochondrial biomarkers, including mitochondrial DNA, citrate synthase and mRNAs of mitochondrial proteins. Transcription analysis further confirmed activation of PPAR{alpha}-associated genes, in addition to genes related to mitochondrial biogenesis and function. Analysis of mitochondrial respiration revealed that the capacity of both electron transport and oxidative phosphorylation were increased. These effects coincided with activation of the stress related factor, ERK1/2, and mTOR. The protein level and phosphorylation of the downstream mTOR actors eIF4G and 4E-BP1 were induced. In summary, TTA increases mitochondrial respiration by inducing hypertrophy and mitochondrial biogenesis in rat hepatocytes, via adaptive regulation of PPARs as well as mTOR.

  7. Acute exposure of Drosophila melanogaster to paraquat causes oxidative stress and mitochondrial dysfunction.

    PubMed

    Hosamani, Ravikumar

    2013-05-01

    Paraquat (PQ; 1, 1'-dimethyl-4-4'-bipyridinium), an herbicide and model neurotoxicant, is identified to be one of the prime risk factors in Parkinson's disease (PD). In the Drosophila system, PQ is commonly used to measure acquired resistance against oxidative stress (PQ resistance test). Despite this, under acute PQ exposure, data on the oxidative stress response and associated impact on mitochondria among flies is limited. Accordingly, in this study, we measured markers of oxidative stress and mitochondrial dysfunctions among adult male flies (8-10 days old) exposed to varying concentrations of PQ (10, 20, and 40 mM in 5% sucrose solution) employing a conventional filter disc method for 24 h. PQ exposure resulted in significant elevation in the levels of oxidative stress biomarkers (malondialdehyde: 43% increase: hydroperoxide: 32-39% increase), with concomitant enhancement in reduced glutathione and total thiol levels in cytosol. Higher activity of antioxidant enzymes were also evident along with increased free iron levels. Furthermore, PQ exposure caused a concentration-dependent increase in mitochondrial superoxide generation and activity of manganese-superoxide dismutase (Mn-SOD). The activity levels of complex I-III, complex II-III, and Mg+2 adinosine triphosphatase (ATPase) were also decreased significantly. A robust diminution in the activity of succinate dehydrogenase and moderate decline in the citrate synthase activity suggested a specific effect on citric acid cycle enzymes. Collectively, these data suggest that acute PQ exposure causes significant oxidative stress and mitochondrial dysfunction among flies in vivo. It is suggested that in various experimental settings, while conducting the "PQ resistance stress test" incorporation of selected biochemical end points is likely to enhance the quality of the data.

  8. A mitochondrial superoxide theory for oxidative stress diseases and aging.

    PubMed

    Indo, Hiroko P; Yen, Hsiu-Chuan; Nakanishi, Ikuo; Matsumoto, Ken-Ichiro; Tamura, Masato; Nagano, Yumiko; Matsui, Hirofumi; Gusev, Oleg; Cornette, Richard; Okuda, Takashi; Minamiyama, Yukiko; Ichikawa, Hiroshi; Suenaga, Shigeaki; Oki, Misato; Sato, Tsuyoshi; Ozawa, Toshihiko; Clair, Daret K St; Majima, Hideyuki J

    2015-01-01

    Fridovich identified CuZnSOD in 1969 and manganese superoxide dismutase (MnSOD) in 1973, and proposed "the Superoxide Theory," which postulates that superoxide (O2 (•-)) is the origin of most reactive oxygen species (ROS) and that it undergoes a chain reaction in a cell, playing a central role in the ROS producing system. Increased oxidative stress on an organism causes damage to cells, the smallest constituent unit of an organism, which can lead to the onset of a variety of chronic diseases, such as Alzheimer's, Parkinson's, amyotrophic lateral sclerosis and other neurological diseases caused by abnormalities in biological defenses or increased intracellular reactive oxygen levels. Oxidative stress also plays a role in aging. Antioxidant systems, including non-enzyme low-molecular-weight antioxidants (such as, vitamins A, C and E, polyphenols, glutathione, and coenzyme Q10) and antioxidant enzymes, fight against oxidants in cells. Superoxide is considered to be a major factor in oxidant toxicity, and mitochondrial MnSOD enzymes constitute an essential defense against superoxide. Mitochondria are the major source of superoxide. The reaction of superoxide generated from mitochondria with nitric oxide is faster than SOD catalyzed reaction, and produces peroxynitrite. Thus, based on research conducted after Fridovich's seminal studies, we now propose a modified superoxide theory; i.e., superoxide is the origin of reactive oxygen and nitrogen species (RONS) and, as such, causes various redox related diseases and aging.

  9. The mechanisms of fatty acid-induced proton permeability of the inner mitochondrial membrane.

    PubMed

    Wojtczak, L; Wieckowski, M R

    1999-10-01

    Nonesterified long-chain fatty acids have long been known as uncouplers of oxidative phosphorylation. They are efficient protonophores in the inner mitochondrial membrane but not so in artificial phospholipid membranes. In the un-ionized form, they undergo a rapid spontaneous transbilayer movement (flip-flop). However, the transbilayer passage of the dissociated (anionic) form is hindered by the negatively charged hydrophilic carboxylic group. In the inner mitochondrial membrane, the transfer of fatty acid anions is mediated by the adenine nucleotide translocase, the dicarboxylate carrier, and the glutamate/aspartate carrier. As a result, the passage of protons and electric charges is a concerted effect of the spontaneous flip-flop of the undissociated (protonated) form in one direction and carrier-facilitated transfer of the ionized (deprotonated) form in the other direction. In addition, fatty acids also promote opening of the mitochondrial permeability transition pore, presumably due to their interaction with one of its constituents, the adenine nucleotide translocase, thus forming an additional route for dissipation of the proton gradient. Structural prerequisites for these proton-conducting mechanisms are (1) a weakly ionized carboxylic group and (2) a hydrocarbon chain of appropriate length without substituents limiting its mobility and hydrophobicity.

  10. Stimulation of fatty acid oxidation by a 3-thia fatty acid reduces triacylglycerol secretion in cultured rat hepatocytes.

    PubMed

    Skrede, S; Bremer, J; Berge, R K; Rustan, A C

    1994-08-01

    The present work shows that when mitochondrial beta-oxidation is stimulated by the hypolipemic, non-beta-oxidizable fatty acid analogue tetradecylthioacetic acid, there is a decrease in the secretion of triacylglycerol in cultured rat hepatocytes. In order to study the effects of tetradecylthioacetic acid in cells with different fatty acid oxidation rates, cells were grown without or with L-carnitine supplement or with addition of the beta-oxidation inhibitor L-aminocarnitine. In cells grown without and with L-carnitine in the medium, the oxidation of [1-14C]oleic acid was stimulated by tetradecylthioacetic acid, whereas it was not significantly changed by palmitic acid. In cells grown with L-aminocarnitine, oxidation of [1-14C]oleic acid was almost abolished both in the absence and in presence of tetradecylthioacetic acid. The effect of tetradecylthioacetic acid and palmitic acid on incorporation of [1-14C]oleic acid into triacylglycerol was similar under all conditions. In the presence of L-carnitine, secretion of oleic acid-labeled triacylglycerol was reduced significantly more by tetradecylthioacetic acid than by palmitic acid. The effects of tetradecylthioacetic acid and palmitic acid on secretion of oleic acid-labeled triacylglycerol were reversed in cells grown with L-aminocarnitine, where palmitic acid was the stronger inhibitor. These results were substantiated by determination of mass of triacylglycerol secreted. It is concluded that tetradecylthioacetic acid reduces secretion of triacylglycerol from rat hepatocytes mainly by acutely stimulating fatty acid oxidation.

  11. Nucleic acid oxidation: an early feature of Alzheimer's disease.

    PubMed

    Bradley-Whitman, Melissa A; Timmons, Michael D; Beckett, Tina L; Murphy, Michael P; Lynn, Bert C; Lovell, Mark A

    2014-01-01

    Studies of oxidative damage during the progression of Alzheimer's disease (AD) suggest its central role in disease pathogenesis. To investigate levels of nucleic acid oxidation in both early and late stages of AD, levels of multiple base adducts were quantified in nuclear and mitochondrial DNA from the superior and middle temporal gyri (SMTG), inferior parietal lobule (IPL), and cerebellum (CER) of age-matched normal control subjects, subjects with mild cognitive impairment, preclinical AD, late-stage AD, and non-AD neurological disorders (diseased control; DC) using gas chromatography/mass spectrometry. Median levels of multiple DNA adducts in nuclear and mitochondrial DNA were significantly (p ≤ 0.05) elevated in the SMTG, IPL, and CER in multiple stages of AD and in DC subjects. Elevated levels of fapyguanine and fapyadenine in mitochondrial DNA suggest a hypoxic environment early in the progression of AD and in DC subjects. Overall, these data suggest that oxidative damage is an early event not only in the pathogenesis of AD but is also present in neurodegenerative diseases in general. Levels of oxidized nucleic acids in nDNA and mtDNA were found to be significantly elevated in mild cognitive impairment (MCI), preclinical Alzheimer's disease (PCAD), late-stage AD (LAD), and a pooled diseased control group (DC) of frontotemporal dementia (FTD) and dementia with Lewy bodies (DLB) subjects compared to normal control (NC) subjects. Nucleic acid oxidation peaked early in disease progression and remained elevated. The study suggests nucleic acid oxidation is a general event in neurodegeneration.

  12. Defects in Mitochondrial Fatty Acid Synthesis Result in Failure of Multiple Aspects of Mitochondrial Biogenesis in Saccharomyces cerevisiae

    PubMed Central

    Kursu, V. A. Samuli; Pietikäinen, Laura P.; Fontanesi, Flavia; Aaltonen, Mari J.; Suomi, Fumi; Nair, Remya Raghavan; Schonauer, Melissa S.; Dieckmann, Carol L.; Barrientos, Antoni; Hiltunen, J. Kalervo; Kastaniotis, Alexander J.

    2014-01-01

    Summary Mitochondrial fatty acid synthesis (mtFAS) shares acetyl-CoA with the Krebs cycle as a common substrate and is required for the production of octanoic acid (C8) precursors of lipoic acid (LA) in mitochondria. MtFAS is a conserved pathway essential for respiration. In a genetic screen in Saccharomyces cerevisiae designed to further elucidate the physiological role of mtFAS, we isolated mutants with defects in mitochondrial post-translational gene expression processes, indicating a novel link to mitochondrial gene expression and respiratory chain biogenesis. In our ensuing analysis, we show that mtFAS, but not lipoylation per se, is required for respiratory competence. We demonstrate that mtFAS is required for mRNA splicing, mitochondrial translation and respiratory complex assembly, and provide evidence that not LA per se, but fatty acids longer than C8 play a role in these processes. We also show that mtFAS- and LA-deficient strains suffer from a mild heme deficiency that may contribute to the respiratory complex assembly defect. Based on our data and previously published information, we propose a model implicating mtFAS as a sensor for mitochondrial acetyl-CoA availability and a coordinator of nuclear and mitochondrial gene expression by adapting the mitochondrial compartment to changes in the metabolic status of the cell. PMID:24102902

  13. Mitochondrial metabolism mediates oxidative stress and inflammation in fatty liver

    PubMed Central

    Satapati, Santhosh; Kucejova, Blanka; Duarte, Joao A.G.; Fletcher, Justin A.; Reynolds, Lacy; Sunny, Nishanth E.; He, Tianteng; Nair, L. Arya; Livingston, Kenneth; Fu, Xiaorong; Merritt, Matthew E.; Sherry, A. Dean; Malloy, Craig R.; Shelton, John M.; Lambert, Jennifer; Parks, Elizabeth J.; Corbin, Ian; Magnuson, Mark A.; Browning, Jeffrey D.; Burgess, Shawn C.

    2015-01-01

    Mitochondria are critical for respiration in all tissues; however, in liver, these organelles also accommodate high-capacity anaplerotic/cataplerotic pathways that are essential to gluconeogenesis and other biosynthetic activities. During nonalcoholic fatty liver disease (NAFLD), mitochondria also produce ROS that damage hepatocytes, trigger inflammation, and contribute to insulin resistance. Here, we provide several lines of evidence indicating that induction of biosynthesis through hepatic anaplerotic/cataplerotic pathways is energetically backed by elevated oxidative metabolism and hence contributes to oxidative stress and inflammation during NAFLD. First, in murine livers, elevation of fatty acid delivery not only induced oxidative metabolism, but also amplified anaplerosis/cataplerosis and caused a proportional rise in oxidative stress and inflammation. Second, loss of anaplerosis/cataplerosis via genetic knockdown of phosphoenolpyruvate carboxykinase 1 (Pck1) prevented fatty acid–induced rise in oxidative flux, oxidative stress, and inflammation. Flux appeared to be regulated by redox state, energy charge, and metabolite concentration, which may also amplify antioxidant pathways. Third, preventing elevated oxidative metabolism with metformin also normalized hepatic anaplerosis/cataplerosis and reduced markers of inflammation. Finally, independent histological grades in human NAFLD biopsies were proportional to oxidative flux. Thus, hepatic oxidative stress and inflammation are associated with elevated oxidative metabolism during an obesogenic diet, and this link may be provoked by increased work through anabolic pathways. PMID:26571396

  14. Mitochondrial division ensures the survival of postmitotic neurons by suppressing oxidative damage.

    PubMed

    Kageyama, Yusuke; Zhang, Zhongyan; Roda, Ricardo; Fukaya, Masahiro; Wakabayashi, Junko; Wakabayashi, Nobunao; Kensler, Thomas W; Reddy, P Hemachandra; Iijima, Miho; Sesaki, Hiromi

    2012-05-14

    Mitochondria divide and fuse continuously, and the balance between these two processes regulates mitochondrial shape. Alterations in mitochondrial dynamics are associated with neurodegenerative diseases. Here we investigate the physiological and cellular functions of mitochondrial division in postmitotic neurons using in vivo and in vitro gene knockout for the mitochondrial division protein Drp1. When mouse Drp1 was deleted in postmitotic Purkinje cells in the cerebellum, mitochondrial tubules elongated due to excess fusion, became large spheres due to oxidative damage, accumulated ubiquitin and mitophagy markers, and lost respiratory function, leading to neurodegeneration. Ubiquitination of mitochondria was independent of the E3 ubiquitin ligase parkin in Purkinje cells lacking Drp1. Treatment with antioxidants rescued mitochondrial swelling and cell death in Drp1KO Purkinje cells. Moreover, hydrogen peroxide converted elongated tubules into large spheres in Drp1KO fibroblasts. Our findings suggest that mitochondrial division serves as a quality control mechanism to suppress oxidative damage and thus promote neuronal survival.

  15. Mitochondrial membrane potential: a novel biomarker of oxidative environmental stress.

    PubMed Central

    Vayssier-Taussat, Muriel; Kreps, Sarah E; Adrie, Christophe; Dall'Ava, Josette; Christiani, David; Polla, Barbara S

    2002-01-01

    Epidemiologic analyses, traditionally based on long-term cohort or case-control studies, provide retrospective causal associations between exposure to a particular environmental stressor and an exposure-related disease end point. Recent research initiatives have propelled a shift toward exploring molecular epidemiology and molecular biological markers (biomarkers) as a means of providing more immediate, quantitative risk assessment of potentially deleterious environmental exposures. We compared, in normal human monocytes isolated from the blood of healthy donors, variations in Hsp70 expression and mitochondrial membrane potential (delta psi m) in response to exposure to either tobacco smoke or gamma-irradiation, two models for environmentally mediated oxidant exposure. On the basis of its mechanistic specificity for oxidants and little baseline variation in cells from distinct individuals, we propose that delta psi m represents a selective in vitro and in vivo biomarker for oxidant exposure. delta psi m may be used to gauge risks associated with oxidant-mediated air pollution and radiation. PMID:11882482

  16. Downregulation of peroxiredoxin-3 by hydrophobic bile acid induces mitochondrial dysfunction and cellular senescence in human trophoblasts

    PubMed Central

    Wu, Wei-Bin; Menon, Ramkumar; Xu, Yue-Ying; Zhao, Jiu-Ru; Wang, Yan-Lin; Liu, Yuan; Zhang, Hui-Juan

    2016-01-01

    Intrahepatic cholestasis of pregnancy (ICP) is a pregnancy-specific disorder characterised by raised bile acids in foetal-maternal circulation, which threatens perinatal health. During the progression of ICP, the effect of oxidative stress is underscored. Peroxiredoxin-3 (PRDX3) is a mitochondrial antioxidant enzyme that is crucial to balance intracellular oxidative stress. However, the role of PRDX3 in placental trophoblast cells under ICP is not fully understood. We demonstrated that the level of PRDX3 was downregulated in ICP placentas as well as bile acids–treated trophoblast cells and villous explant in vitro. Toxic levels of bile acids and PRDX3 knockdown induced oxidative stress and mitochondrial dysfunction in trophoblast cells. Moreover, silencing of PRDX3 in trophoblast cell line HTR8/SVneo induced growth arrest and cellular senescence via activation of p38-mitogen-activated protein kinase (MAPK) and induction of p21WAF1/CIP and p16INK4A. Additionally, enhanced cellular senescence, determined by senescence-associated beta-galactosidase staining, was obviously attenuated by p38-MAPK inhibitor SB203580. Our data determined that exposure to bile acid decreased PRDX3 level in human trophoblasts. PRDX3 protected trophoblast cells against mitochondrial dysfunction and cellular senescence induced by oxidative stress. Our results suggest that decreased PRDX3 by excessive bile acids in trophoblasts plays a critical role in the pathogenesis and progression of ICP. PMID:27958341

  17. Chronic Arsenic Exposure-Induced Oxidative Stress is Mediated by Decreased Mitochondrial Biogenesis in Rat Liver.

    PubMed

    Prakash, Chandra; Kumar, Vijay

    2016-09-01

    The present study was executed to study the effect of chronic arsenic exposure on generation of mitochondrial oxidative stress and biogenesis in rat liver. Chronic sodium arsenite treatment (25 ppm for 12 weeks) decreased mitochondrial complexes activity in rat liver. There was a decrease in mitochondrial superoxide dismutase (MnSOD) activity in arsenic-treated rats that might be responsible for increased protein and lipid oxidation as observed in our study. The messenger RNA (mRNA) expression of mitochondrial and nuclear-encoded subunits of complexes I (ND1 and ND2) and IV (COX I and COX IV) was downregulated in arsenic-treated rats only. The protein and mRNA expression of MnSOD was reduced suggesting increased mitochondrial oxidative damage after arsenic treatment. There was activation of Bax and caspase-3 followed by release of cytochrome c from mitochondria suggesting induction of apoptotic pathway under oxidative stress. The entire phenomenon was associated with decrease in mitochondrial biogenesis as evident by decreased protein and mRNA expression of nuclear respiratory factor 1 (NRF-1), nuclear respiratory factor 2 (NRF-2), peroxisome proliferator activator receptor gamma-coactivator 1α (PGC-1α), and mitochondrial transcription factor A (Tfam) in arsenic-treated rat liver. The results of the present study indicate that arsenic-induced mitochondrial oxidative stress is associated with decreased mitochondrial biogenesis in rat liver that may present one of the mechanisms for arsenic-induced hepatotoxicity.

  18. Mitochondrial alterations and oxidative stress in an acute transient mouse model of muscle degeneration: implications for muscular dystrophy and related muscle pathologies.

    PubMed

    Ramadasan-Nair, Renjini; Gayathri, Narayanappa; Mishra, Sudha; Sunitha, Balaraju; Mythri, Rajeswara Babu; Nalini, Atchayaram; Subbannayya, Yashwanth; Harsha, Hindalahalli Chandregowda; Kolthur-Seetharam, Ullas; Srinivas Bharath, Muchukunte Mukunda

    2014-01-03

    Muscular dystrophies (MDs) and inflammatory myopathies (IMs) are debilitating skeletal muscle disorders characterized by common pathological events including myodegeneration and inflammation. However, an experimental model representing both muscle pathologies and displaying most of the distinctive markers has not been characterized. We investigated the cardiotoxin (CTX)-mediated transient acute mouse model of muscle degeneration and compared the cardinal features with human MDs and IMs. The CTX model displayed degeneration, apoptosis, inflammation, loss of sarcolemmal complexes, sarcolemmal disruption, and ultrastructural changes characteristic of human MDs and IMs. Cell death caused by CTX involved calcium influx and mitochondrial damage both in murine C2C12 muscle cells and in mice. Mitochondrial proteomic analysis at the initial phase of degeneration in the model detected lowered expression of 80 mitochondrial proteins including subunits of respiratory complexes, ATP machinery, fatty acid metabolism, and Krebs cycle, which further decreased in expression during the peak degenerative phase. The mass spectrometry (MS) data were supported by enzyme assays, Western blot, and histochemistry. The CTX model also displayed markers of oxidative stress and a lowered glutathione reduced/oxidized ratio (GSH/GSSG) similar to MDs, human myopathies, and neurogenic atrophies. MS analysis identified 6 unique oxidized proteins from Duchenne muscular dystrophy samples (n = 6) (versus controls; n = 6), including two mitochondrial proteins. Interestingly, these mitochondrial proteins were down-regulated in the CTX model thereby linking oxidative stress and mitochondrial dysfunction. We conclude that mitochondrial alterations and oxidative damage significantly contribute to CTX-mediated muscle pathology with implications for human muscle diseases.

  19. Fatty Acid Oxidation in Cardiac and Skeletal Muscle Mitochondria is Unaffected by Deletion of CD36

    PubMed Central

    King, Kristen L.; Stanley, William C.; Rosca, Mariana; Kerner, Janos; Hoppel, Charles L.; Febbraio, Maria

    2009-01-01

    Recent studies found that the plasma membrane fatty acid transport protein CD36 also resides in mitochondrial membranes in cardiac and skeletal muscle. Pharmacological studies suggest that CD36 may play an essential role in mitochondrial fatty acid oxidation. We isolated cardiac and skeletal muscle mitochondria from wild type and CD36 knock-out mice. There were no differences between wild type and CD36 knock-out mice in mitochondrial respiration with palmitoyl-CoA, palmitoyl-carnitine or glutamate as substrate. We investigated a potential alternative role for CD36 in mitochondria, ie. the export of fatty acids generated in the matrix. Palmitate export was not different between wild type and CD36 knock out mice. Taken together, CD36 does not appear to play an essential role in mitochondrial uptake of fatty acids or export of fatty acid anions. PMID:17904092

  20. Diabetic Polyneuropathy in Type 2 Diabetes Mellitus: Inflammation, Oxidative Stress, and Mitochondrial Function

    PubMed Central

    Román-Pintos, Luis Miguel; Villegas-Rivera, Geannyne; Rodríguez-Carrizalez, Adolfo Daniel; Cardona-Muñoz, Ernesto Germán

    2016-01-01

    Diabetic polyneuropathy (DPN) is defined as peripheral nerve dysfunction. There are three main alterations involved in the pathologic changes of DPN: inflammation, oxidative stress, and mitochondrial dysfunction. Inflammation induces activation of nuclear factor kappa B, activator protein 1, and mitogen-activated protein kinases. Oxidative stress induced by hyperglycemia is mediated by several identified pathways: polyol, hexosamine, protein kinase C, advanced glycosylation end-products, and glycolysis. In addition, mitochondrial dysfunction accounts for most of the production of reactive oxygen and nitrosative species. These free radicals cause lipid peroxidation, protein modification, and nucleic acid damage, to finally induce axonal degeneration and segmental demyelination. The prevalence of DPN ranges from 2.4% to 78.8% worldwide, depending on the diagnostic method and the population assessed (hospital-based or outpatients). Risk factors include age, male gender, duration of diabetes, uncontrolled glycaemia, height, overweight and obesity, and insulin treatment. Several diagnostic methods have been developed, and composite scores combined with nerve conduction studies are the most reliable to identify early DPN. Treatment should be directed to improve etiologic factors besides reducing symptoms; several approaches have been evaluated to reduce neuropathic impairments and improve nerve conduction, such as oral antidiabetics, statins, and antioxidants (alpha-lipoic acid, ubiquinone, and flavonoids). PMID:28058263

  1. Diabetic Polyneuropathy in Type 2 Diabetes Mellitus: Inflammation, Oxidative Stress, and Mitochondrial Function.

    PubMed

    Román-Pintos, Luis Miguel; Villegas-Rivera, Geannyne; Rodríguez-Carrizalez, Adolfo Daniel; Miranda-Díaz, Alejandra Guillermina; Cardona-Muñoz, Ernesto Germán

    2016-01-01

    Diabetic polyneuropathy (DPN) is defined as peripheral nerve dysfunction. There are three main alterations involved in the pathologic changes of DPN: inflammation, oxidative stress, and mitochondrial dysfunction. Inflammation induces activation of nuclear factor kappa B, activator protein 1, and mitogen-activated protein kinases. Oxidative stress induced by hyperglycemia is mediated by several identified pathways: polyol, hexosamine, protein kinase C, advanced glycosylation end-products, and glycolysis. In addition, mitochondrial dysfunction accounts for most of the production of reactive oxygen and nitrosative species. These free radicals cause lipid peroxidation, protein modification, and nucleic acid damage, to finally induce axonal degeneration and segmental demyelination. The prevalence of DPN ranges from 2.4% to 78.8% worldwide, depending on the diagnostic method and the population assessed (hospital-based or outpatients). Risk factors include age, male gender, duration of diabetes, uncontrolled glycaemia, height, overweight and obesity, and insulin treatment. Several diagnostic methods have been developed, and composite scores combined with nerve conduction studies are the most reliable to identify early DPN. Treatment should be directed to improve etiologic factors besides reducing symptoms; several approaches have been evaluated to reduce neuropathic impairments and improve nerve conduction, such as oral antidiabetics, statins, and antioxidants (alpha-lipoic acid, ubiquinone, and flavonoids).

  2. Trimethylamine N-oxide impairs pyruvate and fatty acid oxidation in cardiac mitochondria.

    PubMed

    Makrecka-Kuka, Marina; Volska, Kristine; Antone, Unigunde; Vilskersts, Reinis; Grinberga, Solveiga; Bandere, Dace; Liepinsh, Edgars; Dambrova, Maija

    2017-02-05

    Increased plasma concentration of trimethylamine N-oxide (TMAO), a proatherogenic metabolite, has been linked to adverse cardiovascular outcomes; however, it remains unclear whether TMAO is a biomarker or whether it induces direct detrimental cardiovascular effects. Because altered cardiac energy metabolism and mitochondrial dysfunction play crucial roles in the development of cardiovascular diseases, we hypothesized that increased TMAO concentration may alter mitochondrial energy metabolism. The aim of the present study was to determine the effects of TMAO on cardiac mitochondrial energy metabolism. Acute exposure of cardiac fibers to TMAO decreased LEAK (substrate-dependent) and OXPHOS (oxidative phosphorylation-dependent) mitochondrial respiration with pyruvate and impaired substrate flux via pyruvate dehydrogenase. The administration of TMAO at a dose of 120mg/kg for 8 weeks increased TMAO concentration in plasma and cardiac tissues 22-23 times to about 15μM and 11nmol/g, respectively. Long-term TMAO administration decreased mitochondrial LEAK state respiration with pyruvate by 30% without affecting OXPHOS state respiration. However, no significant changes in mitochondrial reactive oxygen species production were observed after acute exposure of cardiac fibers to TMAO under physiological conditions. In addition, both long-term TMAO administration and acute exposure to TMAO decreased respiration with palmitoyl-CoA indicating impaired β-oxidation. Taken together, our results demonstrate that increased TMAO concentration impairs pyruvate and fatty acid oxidation in cardiac mitochondria. Thus, the accumulation of TMAO in cardiac tissues leads to disturbances in energy metabolism that can increase the severity of cardiovascular events.

  3. Titanium dioxide nanoparticles induce strong oxidative stress and mitochondrial damage in glial cells.

    PubMed

    Huerta-García, Elizabeth; Pérez-Arizti, José Antonio; Márquez-Ramírez, Sandra Gissela; Delgado-Buenrostro, Norma Laura; Chirino, Yolanda Irasema; Iglesias, Gisela Gutiérrez; López-Marure, Rebeca

    2014-08-01

    Titanium dioxide nanoparticles (TiO2 NPs) are widely used in the chemical, electrical, and electronic industries. TiO2 NPs can enter directly into the brain through the olfactory bulb and can be deposited in the hippocampus region; therefore, we determined the toxic effect of TiO2 NPs on rat and human glial cells, C6 and U373, respectively. We evaluated some events related to oxidative stress: (1) redox-signaling mechanisms by oxidation of 2',7'-dichlorodihydrofluorescein diacetate; (2) peroxidation of lipids by cis-parinaric acid; (3) antioxidant enzyme expression by PCR in real time; and (4) mitochondrial damage by MitoTracker Green FM staining and Rh123. TiO2 NPs induced a strong oxidative stress in both glial cell lines by mediating changes in the cellular redox state and lipid peroxidation associated with a rise in the expression of glutathione peroxidase, catalase, and superoxide dismutase 2. TiO2 NPs also produced morphological changes, damage of mitochondria, and an increase in mitochondrial membrane potential, indicating toxicity. TiO2 NPs had a cytotoxic effect on glial cells; however, more in vitro and in vivo studies are required to ascertain that exposure to TiO2 NPs can cause brain injury and be hazardous to health.

  4. Carotid body O2 chemoreception and mitochondrial oxidative phosphorylation.

    PubMed

    Mulligan, E; Lahiri, S; Storey, B T

    1981-08-01

    The effect on carotid chemoreceptor afferents of oligomycin, an inhibitor of mitochondrial oxidative phosphorylation that does not affect energy conservation, was studied in 20 cats that were anesthetized, paralyzed, and artificially ventilated. Responses of single or a few chemoreceptor afferents to changes in arterial O2 tension (PaO2) at constant arterial CO2 tension were recorded. In addition, responses to nicotine, cyanide, and antimycin A or carbonyl cyanide p-tri-fluoromethoxyphenylhydrazone (FCCP) were tested in normoxia. Oligomycin (50-500 microgram) was administered by close intra-arterial injection, and the same tests were repeated at timed intervals. Initially, oligomycin caused vigorous stimulation of carotid chemoreceptor activity. Subsequently, although the afferent fibers were still active and could be vigorously stimulated by nicotine, they no longer responded to changes in PaO2 or to doses of cyanide, antimycin A, or FCCP. These results separate stimulation of chemoreceptor afferents by hypoxia and metabolic inhibitors and uncouplers from that by nicotine and suggest that intact oxidative phosphorylation, required for maintenance of the intracellular high-energy phosphate levels, forms the basis of O2 chemoreception in the carotid body.

  5. Detection and quantification of protein adduction by electrophilic fatty acids: mitochondrial generation of fatty acid nitroalkene derivatives.

    PubMed

    Schopfer, F J; Batthyany, C; Baker, P R S; Bonacci, G; Cole, M P; Rudolph, V; Groeger, A L; Rudolph, T K; Nadtochiy, S; Brookes, P S; Freeman, B A

    2009-05-01

    Nitroalkene fatty acid derivatives manifest a strong electrophilic nature, are clinically detectable, and induce multiple transcriptionally regulated anti-inflammatory responses. At present, the characterization and quantification of endogenous electrophilic lipids are compromised by their Michael addition with protein and small-molecule nucleophilic targets. Herein, we report a trans-nitroalkylation reaction of nitro-fatty acids with beta-mercaptoethanol (BME) and apply this reaction to the unbiased identification and quantification of reaction with nucleophilic targets. Trans-nitroalkylation yields are maximal at pH 7 to 8 and occur with physiological concentrations of target nucleophiles. This reaction is also amenable to sensitive mass spectrometry-based quantification of electrophilic fatty acid-protein adducts upon electrophoretic resolution of proteins. In-gel trans-nitroalkylation reactions also permit the identification of protein targets without the bias and lack of sensitivity of current proteomic approaches. Using this approach, it was observed that fatty acid nitroalkenes are rapidly metabolized in vivo by a nitroalkene reductase activity and mitochondrial beta-oxidation, yielding a variety of electrophilic and nonelectrophilic products that could be structurally characterized upon BME-based trans-nitroalkylation reaction. This strategy was applied to the detection and quantification of fatty acid nitration in mitochondria in response to oxidative inflammatory conditions induced by myocardial ischemia-reoxygenation.

  6. Activating HSP72 in rodent skeletal muscle increases mitochondrial number and oxidative capacity and decreases insulin resistance.

    PubMed

    Henstridge, Darren C; Bruce, Clinton R; Drew, Brian G; Tory, Kálmán; Kolonics, Attila; Estevez, Emma; Chung, Jason; Watson, Nadine; Gardner, Timothy; Lee-Young, Robert S; Connor, Timothy; Watt, Matthew J; Carpenter, Kevin; Hargreaves, Mark; McGee, Sean L; Hevener, Andrea L; Febbraio, Mark A

    2014-06-01

    Induction of heat shock protein (HSP)72 protects against obesity-induced insulin resistance, but the underlying mechanisms are unknown. Here, we show that HSP72 plays a pivotal role in increasing skeletal muscle mitochondrial number and oxidative metabolism. Mice overexpressing HSP72 in skeletal muscle (HSP72Tg) and control wild-type (WT) mice were fed either a chow or high-fat diet (HFD). Despite a similar energy intake when HSP72Tg mice were compared with WT mice, the HFD increased body weight, intramuscular lipid accumulation (triacylglycerol and diacylglycerol but not ceramide), and severe glucose intolerance in WT mice alone. Whole-body VO2, fatty acid oxidation, and endurance running capacity were markedly increased in HSP72Tg mice. Moreover, HSP72Tg mice exhibited an increase in mitochondrial number. In addition, the HSP72 coinducer BGP-15, currently in human clinical trials for type 2 diabetes, also increased mitochondrial number and insulin sensitivity in a rat model of type 2 diabetes. Together, these data identify a novel role for activation of HSP72 in skeletal muscle. Thus, the increased oxidative metabolism associated with activation of HSP72 has potential clinical implications not only for type 2 diabetes but also for other disorders where mitochondrial function is compromised.

  7. Activating HSP72 in Rodent Skeletal Muscle Increases Mitochondrial Number and Oxidative Capacity and Decreases Insulin Resistance

    PubMed Central

    Henstridge, Darren C.; Bruce, Clinton R.; Drew, Brian G.; Tory, Kálmán; Kolonics, Attila; Estevez, Emma; Chung, Jason; Watson, Nadine; Gardner, Timothy; Lee-Young, Robert S.; Connor, Timothy; Watt, Matthew J.; Carpenter, Kevin; Hargreaves, Mark; McGee, Sean L.; Hevener, Andrea L.; Febbraio, Mark A.

    2014-01-01

    Induction of heat shock protein (HSP)72 protects against obesity-induced insulin resistance, but the underlying mechanisms are unknown. Here, we show that HSP72 plays a pivotal role in increasing skeletal muscle mitochondrial number and oxidative metabolism. Mice overexpressing HSP72 in skeletal muscle (HSP72Tg) and control wild-type (WT) mice were fed either a chow or high-fat diet (HFD). Despite a similar energy intake when HSP72Tg mice were compared with WT mice, the HFD increased body weight, intramuscular lipid accumulation (triacylglycerol and diacylglycerol but not ceramide), and severe glucose intolerance in WT mice alone. Whole-body VO2, fatty acid oxidation, and endurance running capacity were markedly increased in HSP72Tg mice. Moreover, HSP72Tg mice exhibited an increase in mitochondrial number. In addition, the HSP72 coinducer BGP-15, currently in human clinical trials for type 2 diabetes, also increased mitochondrial number and insulin sensitivity in a rat model of type 2 diabetes. Together, these data identify a novel role for activation of HSP72 in skeletal muscle. Thus, the increased oxidative metabolism associated with activation of HSP72 has potential clinical implications not only for type 2 diabetes but also for other disorders where mitochondrial function is compromised. PMID:24430435

  8. BGP-15 Protects against Oxidative Stress- or Lipopolysaccharide-Induced Mitochondrial Destabilization and Reduces Mitochondrial Production of Reactive Oxygen Species

    PubMed Central

    Sumegi, Katalin; Fekete, Katalin; Antus, Csenge; Debreceni, Balazs; Hocsak, Eniko; Gallyas, Ferenc; Sumegi, Balazs; Szabo, Aliz

    2017-01-01

    Reactive oxygen species (ROS) play a critical role in the progression of mitochondria-related diseases. A novel insulin sensitizer drug candidate, BGP-15, has been shown to have protective effects in several oxidative stress-related diseases in animal and human studies. In this study, we investigated whether the protective effects of BGP-15 are predominantly via preserving mitochondrial integrity and reducing mitochondrial ROS production. BGP-15 was found to accumulate in the mitochondria, protect against ROS-induced mitochondrial depolarization and attenuate ROS-induced mitochondrial ROS production in a cell culture model, and also reduced ROS production predominantly at the complex I-III system in isolated mitochondria. At physiologically relevant concentrations, BGP-15 protected against hydrogen peroxide-induced cell death by reducing both apoptosis and necrosis. Additionally, it attenuated bacterial lipopolysaccharide (LPS)-induced collapse of mitochondrial membrane potential and ROS production in LPS-sensitive U-251 glioma cells, suggesting that BGP-15 may have a protective role in inflammatory diseases. However, BGP-15 did not have any antioxidant effects as shown by in vitro chemical and cell culture systems. These data suggest that BGP-15 could be a novel mitochondrial drug candidate for the prevention of ROS-related and inflammatory disease progression. PMID:28046125

  9. Resolution of mitochondrial oxidant stress improves aged-cardiovascular performance

    PubMed Central

    Owada, Takashi; Yamauchi, Hiroyuki; Miura, Shunsuke; Machii, Hirofumi; Takeishi, Yasuchika

    2017-01-01

    Background Senescence is a major factor that increases oxidative stress in mitochondria, which contributes toward the pathogenesis of heart disease. However, the effect of antioxidant therapy on cardiac mitochondria in aged-cardiac performance remains elusive. Objectives We postulated that the mitochondrial targeting of superoxide scavenging would have benefits in the aged heart. Methods and results Generation of superoxide in the mitochondria and nicotinamide adenine dinucleotide phosphate oxidase activity increased in the heart of old mice compared with that in young mice. In old mice treated with a mitochondria-targeted antioxidant MitoTEMPO (180 µg/kg/day, 28 days) co-infusion using a subcutaneously implanted minipump, levels of superoxide in the mitochondria and nicotinamide adenine dinucleotide phosphate oxidase activity as well as hydrogen peroxide decreased markedly in cardiomyocytes. Treatment with MitoTEMPO in old mice improved the systolic and diastolic function assessed by echocardiography. Endothelium-dependent vasodilation in isolated coronary arteries and endothelial nitric-oxide synthase phosphorylation were impaired in old mice compared with that in young mice and were improved by MitoTEMPO treatment. Mitochondria from the old mice myocardium showed lower rates of complex I-dependent and II-dependent respiration compared with that from young mice. Supplementation of MitoTEMPO in old mice improved the respiration rates and efficiency of ATP generation in mitochondria to a level similar to that of young mice. Conclusion Resolution of oxidative stress in mitochondria by MitoTEMPO in old mice restored cardiac function and the capacity of coronary vasodilation to the same magnitude observed in young mice. An antioxidant strategy targeting mitochondria could have a therapeutic benefit in heart disease with senescence. PMID:27740971

  10. Thiol-based antioxidants elicit mitochondrial oxidation via respiratory complex III

    PubMed Central

    Beaudoin, Jessica N.; Ponnuraj, Nagendraprabhu; DiLiberto, Stephen J.; Hanafin, William P.; Kenis, Paul J. A.; Gaskins, H. Rex

    2015-01-01

    Excessive oxidation is widely accepted as a precursor to deleterious cellular function. On the other hand, an awareness of the role of reductive stress as a similar pathological insult is emerging. Here we report early dynamic changes in compartmentalized glutathione (GSH) redox potentials in living cells in response to exogenously supplied thiol-based antioxidants. Noninvasive monitoring of intracellular thiol-disulfide exchange via a genetically encoded biosensor targeted to cytosol and mitochondria revealed unexpectedly rapid oxidation of the mitochondrial matrix in response to GSH ethyl ester or N-acetyl-l-cysteine. Oxidation of the probe occurred within seconds in a concentration-dependent manner and was attenuated with the membrane-permeable ROS scavenger tiron. In contrast, the cytosolic sensor did not respond to similar treatments. Surprisingly, the immediate mitochondrial oxidation was not abrogated by depolarization of mitochondrial membrane potential or inhibition of mitochondrial GSH uptake. After detection of elevated levels of mitochondrial ROS, we systematically inhibited multisubunit protein complexes of the mitochondrial respiratory chain and determined that respiratory complex III is a downstream target of thiol-based compounds. Disabling complex III with myxothiazol completely blocked matrix oxidation induced with GSH ethyl ester or N-acetyl-l-cysteine. Our findings provide new evidence of a functional link between exogenous thiol-containing antioxidants and mitochondrial respiration. PMID:25994788

  11. Perm1 enhances mitochondrial biogenesis, oxidative capacity, and fatigue resistance in adult skeletal muscle.

    PubMed

    Cho, Yoshitake; Hazen, Bethany C; Gandra, Paulo G; Ward, Samuel R; Schenk, Simon; Russell, Aaron P; Kralli, Anastasia

    2016-02-01

    Skeletal muscle mitochondrial content and oxidative capacity are important determinants of muscle function and whole-body health. Mitochondrial content and function are enhanced by endurance exercise and impaired in states or diseases where muscle function is compromised, such as myopathies, muscular dystrophies, neuromuscular diseases, and age-related muscle atrophy. Hence, elucidating the mechanisms that control muscle mitochondrial content and oxidative function can provide new insights into states and diseases that affect muscle health. In past studies, we identified Perm1 (PPARGC1- and ESRR-induced regulator, muscle 1) as a gene induced by endurance exercise in skeletal muscle, and regulating mitochondrial oxidative function in cultured myotubes. The capacity of Perm1 to regulate muscle mitochondrial content and function in vivo is not yet known. In this study, we use adeno-associated viral (AAV) vectors to increase Perm1 expression in skeletal muscles of 4-wk-old mice. Compared to control vector, AAV1-Perm1 leads to significant increases in mitochondrial content and oxidative capacity (by 40-80%). Moreover, AAV1-Perm1-transduced muscles show increased capillary density and resistance to fatigue (by 33 and 31%, respectively), without prominent changes in fiber-type composition. These findings suggest that Perm1 selectively regulates mitochondrial biogenesis and oxidative function, and implicate Perm1 in muscle adaptations that also occur in response to endurance exercise.

  12. 3-Nitropropionic Acid is a Suicide Inhibitor of MitochondrialRespiration that, Upon Oxidation by Complex II, Forms a Covalent AdductWith a Catalytic Base Arginine in the Active Site of the Enzyme

    SciTech Connect

    Huang, Li-shar; Sun, Gang; Cobessi, David; Wang, Andy C.; Shen,John T.; Tung, Eric Y.; Anderson, Vernon E.; Berry, Edward A.

    2005-12-01

    We report three new structures of mitochondrial respiratory Complex II (succinate ubiquinone oxidoreductase, E.C. 1.3.5.1) at up to 2.1 {angstrom} resolution, with various inhibitors. The structures define the conformation of the bound inhibitors and suggest the residues involved in substrate binding and catalysis at the dicarboxylate site. In particular they support the role of Arg297 as a general base catalyst accepting a proton in the dehydrogenation of succinate. The dicarboxylate ligand in oxaloacetate-containing crystals appears to be the same as that reported for Shewanella flavocytochrome c treated with fumarate. The plant and fungal toxin 3-nitropropionic acid, an irreversible inactivator of succinate dehydrogenase, forms a covalent adduct with the side chain of Arg297. The modification eliminates a trypsin cleavage site in the flavoprotein, and tandem mass spectroscopic analysis of the new fragment shows the mass of Arg 297 to be increased by 83 Da and to have potential of losing 44 Da, consistent with decarboxylation, during fragmentation.

  13. 3-Nitropropionic acid induces autophagy by forming mitochondrial permeability transition pores rather than activatiing the mitochondrial fission pathway

    PubMed Central

    Solesio, Maria E; Saez-Atienzar, Sara; Jordan, Joaquin; Galindo, Maria F

    2013-01-01

    BACKGROUND AND PURPOSE Huntington's disease is a neurodegenerative process associated with mitochondrial alterations. Inhibitors of the electron–transport channel complex II, such as 3-nitropropionic acid (3NP), are used to study the molecular and cellular pathways involved in this disease. We studied the effect of 3NP on mitochondrial morphology and its involvement in macrophagy. EXPERIMENTAL APPROACH Pharmacological and biochemical methods were used to characterize the effects of 3NP on autophagy and mitochondrial morphology. SH-SY5Y cells were transfected with GFP-LC3, GFP-Drp1 or GFP-Bax to ascertain their role and intracellular localization after 3NP treatment using confocal microscopy. KEY RESULTS Untreated SH-SY5Y cells presented a long, tubular and filamentous net of mitochondria. After 3NP (5 mM) treatment, mitochondria became shorter and rounder. 3NP induced formation of mitochondrial permeability transition pores, both in cell cultures and in isolated liver mitochondria, and this process was inhibited by cyclosporin A. Participation of the mitochondrial fission pathway was excluded because 3NP did not induce translocation of the dynamin-related protein 1 (Drp1) to the mitochondria. The Drp1 inhibitor Mdivi-1 did not affect the observed changes in mitochondrial morphology. Finally, scavengers of reactive oxygen species failed to prevent mitochondrial alterations, while cyclosporin A, but not Mdivi-1, prevented the generation of ROS. CONCLUSIONS AND IMPLICATIONS There was a direct correlation between formation of mitochondrial permeability transition pores and autophagy induced by 3NP treatment. Activation of autophagy preceded the apoptotic process and was mediated, at least partly, by formation of reactive oxygen species and mitochondrial permeability transition pores. LINKED ARTICLE This article is commented on by González-Polo et al., pp. 60–62 of this issue. To view this commentary visit http://dx.doi.org/10.1111/j.1476-5381.2012.02203.x PMID

  14. Lycopene Prevents Amyloid [Beta]-Induced Mitochondrial Oxidative Stress and Dysfunctions in Cultured Rat Cortical Neurons.

    PubMed

    Qu, Mingyue; Jiang, Zheng; Liao, Yuanxiang; Song, Zhenyao; Nan, Xinzhong

    2016-06-01

    Brains affected by Alzheimer's disease (AD) show a large spectrum of mitochondrial alterations at both morphological and genetic level. The causal link between β-amyloid (Aβ) and mitochondrial dysfunction has been established in cellular models of AD. We observed previously that lycopene, a member of the carotenoid family of phytochemicals, could counteract neuronal apoptosis and cell damage induced by Aβ and other neurotoxic substances, and that this neuroprotective action somehow involved the mitochondria. The present study aims to investigate the effects of lycopene on mitochondria in cultured rat cortical neurons exposed to Aβ. It was found that lycopene attenuated Aβ-induced oxidative stress, as evidenced by the decreased intracellular reactive oxygen species generation and mitochondria-derived superoxide production. Additionally, lycopene ameliorated Aβ-induced mitochondrial morphological alteration, opening of the mitochondrial permeability transition pores and the consequent cytochrome c release. Lycopene also improved mitochondrial complex activities and restored ATP levels in Aβ-treated neuron. Furthermore, lycopene prevented mitochondrial DNA damages and improved the protein level of mitochondrial transcription factor A in mitochondria. Those results indicate that lycopene protects mitochondria against Aβ-induced damages, at least in part by inhibiting mitochondrial oxidative stress and improving mitochondrial function. These beneficial effects of lycopene may account for its protection against Aβ-induced neurotoxicity.

  15. Oxidative stress, cardiolipin and mitochondrial dysfunction in nonalcoholic fatty liver disease

    PubMed Central

    Paradies, Giuseppe; Paradies, Valeria; Ruggiero, Francesca M; Petrosillo, Giuseppe

    2014-01-01

    Nonalcoholic fatty liver disease (NAFLD) is today considered the most common form of chronic liver disease, affecting a high proportion of the population worldwide. NAFLD encompasses a large spectrum of liver damage, ranging from simple steatosis to steatohepatitis, advanced fibrosis and cirrhosis. Obesity, hyperglycemia, type 2 diabetes and hypertriglyceridemia are the most important risk factors. The pathogenesis of NAFLD and its progression to fibrosis and chronic liver disease is still unknown. Accumulating evidence indicates that mitochondrial dysfunction plays a key role in the physiopathology of NAFLD, although the mechanisms underlying this dysfunction are still unclear. Oxidative stress is considered an important factor in producing lethal hepatocyte injury associated with NAFLD. Mitochondrial respiratory chain is the main subcellular source of reactive oxygen species (ROS), which may damage mitochondrial proteins, lipids and mitochondrial DNA. Cardiolipin, a phospholipid located at the level of the inner mitochondrial membrane, plays an important role in several reactions and processes involved in mitochondrial bioenergetics as well as in mitochondrial dependent steps of apoptosis. This phospholipid is particularly susceptible to ROS attack. Cardiolipin peroxidation has been associated with mitochondrial dysfunction in multiple tissues in several physiopathological conditions, including NAFLD. In this review, we focus on the potential roles played by oxidative stress and cardiolipin alterations in mitochondrial dysfunction associated with NAFLD. PMID:25339807

  16. Oxidative stress, cardiolipin and mitochondrial dysfunction in nonalcoholic fatty liver disease.

    PubMed

    Paradies, Giuseppe; Paradies, Valeria; Ruggiero, Francesca M; Petrosillo, Giuseppe

    2014-10-21

    Nonalcoholic fatty liver disease (NAFLD) is today considered the most common form of chronic liver disease, affecting a high proportion of the population worldwide. NAFLD encompasses a large spectrum of liver damage, ranging from simple steatosis to steatohepatitis, advanced fibrosis and cirrhosis. Obesity, hyperglycemia, type 2 diabetes and hypertriglyceridemia are the most important risk factors. The pathogenesis of NAFLD and its progression to fibrosis and chronic liver disease is still unknown. Accumulating evidence indicates that mitochondrial dysfunction plays a key role in the physiopathology of NAFLD, although the mechanisms underlying this dysfunction are still unclear. Oxidative stress is considered an important factor in producing lethal hepatocyte injury associated with NAFLD. Mitochondrial respiratory chain is the main subcellular source of reactive oxygen species (ROS), which may damage mitochondrial proteins, lipids and mitochondrial DNA. Cardiolipin, a phospholipid located at the level of the inner mitochondrial membrane, plays an important role in several reactions and processes involved in mitochondrial bioenergetics as well as in mitochondrial dependent steps of apoptosis. This phospholipid is particularly susceptible to ROS attack. Cardiolipin peroxidation has been associated with mitochondrial dysfunction in multiple tissues in several physiopathological conditions, including NAFLD. In this review, we focus on the potential roles played by oxidative stress and cardiolipin alterations in mitochondrial dysfunction associated with NAFLD.

  17. Increased fatty acid unsaturation and production of arachidonic acid by homologous over-expression of the mitochondrial malic enzyme in Mortierella alpina.

    PubMed

    Hao, Guangfei; Chen, Haiqin; Du, Kai; Huang, Xiaoyun; Song, Yuanda; Gu, Zhennan; Wang, Lei; Zhang, Hao; Chen, Wei; Chen, Yong Q

    2014-09-01

    Malic enzyme (ME) catalyses the oxidative decarboxylation of L-malate to pyruvate and provides NADPH for intracellular metabolism, such as fatty acid synthesis. Here, the mitochondrial ME (mME) gene from Mortierella alpina was homologously over-expressed. Compared with controls, fungal arachidonic acid (ARA; 20:4 n-6) content increased by 60 % without affecting the total fatty acid content. Our results suggest that enhancing mME activity may be an effective mean to increase industrial production of ARA in M. alpina.

  18. Photosynthetic water oxidation vs. mitochondrial oxygen reduction: distinct mechanistic parallels.

    PubMed

    Silverstein, Todd P

    2011-08-01

    The photosynthetic oxygen evolving complex (PSII-OEC) and the mitochondrial cytochrome c oxidase (CcO) not only catalyze anti-parallel reactions (the OEC oxidizes water to dioxygen, whereas CcO reduces dioxygen to water), they also share a number of uncanny molecular and mechanistic similarities. Both feature a redox-active polymetallic cluster that includes a key tyrosine, and both utilize a two-phase mechanism. In one phase the polymetallic cluster undergoes four sequential one-electron transfers: In the PSII-OEC, four successive photooxidations of the photosystem II reaction center P680 (to P680(+)) allows acceptance of 4 × 1e- from the Mn(4)Ca cluster; in CcO, four reduced cytochrome c Fe(2+) cations donate 4 × 1e- to the bimetallic center. In the second phase for each enzyme, the polymetallic cluster undergoes a single four-electron transfer with the O(2)/2 H(2)O redox couple. Intriguing mechanistic similarities between these two complex redox enzymes first delineated over a decade ago by Hoganson/Proshlyakov/Babcock et al. are updated and expanded in this article.

  19. Mitochondrial Oxidative Phosphorylation System (OXPHOS) Deficits in Schizophrenia

    PubMed Central

    Bergman, Oded

    2016-01-01

    Mitochondria are key players in the generation and regulation of cellular bioenergetics, producing the majority of adenosine triphosphate molecules by the oxidative phosphorylation system (OXPHOS). Linked to numerous signaling pathways and cellular functions, mitochondria, and OXPHOS in particular, are involved in neuronal development, connectivity, plasticity, and differentiation. Impairments in a variety of mitochondrial functions have been described in different general and psychiatric disorders, including schizophrenia (SCZ), a severe, chronic, debilitating illness that heavily affects the lives of patients and their families. This article reviews findings emphasizing the role of OXPHOS in the pathophysiology of SCZ. Evidence accumulated during the past few decades from imaging, transcriptomic, proteomic, and metabolomic studies points at OXPHOS deficit involvement in SCZ. Abnormalities have been reported in high-energy phosphates generated by the OXPHOS, in the activity of its complexes and gene expression, primarily of complex I (CoI). In addition, cellular signaling such as cAMP/protein kinase A (PKA) and Ca+2, neuronal development, connectivity, and plasticity have been linked to OXPHOS function and are reported to be impaired in SCZ. Finally, CoI has been shown as a site of interaction for both dopamine (DA) and antipsychotic drugs, further substantiating its role in the pathology of SCZ. Understanding the role of mitochondria and the OXPHOS in particular may encourage new insights into the pathophysiology and etiology of this debilitating disorder. PMID:27412728

  20. THE PRESENCE OF THE OVARY PREVENTS HEPATIC MITOCHONDRIAL OXIDATIVE STRESS IN YOUNG AND AGED FEMALE MICE THROUGH GLUTATHIONE PEROXIDASE 1

    PubMed Central

    Valencia, Ana P.; Schappal, Anna E.; Morris, E. Matthew; Thyfault, John P.; Lowe, Dawn A.; Spangenburg, Espen E.

    2016-01-01

    Background For unknown reasons a woman’s risk for developing the Metabolic Syndrome (MetS) increases dramatically with age and/or loss of ovarian function. The MetS is characterized by hepatic insulin resistance (IR), which is strongly associated with intrahepatic lipid (IHL) accumulation, mitochondrial dysfunction, and oxidative stress. Although circumstantial evidence suggests that the endocrine function of the ovary can directly impact hepatic mitochondrial function, this hypothesis remains untested. Thus, the purpose of this study was to assess the influence of age and secretory function of the ovary on mechanisms that regulate hepatic mitochondrial function. Methods Adult (10 week-old) and aged (88 week-old) female C57BL/6 mice were separated into two groups to undergo bilateral ovariectomy (OVX) or control surgery (SHAM). Eight weeks after surgery hepatic tissue was removed for measurements of total IHL and fatty acid species within hepatic triglycerides, mitochondrial function, and reactive oxygen species (ROS) production. Results Hepatic IHL content was not affected by OVX, but was increased by age. OVX had no effect on mitochondrial respiration, however, hepatic mitochondria from aged mice had lower O2 consumption, lower complex IV and higher complex I content. Mitochondrial H2O2 production was highest in OVX groups and exacerbated by age, while mitochondrial lipid peroxidation was highest in the aged mice and exacerbated by OVX. Regardless of age, OVX resulted in lower mitochondrial content of antioxidant glutathione peroxidase 1 (Gpx1). Isolated liver tissue from a sub-set of animals were acutely treated with conditioned ovarian media which increased Gpx1 mRNA expression compared to vehicle treated liver tissue. Conclusion Ovarian secretory function is necessary for the maintenance of hepatic ROS buffering capacity in the mitochondria, while age significantly influences mitochondrial respiration. These data suggest that when age is coupled with loss of

  1. An update on the role of mitochondrial α-ketoglutarate dehydrogenase in oxidative stress

    PubMed Central

    Starkov, Anatoly A.

    2012-01-01

    The activity of mitochondrial alpha-ketoglutarate dehydrogenase complex (KGDHC) is severely reduced in human pathologies where oxidative stress is traditionally thought to play an important role, such as familial and sporadic forms of Alzheimer's disease and other age-related neurodegenerative diseases. This minireview is focused on substantial data that were accumulated over the last 2 decades to support the concept that KGDHC can be a primary mitochondrial target of oxidative stress and at the same time a key contributor to it by producing reactive oxygen species. This article is part of a Special Issue entitled ‘Mitochondrial function’. PMID:22820180

  2. EFFECT OF PERILLA FRUTESCENS EXTRACTS AND ROSMARINIC ACID ON RAT HEART MITOCHONDRIAL FUNCTIONS.

    PubMed

    Raudone, Lina; Burdulis, Deividas; Raudonis, Raimondas; Janulis, Valdimaras; Jankauskiene, Laima; Viskelis, Pranas; Trumbeckaite, Sonata

    2016-01-01

    Perilla frutescens L. due to its aromatic, antibacterial, anti-inflammatory and antioxidant traits has been traditionally used as medicinal plant in Eastern Asia. Alterations of mitochondria are interconnected with many chronic diseases. Bioactives of herbal extracts can modulate mitochondrial effects and be beneficial in prevention of mitochondrial related chronic diseases. Direct effects of the red-leaf form P. frutescens extract (PFE) and the green-leaf form P. frutescens var. crispa f. viridis extract (PCE) were evaluated investigating activities on the oxidative phosphorylation and antioxidant activity in the rat heart mitochondria in vitro. HPLC-MS analysis was applied for the identification of phenolic compounds. Cell with a Clark-type oxygen electrode was used for mitochondrial respiration measurement. The generation of reactive oxygen species was estimated in isolated rat heart mitochondria and determined fluorimetrically. State 3 respiration rate was not affected by lower concentrations, however, it was inhibited at higher concentrations by 22-70% for PFE and by 45-55% for PCE. PFE containing anthocyanins induced the concentration-dependent stimulation (by 23-76%) of the State 4 respiration rate after addition of cytochrome c due to reducing properties. Significant reduction of H₂O₂ pro- duction was observed with investigated concentrations of rosmarinic acid and both perilla extracts. Our results demonstrate that the effect of PFE and PCE extracts on rat heart mitochondria depend on the qualitative characteristics of complex of biologically active compounds. Selective effects on mitochondrial function could enable the regulation of apoptosis or another mechanisms occurring in cells.

  3. Concentration dependent effect of calcium on brain mitochondrial bioenergetics and oxidative stress parameters

    PubMed Central

    Pandya, Jignesh D.; Nukala, Vidya N.; Sullivan, Patrick G.

    2013-01-01

    Mitochondrial dysfunction following traumatic brain and spinal cord injury (TBI and SCI) plays a pivotal role in the development of secondary pathophysiology and subsequent neuronal cell death. Previously, we demonstrated a loss of mitochondrial bioenergetics in the first 24 h following TBI and SCI initiates a rapid and extensive necrotic event at the primary site of injury. Within the mitochondrial derived mechanisms, the cross talk and imbalance amongst the processes of excitotoxicity, Ca2+ cycling/overload, ATP synthesis, free radical production and oxidative damage ultimately lead to mitochondrial damage followed by neuronal cell death. Mitochondria are one of the important organelles that regulate intracellular calcium (Ca2+) homeostasis and are equipped with a tightly regulated Ca2+ transport system. However, owing to the lack of consensus and the link between downstream effects of calcium in published literature, we undertook a systematic in vitro study for measuring concentration dependent effects of calcium (100–1000 nmols/mg mitochondrial protein) on mitochondrial respiration, enzyme activities, reactive oxygen/nitrogen species (ROS/RNS) generation, membrane potential (ΔΨ) and oxidative damage markers in isolated brain mitochondria. We observed a dose- and time-dependent inhibition of mitochondrial respiration by calcium without influencing mitochondrial pyruvate dehydrogenase complex (PDHC) and NADH dehydrogenase (Complex I) enzyme activities. We observed dose-dependent decreased production of hydrogen peroxide and total ROS/RNS species generation by calcium and no significant changes in protein and lipid oxidative damage markers. These results may shed new light on the prevailing dogma of the direct effects of calcium on mitochondrial bioenergetics, free radical production and oxidative stress parameters that are primary regulatory mitochondrial mechanisms following neuronal injury. PMID:24385963

  4. Mitochondrial ferritin limits oxidative damage regulating mitochondrial iron availability: hypothesis for a protective role in Friedreich ataxia

    PubMed Central

    Campanella, Alessandro; Rovelli, Elisabetta; Santambrogio, Paolo; Cozzi, Anna; Taroni, Franco; Levi, Sonia

    2009-01-01

    Mitochondrial ferritin (FtMt) is a nuclear-encoded iron-sequestering protein that specifically localizes in mitochondria. In mice it is highly expressed in cells characterized by high-energy consumption, while is undetectable in iron storage tissues like liver and spleen. FtMt expression in mammalian cells was shown to cause a shift of iron from cytosol to mitochondria, and in yeast it rescued the defects associated with frataxin deficiency. To study the role of FtMt in oxidative damage, we analyzed the effect of its expression in HeLa cells after incubation with H2O2 and Antimycin A, and after a long-term growth in glucose-free media that enhances mitochondrial respiratory activity. FtMt reduced the level of reactive oxygen species (ROS), increased the level of adenosine 5'triphosphate and the activity of mitochondrial Fe-S enzymes, and had a positive effect on cell viability. Furthermore, FtMt expression reduces the size of cytosolic and mitochondrial labile iron pools. In cells grown in glucose-free media, FtMt level was reduced owing to faster degradation rate, however it still protected the activity of mitochondrial Fe-S enzymes without affecting the cytosolic iron status. In addition, FtMt expression in fibroblasts from Friedreich ataxia (FRDA) patients prevented the formation of ROS and partially rescued the impaired activity of mitochondrial Fe-S enzymes, caused by frataxin deficiency. These results indicate that the primary function of FtMt involves the control of ROS formation through the regulation of mitochondrial iron availability. They are consistent with the expression pattern of FtMt observed in mouse tissues, suggesting a FtMt protective role in cells characterized by defective iron homeostasis and respiration, such as in FRDA. PMID:18815198

  5. Metabolic interplay between glycolysis and mitochondrial oxidation: The reverse Warburg effect and its therapeutic implication

    PubMed Central

    Lee, Minjong; Yoon, Jung-Hwan

    2015-01-01

    Aerobic glycolysis, i.e., the Warburg effect, may contribute to the aggressive phenotype of hepatocellular carcinoma. However, increasing evidence highlights the limitations of the Warburg effect, such as high mitochondrial respiration and low glycolysis rates in cancer cells. To explain such contradictory phenomena with regard to the Warburg effect, a metabolic interplay between glycolytic and oxidative cells was proposed, i.e., the “reverse Warburg effect”. Aerobic glycolysis may also occur in the stromal compartment that surrounds the tumor; thus, the stromal cells feed the cancer cells with lactate and this interaction prevents the creation of an acidic condition in the tumor microenvironment. This concept provides great heterogeneity in tumors, which makes the disease difficult to cure using a single agent. Understanding metabolic flexibility by lactate shuttles offers new perspectives to develop treatments that target the hypoxic tumor microenvironment and overcome the limitations of glycolytic inhibitors. PMID:26322173

  6. Antioxidant enzyme activities and mitochondrial fatty acids in pulmonary hypertension syndrome (PHS) in broilers.

    PubMed

    Iqbal, M; Cawthon, D; Beers, K; Wideman, R F; Bottje, W G

    2002-02-01

    Major objectives of this study were to assess antioxidant protection and fatty acid profile in lung mitochondria and whole liver in broilers with pulmonary hypertension syndrome [(PHS; with and without high dietary vitamin E (VE)] (Experiment 1) and in broilers that did not develop PHS but were genetically selected (S) or not selected (NS) for resistance to PHS (Experiment 2). In Experiment 1, lung mitochondrial glutathione peroxidase (GSH-Px) activity was elevated in broilers with PHS compared to controls, broilers fed high VE, and broilers fed high VE with PHS (VE-PHS), but there were no differences in GSH reductase (GSH-Rd) among groups. In liver tissue, GSH-Px was also elevated by PHS but was lower in VE and VE-PHS groups than in controls. There were no differences in liver GSH-Rd, superoxide dismutase (SOD), or gamma-glutamylcysteine synthetase (gamma-GCS) activities with the exception that gamma-GCS was higher in the VE-PHS group than in the other groups. In Experiment 2, S lung mitochondria exhibited lower GSH-Px and higher GSH-Rd compared to NS broilers. In the liver, there were no differences in GSH-Px, GSH-Rd, or gamma-GCS, but SOD was lower in S compared to the NS broilers. High VE increased the percentage of saturated fatty acids and decreased the percentage of unsaturated fatty acids in lung mitochondria in Experiment 1; there were no differences in fatty acid content between S and NS mitochondria in Experiment 2. Thus, it appears that GSH recycling enzyme activities are affected by PHS and high VE presumably in response to differences in oxidative stress and that genetic resistance to PHS is associated with an inherently better capability to metabolize oxidants in lung mitochondria. The increase in saturation of lung mitochondrial fatty acids with high dietary VE would presumably make them more resistant to oxidative stress and, thus, reduce the level of PHS-induced oxidative stress.

  7. The potential for mitochondrial fat oxidation in human skeletal muscle influences whole body fat oxidation during low-intensity exercise.

    PubMed

    Sahlin, K; Mogensen, M; Bagger, M; Fernström, M; Pedersen, P K

    2007-01-01

    The purpose of this study was to investigate fatty acid (FA) oxidation in isolated mitochondrial vesicles (mit) and its relation to training status, fiber type composition, and whole body FA oxidation. Trained (Vo(2 peak) 60.7 +/- 1.6, n = 8) and untrained subjects (39.5 +/- 2.0 ml.min(-1).kg(-1), n = 5) cycled at 40, 80, and 120 W, and whole body relative FA oxidation was assessed from respiratory exchange ratio (RER). Mit were isolated from muscle biopsies, and maximal ADP stimulated respiration was measured with carbohydrate-derived substrate [pyruvate + malate (Pyr)] and FA-derived substrate [palmitoyl-l-carnitine + malate (PC)]. Fiber type composition was determined from analysis of myosin heavy-chain (MHC) composition. The rate of mit oxidation was lower with PC than with Pyr, and the ratio between PC and Pyr oxidation (MFO) varied greatly between subjects (49-93%). MFO was significantly correlated to muscle fiber type distribution, i.e., %MHC I (r = 0.62, P = 0.03), but was not different between trained (62 +/- 5%) and untrained subjects (72 +/- 2%). MFO was correlated to RER during submaximal exercise at 80 (r = -0.62, P = 0.02) and 120 W (r = -0.71, P = 0.007) and interpolated 35% Vo(2 peak) (r = -0.74, P = 0.004). ADP sensitivity of mit respiration was significantly higher with PC than with Pyr. It is concluded that MFO is influenced by fiber type composition but not by training status. The inverse correlation between RER and MFO implies that intrinsic mit characteristics are of importance for whole body FA oxidation during low-intensity exercise. The higher ADP sensitivity with PC than that with Pyr may influence fuel utilization at low rate of respiration.

  8. Impaired mitochondrial oxidative phosphorylation in the peroxisomal disease X-linked adrenoleukodystrophy.

    PubMed

    López-Erauskin, J; Galino, J; Ruiz, M; Cuezva, J M; Fabregat, I; Cacabelos, D; Boada, J; Martínez, J; Ferrer, I; Pamplona, R; Villarroya, F; Portero-Otín, M; Fourcade, S; Pujol, A

    2013-08-15

    X-linked adrenoleukodystrophy (X-ALD) is an inherited metabolic disorder of the nervous system characterized by axonopathy in spinal cords and/or cerebral demyelination, adrenal insufficiency and accumulation of very long-chain fatty acids (VLCFAs) in plasma and tissues. The disease is caused by malfunction of the ABCD1 gene, which encodes a peroxisomal transporter of VLCFAs or VLCFA-CoA. In the mouse, Abcd1 loss causes late onset axonal degeneration in the spinal cord, associated with locomotor disability resembling the most common phenotype in patients, adrenomyeloneuropathy. We have formerly shown that an excess of the VLCFA C26:0 induces oxidative damage, which underlies the axonal degeneration exhibited by the Abcd1(-) mice. In the present study, we sought to investigate the noxious effects of C26:0 on mitochondria function. Our data indicate that in X-ALD patients' fibroblasts, excess of C26:0 generates mtDNA oxidation and specifically impairs oxidative phosphorylation (OXPHOS) triggering mitochondrial ROS production from electron transport chain complexes. This correlates with impaired complex V phosphorylative activity, as visualized by high-resolution respirometry on spinal cord slices of Abcd1(-) mice. Further, we identified a marked oxidation of key OXPHOS system subunits in Abcd1(-) mouse spinal cords at presymptomatic stages. Altogether, our results illustrate some of the mechanistic intricacies by which the excess of a fatty acid targeted to peroxisomes activates a deleterious process of oxidative damage to mitochondria, leading to a multifaceted dysfunction of this organelle. These findings may be of relevance for patient management while unveiling novel therapeutic targets for X-ALD.

  9. Enhancement of Muscle Mitochondrial Oxidative Capacity and Alterations in Insulin Action Are Lipid Species Dependent

    PubMed Central

    Turner, Nigel; Hariharan, Krit; TidAng, Jennifer; Frangioudakis, Georgia; Beale, Susan M.; Wright, Lauren E.; Zeng, Xiao Yi; Leslie, Simon J.; Li, Jing-Ya; Kraegen, Edward W.; Cooney, Gregory J.; Ye, Ji-Ming

    2009-01-01

    OBJECTIVE Medium-chain fatty acids (MCFAs) have been reported to be less obesogenic than long-chain fatty acids (LCFAs); however, relatively little is known regarding their effect on insulin action. Here, we examined the tissue-specific effects of MCFAs on lipid metabolism and insulin action. RESEARCH DESIGN AND METHODS C57BL6/J mice and Wistar rats were fed either a low-fat control diet or high-fat diets rich in MCFAs or LCFAs for 4–5 weeks, and markers of mitochondrial oxidative capacity, lipid levels, and insulin action were measured. RESULTS Mice fed the MCFA diet displayed reduced adiposity and better glucose tolerance than LCFA-fed animals. In skeletal muscle, triglyceride levels were increased by the LCFA diet (77%, P < 0.01) but remained at low-fat diet control levels in the MCFA-fed animals. The LCFA diet increased (20–50%, P < 0.05) markers of mitochondrial metabolism in muscle compared with low-fat diet–fed controls; however; the increase in oxidative capacity was substantially greater in MCFA-fed animals (50–140% versus low-fat–fed controls, P < 0.01). The MCFA diet induced a greater accumulation of liver triglycerides than the LCFA diet, likely due to an upregulation of several lipogenic enzymes. In rats, isocaloric feeding of MCFA or LCFA high-fat diets induced hepatic insulin resistance to a similar degree; however, insulin action was preserved at the level of low-fat diet–fed controls in muscle and adipose from MCFA-fed animals. CONCLUSIONS MCFAs reduce adiposity and preserve insulin action in muscle and adipose, despite inducing steatosis and insulin resistance in the liver. Dietary supplementation with MCFAs may therefore be beneficial for preventing obesity and peripheral insulin resistance. PMID:19720794

  10. XPD localizes in mitochondria and protects the mitochondrial genome from oxidative DNA damage.

    PubMed

    Liu, Jing; Fang, Hongbo; Chi, Zhenfen; Wu, Zan; Wei, Di; Mo, Dongliang; Niu, Kaifeng; Balajee, Adayabalam S; Hei, Tom K; Nie, Linghu; Zhao, Yongliang

    2015-06-23

    Xeroderma pigmentosum group D (XPD/ERCC2) encodes an ATP-dependent helicase that plays essential roles in both transcription and nucleotide excision repair of nuclear DNA, however, whether or not XPD exerts similar functions in mitochondria remains elusive. In this study, we provide the first evidence that XPD is localized in the inner membrane of mitochondria, and cells under oxidative stress showed an enhanced recruitment of XPD into mitochondrial compartment. Furthermore, mitochondrial reactive oxygen species production and levels of oxidative stress-induced mitochondrial DNA (mtDNA) common deletion were significantly elevated, whereas capacity for oxidative damage repair of mtDNA was markedly reduced in both XPD-suppressed human osteosarcoma (U2OS) cells and XPD-deficient human fibroblasts. Immunoprecipitation-mass spectrometry analysis was used to identify interacting factor(s) with XPD and TUFM, a mitochondrial Tu translation elongation factor was detected to be physically interacted with XPD. Similar to the findings in XPD-deficient cells, mitochondrial common deletion and oxidative damage repair capacity in U2OS cells were found to be significantly altered after TUFM knock-down. Our findings clearly demonstrate that XPD plays crucial role(s) in protecting mitochondrial genome stability by facilitating an efficient repair of oxidative DNA damage in mitochondria.

  11. Maximal oxidative capacity during exercise is associated with skeletal muscle fuel selection and dynamic changes in mitochondrial protein acetylation

    PubMed Central

    Overmyer, Katherine A.; Evans, Charles R.; Qi, Nathan R.; Minogue, Catherine E.; Carson, Joshua J.; Chermside-Scabbo, Christopher J.; Koch, Lauren G.; Britton, Steven L.; Pagliarini, David J.; Coon, Joshua J.; Burant, Charles F.

    2015-01-01

    Summary Maximal exercise-associated oxidative capacity is strongly correlated with health and longevity in humans. Rats selectively bred for high running capacity (HCR) have improved metabolic health and are longer-lived than their low capacity counterparts (LCR). Using metabolomic and proteomic profiling, we show that HCR efficiently oxidize fatty acids (FA) and branched-chain amino acid (BCAA), sparing glycogen and reducing accumulation of short- and medium-chain acylcarnitines. HCR mitochondria have reduced acetylation of mitochondrial proteins within oxidative pathways at rest, and there is rapid protein deacetylation with exercise, which is greater in HCR than LCR. Fluxomic analysis of valine degradation with exercise demonstrates a functional role of differential protein acetylation in HCR and LCR. Our data suggest efficient FA and BCAA utilization contribute to high intrinsic exercise capacity and the health and longevity benefits associated with enhanced fitness. PMID:25738461

  12. Oxidative stress, mitochondrial perturbations and fetal programming of renal disease induced by maternal smoking.

    PubMed

    Stangenberg, Stefanie; Nguyen, Long T; Chen, Hui; Al-Odat, Ibrahim; Killingsworth, Murray C; Gosnell, Martin E; Anwer, Ayad G; Goldys, Ewa M; Pollock, Carol A; Saad, Sonia

    2015-07-01

    An adverse in-utero environment is increasingly recognized to predispose to chronic disease in adulthood. Maternal smoking remains the most common modifiable adverse in-utero exposure leading to low birth weight, which is strongly associated with chronic kidney disease (CKD) in later life. In order to investigate underlying mechanisms for such susceptibility, female Balb/c mice were sham or cigarette smoke-exposed (SE) for 6 weeks before mating, throughout gestation and lactation. Offspring kidneys were examined for oxidative stress, expression of mitochondrial proteins, mitochondrial structure as well as renal functional parameters on postnatal day 1, day 20 (weaning) and week 13 (adult age). From birth throughout adulthood, SE offspring had increased renal levels of mitochondrial-derived reactive oxygen species (ROS), which left a footprint on DNA with increased 8-hydroxydeoxyguanosin (8-OHdG) in kidney tubular cells. Mitochondrial structural abnormalities were seen in SE kidneys at day 1 and week 13 along with a reduction in oxidative phosphorylation (OXPHOS) proteins and activity of mitochondrial antioxidant Manganese superoxide dismutase (MnSOD). Smoke exposure also resulted in increased mitochondrial DNA copy number (day 1-week 13) and lysosome density (day 1 and week 13). The appearance of mitochondrial defects preceded the onset of albuminuria at week 13. Thus, mitochondrial damage caused by maternal smoking may play an important role in development of CKD at adult life.

  13. Oxidants and not alkylating agents induce rapid mtDNA loss and mitochondrial dysfunction

    PubMed Central

    Furda, Amy M.; Marrangoni, Adele M.; Lokshin, Anna; Van Houten, Bennett

    2013-01-01

    Mitochondrial DNA (mtDNA) is essential for proper mitochondrial function and encodes 22 tRNAs, 2 rRNAs and 13 polypeptides that make up subunits of complex I, III, IV, in the electron transport chain and complex V, the ATP synthase. Although mitochondrial dysfunction has been implicated in processes such as premature aging, neurodegeneration, and cancer, it has not been shown whether persistent mtDNA damage causes a loss of oxidative phosphorylation. We addressed this question by treating mouse embryonic fibroblasts with either hydrogen peroxide (H2O2) or the alkylating agent methyl methanesulfonate (MMS) and measuring several endpoints, including mtDNA damage and repair rates using QPCR, levels of mitochondrial- and nuclear-encoded proteins using antibody analysis, and a pharmacologic profile of mitochondria using the Seahorse Extracellular Flux Analyzer. We show that a 60 min treatment with H2O2 causes persistent mtDNA lesions, mtDNA loss, decreased levels of a nuclear-encoded mitochondrial subunit, a loss of ATP-linked oxidative phosphorylation and a loss of total reserve capacity. Conversely, a 60 min treatment with 2 mM MMS causes persistent mtDNA lesions but no mtDNA loss, no decrease in levels of a nuclear-encoded mitochondrial subunit, and no mitochondrial dysfunction. These results suggest that persistent mtDNA damage is not sufficient to cause mitochondrial dysfunction. PMID:22766155

  14. Melatonin improves age-induced fertility decline and attenuates ovarian mitochondrial oxidative stress in mice

    PubMed Central

    Song, Chao; Peng, Wei; Yin, Songna; Zhao, Jiamin; Fu, Beibei; Zhang, Jingcheng; Mao, Tingchao; Wu, Haibo; Zhang, Yong

    2016-01-01

    Increasing evidence shows that melatonin protected against age-related mitochondrial oxidative damage. However, the protective effects of melatonin against ovarian aging has not been explored. Young Kunming females (aged 2–3 months) were fed with melatonin added to drinking water for 6 or 12 months (mo). We found that long-term (12 mo) melatonin treatment significantly reduced ovarian aging, as indicated by substantial increases in litter size, pool of follicles, and telomere length as well as oocyte quantity and quality. Melatonin treatment suppressed ovarian mitochondrial oxidative damage by decreasing mitochondrial reactive oxygen species (mROS) generation, inhibiting apoptosis, repressing collapse of mitochondrial membrane potential and preserving respiratory chain complex activities. Female mice fed with melatonin had enhanced mitochondrial antioxidant activities, thus reducing the risk of mitochondrial oxidative damage cause by free radicals. Notably, melatonin treatment enhanced SIRT3 activity but not the protein expression level, and increased the binding affinity of FoxO3a to the promoters of both superoxide dismutase 2 (SOD2) and catalase (CAT). In conclusion, melatonin exerted protection against aging-induced fertility decline and maintenance of mitochondrial redox balance. PMID:27731402

  15. [Cyclosporin A causes oxidative stress and mitochondrial dysfunction in renal tubular cells].

    PubMed

    Pérez de Hornedo, J; de Arriba, G; Calvino, M; Benito, S; Parra, T

    2007-01-01

    Reactive oxygen species (ROS) have been implicated in cyclosporin A (CsA) nephrotoxicity. As mitochondria are one of the main sources of ROS in cells, we evaluated the role of CsA in mitochondrial structure and function in LLC-PK1 cells. We incubated cells with CsA 1 microM for 24 hours and studies were performed with flow citometry and confocal microscopy. We studied mitochondrial NAD(P)H content, superoxide anion (O2.-) production (MitoSOX Red), oxidation of cardiolipin of inner mitochondrial membrane (NAO) and mitochondrial membrane potential (DIOC2(3)). Also we analyzed the intracellular ROS synthesis (H2DCF-DA) and reduced glutation (GSH) of cells. Our results showed that CsA decreased NAD(P)H and membrane potential, and increased O2.- in mitochondria. CsA also provoked oxidation of cardiolipin. Furthermore, CsA increased intracellular ROS production and decreased GSH content. These results suggest that CsA has crucial effects in mitochondria. CsA modified mitochondrial physiology through the decrease of antioxidant mitochondrial compounds as NAD(P)H and the dissipation of mitochondrial membrane potential and increase of oxidants as O2.-. Also, CsA alters lipidic structure of inner mitochondrial membrane through the oxidation of cardiolipin. These effects trigger a chain of events that favour intracellular synthesis of ROS and depletion of GSH that can compromise cellular viability. Nephrotoxic cellular effects of CsA can be explained, at least in part, through its influence on mitochondrial functionalism.

  16. Fatty acid synthase inhibitors induce apoptosis in non-tumorigenic melan-a cells associated with inhibition of mitochondrial respiration.

    PubMed

    Rossato, Franco A; Zecchin, Karina G; La Guardia, Paolo G; Ortega, Rose M; Alberici, Luciane C; Costa, Rute A P; Catharino, Rodrigo R; Graner, Edgard; Castilho, Roger F; Vercesi, Aníbal E

    2014-01-01

    The metabolic enzyme fatty acid synthase (FASN) is responsible for the endogenous synthesis of palmitate, a saturated long-chain fatty acid. In contrast to most normal tissues, a variety of human cancers overexpress FASN. One such cancer is cutaneous melanoma, in which the level of FASN expression is associated with tumor invasion and poor prognosis. We previously reported that two FASN inhibitors, cerulenin and orlistat, induce apoptosis in B16-F10 mouse melanoma cells via the intrinsic apoptosis pathway. Here, we investigated the effects of these inhibitors on non-tumorigenic melan-a cells. Cerulenin and orlistat treatments were found to induce apoptosis and decrease cell proliferation, in addition to inducing the release of mitochondrial cytochrome c and activating caspases-9 and -3. Transfection with FASN siRNA did not result in apoptosis. Mass spectrometry analysis demonstrated that treatment with the FASN inhibitors did not alter either the mitochondrial free fatty acid content or composition. This result suggests that cerulenin- and orlistat-induced apoptosis events are independent of FASN inhibition. Analysis of the energy-linked functions of melan-a mitochondria demonstrated the inhibition of respiration, followed by a significant decrease in mitochondrial membrane potential (ΔΨm) and the stimulation of superoxide anion generation. The inhibition of NADH-linked substrate oxidation was approximately 40% and 61% for cerulenin and orlistat treatments, respectively, and the inhibition of succinate oxidation was approximately 46% and 52%, respectively. In contrast, no significant inhibition occurred when respiration was supported by the complex IV substrate N,N,N',N'-tetramethyl-p-phenylenediamine (TMPD). The protection conferred by the free radical scavenger N-acetyl-cysteine indicates that the FASN inhibitors induced apoptosis through an oxidative stress-associated mechanism. In combination, the present results demonstrate that cerulenin and orlistat induce

  17. Fatty Acid Synthase Inhibitors Induce Apoptosis in Non-Tumorigenic Melan-A Cells Associated with Inhibition of Mitochondrial Respiration

    PubMed Central

    Rossato, Franco A.; Zecchin, Karina G.; La Guardia, Paolo G.; Ortega, Rose M.; Alberici, Luciane C.; Costa, Rute A. P.; Catharino, Rodrigo R.; Graner, Edgard; Castilho, Roger F.; Vercesi, Aníbal E.

    2014-01-01

    The metabolic enzyme fatty acid synthase (FASN) is responsible for the endogenous synthesis of palmitate, a saturated long-chain fatty acid. In contrast to most normal tissues, a variety of human cancers overexpress FASN. One such cancer is cutaneous melanoma, in which the level of FASN expression is associated with tumor invasion and poor prognosis. We previously reported that two FASN inhibitors, cerulenin and orlistat, induce apoptosis in B16-F10 mouse melanoma cells via the intrinsic apoptosis pathway. Here, we investigated the effects of these inhibitors on non-tumorigenic melan-a cells. Cerulenin and orlistat treatments were found to induce apoptosis and decrease cell proliferation, in addition to inducing the release of mitochondrial cytochrome c and activating caspases-9 and -3. Transfection with FASN siRNA did not result in apoptosis. Mass spectrometry analysis demonstrated that treatment with the FASN inhibitors did not alter either the mitochondrial free fatty acid content or composition. This result suggests that cerulenin- and orlistat-induced apoptosis events are independent of FASN inhibition. Analysis of the energy-linked functions of melan-a mitochondria demonstrated the inhibition of respiration, followed by a significant decrease in mitochondrial membrane potential (ΔΨm) and the stimulation of superoxide anion generation. The inhibition of NADH-linked substrate oxidation was approximately 40% and 61% for cerulenin and orlistat treatments, respectively, and the inhibition of succinate oxidation was approximately 46% and 52%, respectively. In contrast, no significant inhibition occurred when respiration was supported by the complex IV substrate N,N,N′,N′-tetramethyl-p-phenylenediamine (TMPD). The protection conferred by the free radical scavenger N-acetyl-cysteine indicates that the FASN inhibitors induced apoptosis through an oxidative stress-associated mechanism. In combination, the present results demonstrate that cerulenin and orlistat

  18. Indirubin-3'-oxime impairs mitochondrial oxidative phosphorylation and prevents mitochondrial permeability transition induction

    SciTech Connect

    Varela, Ana T.; Gomes, Ana P.; Simoes, Anabela M.; Teodoro, Joao S.; Duarte, Filipe V.; Rolo, Anabela P.; Palmeira, Carlos M.

    2008-12-01

    Indirubin, a red colored 3,2'-bisindole isomer, is a component of Indigo naturalis and is an active ingredient used in traditional Chinese medicine for the treatment of chronic diseases. The family of indirubin derivatives, such as indirubin-3'-oxime, has been suggested for various therapeutic indications. However, potential toxic interactions such as indirubin effects on mitochondrial bioenergetics are still unknown. This study evaluated the action of indirubin-3'-oxime on the function of isolated rat liver mitochondria contributing to a better understanding of the biochemical mechanisms underlying the multiple effects of indirubin. Indirubin-3'-oxime incubated with isolated rat liver mitochondria, at concentrations above 10{mu}M, significantly depresses the phosphorylation efficiency of mitochondria as inferred from the decrease in the respiratory control and ADP/O ratios, the perturbations in mitochondrial membrane potential and in the phosphorylative cycle induced by ADP. Furthermore, indirubin-3'-oxime at up to 25{mu}M stimulates the rate of state 4 respiration and inhibits state 3 respiration. The increased lag phase of repolarization was associated with a direct inhibition of the mitochondrial ATPase. Indirubin-3'-oxime significantly inhibited the activity of complex II and IV thus explaining the decreased FCCP-stimulated mitochondrial respiration. Mitochondria pre-incubated with indirubin-3'-oxime exhibits decreased susceptibility to calcium-induced mitochondrial permeability transition. This work shows for the first time multiple effects of indirubin-3'-oxime on mitochondrial bioenergetics thus indicating a potential mechanism for indirubin-3'-oxime effects on cell function.

  19. The Loss Of Macrophage Fatty Acid Oxidation Does Not Potentiate Systemic Metabolic Dysfunction.

    PubMed

    Gonzalez-Hurtado, Elsie; Lee, Jieun; Choi, Joseph; Selen Alpergin, Ebru S; Collins, Samuel L; Horton, Maureen R; Wolfgang, Michael J

    2017-02-21

    Fatty acid oxidation in macrophages has been suggested to play a causative role in high-fat diet-induced metabolic dysfunction, particularly in the etiology of adipose driven insulin resistance. To understand the contribution of macrophage fatty acid oxidation directly to metabolic dysfunction in high-fat diet-induced obesity, we generated mice with a myeloid-specific knockout of carnitine palmitoyltransferase 2 (CPT2 Mϕ-KO), an obligate step in mitochondrial long-chain fatty acid oxidation. While fatty acid oxidation was clearly induced upon IL-4 stimulation, fatty acid oxidation deficient CPT2 Mϕ-KO bone marrow derived macrophages (BMDM) displayed canonical markers of M2 polarization following IL-4 stimulation in vitro. In addition, loss of macrophage fatty acid oxidation in vivo did not alter the progression of high-fat diet induced obesity, inflammation, macrophage polarization, oxidative stress, or glucose intolerance. These data suggest that although alternatively activated macrophages up-regulate fatty acid oxidation, fatty acid oxidation is dispensable for macrophage polarization and high-fat diet-induced metabolic dysfunction. Macrophage fatty acid oxidation likely plays a correlative rather than causative role in systemic metabolic dysfunction.

  20. An Acidity Scale for Binary Oxides.

    ERIC Educational Resources Information Center

    Smith, Derek W.

    1987-01-01

    Discusses the classification of binary oxides as acidic, basic, or amphoteric. Demonstrates how a numerical scale for acidity/basicity of binary oxides can be constructed using thermochemical data for oxoacid salts. Presents the calculations derived from the data that provide the numeric scale values. (TW)

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

    PubMed

    Mordas, Amelia; Tokatlidis, Kostas

    2015-08-18

    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.

  2. The MIA Pathway: A Key Regulator of Mitochondrial Oxidative Protein Folding and Biogenesis

    PubMed Central

    2015-01-01

    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. PMID:26214018

  3. Effect of endogenous nitric oxide on mitochondrial respiration of rat hepatocytes in vitro and in vivo

    SciTech Connect

    Stadler, J.; Curran, R.D.; Ochoa, J.B.; Harbrecht, B.G.; Hoffman, R.A.; Simmons, R.L.; Billiar, T.R. )

    1991-02-01

    Nitric oxide, a highly reactive radical, was recently identified as an intermediate of L-arginine metabolism in mammalian cells. We have shown that nitric oxide synthesis is induced in vitro in cultured hepatocytes by supernatants from activated Kupffer cells or in vivo by injecting rats with nonviable Corynebacterium parvum. In both cases, nitric oxide biosynthesis in hepatocytes was associated with suppression of total protein synthesis. This study attempts to determine the effect of nitric oxide biosynthesis on the activity of specific hepatocytic mitochondrial enzymes and to determine whether inhibition of protein synthesis is caused by suppression of energy metabolism. Exposure of hepatocytes to supernatants from activated Kupffer cells led to a 30% decrease of aconitase (Krebs cycle) and complex I (mitochondrial electron transport chain) activity. Using NG-monomethyl-L-arginine, an inhibitor of nitric oxide synthesis, we demonstrated that the inhibition of mitochondrial aconitase activity was due, in part, to the action of nitric oxide. In contrast, in vivo nitric oxide synthesis of hepatocytes from Corynebacterium parvum-treated animals had no effect on mitochondrial respiration. This suggests that inhibition of protein synthesis by nitric oxide is not likely to be mediated by inhibition of energy metabolism.

  4. Sab (Sh3bp5) dependence of JNK mediated inhibition of mitochondrial respiration in palmitic acid induced hepatocyte lipotoxicity

    PubMed Central

    Win, Sanda; Than, Tin Aung; Le, Bao Han Allison; García-Ruiz, Carmen; Fernandez-Checa, Jose C; Kaplowitz, Neil

    2015-01-01

    Background & Aims Sustained JNK activation by saturated fatty acids plays a role in lipotoxicity and the pathogenesis of NASH. We have reported that the interaction of JNK with mitochondrial Sab leads to inhibition of respiration, increased ROS, cell death and hepatotoxicity. We tested whether this pathway underlies palmitic acid (PA)-induced lipotoxicity in hepatocytes. Methods Primary mouse hepatocytes from adeno-shlacZ or adeno-shSab treated mice and Huh7 cells were used. Results In PMH, PA dose dependently up to 1mM stimulated oxygen consumption rate (OCR) due to mitochondrial β-oxidation. At ≥ 1.5mM, PA gradually reduced OCR, followed by cell death. Inhibition of JNK, caspases or treatment with antioxidant butylated hydroxyanisole (BHA) protected PMH against cell death. Sab knockdown or a membrane permeable Sab blocking peptide prevented PA-induced mitochondrial impairment, but inhibited only the late phase of both JNK activation (beyond 4 hours) and cell death. PA increased P-PERK and downstream target CHOP in PMH but failed to activate the IRE-1α arm of the UPR. However, Sab silencing did not affect PA-induced PERK activation. Conversely, specific inhibition of PERK prevented JNK activation and cell death, indicating a major role upstream of JNK activation. Conclusions The effect of P-JNK on mitochondria plays a key role in PA-mediated lipotoxicity. The interplay of P-JNK with mitochondrial Sab leads to impaired respiration, ROS production, sustained JNK activation, and apoptosis. PMID:25666017

  5. Curcumin prevents mitochondrial dynamics disturbances in early 5/6 nephrectomy: Relation to oxidative stress and mitochondrial bioenergetics.

    PubMed

    Aparicio-Trejo, Omar Emiliano; Tapia, Edilia; Molina-Jijón, Eduardo; Medina-Campos, Omar Noel; Macías-Ruvalcaba, Norma Angélica; León-Contreras, Juan Carlos; Hernández-Pando, Rogelio; García-Arroyo, Fernando E; Cristóbal, Magdalena; Sánchez-Lozada, Laura Gabriela; Pedraza-Chaverri, José

    2016-11-01

    Five-sixths nephrectomy (5/6NX) is a widely used model to study the mechanisms leading to renal damage in chronic kidney disease (CKD). However, early alterations on renal function, mitochondrial dynamics, and oxidative stress have not been explored yet. Curcumin is an antioxidant that has shown nephroprotection in 5/6NX-induced renal damage. The aim of this study was to explore the effect of curcumin on early mitochondrial alterations induced by 5/6NX in rats. In isolated mitochondria, 5/6NX-induced hydrogen peroxide production was associated with decreased activity of complexes I and V, decreased activity of antioxidant enzymes, alterations in oxygen consumption and increased MDA-protein adducts. In addition, it was found that 5/6NX shifted mitochondrial dynamics to fusion, which was evidenced by increased optic atrophy 1 and mitofusin 1 (Mfn1) and decreased fission 1 and dynamin-related protein 1 expressions. These data were confirmed by morphological analysis and immunoelectron microscopy of Mfn-1. All the above-described mechanisms were prevented by curcumin. Also, it was found that curcumin prevented renal dysfunction by improving renal blood flow and the total antioxidant capacity induced by 5/6NX. Moreover, in glomeruli and proximal tubules 5/6NX-induced superoxide anion production by uncoupled nitric oxide synthase (NOS) and nicotinamide adenine dinucleotide phosphate oxidase (NOX) dependent way, this latter was associated with increased phosphorylation of serine 304 of p47phox subunit of NOX. In conclusion, this study shows that curcumin pretreatment decreases early 5/6NX-induced altered mitochondrial dynamics, bioenergetics, and oxidative stress, which may be associated with the preservation of renal function. © 2016 BioFactors, 00(00):000000, 2016.

  6. Therapeutic Strategies for Mitochondrial Dysfunction and Oxidative Stress in Age-Related Metabolic Disorders.

    PubMed

    Bhatti, J S; Kumar, S; Vijayan, M; Bhatti, G K; Reddy, P H

    2017-01-01

    Mitochondria are complex, intercellular organelles present in the cells and are involved in multiple roles including ATP formation, free radicals generation and scavenging, calcium homeostasis, cellular differentiation, and cell death. Many studies depicted the involvement of mitochondrial dysfunction and oxidative damage in aging and pathogenesis of age-related metabolic disorders and neurodegenerative diseases. Remarkable advancements have been made in understanding the structure, function, and physiology of mitochondria in metabolic disorders such as diabetes, obesity, cardiovascular diseases, and stroke. Further, much progress has been done in the improvement of therapeutic strategies, including lifestyle interventions, pharmacological, and mitochondria-targeted therapeutic approaches. These strategies were mainly focused to reduce the mitochondrial dysfunction caused by oxidative stress and to retain the mitochondrial health in various diseases. In this chapter, we have highlighted the involvement of mitochondrial dysfunction in the pathophysiology of various disorders and recent progress in the development of mitochondria-targeted molecules as therapeutic measures for metabolic disorders.

  7. Mitochondrial ROS regulate oxidative damage and mitophagy but not age-related muscle fiber atrophy

    PubMed Central

    Sakellariou, Giorgos K.; Pearson, Timothy; Lightfoot, Adam P.; Nye, Gareth A.; Wells, Nicola; Giakoumaki, Ifigeneia I.; Vasilaki, Aphrodite; Griffiths, Richard D.; Jackson, Malcolm J.; McArdle, Anne

    2016-01-01

    Age-related loss of skeletal muscle mass and function is a major contributor to morbidity and has a profound effect on the quality of life of older people. The potential role of age-dependent mitochondrial dysfunction and cumulative oxidative stress as the underlying cause of muscle aging remains a controversial topic. Here we show that the pharmacological attenuation of age-related mitochondrial redox changes in muscle with SS31 is associated with some improvements in oxidative damage and mitophagy in muscles of old mice. However, this treatment failed to rescue the age-related muscle fiber atrophy associated with muscle atrophy and weakness. Collectively, these data imply that the muscle mitochondrial redox environment is not a key regulator of muscle fiber atrophy during sarcopenia but may play a key role in the decline of mitochondrial organelle integrity that occurs with muscle aging. PMID:27681159

  8. Aluminium induced oxidative stress results in decreased mitochondrial biogenesis via modulation of PGC-1α expression.

    PubMed

    Sharma, Deep Raj; Sunkaria, Aditya; Wani, Willayat Yousuf; Sharma, Reeta Kumari; Kandimalla, Ramesh J L; Bal, Amanjit; Gill, Kiran Dip

    2013-12-01

    The present investigation was carried out to elucidate a possible molecular mechanism related to the effects of aluminium-induced oxidative stress on various mitochondrial respiratory complex subunits with special emphasis on the role of Peroxisome proliferator activated receptor gamma co-activator 1α (PGC-1α) and its downstream targets i.e. Nuclear respiratory factor-1(NRF-1), Nuclear respiratory factor-2(NRF-2) and Mitochondrial transcription factor A (Tfam) in mitochondrial biogenesis. Aluminium lactate (10mg/kgb.wt./day) was administered intragastrically to rats for 12 weeks. After 12 weeks of exposure, we found an increase in ROS levels, mitochondrial DNA oxidation and decrease in citrate synthase activity in the Hippocampus (HC) and Corpus striatum (CS) regions of rat brain. On the other hand, there was a decrease in the mRNA levels of the mitochondrial encoded subunits-NADH dehydrogenase (ND) subunits i.e. ND1, ND2, ND3, Cytochrome b (Cytb), Cytochrome oxidase (COX) subunits i.e. COX1, COX3, ATP synthase (ATPase) subunit 6 along with reduced expression of nuclear encoded subunits COX4, COX5A, COX5B of Electron transport chain (ETC). Besides, a decrease in mitochondrial DNA copy number and mitochondrial content in both regions of rat brain was observed. The PGC-1α was down-regulated in aluminium treated rats along with NRF-1, NRF-2 and Tfam, which act downstream from PGC-1α in aluminium treated rats. Electron microscopy results revealed a significant increase in the mitochondrial swelling, loss of cristae, chromatin condensation and decreases in mitochondrial number in case of aluminium treated rats as compared to control. So, PGC-1α seems to be a potent target for aluminium neurotoxicity, which makes it an almost ideal target to control or limit the damage that has been associated with the defective mitochondrial function seen in neurodegenerative diseases.

  9. Protective effect of montelukast against quinolinic acid/malonic acid induced neurotoxicity: possible behavioral, biochemical, mitochondrial and tumor necrosis factor-α level alterations in rats.

    PubMed

    Kalonia, H; Kumar, P; Kumar, A; Nehru, B

    2010-11-24

    The present study has been designed to explore the protective effect of montelukast (leukotriene receptor antagonist) against intrastriatal quinolinic acid (QA; 300 nmol) and malonic acid (MA; 6 μmol) induced Huntington's like symptoms in rats. Quinolinic acid has been reported to induce excitotoxicity by stimulating the N-methyl-D-aspartate receptor, causing calcium overload which in turn leads to the neurodegeneration. On the other hand, MA, being a reversible inhibitor of mitochondrial enzyme complex-II, leads to energy crisis and free radical generation. Recent studies have reported the therapeutic potential of leukotriene receptor antagonists in different neurodegenerative disorders. However, their exact role is yet to be established. The present study accordingly, is an attempt to investigate the effect of montelukast against QA and MA induced behavioral, biochemical and molecular alterations in rat striatum. Oxidative stress, mitochondrial enzyme complex and tumor necrosis factor-alpha (TNF-α) were evaluated on day 21st and 14th post intrastriatal QA and MA treatment, respectively. Findings of the present study demonstrate significant alteration in the locomotor activity and motor coordination as well as oxidative burden (increased lipid peroxidation, nitrite concentration and decreased endogenous antioxidants), mitochondrial enzyme complex (I, II and IV) activities and TNF-α level, in both intrastriatal QA and MA treated animals. Further, montelukast (0.4, 0.8 mg/kg p.o.) treatment for 21 and 14 days respectively, attenuated the behavioral alterations, oxidative stress, mitochondrial dysfunction and TNF-α level in these models of Huntington's disease in a significant manner. In conclusion, the present study emphasizes the neuroprotective potential of montelukast in the therapeutic management of Huntington like symptoms.

  10. Impaired Mitochondrial Energy Production Causes Light-Induced Photoreceptor Degeneration Independent of Oxidative Stress.

    PubMed

    Jaiswal, Manish; Haelterman, Nele A; Sandoval, Hector; Xiong, Bo; Donti, Taraka; Kalsotra, Auinash; Yamamoto, Shinya; Cooper, Thomas A; Graham, Brett H; Bellen, Hugo J

    2015-07-01

    Two insults often underlie a variety of eye diseases including glaucoma, optic atrophy, and retinal degeneration--defects in mitochondrial function and aberrant Rhodopsin trafficking. Although mitochondrial defects are often associated with oxidative stress, they have not been linked to Rhodopsin trafficking. In an unbiased forward genetic screen designed to isolate mutations that cause photoreceptor degeneration, we identified mutations in a nuclear-encoded mitochondrial gene, ppr, a homolog of human LRPPRC. We found that ppr is required for protection against light-induced degeneration. Its function is essential to maintain membrane depolarization of the photoreceptors upon repetitive light exposure, and an impaired phototransduction cascade in ppr mutants results in excessive Rhodopsin1 endocytosis. Moreover, loss of ppr results in a reduction in mitochondrial RNAs, reduced electron transport chain activity, and reduced ATP levels. Oxidative stress, however, is not induced. We propose that the reduced ATP level in ppr mutants underlies the phototransduction defect, leading to increased Rhodopsin1 endocytosis during light exposure, causing photoreceptor degeneration independent of oxidative stress. This hypothesis is bolstered by characterization of two other genes isolated in the screen, pyruvate dehydrogenase and citrate synthase. Their loss also causes a light-induced degeneration, excessive Rhodopsin1 endocytosis and reduced ATP without concurrent oxidative stress, unlike many other mutations in mitochondrial genes that are associated with elevated oxidative stress and light-independent photoreceptor demise.

  11. Crystal structures of oxidized and reduced forms of human mitochondrial thioredoxin 2

    PubMed Central

    Smeets, Aude; Evrard, Christine; Landtmeters, Marie; Marchand, Cécile; Knoops, Bernard; Declercq, Jean-Paul

    2005-01-01

    Mammalian thioredoxin 2 is a mitochondrial isoform of highly evolutionary conserved thioredoxins. Thioredoxins are small ubiquitous protein–disulfide oxidoreductases implicated in a large variety of biological functions. In mammals, thioredoxin 2 is encoded by a nuclear gene and is targeted to mitochondria by a N-terminal mitochondrial presequence. Recently, mitochondrial thioredoxin 2 was shown to interact with components of the mitochondrial respiratory chain and to play a role in the control of mitochondrial membrane potential, regulating mitochondrial apoptosis signaling pathway. Here we report the first crystal structures of a mammalian mitochondrial thioredoxin 2. Crystal forms of reduced and oxidized human thioredoxin 2 are described at 2.0 and 1.8 Å resolution. Though the folding is rather similar to that of human cytosolic/nuclear thioredoxin 1, important differences are observed during the transition between the oxidized and the reduced states of human thioredoxin 2, compared with human thioredoxin 1. In spite of the absence of the Cys residue implicated in dimer formation in human thioredoxin 1, dimerization still occurs in the crystal structure of human thioredoxin 2, mainly mediated by hydrophobic contacts, and the dimers are associated to form two-dimensional polymers. Interestingly, the structure of human thioredoxin 2 reveals possible interaction domains with human peroxiredoxin 5, a substrate protein of human thioredoxin 2 in mitochondria. PMID:16195549

  12. Effect of Syzygium cumini and Bauhinia forficata aqueous-leaf extracts on oxidative and mitochondrial parameters in vitro

    PubMed Central

    Ecker, Assis; Araujo Vieira, Francielli; de Souza Prestes, Alessandro; Mulling dos Santos, Matheus; Ramos, Angelica; Dias Ferreira, Rafael; Teixeira de Macedo, Gabriel; Vargas Klimaczewski, Claudia; Lopes Seeger, Rodrigo; Teixeira da Rocha, João Batista; de Vargas Barbosa, Nilda B.

    2015-01-01

    Aqueous-leaf extract of Syzygium cumini and Bauhinia forficata are traditionally used in the treatment of diabetes and cancer, especially in South America, Africa, and Asia. In this study, we analyzed the effects of these extracts on oxidative and mitochondrial parameters in vitro, as well as their protective activities against toxic agents. Phytochemical screenings of the extracts were carried out by HPLC analysis. The in vitro antioxidant capacities were compared by DPPH radical scavenging and Fe2+ chelating activities. Mitochondrial parameters observed were swelling, lipid peroxidation and dehydrogenase activity. The major chemical constituent of S. cumini was rutin. In B. forficata were predominant quercetin and gallic acid. S. cumini reduced DPPH radical more than B. forficata, and showed iron chelating activity at all tested concentrations, while B. forficata had not similar property. In mitochondria, high concentrations of B. forficata alone induced a decrease in mitochondrial dehydrogenase activity, but low concentrations of this extract prevented the effect induced by Fe2++H2O2. This was also observed with high concentrations of S. cumini. Both extracts partially prevented the lipid peroxidation induced by Fe2+/citrate. S. cumini was effective against mitochondrial swelling induced by Ca2+, while B. forficata alone induced swelling more than Ca2+. This study suggests that leaf extract of S. cumini might represent a useful therapeutic for the treatment of diseases related with mitochondrial dysfunctions. On the other hand, the consumption of B. forficata should be avoided because mitochondrial damages were observed, and this possibly may pose risk to human health. PMID:27152111

  13. Effect of Syzygium cumini and Bauhinia forficata aqueous-leaf extracts on oxidative and mitochondrial parameters in vitro.

    PubMed

    Ecker, Assis; Araujo Vieira, Francielli; de Souza Prestes, Alessandro; Mulling Dos Santos, Matheus; Ramos, Angelica; Dias Ferreira, Rafael; Teixeira de Macedo, Gabriel; Vargas Klimaczewski, Claudia; Lopes Seeger, Rodrigo; Teixeira da Rocha, João Batista; de Vargas Barbosa, Nilda B

    2015-01-01

    Aqueous-leaf extract of Syzygium cumini and Bauhinia forficata are traditionally used in the treatment of diabetes and cancer, especially in South America, Africa, and Asia. In this study, we analyzed the effects of these extracts on oxidative and mitochondrial parameters in vitro, as well as their protective activities against toxic agents. Phytochemical screenings of the extracts were carried out by HPLC analysis. The in vitro antioxidant capacities were compared by DPPH radical scavenging and Fe(2+) chelating activities. Mitochondrial parameters observed were swelling, lipid peroxidation and dehydrogenase activity. The major chemical constituent of S. cumini was rutin. In B. forficata were predominant quercetin and gallic acid. S. cumini reduced DPPH radical more than B. forficata, and showed iron chelating activity at all tested concentrations, while B. forficata had not similar property. In mitochondria, high concentrations of B. forficata alone induced a decrease in mitochondrial dehydrogenase activity, but low concentrations of this extract prevented the effect induced by Fe(2+)+H2O2. This was also observed with high concentrations of S. cumini. Both extracts partially prevented the lipid peroxidation induced by Fe(2+)/citrate. S. cumini was effective against mitochondrial swelling induced by Ca(2+), while B. forficata alone induced swelling more than Ca(2+). This study suggests that leaf extract of S. cumini might represent a useful therapeutic for the treatment of diseases related with mitochondrial dysfunctions. On the other hand, the consumption of B. forficata should be avoided because mitochondrial damages were observed, and this possibly may pose risk to human health.

  14. Aluminium induced oxidative stress results in decreased mitochondrial biogenesis via modulation of PGC-1α expression

    SciTech Connect

    Sharma, Deep Raj; Sunkaria, Aditya; Wani, Willayat Yousuf; Sharma, Reeta Kumari; Kandimalla, Ramesh J.L.; Bal, Amanjit; Gill, Kiran Dip

    2013-12-01

    The present investigation was carried out to elucidate a possible molecular mechanism related to the effects of aluminium-induced oxidative stress on various mitochondrial respiratory complex subunits with special emphasis on the role of Peroxisome proliferator activated receptor gamma co-activator 1α (PGC-1α) and its downstream targets i.e. Nuclear respiratory factor-1(NRF-1), Nuclear respiratory factor-2(NRF-2) and Mitochondrial transcription factor A (Tfam) in mitochondrial biogenesis. Aluminium lactate (10 mg/kg b.wt./day) was administered intragastrically to rats for 12 weeks. After 12 weeks of exposure, we found an increase in ROS levels, mitochondrial DNA oxidation and decrease in citrate synthase activity in the Hippocampus (HC) and Corpus striatum (CS) regions of rat brain. On the other hand, there was a decrease in the mRNA levels of the mitochondrial encoded subunits–NADH dehydrogenase (ND) subunits i.e. ND1, ND2, ND3, Cytochrome b (Cytb), Cytochrome oxidase (COX) subunits i.e. COX1, COX3, ATP synthase (ATPase) subunit 6 along with reduced expression of nuclear encoded subunits COX4, COX5A, COX5B of Electron transport chain (ETC). Besides, a decrease in mitochondrial DNA copy number and mitochondrial content in both regions of rat brain was observed. The PGC-1α was down-regulated in aluminium treated rats along with NRF-1, NRF-2 and Tfam, which act downstream from PGC-1α in aluminium treated rats. Electron microscopy results revealed a significant increase in the mitochondrial swelling, loss of cristae, chromatin condensation and decreases in mitochondrial number in case of aluminium treated rats as compared to control. So, PGC-1α seems to be a potent target for aluminium neurotoxicity, which makes it an almost ideal target to control or limit the damage that has been associated with the defective mitochondrial function seen in neurodegenerative diseases. - Highlights: • Aluminium decreases the mRNA levels of mitochondrial and nuclear encoded

  15. Alteration of Fatty-Acid-Metabolizing Enzymes Affects Mitochondrial Form and Function in Hereditary Spastic Paraplegia

    PubMed Central

    Tesson, Christelle; Nawara, Magdalena; Salih, Mustafa A.M.; Rossignol, Rodrigue; Zaki, Maha S.; Al Balwi, Mohammed; Schule, Rebecca; Mignot, Cyril; Obre, Emilie; Bouhouche, Ahmed; Santorelli, Filippo M.; Durand, Christelle M.; Oteyza, Andrés Caballero; El-Hachimi, Khalid H.; Al Drees, Abdulmajeed; Bouslam, Naima; Lamari, Foudil; Elmalik, Salah A.; Kabiraj, Mohammad M.; Seidahmed, Mohammed Z.; Esteves, Typhaine; Gaussen, Marion; Monin, Marie-Lorraine; Gyapay, Gabor; Lechner, Doris; Gonzalez, Michael; Depienne, Christel; Mochel, Fanny; Lavie, Julie; Schols, Ludger; Lacombe, Didier; Yahyaoui, Mohamed; Al Abdulkareem, Ibrahim; Zuchner, Stephan; Yamashita, Atsushi; Benomar, Ali; Goizet, Cyril; Durr, Alexandra; Gleeson, Joseph G.; Darios, Frederic; Brice, Alexis; Stevanin, Giovanni

    2012-01-01

    Hereditary spastic paraplegia (HSP) is considered one of the most heterogeneous groups of neurological disorders, both clinically and genetically. The disease comprises pure and complex forms that clinically include slowly progressive lower-limb spasticity resulting from degeneration of the corticospinal tract. At least 48 loci accounting for these diseases have been mapped to date, and mutations have been identified in 22 genes, most of which play a role in intracellular trafficking. Here, we identified mutations in two functionally related genes (DDHD1 and CYP2U1) in individuals with autosomal-recessive forms of HSP by using either the classical positional cloning or a combination of whole-genome linkage mapping and next-generation sequencing. Interestingly, three subjects with CYP2U1 mutations presented with a thin corpus callosum, white-matter abnormalities, and/or calcification of the basal ganglia. These genes code for two enzymes involved in fatty-acid metabolism, and we have demonstrated in human cells that the HSP pathophysiology includes alteration of mitochondrial architecture and bioenergetics with increased oxidative stress. Our combined results focus attention on lipid metabolism as a critical HSP pathway with a deleterious impact on mitochondrial bioenergetic function. PMID:23176821

  16. Mitochondrial dysfunction and oxidative stress in metabolic disorders - A step towards mitochondria based therapeutic strategies.

    PubMed

    Bhatti, Jasvinder Singh; Bhatti, Gurjit Kaur; Reddy, P Hemachandra

    2016-11-09

    Mitochondria are the powerhouses of the cell and are involved in essential functions of the cell, including ATP production, intracellular Ca(2+) regulation, reactive oxygen species production & scavenging, regulation of apoptotic cell death and activation of the caspase family of proteases. Mitochondrial dysfunction and oxidative stress are largely involved in aging, cancer, age-related neurodegenerative and metabolic syndrome. In the last decade, tremendous progress has been made in understanding mitochondrial structure, function and their physiology in metabolic syndromes such as diabetes, obesity, stroke and hypertension, and heart disease. Further, progress has also been made in developing therapeutic strategies, including lifestyle interventions (healthy diet and regular exercise), pharmacological strategies and mitochondria-targeted approaches. These strategies were mainly focused to reduce mitochondrial dysfunction and oxidative stress and to maintain mitochondrial quality in metabolic syndromes. The purpose of our article is to highlight the recent progress on the mitochondrial role in metabolic syndromes and also summarize the progress of mitochondria-targeted molecules as therapeutic targets to treat metabolic syndromes. This article is part of a Special Issue entitled: Oxidative Stress and Mitochondrial Quality in Diabetes/Obesity and Critical Illness Spectrum of Diseases - edited by P. Hemachandra Reddy.

  17. FAT/CD36 is located on the outer mitochondrial membrane, upstream of long-chain acyl-CoA synthetase, and regulates palmitate oxidation.

    PubMed

    Smith, Brennan K; Jain, Swati S; Rimbaud, Stéphanie; Dam, Aaron; Quadrilatero, Joe; Ventura-Clapier, Renée; Bonen, Arend; Holloway, Graham P

    2011-07-01

    FAT/CD36 (fatty acid translocase/Cluster of Differentiation 36), a plasma membrane fatty-acid transport protein, has been found on mitochondrial membranes; however, it remains unclear where FAT/CD36 resides on this organelle or its functional role within mitochondria. In the present study, we demonstrate, using several different approaches, that in skeletal muscle FAT/CD36 resides on the OMM (outer mitochondrial membrane). To determine the functional role of mitochondrial FAT/CD36 in this tissue, we determined oxygen consumption rates in permeabilized muscle fibres in WT (wild-type) and FAT/CD36-KO (knockout) mice using a variety of substrates. Despite comparable muscle mitochondrial content, as assessed by unaltered mtDNA (mitochondrial DNA), citrate synthase, β-hydroxyacyl-CoA dehydrogenase, cytochrome c oxidase complex IV and respiratory capacities [maximal OXPHOS (oxidative phosphorylation) respiration] in WT and KO mice, palmitate-supported respiration was 34% lower in KO animals. In contrast, palmitoyl-CoA-supported respiration was unchanged. These results indicate that FAT/CD36 is key for palmitate-supported respiration. Therefore we propose a working model of mitochondrial fatty-acid transport, in which FAT/CD36 is positioned on the OMM, upstream of long-chain acyl-CoA synthetase, thereby contributing to the regulation of mitochondrial fatty-acid transport. We further support this model by providing evidence that FAT/CD36 is not located in mitochondrial contact sites, and therefore does not directly interact with carnitine palmitoyltransferase-I as original proposed.

  18. Palmitoleic acid induces the cardiac mitochondrial membrane permeability transition despite the presence of L-carnitine.

    PubMed

    Oyanagi, Eri; Uchida, Masataka; Miyakawa, Takeshi; Miyachi, Motohiko; Yamaguchi, Hidetaka; Nagami, Kuniatsu; Utsumi, Kozo; Yano, Hiromi

    Although palmitoleic acid (C16:1) is associated with arrhythmias, and increases in an age-dependent matter, the effects of L-carnitine, which is essential for the transport of long-chain fatty acids into the mitochondria, are unclear. It has been postulated that L-carnitine may attenuate palmitate (C16:0)-induced mitochondrial dysfunction and the apoptosis of cardiomyocytes. The aim of this study was to elucidate the activity of L-carnitine in the prevention of the palmitoleic acid-induced mitochondrial membrane permeability transition and cytochrome c release using isolated cardiac mitochondria from rats. Palmitoleoyl-CoA-induced mitochondrial respiration was not accelerated by L-carnitine treatment, and this respiration was slightly inhibited by oligomycin, which is an inhibitor of ATP synthase. Despite pretreatment with L-carnitine, the mitochondrial membrane potential decreased and mitochondrial swelling was induced by palmitoleoyl-CoA. In the presence of a combination of L-carnitine and tiron, a free radical scavenger, there was attenuated mitochondrial swelling and cytochrome c release following palmitoleoyl-CoA treatment. We concluded that palmitoleic acid, but not palmitate, induces the cardiac mitochondrial membrane permeability transition despite the presence of L-carnitine.

  19. Liver fatty acid binding protein is required for high rates of hepatic fatty acid oxidation but not for the action of PPARalpha in fasting mice.

    PubMed

    Erol, Erdal; Kumar, Leena S; Cline, Gary W; Shulman, Gerald I; Kelly, Daniel P; Binas, Bert

    2004-02-01

    Liver fatty acid binding protein (L-FABP) has been proposed to limit the availability of long-chain fatty acids (LCFA) for oxidation and for peroxisome proliferator-activated receptor alpha (PPAR-alpha), a fatty acid binding transcription factor that determines the capacity of hepatic fatty acid oxidation. Here, we used L-FABP null mice to test this hypothesis. Under fasting conditions, this mutation reduced beta-hydroxybutyrate (BHB) plasma levels as well as BHB release and palmitic acid oxidation by isolated hepatocytes. However, the capacity for ketogenesis was not reduced: BHB plasma levels were restored by octanoate injection; BHB production and palmitic acid oxidation were normal in liver homogenates; and hepatic expression of key PPAR-alpha target (MCAD, mitochondrial HMG CoA synthase, ACO, CYP4A3) and other (CPT1, LCAD) genes of mitochondrial and extramitochondrial LCFA oxidation and ketogenesis remained at wild-type levels. During standard diet, mitochondrial HMG CoA synthase mRNA was selectively reduced in L-FABP null liver. These results suggest that under fasting conditions, hepatic L-FABP contributes to hepatic LCFA oxidation and ketogenesis by a nontranscriptional mechanism, whereas L-FABP can activate ketogenic gene expression in fed mice. Thus, the mechanisms whereby L-FABP affects fatty acid oxidation may vary with physiological condition.

  20. Polyunsaturated Fatty Acids Attenuate Diet Induced Obesity and Insulin Resistance, Modulating Mitochondrial Respiratory Uncoupling in Rat Skeletal Muscle

    PubMed Central

    Bergamo, Paolo; De Filippo, Chiara; Mattace Raso, Giuseppina; Gifuni, Giorgio; Putti, Rosalba; Moni, Bottu Heleena; Canani, Roberto Berni; Meli, Rosaria; Mollica, Maria Pina

    2016-01-01

    Objectives Omega (ω)-3 polyunsaturated fatty acids (PUFA) are dietary compounds able to attenuate insulin resistance. Anyway, the precise actions of ω-3PUFAs in skeletal muscle are overlooked. We hypothesized that PUFAs, modulating mitochondrial function and efficiency, would ameliorate pro-inflammatory and pro-oxidant signs of nutritionally induced obesity. Study Design To this aim, rats were fed a control diet (CD) or isocaloric high fat diets containing either ω-3 PUFA (FD) or lard (LD) for 6 weeks. Results FD rats showed lower weight, lipid gain and energy efficiency compared to LD-fed animals, showing higher energy expenditure and O2 consumption/CO2 production. Serum lipid profile and pro-inflammatory parameters in FD-fed animals were reduced compared to LD. Accordingly, FD rats exhibited a higher glucose tolerance revealed by an improved glucose and insulin tolerance tests compared to LD, accompanied by a restoration of insulin signalling in skeletal muscle. PUFAs increased lipid oxidation and reduced energy efficiency in subsarcolemmal mitochondria, and increase AMPK activation, reducing both endoplasmic reticulum and oxidative stress. Increased mitochondrial respiration was related to an increased mitochondriogenesis in FD skeletal muscle, as shown by the increase in PGC1-α and -β. Conclusions our data strengthened the association of high dietary ω3-PUFA intake with reduced mitochondrial energy efficiency in the skeletal muscle. PMID:26901315

  1. Oxidative stress–induced mitochondrial dysfunction drives inflammation and airway smooth muscle remodeling in patients with chronic obstructive pulmonary disease

    PubMed Central

    Wiegman, Coen H.; Michaeloudes, Charalambos; Haji, Gulammehdi; Narang, Priyanka; Clarke, Colin J.; Russell, Kirsty E.; Bao, Wuping; Pavlidis, Stelios; Barnes, Peter J.; Kanerva, Justin; Bittner, Anton; Rao, Navin; Murphy, Michael P.; Kirkham, Paul A.; Chung, Kian Fan; Adcock, Ian M.; Brightling, Christopher E.; Davies, Donna E.; Finch, Donna K.; Fisher, Andrew J.; Gaw, Alasdair; Knox, Alan J.; Mayer, Ruth J.; Polkey, Michael; Salmon, Michael; Singh, David

    2015-01-01

    Background Inflammation and oxidative stress play critical roles in patients with chronic obstructive pulmonary disease (COPD). Mitochondrial oxidative stress might be involved in driving the oxidative stress–induced pathology. Objective We sought to determine the effects of oxidative stress on mitochondrial function in the pathophysiology of airway inflammation in ozone-exposed mice and human airway smooth muscle (ASM) cells. Methods Mice were exposed to ozone, and lung inflammation, airway hyperresponsiveness (AHR), and mitochondrial function were determined. Human ASM cells were isolated from bronchial biopsy specimens from healthy subjects, smokers, and patients with COPD. Inflammation and mitochondrial function in mice and human ASM cells were measured with and without the presence of the mitochondria-targeted antioxidant MitoQ. Results Mice exposed to ozone, a source of oxidative stress, had lung inflammation and AHR associated with mitochondrial dysfunction and reflected by decreased mitochondrial membrane potential (ΔΨm), increased mitochondrial oxidative stress, and reduced mitochondrial complex I, III, and V expression. Reversal of mitochondrial dysfunction by the mitochondria-targeted antioxidant MitoQ reduced inflammation and AHR. ASM cells from patients with COPD have reduced ΔΨm, adenosine triphosphate content, complex expression, basal and maximum respiration levels, and respiratory reserve capacity compared with those from healthy control subjects, whereas mitochondrial reactive oxygen species (ROS) levels were increased. Healthy smokers were intermediate between healthy nonsmokers and patients with COPD. Hydrogen peroxide induced mitochondrial dysfunction in ASM cells from healthy subjects. MitoQ and Tiron inhibited TGF-β–induced ASM cell proliferation and CXCL8 release. Conclusions Mitochondrial dysfunction in patients with COPD is associated with excessive mitochondrial ROS levels, which contribute to enhanced inflammation and cell

  2. Cellular Dysfunction in Diabetes as Maladaptive Response to Mitochondrial Oxidative Stress

    PubMed Central

    Naudi, Alba; Jove, Mariona; Ayala, Victoria; Cassanye, Anna; Serrano, Jose; Gonzalo, Hugo; Boada, Jordi; Prat, Joan; Portero-Otin, Manuel; Pamplona, Reinald

    2012-01-01

    Oxidative stress has been implicated in diabetes long-term complications. In this paper, we summarize the growing evidence suggesting that hyperglycemia-induced overproduction of superoxide by mitochondrial electron transport chain triggers a maladaptive response by affecting several metabolic and signaling pathways involved in the pathophysiology of cellular dysfunction and diabetic complications. In particular, it is our goal to describe physiological mechanisms underlying the mitochondrial free radical production and regulation to explain the oxidative stress derived from a high intracellular glucose concentration and the resulting maladaptive response that leads to a cellular dysfunction and pathological state. Finally, we outline potential therapies for diabetes focused to the prevention of mitochondrial oxidative damage. PMID:22253615

  3. Glyceraldehyde-3-phosphate Dehydrogenase (GAPDH) Aggregation Causes Mitochondrial Dysfunction during Oxidative Stress-induced Cell Death*

    PubMed Central

    Itakura, Masanori; Kubo, Takeya; Kaneshige, Akihiro; Harada, Naoki; Izawa, Takeshi; Azuma, Yasu-Taka; Kuwamura, Mitsuru; Yamaji, Ryouichi; Takeuchi, Tadayoshi

    2017-01-01

    Glycolytic glyceraldehyde-3-phosphate dehydrogenase (GAPDH) is a multifunctional protein that also mediates cell death under oxidative stress. We reported previously that the active-site cysteine (Cys-152) of GAPDH plays an essential role in oxidative stress-induced aggregation of GAPDH associated with cell death, and a C152A-GAPDH mutant rescues nitric oxide (NO)-induced cell death by interfering with the aggregation of wild type (WT)-GAPDH. However, the detailed mechanism underlying GAPDH aggregate-induced cell death remains elusive. Here we report that NO-induced GAPDH aggregation specifically causes mitochondrial dysfunction. First, we observed a correlation between NO-induced GAPDH aggregation and mitochondrial dysfunction, when GAPDH aggregation occurred at mitochondria in SH-SY5Y cells. In isolated mitochondria, aggregates of WT-GAPDH directly induced mitochondrial swelling and depolarization, whereas mixtures containing aggregates of C152A-GAPDH reduced mitochondrial dysfunction. Additionally, treatment with cyclosporin A improved WT-GAPDH aggregate-induced swelling and depolarization. In doxycycline-inducible SH-SY5Y cells, overexpression of WT-GAPDH augmented NO-induced mitochondrial dysfunction and increased mitochondrial GAPDH aggregation, whereas induced overexpression of C152A-GAPDH significantly suppressed mitochondrial impairment. Further, NO-induced cytochrome c release into the cytosol and nuclear translocation of apoptosis-inducing factor from mitochondria were both augmented in cells overexpressing WT-GAPDH but ameliorated in C152A-GAPDH-overexpressing cells. Interestingly, GAPDH aggregates induced necrotic cell death via a permeability transition pore (PTP) opening. The expression of either WT- or C152A-GAPDH did not affect other cell death pathways associated with protein aggregation, such as proteasome inhibition, gene expression induced by endoplasmic reticulum stress, or autophagy. Collectively, these results suggest that NO-induced GAPDH

  4. AMPK controls exercise endurance, mitochondrial oxidative capacity, and skeletal muscle integrity.

    PubMed

    Lantier, Louise; Fentz, Joachim; Mounier, Rémi; Leclerc, Jocelyne; Treebak, Jonas T; Pehmøller, Christian; Sanz, Nieves; Sakakibara, Iori; Saint-Amand, Emmanuelle; Rimbaud, Stéphanie; Maire, Pascal; Marette, André; Ventura-Clapier, Renée; Ferry, Arnaud; Wojtaszewski, Jørgen F P; Foretz, Marc; Viollet, Benoit

    2014-07-01

    AMP-activated protein kinase (AMPK) is a sensor of cellular energy status that plays a central role in skeletal muscle metabolism. We used skeletal muscle-specific AMPKα1α2 double-knockout (mdKO) mice to provide direct genetic evidence of the physiological importance of AMPK in regulating muscle exercise capacity, mitochondrial function, and contraction-stimulated glucose uptake. Exercise performance was significantly reduced in the mdKO mice, with a reduction in maximal force production and fatigue resistance. An increase in the proportion of myofibers with centralized nuclei was noted, as well as an elevated expression of interleukin 6 (IL-6) mRNA, possibly consistent with mild skeletal muscle injury. Notably, we found that AMPKα1 and AMPKα2 isoforms are dispensable for contraction-induced skeletal muscle glucose transport, except for male soleus muscle. However, the lack of skeletal muscle AMPK diminished maximal ADP-stimulated mitochondrial respiration, showing an impairment at complex I. This effect was not accompanied by changes in mitochondrial number, indicating that AMPK regulates muscle metabolic adaptation through the regulation of muscle mitochondrial oxidative capacity and mitochondrial substrate utilization but not baseline mitochondrial muscle content. Together, these results demonstrate that skeletal muscle AMPK has an unexpected role in the regulation of mitochondrial oxidative phosphorylation that contributes to the energy demands of the exercising muscle.-Lantier, L., Fentz, J., Mounier, R., Leclerc, J., Treebak, J. T., Pehmøller, C., Sanz, N., Sakakibara, I., Saint-Amand, E., Rimbaud, S., Maire, P., Marette, A., Ventura-Clapier, R., Ferry, A., Wojtaszewski, J. F. P., Foretz, M., Viollet, B. AMPK controls exercise endurance, mitochondrial oxidative capacity, and skeletal muscle integrity.

  5. Deoxycholic acid modulates cell death signaling through changes in mitochondrial membrane properties[S

    PubMed Central

    Sousa, Tânia; Castro, Rui E.; Pinto, Sandra N.; Coutinho, Ana; Lucas, Susana D.; Moreira, Rui; Rodrigues, Cecília M. P.; Prieto, Manuel; Fernandes, Fábio

    2015-01-01

    Cytotoxic bile acids, such as deoxycholic acid (DCA), are responsible for hepatocyte cell death during intrahepatic cholestasis. The mechanisms responsible for this effect are unclear, and recent studies conflict, pointing to either a modulation of plasma membrane structure or mitochondrial-mediated toxicity through perturbation of mitochondrial outer membrane (MOM) properties. We conducted a comprehensive comparative study of the impact of cytotoxic and cytoprotective bile acids on the membrane structure of different cellular compartments. We show that DCA increases the plasma membrane fluidity of hepatocytes to a minor extent, and that this effect is not correlated with the incidence of apoptosis. Additionally, plasma membrane fluidity recovers to normal values over time suggesting the presence of cellular compensatory mechanisms for this perturbation. Colocalization experiments in living cells confirmed the presence of bile acids within mitochondrial membranes. Experiments with active isolated mitochondria revealed that physiologically active concentrations of DCA change MOM order in a concentration- and time-dependent manner, and that these changes preceded the mitochondrial permeability transition. Importantly, these effects are not observed on liposomes mimicking MOM lipid composition, suggesting that DCA apoptotic activity depends on features of mitochondrial membranes that are absent in protein-free mimetic liposomes, such as the double-membrane structure, lipid asymmetry, or mitochondrial protein environment. In contrast, the mechanism of action of cytoprotective bile acids is likely not associated with changes in cellular membrane structure. PMID:26351365

  6. Deoxycholic acid modulates cell death signaling through changes in mitochondrial membrane properties.

    PubMed

    Sousa, Tânia; Castro, Rui E; Pinto, Sandra N; Coutinho, Ana; Lucas, Susana D; Moreira, Rui; Rodrigues, Cecília M P; Prieto, Manuel; Fernandes, Fábio

    2015-11-01

    Cytotoxic bile acids, such as deoxycholic acid (DCA), are responsible for hepatocyte cell death during intrahepatic cholestasis. The mechanisms responsible for this effect are unclear, and recent studies conflict, pointing to either a modulation of plasma membrane structure or mitochondrial-mediated toxicity through perturbation of mitochondrial outer membrane (MOM) properties. We conducted a comprehensive comparative study of the impact of cytotoxic and cytoprotective bile acids on the membrane structure of different cellular compartments. We show that DCA increases the plasma membrane fluidity of hepatocytes to a minor extent, and that this effect is not correlated with the incidence of apoptosis. Additionally, plasma membrane fluidity recovers to normal values over time suggesting the presence of cellular compensatory mechanisms for this perturbation. Colocalization experiments in living cells confirmed the presence of bile acids within mitochondrial membranes. Experiments with active isolated mitochondria revealed that physiologically active concentrations of DCA change MOM order in a concentration- and time-dependent manner, and that these changes preceded the mitochondrial permeability transition. Importantly, these effects are not observed on liposomes mimicking MOM lipid composition, suggesting that DCA apoptotic activity depends on features of mitochondrial membranes that are absent in protein-free mimetic liposomes, such as the double-membrane structure, lipid asymmetry, or mitochondrial protein environment. In contrast, the mechanism of action of cytoprotective bile acids is likely not associated with changes in cellular membrane structure.

  7. Role of oxidative DNA damage in mitochondrial dysfunction and Huntington's disease pathogenesis.

    PubMed

    Ayala-Peña, Sylvette

    2013-09-01

    Huntington's disease (HD) is a neurodegenerative disorder with an autosomal dominant expression pattern and typically a late-onset appearance. HD is a movement disorder with a heterogeneous phenotype characterized by involuntary dance-like gait, bioenergetic deficits, motor impairment, and cognitive and psychiatric deficits. Compelling evidence suggests that increased oxidative stress and mitochondrial dysfunction may underlie HD pathogenesis. However, the exact mechanisms underlying mutant huntingtin-induced neurological toxicity remain unclear. The objective of this paper is to review recent literature regarding the role of oxidative DNA damage in mitochondrial dysfunction and HD pathogenesis.

  8. The Mechanism of High Pressure Oxidations of Aliphatic Acids.

    DTIC Science & Technology

    ACETIC ACID , *OXIDATION), (*CARBOXYLIC ACIDS, OXIDATION), CHROMIUM ALLOYS, REACTION KINETICS, COPPER ALLOYS, NICKEL ALLOYS, TEMPERATURE, HIGH PRESSURE, CATALYSTS, GAS CHROMATOGRAPHY, VOLUMETRIC ANALYSIS, THESES

  9. Comparison of mammalian mitochondrial ribosomal ribonucleic acid from different species.

    PubMed

    Mitra, R S; Bartoov, B; Monahan, J; Freeman, K B

    1972-08-01

    Mitochondrial ribosomal RNA species from mouse L cells, rat liver, rat hepatoma, hamster BHK-21 cells and human KB cells were examined by electrophoresis on polyacrylamide-agarose gels and sedimentation in sucrose density gradients. The S(E) (electrophoretic mobility) and S values of mitochondrial rRNA of all species were highly dependent on temperature and ionic strength of the medium; the S(E) values increased and the S values decreased with an increase in temperature at a low ionic strength. At an ionic strength of 0.3 at 23-25 degrees C or an ionic strength of 0.01 at 3-4 degrees C the S and S(E) values were almost the same being about 16.2-18.0 and 12.3-13.6 for human and mouse mitochondrial rRNA. The molecular weights under these conditions were calculated to be 3.8x10(5)-4.3x10(5) and 5.9x10(5)-6.8x10(5), depending on the technique used. At 25 degrees C in buffers of low ionic strength mouse mitochondrial rRNA species had a lower electrophoretic mobility than those of human and hamster. Under these conditions the smaller mitochondrial rRNA species of hamster had a lower electrophoretic mobility than that of human but the larger component had an identical mobility. Mouse and rat mitochondrial rRNA species had identical electrophoretic mobilities. Complex differences between human and mouse mitochondrial rRNA species were observed on sedimentation in sucrose density gradients under various conditions of temperature and ionic strength. Mouse L-cell mitochondrial rRNA was eluted after cytoplasmic rRNA on a column of methylated albumin-kieselguhr.

  10. Proteomic analysis of pRb loss highlights a signature of decreased mitochondrial oxidative phosphorylation

    PubMed Central

    Nicolay, Brandon N.; Danielian, Paul S.; Kottakis, Filippos; Lapek, John D.; Sanidas, Ioannis; Miles, Wayne O.; Dehnad, Mantre; Tschöp, Katrin; Gierut, Jessica J.; Manning, Amity L.; Morris, Robert; Haigis, Kevin; Bardeesy, Nabeel; Lees, Jacqueline A.; Haas, Wilhelm; Dyson, Nicholas J.

    2015-01-01

    The retinoblastoma tumor suppressor (pRb) protein associates with chromatin and regulates gene expression. Numerous studies have identified Rb-dependent RNA signatures, but the proteomic effects of Rb loss are largely unexplored. We acutely ablated Rb in adult mice and conducted a quantitative analysis of RNA and proteomic changes in the colon and lungs, where RbKO was sufficient or insufficient to induce ectopic proliferation, respectively. As expected, RbKO caused similar increases in classic pRb/E2F-regulated transcripts in both tissues, but, unexpectedly, their protein products increased only in the colon, consistent with its increased proliferative index. Thus, these protein changes induced by Rb loss are coupled with proliferation but uncoupled from transcription. The proteomic changes in common between RbKO tissues showed a striking decrease in proteins with mitochondrial functions. Accordingly, RB1 inactivation in human cells decreased both mitochondrial mass and oxidative phosphorylation (OXPHOS) function. RBKO cells showed decreased mitochondrial respiratory capacity and the accumulation of hypopolarized mitochondria. Additionally, RB/Rb loss altered mitochondrial pyruvate oxidation from 13C-glucose through the TCA cycle in mouse tissues and cultured cells. Consequently, RBKO cells have an enhanced sensitivity to mitochondrial stress conditions. In summary, proteomic analyses provide a new perspective on Rb/RB1 mutation, highlighting the importance of pRb for mitochondrial function and suggesting vulnerabilities for treatment. PMID:26314710

  11. The effect of valinomycin in fibroblasts from patients with fatty acid oxidation disorders

    SciTech Connect

    Ndukwe Erlingsson, Uzochi Chimdinma; Iacobazzi, Francesco; Liu, Aiping; Ardon, Orly; Pasquali, Marzia; Longo, Nicola

    2013-08-09

    Highlights: •Valinomycin can cause mitochondrial stress and stimulate fatty acid oxidation. •Cells with VLCAD deficiency fail to increase fatty acid oxidation in response to valinomycin. •Response to valinomycin can help in the diagnosis of VLCAD deficiency. -- Abstract: Disorders of the carnitine cycle and of the beta oxidation spiral impair the ability to obtain energy from fats at time of fasting and stress. This can result in hypoketotic hypoglycemia, cardiomyopathy, cardiac arrhythmia and other chronic medical problems. The in vitro study of fibroblasts from patients with these conditions is impaired by their limited oxidative capacity. Here we evaluate the capacity of valinomycin, a potassium ionophore that increases mitochondrial respiration, to increase the oxidation of fatty acids in cells from patients with inherited fatty acid oxidation defects. The addition of valinomycin to fibroblasts decreased the accumulation of the lipophilic cation tetraphenylphosphonium (TPP{sup +}) at low concentrations due to the dissipation of the mitochondrial membrane potential. At higher doses, valinomycin increased TPP{sup +} accumulation due to the increased potassium permeability of the plasma membrane and subsequent cellular hyperpolarization. The incubation of normal fibroblasts with valinomycin increased [{sup 14}C]-palmitate oxidation (measured as [{sup 14}C]O{sub 2} release) in a dose-dependent manner. By contrast, valinomycin failed to increase palmitate oxidation in fibroblasts from patients with very long chain acyl CoA dehydrogenase (VLCAD) deficiency. This was not observed in fibroblasts from patients heterozygous for this condition. These results indicate that valinomycin can increase fatty acid oxidation in normal fibroblasts and could be useful to differentiate heterozygotes from patients affected with VLCAD deficiency.

  12. Impaired cerebral mitochondrial oxidative phosphorylation function in a rat model of ventricular fibrillation and cardiopulmonary resuscitation.

    PubMed

    Jiang, Jun; Fang, Xiangshao; Fu, Yue; Xu, Wen; Jiang, Longyuan; Huang, Zitong

    2014-01-01

    Postcardiac arrest brain injury significantly contributes to mortality and morbidity in patients suffering from cardiac arrest (CA). Evidence that shows that mitochondrial dysfunction appears to be a key factor in tissue damage after ischemia/reperfusion is accumulating. However, limited data are available regarding the cerebral mitochondrial dysfunction during CA and cardiopulmonary resuscitation (CPR) and its relationship to the alterations of high-energy phosphate. Here, we sought to identify alterations of mitochondrial morphology and oxidative phosphorylation function as well as high-energy phosphates during CA and CPR in a rat model of ventricular fibrillation (VF). We found that impairment of mitochondrial respiration and partial depletion of adenosine triphosphate (ATP) and phosphocreatine (PCr) developed in the cerebral cortex and hippocampus following a prolonged cardiac arrest. Optimal CPR might ameliorate the deranged phosphorus metabolism and preserve mitochondrial function. No obvious ultrastructural abnormalities of mitochondria have been found during CA. We conclude that CA causes cerebral mitochondrial dysfunction along with decay of high-energy phosphates, which would be mitigated with CPR. This study may broaden our understanding of the pathogenic processes underlying global cerebral ischemic injury and provide a potential therapeutic strategy that aimed at preserving cerebral mitochondrial function during CA.

  13. Regulation of Skeletal Muscle Oxidative Capacity and Insulin Signaling by the Mitochondrial Rhomboid Protease PARL

    PubMed Central

    Civitarese, Anthony E.; MacLean, Paul S.; Carling, Stacy; Kerr-Bayles, Lyndal; McMillan, Ryan P.; Pierce, Anson; Becker, Thomas C.; Moro, Cedric; Finlayson, Jean; Lefort, Natalie; Newgard, Christopher B.; Mandarino, Lawrence; Cefalu, William; Walder, Ken; Collier, Greg R.; Hulver, Matthew W.; Smith, Steven R.; Ravussin, Eric

    2010-01-01

    SUMMARY Type 2 diabetes Mellitus (T2DM) and aging are characterized by insulin resistance, lower mitochondrial density and function and increased production of reactive oxygen species (ROS). In lower organisms continuous remodeling critically maintains the function and life cycle of mitochondria, in part by the protease pcp1 (PARL ortholog). We therefore examined whether variation in PARL protein content is associated with mitochondrial abnormalities and insulin resistance. Relative to healthy, young individuals (23±1y), PARL mRNA and mitochondrial mass were both reduced in elderly subjects (64.4±1.2 y; 51% and 44% respectively) and in subjects with T2DM (51.8±3 y; 31% and 41% respectively; all p<0.05). Muscle knock-down of PARL in mice resulted in lower mitochondrial content (−31±3%, p<0.05), lower OPA1 and PGC1α protein levels and impaired insulin signaling. Furthermore, mitochondrial cristae were malformed and resulted in elevated in vivo oxidative stress. Adenoviral suppression of PARL protein in healthy myotubes lowered mitochondrial mass (−33±8%), insulin stimulated glycogen synthesis (−33±9%) and increased ROS production (2-fold) (all p<0.05). We propose that lower PARL expression may contribute to the mitochondrial abnormalities seen in aging and T2DM. PMID:20444421

  14. Mitochondrial oxidative stress is the achille's heel of melanoma cells resistant to Braf-mutant inhibitor

    PubMed Central

    André, Fanny; Jonneaux, Aurélie; Scalbert, Camille; Garçon, Guillaume; Malet-Martino, Myriam; Balayssac, Stéphane; Rocchi, Stephane; Savina, Ariel; Formstecher, Pierre; Mortier, Laurent; Kluza, Jérome; Marchetti, Philippe

    2013-01-01

    Vemurafenib/PLX4032, a selective inhibitor of mutant BRAFV600E, constitutes a paradigm shift in melanoma therapy. Unfortunately, acquired resistance, which unavoidably occurs, represents one major limitation to clinical responses. Recent studies have highlighted that vemurafenib activated oxidative metabolism in BRAFV600E melanomas expressing PGC1α. However, the oxidative state of melanoma resistant to BRAF inhibitors is unknown. We established representative in vitro and in vivo models of human melanoma resistant to vemurafenib including primary specimens derived from melanoma patients. Firstly, our study reveals that vemurafenib increased mitochondrial respiration and ROS production in BRAFV600E melanoma cell lines regardless the expression of PGC1α. Secondly, melanoma cells that have acquired resistance to vemurafenib displayed intrinsically high rates of mitochondrial respiration associated with elevated mitochondrial oxidative stress irrespective of the presence of vemurafenib. Thirdly, the elevated ROS level rendered vemurafenib-resistant melanoma cells prone to cell death induced by pro-oxidants including the clinical trial drug, elesclomol. Based on these observations, we propose that the mitochondrial oxidative signature of resistant melanoma constitutes a novel opportunity to overcome resistance to BRAF inhibition. PMID:24161908

  15. Mitochondrial Dysfunction Is Involved in the Toxic Activity of Boric Acid against Saprolegnia

    PubMed Central

    Ali, Shimaa E.; Thoen, Even; Evensen, Øystein; Wiik-Nielsen, Jannicke; Gamil, Amr A. A.; Skaar, Ida

    2014-01-01

    There has been a significant increase in the incidence of Saprolegnia infections over the past decades, especially after the banning of malachite green. Very often these infections are associated with high economic losses in salmonid farms and hatcheries. The use of boric acid to control the disease has been investigated recently both under in vitro and in vivo conditions, however its possible mode of action against fish pathogenic Saprolegnia is not known. In this study, we have explored the transformation in Saprolegnia spores/hyphae after exposure to boric acid (1 g/L) over a period 4–24 h post treatment. Using transmission electron microscopy (TEM), early changes in Saprolegnia spores were detected. Mitochondrial degeneration was the most obvious sign observed following 4 h treatment in about 20% of randomly selected spores. We also investigated the effect of the treatment on nuclear division, mitochondrial activity and function using confocal laser scanning microscopy (CLSM). Fluorescence microscopy was also used to test the effect of treatment on mitochondrial membrane potential and formation of reactive oxygen species. Additionally, the viability and proliferation of treated spores that correlated to mitochondrial enzymatic activity were tested using an MTS assay. All obtained data pointed towards changes in the mitochondrial structure, membrane potential and enzymatic activity following treatment. We have found that boric acid has no effect on the integrity of membranes of Saprolegnia spores at concentrations tested. It is therefore likely that mitochondrial dysfunction is involved in the toxic activity of boric acid against Saprolegnia spp. PMID:25354209

  16. Mitochondrial dysfunction is involved in the toxic activity of boric acid against Saprolegnia.

    PubMed

    Ali, Shimaa E; Thoen, Even; Evensen, Øystein; Wiik-Nielsen, Jannicke; Gamil, Amr A A; Skaar, Ida

    2014-01-01

    There has been a significant increase in the incidence of Saprolegnia infections over the past decades, especially after the banning of malachite green. Very often these infections are associated with high economic losses in salmonid farms and hatcheries. The use of boric acid to control the disease has been investigated recently both under in vitro and in vivo conditions, however its possible mode of action against fish pathogenic Saprolegnia is not known. In this study, we have explored the transformation in Saprolegnia spores/hyphae after exposure to boric acid (1 g/L) over a period 4-24 h post treatment. Using transmission electron microscopy (TEM), early changes in Saprolegnia spores were detected. Mitochondrial degeneration was the most obvious sign observed following 4 h treatment in about 20% of randomly selected spores. We also investigated the effect of the treatment on nuclear division, mitochondrial activity and function using confocal laser scanning microscopy (CLSM). Fluorescence microscopy was also used to test the effect of treatment on mitochondrial membrane potential and formation of reactive oxygen species. Additionally, the viability and proliferation of treated spores that correlated to mitochondrial enzymatic activity were tested using an MTS assay. All obtained data pointed towards changes in the mitochondrial structure, membrane potential and enzymatic activity following treatment. We have found that boric acid has no effect on the integrity of membranes of Saprolegnia spores at concentrations tested. It is therefore likely that mitochondrial dysfunction is involved in the toxic activity of boric acid against Saprolegnia spp.

  17. Wet oxidation of salicylic acid solutions.

    PubMed

    Collado, Sergio; Garrido, Laura; Laca, Adriana; Diaz, Mario

    2010-11-15

    Salicylic acid is a frequent pollutant in several industrial wastewaters. Uncatalyzed wet air oxidation, which is a promising technique for the treatment of phenolic effluents, has not been analyzed yet for the removal of salicylic acid. The effect of different conditions of pH (1.3-12.3), pressure (1.0-4.1 MPa), temperature (413-443 K), and initial concentrations (1.45-14.50 mM) on the wet oxidation of salicylate/salicylic acid solutions have here been investigated. The pH value of the reaction media was found to be a key parameter for the rate of the oxidation process with an optimum at pH 3.1, when the concentrations of salicylic acid and salicylate were similar. The oxidation reaction followed pseudofirst-order kinetics with respect to salicylic acid and 0.82 order with respect to dissolved oxygen. Additionally, the evolution of the color during the wet oxidation was analyzed and discussed in relation with the formation of intermediate compounds. Then, a reaction pathway for the noncatalytic wet oxidation of the salicylic acid was proposed.

  18. Glucocorticoids Suppress Mitochondrial Oxidant Production via Upregulation of Uncoupling Protein 2 in Hyperglycemic Endothelial Cells

    PubMed Central

    Szabo, Csaba

    2016-01-01

    Diabetic complications are the leading cause of morbidity and mortality in diabetic patients. Elevated blood glucose contributes to the development of endothelial and vascular dysfunction, and, consequently, to diabetic micro- and macrovascular complications, because it increases the mitochondrial proton gradient and mitochondrial oxidant production. Therapeutic approaches designed to counteract glucose-induced mitochondrial reactive oxygen species (ROS) production in the vasculature are expected to show efficacy against all diabetic complications, but direct pharmacological targeting (scavenging) of mitochondrial oxidants remains challenging due to the high reactivity of some of these oxidant species. In a recent study, we have conducted a medium-throughput cell-based screening of a focused library of well-annotated pharmacologically active compounds and identified glucocorticoids as inhibitors of mitochondrial superoxide production in microvascular endothelial cells exposed to elevated extracellular glucose. The goal of the current study was to investigate the mechanism of glucocorticoids' action. Our findings show that glucocorticoids induce the expression of the mitochondrial UCP2 protein and decrease the mitochondrial potential. UCP2 silencing prevents the protective effect of the glucocorticoids on ROS production. UCP2 induction also increases the oxygen consumption and the “proton leak” in microvascular endothelial cells. Furthermore, glutamine supplementation augments the effect of glucocorticoids via further enhancing the expression of UCP2 at the translational level. We conclude that UCP2 induction represents a novel experimental therapeutic intervention in diabetic vascular complications. While direct repurposing of glucocorticoids may not be possible for the therapy of diabetic complications due to their significant side effects that develop during chronic administration, the UCP2 pathway may be therapeutically targetable by other, glucocorticoid

  19. SIRT1 activation by curcumin pretreatment attenuates mitochondrial oxidative damage induced by myocardial ischemia reperfusion injury.

    PubMed

    Yang, Yang; Duan, Weixun; Lin, Yan; Yi, Wei; Liang, Zhenxing; Yan, Juanjuan; Wang, Ning; Deng, Chao; Zhang, Song; Li, Yue; Chen, Wensheng; Yu, Shiqiang; Yi, Dinghua; Jin, Zhenxiao

    2013-12-01

    Ischemia reperfusion (IR) injury (IRI) is harmful to the cardiovascular system and causes mitochondrial oxidative stress. Silent information regulator 1 (SIRT1), a type of histone deacetylase, contributes to IRI. Curcumin (Cur) is a strong natural antioxidant and is the active component in Curcuma longa; Cur has protective effects against IRI and may regulate the activity of SIRT1. This study was designed to investigate the protective effect of Cur pretreatment on myocardial IRI and to elucidate this potential mechanism. Isolated and in vivo rat hearts and cultured neonatal rat cardiomyocytes were subjected to IR. Prior to this procedure, the hearts or cardiomyocytes were exposed to Cur in the absence or presence of the SIRT1 inhibitor sirtinol or SIRT1 siRNA. Cur conferred a cardioprotective effect, as shown by improved postischemic cardiac function, decreased myocardial infarct size, decreased myocardial apoptotic index, and several biochemical parameters, including the up-regulation of the antiapoptotic protein Bcl2 and the down-regulation of the proapoptotic protein Bax. Sirtinol and SIRT1 siRNA each blocked the Cur-mediated cardioprotection by inhibiting SIRT1 signaling. Cur also resulted in a well-preserved mitochondrial redox potential, significantly elevated mitochondrial superoxide dismutase activity, and decreased formation of mitochondrial hydrogen peroxide and malondialdehyde. These observations indicated that the IR-induced mitochondrial oxidative damage was remarkably attenuated. However, this Cur-elevated mitochondrial function was reversed by sirtinol or SIRT1 siRNA treatment. In summary, our results demonstrate that Cur pretreatment attenuates IRI by reducing IR-induced mitochondrial oxidative damage through the activation of SIRT1 signaling.

  20. Chronic mitochondrial uncoupling treatment prevents acute cold-induced oxidative stress in birds.

    PubMed

    Stier, Antoine; Massemin, Sylvie; Criscuolo, François

    2014-12-01

    Endotherms have evolved two major types of thermogenesis that allow them to actively produce heat in response to cold exposure, either through muscular activity (i.e. shivering thermogenesis) or through futile electro-chemical cycles (i.e. non-shivering thermogenesis). Amongst the latter, mitochondrial uncoupling is of key importance because it is suggested to drive heat production at a low cost in terms of oxidative stress. While this has been experimentally shown in mammals, the oxidative stress consequences of cold exposure and mitochondrial uncoupling are clearly less understood in the other class of endotherms, the birds. We compared metabolic and oxidative stress responses of zebra finches chronically treated with or without a chemical mitochondrial uncoupler (2,4-dinitrophenol: DNP), undergoing an acute (24 h) and a chronic (4 weeks) cold exposure (12 °C). We predicted that control birds should present at least a transient elevation of oxidative stress levels in response to cold exposure. This oxidative stress cost should be more pronounced in control birds than in DNP-treated birds, due to their lower basal uncoupling state. Despite similar increase in metabolism, control birds presented elevated levels of DNA oxidative damage in response to acute (but not chronic) cold exposure, while DNP-treated birds did not. Plasma antioxidant capacity decreased overall in response to chronic cold exposure. These results show that acute cold exposure increases oxidative stress in birds. However, uncoupling mitochondrial functioning appears as a putative compensatory mechanism preventing cold-induced oxidative stress. This result confirms previous observations in mice and underlines non-shivering thermogenesis as a putative key mechanism for endotherms in mounting a response to cold at a low oxidative cost.

  1. Neuroprotective effects of gallic acid against hypoxia/reoxygenation-induced mitochondrial dysfunctions in vitro and cerebral ischemia/reperfusion injury in vivo.

    PubMed

    Sun, Jing; Li, Yun-Zi; Ding, Yin-Hui; Wang, Jin; Geng, Ji; Yang, Huan; Ren, Jie; Tang, Jin-Yan; Gao, Jing

    2014-11-17

    Oxidative stress and mitochondrial dysfunction are frequently implicated in the pathology of secondary neuronal damage following cerebral ischemia/reperfusion. Recent evidence suggests that gallic acid (GA) reverses oxidative stress in rat model of streptozotocin-induced dementia, but the roles and mechanisms of GA on cerebral ischemia/reperfusion injury remain unknown. Here we investigated the potential roles and mechanisms of GA in hypoxia/reoxygenation induced by sodium hydrosulfite (Na2S2O4) in vitro and cerebral ischemia/reperfusion induced by middle cerebral artery occlusion (MCAO) in vivo. 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) assay, 5, 5', 6, 6'-tetrachloro-1, 1', 3, 3'-tetraethylbenzimidazol carbocyanine iodide (JC-1), Dichlorofluorescin diacetate (DCF-DA) and MitoSOX fluorescent assay, Clark-type oxygen electrode, firefly luciferase assay, and calcium-induced mitochondrial swelling were conducted to detect cell death, mitochondrial membrane potential (MMP), intracellular and mitochondrial reactive oxygen species (ROS), oxygen consumption, ATP level, and mitochondrial permeability transition pore (MPTP) viability. We firstly find that modulation of the mitochondrial dysfunction is an important mechanism by GA attenuating hypoxia/reoxygenation insult. To further assess the effects of GA on cerebral ischemia/reperfusion injury, 2, 3, 5-triphenyl-tetrazolium chloride (TTC) staining, dUTP nick-end labeling (TUNEL) assay, and Cytochrome C (Cyt C) release were performed in MCAO rats. The results support that GA is useful against cerebral ischemia/reperfusion injury as a potential protective agent.

  2. N-Acetyl-L-cysteine Protects the Enterocyte against Oxidative Damage by Modulation of Mitochondrial Function

    PubMed Central

    Xiao, Hao; Wu, Miaomiao; Shao, Fangyuan; Guan, Guiping; Huang, Bo

    2016-01-01

    The neonatal small intestine is susceptible to damage caused by oxidative stress. This study aimed to evaluate the protective role of antioxidant N-acetylcysteine (NAC) in intestinal epithelial cells against oxidative damage induced by H2O2. IPEC-J2 cells were cultured in DMEM-H with NAC and H2O2. After 2-day incubation, IPEC-J2 cells were collected for analysis of DNA synthesis, antioxidation capacity, mitochondrial respiration, and cell apoptosis. The results showed that H2O2 significantly decreased (P < 0.05) proliferation rate, mitochondrial respiration, and antioxidation capacity and increased cell apoptosis and the abundance of associated proteins, including cytochrome C, Bcl-XL, cleaved caspase-3, and total caspase-3. NAC supplementation remarkably increased (P < 0.05) proliferation rate, antioxidation capacity, and mitochondrial bioenergetics but decreased cell apoptosis. These findings indicate that NAC might rescue the intestinal injury induced by H2O2. PMID:28003713

  3. The fusogenic lipid phosphatidic acid promotes the biogenesis of mitochondrial outer membrane protein Ugo1

    PubMed Central

    Keller, Michael; Taskin, Asli A.; Horvath, Susanne E.; Guan, Xue Li; Prinz, Claudia; Opalińska, Magdalena; Zorzin, Carina; van der Laan, Martin; Wenk, Markus R.; Schubert, Rolf; Wiedemann, Nils; Holzer, Martin

    2015-01-01

    Import and assembly of mitochondrial proteins depend on a complex interplay of proteinaceous translocation machineries. The role of lipids in this process has been studied only marginally and so far no direct role for a specific lipid in mitochondrial protein biogenesis has been shown. Here we analyzed a potential role of phosphatidic acid (PA) in biogenesis of mitochondrial proteins in Saccharomyces cerevisiae. In vivo remodeling of the mitochondrial lipid composition by lithocholic acid treatment or by ablation of the lipid transport protein Ups1, both leading to an increase of mitochondrial PA levels, specifically stimulated the biogenesis of the outer membrane protein Ugo1, a component of the mitochondrial fusion machinery. We reconstituted the import and assembly pathway of Ugo1 in protein-free liposomes, mimicking the outer membrane phospholipid composition, and found a direct dependency of Ugo1 biogenesis on PA. Thus, PA represents the first lipid that is directly involved in the biogenesis pathway of a mitochondrial membrane protein. PMID:26347140

  4. Carnosic acid attenuates acute ethanol-induced liver injury via a SIRT1/p66Shc-mediated mitochondrial pathway.

    PubMed

    Tian, Xinyao; Hu, Yan; Li, Mingzhu; Xia, Kun; Yin, Jiye; Chen, Juan; Liu, Zhaoqian

    2016-04-01

    Ethanol-induced liver injury is associated with oxidative stress and hepatocyte apoptosis. We previously demonstrated that SIRT1/p66Shc pathway activation attenuates hepatocyte apoptosis in liver ischemia/reperfusion. The current study aimed to investigate whether carnosic acid (CA), a natural antioxidant, can inhibit acute ethanol-induced apoptosis of hepatocytes and to determine the effect of SIRT1/p66Shc on this process. Our results showed that CA pretreatment significantly reduced ethanol-induced histologic damage, serum aminotransferase activity, and oxidative stress in rats. Importantly, CA pretreatment increased SIRT1 expression following ethanol exposure. Furthermore, p66Shc expression was negatively correlated with SIRT1 expression. Consistent with the results demonstrating p66Shc inhibition, CA pretreatment inhibited the release of cytochrome C and apoptosis-inducing factor (AIF) from mitochondria. After exposing L02 cells to ethanol, the increased SIRT1 expression induced by CA was abrogated by pharmacologic SIRT1 inhibition or the use of siRNA against SIRT1. Additionally, SIRT1 inhibition significantly abrogated the suppression of p66Shc expression and mitochondrial translocation induced by CA. Accordingly, CA-induced decreases in the release of cytochrome C and AIF and in mitochondrial apoptosis were nearly abolished by SIRT1 knockdown. These data indicated that CA-activated SIRT1 is protective against ethanol treatment. In summary, CA attenuates acute ethanol-induced liver injury via a SIRT1/p66Shc-mediated mitochondrial pathway.

  5. Protection of mitochondrial and heart function by amino acids after ischemia and cardioplegia.

    PubMed

    Shug, A L; Madsen, D; Dobbie, R; Paulson, D J

    1994-01-01

    The effects of amino acids in protecting against ischemic/reperfusion injury were tested in two experimental models: the isolated perfused rat heart subjected to 21 min of zero flow ischemia (37 degrees) followed by 40 min of reperfusion and the isolated perfused rabbit heart subjected to 300 min of cardioplegic arrest (29 degrees) followed by 60 min of reperfusion. In both cases, the addition of amino acids to the perfusion medium significantly improved the recovery of cardiac contractile function. The protective effects of amino acids were associated with a preservation of mitochondrial respiratory activity. These findings suggest that amino acids by replenishing mitochondrial matrix levels of critical TCA cycle substrates, such as malate, stimulate mitochondrial respiration and thereby enhance the recovery of heart function.

  6. Exenatide improves liver mitochondrial dysfunction and insulin resistance by reducing oxidative stress in high fat diet-induced obese mice.

    PubMed

    Wang, Zixuan; Hou, Lin; Huang, Lanhui; Guo, Jun; Zhou, Xinli

    2017-04-22

    Oxidative stress is associated with obesity and may be accompanied by liver insulin resistance and mitochondrial dysfunction. Decreased mitochondrial respiratory chain enzymatic activities and decreased insulin metabolic signaling may promote these maladaptive changes. In this context, exenatide has been reported to reduce hepatic lipid deposition, improve insulin sensitivity and improve mitochondrial dysfunction. We hypothesized that exenatide would attenuate mitochondrial dysfunction by reducing hepatic lipid deposition, blunting oxidant stress and promoting insulin metabolic signaling in a high fat diet-induced model of obesity and insulin resistance. Sixteen-week-old male C57BL/6 diet-induced obese (DIO) mices and age-matched standard diet (STD) mices were treated with exenatide (10 μg/kg twice a day) for 28 days. Compared with untreated STD mice, untreated DIO mice exhibited deposited excessive lipid in liver and produced the oxidative stress in conjunction with insulin resistance, abnormal hepatic cells and mitochondrial histoarchitecture, mitochondrial dysfunction and reduced organism metabolism. Exenatide reduced hepatic steatosis, decreased oxidative stress, and improved insulin resistance in DIO mice, in concert with improvements in the insulin metabolic signaling, mitochondrial respiratory chain enzymatic activation, adenine nucleotide production, organism metabolism and weight gain. Results support the hypothesis that exenatide reduces hepatic cells and mitochondrial structural anomaly and improves insulin resistance in concert with improvements in insulin sensitivity and mitochondrial function activation, concomitantly with reductions in oxidative stress.

  7. Acetyl-L-carnitine and lipoic acid improve mitochondrial abnormalities and serum levels of liver enzymes in a mouse model of nonalcoholic fatty liver disease.

    PubMed

    Kathirvel, Elango; Morgan, Kengathevy; French, Samuel W; Morgan, Timothy R

    2013-11-01

    Mitochondrial abnormalities are suggested to be associated with the development of nonalcoholic fatty liver. Liver mitochondrial content and function have been shown to improve in oral feeding of acetyl-L-carnitine (ALC) to rodents. Carnitine is involved in the transport of acyl-coenzyme A across the mitochondrial membrane to be used in mitochondrial β-oxidation. We hypothesized that oral administration ALC with the antioxidant lipoic acid (ALC + LA) would benefit nonalcoholic fatty liver. To test our hypothesis, we fed Balb/C mice a standard diet (SF) or SF with ALC + LA or high-fat diet (HF) or HF with ALC + LA for 6 months. Acetyl-L-carnitine and LA were dissolved at 0.2:0.1% (wt/vol) in drinking water, and mice were allowed free access to food and water. Along with physical parameters, insulin resistance (blood glucose, insulin, glucose tolerance), liver function (alanine transaminase [ALT], aspartate transaminase [AST]), liver histology (hematoxylin and eosin), oxidative stress (malondialdehyde), and mitochondrial abnormalities (carbamoyl phosphate synthase 1 and electron microscopy) were done. Compared with SF, HF had higher body, liver, liver-to-body weight ratio, white adipose tissue, ALT, AST, liver fat, oxidative stress, and insulin resistance. Coadministration of ALC + LA to HF animals significantly improved the mitochondrial marker carbamoyl phosphate synthase 1 and the size of the mitochondria in liver. Alanine transaminase and AST levels were decreased. In a nonalcoholic fatty liver mice model, ALC + LA combination improved liver mitochondrial content, size, serum ALT, and AST without significant changes in oxidative stress, insulin resistance, and liver fat accumulation.

  8. Post-translational oxidative modification and inactivation of mitochondrial complex I in epileptogenesis.

    PubMed

    Ryan, Kristen; Backos, Donald S; Reigan, Philip; Patel, Manisha

    2012-08-15

    Mitochondrial oxidative stress and damage have been implicated in the etiology of temporal lobe epilepsy, but whether or not they have a functional impact on mitochondrial processes during epilepsy development (epileptogenesis) is unknown. One consequence of increased steady-state mitochondrial reactive oxygen species levels is protein post-translational modification (PTM). We hypothesize that complex I (CI), a protein complex of the mitochondrial electron transport chain, is a target for oxidant-induced PTMs, such as carbonylation, leading to impaired function during epileptogenesis. The goal of this study was to determine whether oxidative modifications occur and what impact they have on CI enzymatic activity in the rat hippocampus in response to kainate (KA)-induced epileptogenesis. Rats were injected with a single high dose of KA or vehicle and evidence for CI modifications was measured during the acute, latent, and chronic stages of epilepsy. Mitochondrial-specific carbonylation was increased acutely (48 h) and chronically (6 week), coincident with decreased CI activity. Mass spectrometry analysis of immunocaptured CI identified specific metal catalyzed carbonylation to Arg76 within the 75 kDa subunit concomitant with inhibition of CI activity during epileptogenesis. Computational-based molecular modeling studies revealed that Arg76 is in close proximity to the active site of CI and carbonylation of the residue is predicted to induce substantial structural alterations to the protein complex. These data provide evidence for the occurrence of a specific and irreversible oxidative modification of an important mitochondrial enzyme complex critical for cellular bioenergetics during the process of epileptogenesis.

  9. The spectrum of pyruvate oxidation defects in the diagnosis of mitochondrial disorders.

    PubMed

    Sperl, Wolfgang; Fleuren, Leanne; Freisinger, Peter; Haack, Tobias B; Ribes, Antonia; Feichtinger, René G; Rodenburg, Richard J; Zimmermann, Franz A; Koch, Johannes; Rivera, Isabel; Prokisch, Holger; Smeitink, Jan A; Mayr, Johannes A

    2015-05-01

    Pyruvate oxidation defects (PODs) are among the most frequent causes of deficiencies in the mitochondrial energy metabolism and represent a substantial subset of classical mitochondrial diseases. PODs are not only caused by deficiency of subunits of the pyruvate dehydrogenase complex (PDHC) but also by various disorders recently described in the whole pyruvate oxidation route including cofactors, regulation of PDHC and the mitochondrial pyruvate carrier. Our own patients from 2000 to July 2014 and patients identified by a systematic survey of the literature from 1970 to July 2014 with a pyruvate oxidation disorder and a genetically proven defect were included in the study (n=628). Of these defects 74.2% (n=466) belong to PDHC subunits, 24.5% (n=154) to cofactors, 0.5% (n=3) to PDHC regulation and 0.8% (n=5) to mitochondrial pyruvate import. PODs are underestimated in the field of mitochondrial diseases because not all diagnostic centres include biochemical investigations of PDHC in their routine analysis. Cofactor and transport defects can be missed, if pyruvate oxidation is not measured in intact mitochondria routinely. Furthermore deficiency of the X-chromosomal PDHA1 can be biochemically missed depending on the X-inactivation pattern. This is reflected by an increasing number of patients diagnosed recently by genetic high throughput screening approaches. PDHC deficiency including regulation and import affect mainly the glucose dependent central and peripheral nervous system and skeletal muscle. PODs with combined enzyme defects affect also other organs like heart, lung and liver. The spectrum of clinical presentation of PODs is still expanding. PODs are a therapeutically interesting group of mitochondrial diseases since some can be bypassed by ketogenic diet or treated by cofactor supplementation. PDHC kinase inhibition, chaperone therapy and PGC1α stimulation is still a matter of further investigations.

  10. Coenzyme Q10 ameliorates oxidative stress and prevents mitochondrial alteration in ischemic retinal injury.

    PubMed

    Lee, Dongwook; Kim, Keun-Young; Shim, Myoung Sup; Kim, Sang Yeop; Ellisman, Mark H; Weinreb, Robert N; Ju, Won-Kyu

    2014-04-01

    Coenzyme Q10 (CoQ10) acts by scavenging reactive oxygen species for protecting neuronal cells against oxidative stress in neurodegenerative diseases. We tested whether a diet supplemented with CoQ10 ameliorates oxidative stress and mitochondrial alteration, as well as promotes retinal ganglion cell (RGC) survival in ischemic retina induced by intraocular pressure elevation. A CoQ10 significantly promoted RGC survival at 2 weeks after ischemia. Superoxide dismutase 2 (SOD2) and heme oxygenase-1 (HO-1) expression were significantly increased at 12 h after ischemic injury. In contrast, the CoQ10 significantly prevented the upregulation of SOD2 and HO-1 protein expression in ischemic retina. In addition, the CoQ10 significantly blocked activation of astroglial and microglial cells in ischemic retina. Interestingly, the CoQ10 blocked apoptosis by decreasing caspase-3 protein expression in ischemic retina. Bax and phosphorylated Bad (pBad) protein expression were significantly increased in ischemic retina at 12 h. Interestingly, while CoQ10 significantly decreased Bax protein expression in ischemic retina, CoQ10 showed greater increase of pBad protein expression. Of interest, ischemic injury significantly increased mitochondrial transcription factor A (Tfam) protein expression in the retina at 12 h, however, CoQ10 significantly preserved Tfam protein expression in ischemic retina. Interestingly, there were no differences in mitochondrial DNA content among control- or CoQ10-treated groups. Our findings demonstrate that CoQ10 protects RGCs against oxidative stress by modulating the Bax/Bad-mediated mitochondrial apoptotic pathway as well as prevents mitochondrial alteration by preserving Tfam protein expression in ischemic retina. Our results suggest that CoQ10 may provide neuroprotection against oxidative stress-mediated mitochondrial alterations in ischemic retinal injury.

  11. Screening SIRT1 Activators from Medicinal Plants as Bioactive Compounds against Oxidative Damage in Mitochondrial Function

    PubMed Central

    Wang, Yi; Liang, Xinying; Chen, Yaqi; Zhao, Xiaoping

    2016-01-01

    Sirtuin type 1 (SIRT1) belongs to the family of NAD+ dependent histone deacetylases and plays a critical role in cellular metabolism and response to oxidative stress. Traditional Chinese medicines (TCMs), as an important part of natural products, have been reported to exert protective effect against oxidative stress in mitochondria. In this study, we screened SIRT1 activators from TCMs and investigated their activities against mitochondrial damage. 19 activators were found in total by in vitro SIRT1 activity assay. Among those active compounds, four compounds, ginsenoside Rb2, ginsenoside F1, ginsenoside Rc, and schisandrin A, were further studied to validate the SIRT1-activation effects by liquid chromatography-mass spectrometry and confirm their activities against oxidative damage in H9c2 cardiomyocytes exposed to tert-butyl hydroperoxide (t-BHP). The results showed that those compounds enhanced the deacetylated activity of SIRT1, increased ATP content, and inhibited intracellular ROS formation as well as regulating the activity of Mn-SOD. These SIRT1 activators also showed moderate protective effects on mitochondrial function in t-BHP cells by recovering oxygen consumption and increasing mitochondrial DNA content. Our results suggested that those compounds from TCMs attenuated oxidative stress-induced mitochondrial damage in cardiomyocytes through activation of SIRT1. PMID:26981165

  12. Induction of Mitochondrial Dysfunction and Oxidative Stress in Leishmania donovani by Orally Active Clerodane Diterpene

    PubMed Central

    Kathuria, Manoj; Bhattacharjee, Arindam; Sashidhara, Koneni V.; Singh, Suriya Pratap

    2014-01-01

    This study was performed to investigate the mechanistic aspects of cell death induced by a clerodane diterpene (K-09) in Leishmania donovani promastigotes that was previously demonstrated to be safe and orally active against visceral leishmaniasis (VL). K-09 caused depolarization of the mitochondrion and the generation of reactive oxygen species, triggering an apoptotic response in L. donovani promastigotes. Mitochondrial dysfunction subsequently resulted in the release of cytochrome c into the cytosol, impairing ATP production. Oxidative stress caused the depletion of reduced glutathione, while pretreatment with antioxidant N-acetyl cysteine (NAC) was able to abrogate oxidative stress. However, NAC failed to restore the mitochondrial membrane potential or intracellular calcium homeostasis after K-09 treatment, suggesting that the generation of oxidative stress is a downstream event relative to the other events. Caspase-3/-7-like protease activity and genomic DNA fragmentation were observed. Electron microscopy studies revealed gross morphological alterations typical of apoptosis, including severe mitochondrial damage, pyknosis of the nucleus, structural disruption of the mitochondrion-kinetoplast complex, flagellar pocket alterations, and the displacement of organelles. Moreover, an increased number of lipid droplets was detected after K-09 treatment, which is suggestive of altered lipid metabolism. Our results indicate that K-09 induces mitochondrial dysfunction and oxidative stress-mediated apoptotic cell death in L. donovani promastigotes, sharing many features with metazoan apoptosis. These mechanistic insights provide a basis for further investigation toward the development of K-09 as a potential drug candidate for VL. PMID:25070112

  13. Induction of mitochondrial dysfunction and oxidative stress in Leishmania donovani by orally active clerodane diterpene.

    PubMed

    Kathuria, Manoj; Bhattacharjee, Arindam; Sashidhara, Koneni V; Singh, Suriya Pratap; Mitra, Kalyan

    2014-10-01

    This study was performed to investigate the mechanistic aspects of cell death induced by a clerodane diterpene (K-09) in Leishmania donovani promastigotes that was previously demonstrated to be safe and orally active against visceral leishmaniasis (VL). K-09 caused depolarization of the mitochondrion and the generation of reactive oxygen species, triggering an apoptotic response in L. donovani promastigotes. Mitochondrial dysfunction subsequently resulted in the release of cytochrome c into the cytosol, impairing ATP production. Oxidative stress caused the depletion of reduced glutathione, while pretreatment with antioxidant N-acetyl cysteine (NAC) was able to abrogate oxidative stress. However, NAC failed to restore the mitochondrial membrane potential or intracellular calcium homeostasis after K-09 treatment, suggesting that the generation of oxidative stress is a downstream event relative to the other events. Caspase-3/-7-like protease activity and genomic DNA fragmentation were observed. Electron microscopy studies revealed gross morphological alterations typical of apoptosis, including severe mitochondrial damage, pyknosis of the nucleus, structural disruption of the mitochondrion-kinetoplast complex, flagellar pocket alterations, and the displacement of organelles. Moreover, an increased number of lipid droplets was detected after K-09 treatment, which is suggestive of altered lipid metabolism. Our results indicate that K-09 induces mitochondrial dysfunction and oxidative stress-mediated apoptotic cell death in L. donovani promastigotes, sharing many features with metazoan apoptosis. These mechanistic insights provide a basis for further investigation toward the development of K-09 as a potential drug candidate for VL.

  14. Mitochondrial defects and oxidative stress in Alzheimer disease and Parkinson disease.

    PubMed

    Yan, Michael H; Wang, Xinglong; Zhu, Xiongwei

    2013-09-01

    Alzheimer disease (AD) and Parkinson disease (PD) are the two most common age-related neurodegenerative diseases characterized by prominent neurodegeneration in selective neural systems. Although a small fraction of AD and PD cases exhibit evidence of heritability, among which many genes have been identified, the majority are sporadic without known causes. Molecular mechanisms underlying neurodegeneration and pathogenesis of these diseases remain elusive. Convincing evidence demonstrates oxidative stress as a prominent feature in AD and PD and links oxidative stress to the development of neuronal death and neural dysfunction, which suggests a key pathogenic role for oxidative stress in both AD and PD. Notably, mitochondrial dysfunction is also a prominent feature in these diseases, which is likely to be of critical importance in the genesis and amplification of reactive oxygen species and the pathophysiology of these diseases. In this review, we focus on changes in mitochondrial DNA and mitochondrial dynamics, two aspects critical to the maintenance of mitochondrial homeostasis and function, in relationship with oxidative stress in the pathogenesis of AD and PD.

  15. The Heuristic of Form: Mitochondrial Morphology and the Explanation of Oxidative Phosphorylation.

    PubMed

    Matlin, Karl S

    2016-02-01

    In the 1950s and 1960s, the search for the mechanism of oxidative phosphorylation by biochemists paralleled the description of mitochondrial form by George Palade and Fritiof Sjöstrand using electron microscopy. This paper explores the extent to which biochemists studying oxidative phosphorylation took mitochondrial form into account in the formulation of hypotheses, design of experiments, and interpretation of results. By examining experimental approaches employed by the biochemists studying oxidative phosphorylation, and their interactions with Palade, I suggest that use of mitochondrial form as a guide to experimentation and interpretation varied considerably among investigators. Most notably, Peter Mitchell, whose chemiosmotic hypothesis was ultimately the basis of the correct mechanism of oxidative phosphorylation, incorporated crucial aspects of mitochondrial form into his model that others failed to recognize. I discuss these historical observations in terms of the background and training of the biochemists, as well as a proposed heuristic of form, whose use may increase the possibility that biologically meaningful molecular mechanisms will be discovered.

  16. Carboxylic Acid Fullerene (C60) Derivatives Attenuated Neuroinflammatory Responses by Modulating Mitochondrial Dynamics

    NASA Astrophysics Data System (ADS)

    Ye, Shefang; Zhou, Tong; Cheng, Keman; Chen, Mingliang; Wang, Yange; Jiang, Yuanqin; Yang, Peiyan

    2015-05-01

    Fullerene (C60) derivatives, a unique class of compounds with potent antioxidant properties, have been reported to exert a wide variety of biological activities including neuroprotective properties. Mitochondrial dynamics are an important constituent of cellular quality control and function, and an imbalance of the dynamics eventually leads to mitochondria disruption and cell dysfunctions. This study aimed to assess the effects of carboxylic acid C60 derivatives (C60-COOH) on mitochondrial dynamics and elucidate its associated mechanisms in lipopolysaccharide (LPS)-stimulated BV-2 microglial cell model. Using a cell-based functional screening system labeled with DsRed2-mito in BV-2 cells, we showed that LPS stimulation led to excessive mitochondrial fission, increased mitochondrial localization of dynamin-related protein 1 (Drp1), both of which were markedly suppressed by C60-COOH pretreatment. LPS-induced mitochondria reactive oxygen species (ROS) generation and collapse of mitochondrial membrane potential (Δ Ψm) were also significantly inhibited by C60-COOH. Moreover, we also found that C60-COOH pretreatment resulted in the attenuation of LPS-mediated activation of nuclear factor (NF)-κB and mitogen-activated protein kinase (MAPK) signaling, as well as the production of pro-inflammatory mediators. Taken together, these findings demonstrated that carboxylic acid C60 derivatives may exert neuroprotective effects through regulating mitochondrial dynamics and functions in microglial cells, thus providing novel insights into the mechanisms of the neuroprotective properties of carboxylic acid C60 derivatives.

  17. Branched-chain amino acid metabolon: interaction of glutamate dehydrogenase with the mitochondrial branched-chain aminotransferase (BCATm).

    PubMed

    Islam, Mohammad Mainul; Nautiyal, Manisha; Wynn, R Max; Mobley, James A; Chuang, David T; Hutson, Susan M

    2010-01-01

    The catabolic pathway for branched-chain amino acids includes deamination followed by oxidative decarboxylation of the deaminated product branched-chain alpha-keto acids, catalyzed by the mitochondrial branched-chain aminotransferase (BCATm) and branched-chain alpha-keto acid dehydrogenase enzyme complex (BCKDC). We found that BCATm binds to the E1 decarboxylase of BCKDC, forming a metabolon that allows channeling of branched-chain alpha-keto acids from BCATm to E1. The protein complex also contains glutamate dehydrogenase (GDH1), 4-nitrophenylphosphatase domain and non-neuronal SNAP25-like protein homolog 1, pyruvate carboxylase, and BCKDC kinase. GDH1 binds to the pyridoxamine 5'-phosphate (PMP) form of BCATm (PMP-BCATm) but not to the pyridoxal 5'-phosphate-BCATm and other metabolon proteins. Leucine activates GDH1, and oxidative deamination of glutamate is increased further by addition of PMP-BCATm. Isoleucine and valine are not allosteric activators of GDH1, but in the presence of 5'-phosphate-BCATm, they convert BCATm to PMP-BCATm, stimulating GDH1 activity. Sensitivity to ADP activation of GDH1 was unaffected by PMP-BCATm; however, addition of a 3 or higher molar ratio of PMP-BCATm to GDH1 protected GDH1 from GTP inhibition by 50%. Kinetic results suggest that GDH1 facilitates regeneration of the form of BCATm that binds to E1 decarboxylase of the BCKDC, promotes metabolon formation, branched-chain amino acid oxidation, and cycling of nitrogen through glutamate.

  18. Fatty acid oxidation and ketogenesis in astrocytes

    SciTech Connect

    Auestad, N.

    1988-01-01

    Astrocytes were derived from cortex of two-day-old rat brain and grown in primary culture to confluence. The metabolism of the fatty acids, octanoate and palmitate, to CO{sub 2} in oxidative respiration and to the formation of ketone bodies was examined by radiolabeled tracer methodology. The net production of acetoacetate was also determined by measurement of its mass. The enzymes in the ketogenic pathway were examined by measuring enzymic activity and/or by immunoblot analyses. Labeled CO{sub 2} and labeled ketone bodies were produced from the oxidation of fatty acids labeled at carboxy- and {omega}-terminal carbons, indicating that fatty acids were oxidized by {beta}-oxidation. The results from the radiolabeled tracer studies also indicated that a substantial proportion of the {omega}-terminal 4-carbon unit of the fatty acids bypassed the {beta}-ketothiolase step of the {beta}-oxidation pathway. The ({sup 14}C)acetoacetate formed from the (1-{sup 14}C)labeled fatty acids, obligated to pass through the acetyl-CoA pool, contained 50% of the label at carbon 3 and 50% at carbon 1. In contrast, the ({sup 14}C)acetoacetate formed from the ({omega}-1)labeled fatty acids contained 90% of the label at carbon 3 and 10% at carbon 1.

  19. Ca2+ acting at the external side of the inner mitochondrial membrane can stimulate mitochondrial permeability transition induced by phenylarsine oxide.

    PubMed

    Kowaltowski, A J; Castilho, R F

    1997-12-15

    Mitochondrial permeability transition (MPT) induced by the thiol cross-linker phenylarsine oxide (PhAsO) in Ca(2+)-depleted mitochondria incubated in the presence of ruthenium red, an inhibitor of the Ca2+ uniporter, is stimulated by the addition of extramitochondrial Ca2+. The presence of extramitochondrial Ca2+ stimulates the reaction of mitochondrial membrane protein thiol groups with PhAsO. Both Ca(2+)-induced increase in mitochondrial membrane permeabilization and protein thiol group reaction with PhAsO are dependent on time (5-10 min to be complete) and the concentration of Ca2+ (1-25 microM). Mitochondrial permeabilization induced by PhAsO (15 microM) and extramitochondrial Ca2+ is inhibited by ADP, cyclosporin A, dibucaine and Mg2+, while mitochondrial permeabilization induced by high concentrations of PhAsO (60 microM) in the absence of Ca2+ is inhibited only by ADP and cyclosporin A. These results suggest that dibucaine and Mg2+ can inhibit mitochondrial permeabilization by antagonizing the effect of Ca2+ on the mitochondrial membrane. Once mitochondrial permeabilization induced by 15 microM PhAsO and extramitochondrial Ca2+ has already occurred, the addition of the Ca2+ chelator EGTA restores mitochondrial membrane potential (MPT pore closure), suggesting that the presence of Ca2+ is essential for the maintenance of the permeability of the mitochondrial membrane to protons (MPT pore opening). In conclusion, the results presented indicate that low Ca2+ concentrations acting at the external side of the inner mitochondrial membrane can stimulate mitochondrial permeability transition induced by PhAsO, due to increased accessibility of protein thiol groups to the reaction with PhAsO and increased probability of MPT pore opening.

  20. Mechanisms of MDMA (Ecstasy)-Induced Oxidative Stress, Mitochondrial Dysfunction, and Organ Damage

    PubMed Central

    Song, Byoung-Joon; Moon, Kwan-Hoon; Upreti, Vijay V.; Eddington, Natalie D.; Lee, Insong J.

    2010-01-01

    Despite numerous reports about the acute and sub-chronic toxicities caused by MDMA (3,4-methylenedioxymethamphetamine, ecstasy), the underlying mechanism of organ damage is poorly understood. The aim of this review is to present an update of the mechanistic studies on MDMA-mediated organ damage partly caused by increased oxidative/nitrosative stress. Because of the extensive reviews on MDMA-mediated oxidative stress and tissue damage, we specifically focus on the mechanisms and consequences of oxidative-modifications of mitochondrial proteins, leading to mitochondrial dysfunction. We briefly describe a method to systematically identify oxidatively-modified mitochondrial proteins in control and MDMA-exposed rats by using biotin-N-maleimide (biotin-NM) as a sensitive probe for oxidized proteins. We also describe various applications and advantages of this Cys-targeted proteomics method and alternative approaches to overcome potential limitations of this method in studying oxidized proteins from MDMA-exposed tissues. Finally we discuss the mechanism of synergistic drug-interaction between MDMA and other abused substances including alcohol (ethanol) as well as application of this redox-based proteomics method in translational studies for developing effective preventive and therapeutic agents against MDMA-induced organ damage. PMID:20420575

  1. Mechanisms of MDMA (ecstasy)-induced oxidative stress, mitochondrial dysfunction, and organ damage.

    PubMed

    Song, Byoung-Joon; Moon, Kwan-Hoon; Upreti, Vijay V; Eddington, Natalie D; Lee, Insong J

    2010-08-01

    Despite numerous reports about the acute and sub-chronic toxicities caused by MDMA (3,4-methylenedioxymethamphetamine, ecstasy), the underlying mechanism of organ damage is poorly understood. The aim of this review is to present an update of the mechanistic studies on MDMA-mediated organ damage partly caused by increased oxidative/nitrosative stress. Because of the extensive reviews on MDMA-mediated oxidative stress and tissue damage, we specifically focus on the mechanisms and consequences of oxidative-modifications of mitochondrial proteins, leading to mitochondrial dysfunction. We briefly describe a method to systematically identify oxidatively-modified mitochondrial proteins in control and MDMA-exposed rats by using biotin-N-maleimide (biotin-NM) as a sensitive probe for oxidized proteins. We also describe various applications and advantages of this Cys-targeted proteomics method and alternative approaches to overcome potential limitations of this method in studying oxidized proteins from MDMA-exposed tissues. Finally we discuss the mechanism of synergistic drug-interaction between MDMA and other abused substances including alcohol (ethanol) as well as application of this redox-based proteomics method in translational studies for developing effective preventive and therapeutic agents against MDMA-induced organ damage.

  2. Inhibition of NAPDH Oxidase 2 (NOX2) Prevents Oxidative Stress and Mitochondrial Abnormalities Caused by Saturated Fat in Cardiomyocytes.

    PubMed

    Joseph, Leroy C; Barca, Emanuele; Subramanyam, Prakash; Komrowski, Michael; Pajvani, Utpal; Colecraft, Henry M; Hirano, Michio; Morrow, John P

    2016-01-01

    Obesity and high saturated fat intake increase the risk of heart failure and arrhythmias. The molecular mechanisms are poorly understood. We hypothesized that physiologic levels of saturated fat could increase mitochondrial reactive oxygen species (ROS) in cardiomyocytes, leading to abnormalities of calcium homeostasis and mitochondrial function. We investigated the effect of saturated fat on mitochondrial function and calcium homeostasis in isolated ventricular myocytes. The saturated fatty acid palmitate causes a decrease in mitochondrial respiration in cardiomyocytes. Palmitate, but not the monounsaturated fatty acid oleate, causes an increase in both total cellular ROS and mitochondrial ROS. Palmitate depolarizes the mitochondrial inner membrane and causes mitochondrial calcium overload by increasing sarcoplasmic reticulum calcium leak. Inhibitors of PKC or NOX2 prevent mitochondrial dysfunction and the increase in ROS, demonstrating that PKC-NOX2 activation is also required for amplification of palmitate induced-ROS. Cardiomyocytes from mice with genetic deletion of NOX2 do not have palmitate-induced ROS or mitochondrial dysfunction. We conclude that palmitate induces mitochondrial ROS that is amplified by NOX2, causing greater mitochondrial ROS generation and partial depolarization of the mitochondrial inner membrane. The abnormal sarcoplasmic reticulum calcium leak caused by palmitate could promote arrhythmia and heart failure. NOX2 inhibition is a potential therapy for heart disease caused by diabetes or obesity.

  3. Mitochondrial oxidant stress triggers cell death in simulated ischemia-reperfusion.

    PubMed

    Loor, Gabriel; Kondapalli, Jyothisri; Iwase, Hirotaro; Chandel, Navdeep S; Waypa, Gregory B; Guzy, Robert D; Vanden Hoek, Terry L; Schumacker, Paul T

    2011-07-01

    To clarify the relationship between reactive oxygen species (ROS) and cell death during ischemia-reperfusion (I/R), we studied cell death mechanisms in a cellular model of I/R. Oxidant stress during simulated ischemia was detected in the mitochondrial matrix using mito-roGFP, a ratiometric redox sensor, and by Mito-Sox Red oxidation. Reperfusion-induced death was attenuated by over-expression of Mn-superoxide dismutase (Mn-SOD) or mitochondrial phospholipid hydroperoxide glutathione peroxidase (mito-PHGPx), but not by catalase, mitochondria-targeted catalase, or Cu,Zn-SOD. Protection was also conferred by chemically distinct antioxidant compounds, and mito-roGFP oxidation was attenuated by NAC, or by scavenging of residual O(2) during the ischemia (anoxic ischemia). Mitochondrial permeability transition pore (mPTP) oscillation/opening was monitored by real-time imaging of mitochondrial calcein fluorescence. Oxidant stress caused release of calcein to the cytosol during ischemia, a response that was inhibited by chemically diverse antioxidants, anoxia, or over-expression of Mn-SOD or mito-PHGPx. These findings suggest that mitochondrial oxidant stress causes oscillation of the mPTP prior to reperfusion. Cytochrome c release from mitochondria to the cytosol was not detected until after reperfusion, and was inhibited by anoxic ischemia or antioxidant administration during ischemia. Although DNA fragmentation was detected after I/R, no evidence of Bax activation was detected. Over-expression of the anti-apoptotic protein Bcl-X(L) in cardiomyocytes did not confer protection against I/R-induced cell death. Moreover, murine embryonic fibroblasts with genetic depletion of Bax and Bak, or over-expression of Bcl-X(L), failed to show protection against I/R. These findings indicate that mitochondrial ROS during ischemia triggers mPTP activation, mitochondrial depolarization, and cell death during reperfusion through a Bax/Bak-independent cell death pathway. Therefore

  4. Deoxycholic and chenodeoxycholic bile acids induce apoptosis via oxidative stress in human colon adenocarcinoma cells.

    PubMed

    Ignacio Barrasa, Juan; Olmo, Nieves; Pérez-Ramos, Pablo; Santiago-Gómez, Angélica; Lecona, Emilio; Turnay, Javier; Antonia Lizarbe, M

    2011-10-01

    The continuous exposure of the colonic epithelium to high concentrations of bile acids may exert cytotoxic effects and has been related to pathogenesis of colon cancer. A better knowledge of the mechanisms by which bile acids induce toxicity is still required and may be useful for the development of new therapeutic strategies. We have studied the effect of deoxycholic acid (DCA) and chenodeoxycholic acid (CDCA) treatments in BCS-TC2 human colon adenocarcinoma cells. Both bile acids promote cell death, being this effect higher for CDCA. Apoptosis is detected after 30 min-2 h of treatment, as observed by cell detachment, loss of membrane asymmetry, internucleosomal DNA degradation, appearance of mitochondrial transition permeability (MPT), and caspase and Bax activation. At longer treatment times, apoptosis is followed in vitro by secondary necrosis due to impaired mitochondrial activity and ATP depletion. Bile acid-induced apoptosis is a result of oxidative stress with increased ROS generation mainly by activation of plasma membrane enzymes, such as NAD(P)H oxidases and, to a lower extent, PLA2. These effects lead to a loss of mitochondrial potential and release of pro-apoptotic factors to the cytosol, which is confirmed by activation of caspase-9 and -3, but not caspase-8. This initial apoptotic steps promote cleavage of Bcl-2, allowing Bax activation and formation of additional pores in the mitochondrial membrane that amplify the apoptotic signal.

  5. Mitochondrial oxidative stress and dysfunction induced by isoniazid: study on isolated rat liver and brain mitochondria.

    PubMed

    Ahadpour, Morteza; Eskandari, Mohammad Reza; Mashayekhi, Vida; Haj Mohammad Ebrahim Tehrani, Kamaleddin; Jafarian, Iman; Naserzadeh, Parvaneh; Hosseini, Mir-Jamal

    2016-01-01

    Isoniazid (INH or isonicotinic hydrazide) is used for the treatment and prophylaxis of tuberculosis. Liver and brain are two important target organs in INH toxicity. However, the exact mechanisms behind the INH hepatotoxicity or neurotoxicity have not yet been completely understood. Considering the mitochondria as one of the possible molecular targets for INH toxicity, the aim of this study was to evaluate the mechanisms of INH mitochondrial toxicity on isolated mitochondria. Mitochondria were isolated by differential ultracentrifugation from male Sprague-Dawley rats and incubated with different concentrations of INH (25-2000 μM) for the investigation of mitochondrial parameters. The results indicated that INH could interact with mitochondrial respiratory chain and inhibit its activity. Our results showed an elevation in mitochondrial reactive oxygen species (ROS) formation, lipid peroxidation and mitochondrial membrane potential collapse after exposure of isolated liver mitochondria in INH. However, different results were obtained in brain mitochondria. Noteworthy, significant glutathione oxidation, adenosine triphosphate (ATP) depletion and lipid peroxidation were observed in higher concentration of INH, as compared to liver mitochondria. In conclusion, our results suggest that INH may initiate its toxicity in liver mitochondria through interaction with electron transfer chain, lipid peroxidation, mitochondrial membrane potential decline and cytochrome c expulsion which ultimately lead to cell death signaling.

  6. A high-fat diet coordinately downregulates genes required for mitochondrial oxidative phosphorylation in skeletal muscle.

    PubMed

    Sparks, Lauren M; Xie, Hui; Koza, Robert A; Mynatt, Randall; Hulver, Matthew W; Bray, George A; Smith, Steven R

    2005-07-01

    Obesity and type 2 diabetes have been associated with a high-fat diet (HFD) and reduced mitochondrial mass and function. We hypothesized a HFD may affect expression of genes involved in mitochondrial function and biogenesis. To test this hypothesis, we fed 10 insulin-sensitive males an isoenergetic HFD for 3 days with muscle biopsies before and after intervention. Oligonucleotide microarray analysis revealed 297 genes were differentially regulated by the HFD (Bonferonni adjusted P < 0.001). Six genes involved in oxidative phosphorylation (OXPHOS) decreased. Four were members of mitochondrial complex I: NDUFB3, NDUFB5, NDUFS1, and NDUFV1; one was SDHB in complex II and a mitochondrial carrier protein SLC25A12. Peroxisome proliferator-activated receptor gamma coactivator-1 (PGC1) alpha and PGC1beta mRNA were decreased by -20%, P < 0.01, and -25%, P < 0.01, respectively. In a separate experiment, we fed C57Bl/6J mice a HFD for 3 weeks and found that the same OXPHOS and PGC1 mRNAs were downregulated by approximately 90%, cytochrome C and PGC1alpha protein by approximately 40%. Combined, these results suggest a mechanism whereby HFD downregulates genes necessary for OXPHOS and mitochondrial biogenesis. These changes mimic those observed in diabetes and insulin resistance and, if sustained, may result in mitochondrial dysfunction in the prediabetic/insulin-resistant state.

  7. Methylglyoxal induces oxidative stress and mitochondrial dysfunction in osteoblastic MC3T3-E1 cells.

    PubMed

    Suh, K S; Choi, E M; Rhee, S Y; Kim, Y S

    2014-02-01

    Methylglyoxal is a reactive dicarbonyl compound produced by glycolytic processing and identified as a precursor of advanced glycation end products. The elevated methylglyoxal levels in patients with diabetes are believed to contribute to diabetic complications, including bone defects. The objective of this study was to evaluate the effect of methylglyoxal on the function of osteoblastic MC3T3-E1 cells. The data indicated that methylglyoxal decreased osteoblast differentiation and induced osteoblast cytotoxicity. Pretreatment of MC3T3-E1 cells with aminoguanidine (a carbonyl scavenger), Trolox (an antioxidant), and cyclosporin A (a blocker of the mitochondrial permeability transition pore) prevented methylglyoxal-induced cytotoxicity in MC3T3-E1 cells. However, BAPTA/AM (an intracellular Ca(2+) chelator) and dantrolene (an inhibitor of endoplasmic reticulum Ca(2+) release) did not reverse the cytotoxic effect of methylglyoxal. Methylglyoxal increased the formation of intracellular reactive oxygen species, mitochondrial superoxide, and cardiolipin peroxidation in osteoblastic MC3T3-E1 cells. Methylglyoxal also decreased the mitochondrial membrane potential and intracellular ATP and nitric oxide levels, suggesting that carbonyl stress-induced loss of mitochondrial integrity contributes to the cytotoxicity of methylglyoxal. Furthermore, the results demonstrated that methylglyoxal induced protein adduct formation, inactivation of glyoxalase I, and activation of glyoxalase II. Aminoguanidine reversed all aforementioned effects of methylglyoxal. Taken together, these data support the notion that high methylglyoxal concentrations have detrimental effects on osteoblasts through a mechanism involving oxidative stress and mitochondrial dysfunction.

  8. PM2.5-Induced Oxidative Stress and Mitochondrial Damage in the Nasal Mucosa of Rats

    PubMed Central

    Guo, Zhiqiang; Hong, Zhicong; Dong, Weiyang; Deng, Congrui; Zhao, Renwu; Xu, Jian; Zhuang, Guoshun; Zhang, Ruxin

    2017-01-01

    Exposure to PM2.5 (particulate matter ≤2.5 μm) increases the risk of nasal lesions, but the underlying mechanisms, especially the mechanisms leading to mitochondrial damage, are still unclear. Thus, we investigated the in vivo effects of PM2.5 exposure on the inflammatory response, oxidative stress, the enzyme activities of Na+K+-ATPase and Ca2+-ATPase, and the morphology and function of mitochondria in the nasal mucosa of rats. Exposure to PM2.5 occurred through inhalation of a PM2.5 solution aerosol. The results show that the PM2.5 exposure induced increased levels of malondialdehyde (MDA) and levels of proinflammatory mediators, including interleukin 6 (IL-6), IL-8, and tumor necrosis factor-α (TNF-α). These changes were accompanied by decreases in the activities of total superoxide dismutase (T-SOD), Na+K+-ATPase, and Ca2+-ATPase in rat nasal mucosa. PM2.5 significantly affected the expression of specific mitochondrial fission/fusion genes (OPA1, Mfn1, Fis1, and Drp1) in nasal mucosa. These changes were accompanied by abnormal alterations of mitochondrial structures, including mitochondrial swelling, cristae disorder, and even fission resulting from higher doses of PM2.5. Our data shows that oxidative damage, inflammatory response, and mitochondrial dysfunction may be the toxic mechanisms that cause nasal lesions after exposure to PM2.5. PMID:28146064

  9. Variations in mitochondrial membrane potential correlate with malic acid production by natural isolates of Saccharomyces cerevisiae sake strains.

    PubMed

    Oba, Takahiro; Kusumoto, Kenichi; Kichise, Yuki; Izumoto, Eiji; Nakayama, Shunichi; Tashiro, Kosuke; Kuhara, Satoru; Kitagaki, Hiroshi

    2014-08-01

    Research on the relationship between mitochondrial membrane potential and fermentation profile is being intensely pursued because of the potential for developing advanced fermentation technologies. In the present study, we isolated naturally occurring strains of yeast from sake mash that produce high levels of malic acid and demonstrate that variations in mitochondrial membrane potential correlate with malic acid production. To define the underlying biochemical mechanism, we determined the activities of enzymes required for malic acid synthesis and found that pyruvate carboxylase and malate dehydrogenase activities in strains that produce high levels of malic acid were elevated compared with the standard sake strain K901. These results inspired us to hypothesize that decreased mitochondrial membrane potential was responsible for increased malic acid synthesis, and we present data supporting this hypothesis. Thus, the mitochondrial membrane potential of high malic acid producers was lower compared with standard strains. We conclude that mitochondrial membrane potential correlates with malic acid production.

  10. Proinflammatory cytokines differentially regulate adipocyte mitochondrial metabolism, oxidative stress, and dynamics

    PubMed Central

    Hahn, Wendy S.; Kuzmicic, Jovan; Burrill, Joel S.; Donoghue, Margaret A.; Foncea, Rocio; Jensen, Michael D.; Lavandero, Sergio; Arriaga, Edgar A.

    2014-01-01

    Proinflammatory cytokines differentially regulate adipocyte mitochondrial metabolism, oxidative stress, and dynamics. Macrophage infiltration of adipose tissue and the chronic low-grade production of inflammatory cytokines have been mechanistically linked to the development of insulin resistance, the forerunner of type 2 diabetes mellitus. In this study, we evaluated the chronic effects of TNFα, IL-6, and IL-1β on adipocyte mitochondrial metabolism and morphology using the 3T3-L1 model cell system. TNFα treatment of cultured adipocytes led to significant changes in mitochondrial bioenergetics, including increased proton leak, decreased ΔΨm, increased basal respiration, and decreased ATP turnover. In contrast, although IL-6 and IL-1β decreased maximal respiratory capacity, they had no effect on ΔΨm and varied effects on ATP turnover, proton leak, or basal respiration. Only TNFα treatment of 3T3-L1 cells led to an increase in oxidative stress (as measured by superoxide anion production and protein carbonylation) and C16 ceramide synthesis. Treatment of 3T3-L1 adipocytes with cytokines led to decreased mRNA expression of key transcription factors and control proteins implicated in mitochondrial biogenesis, including PGC-1α and eNOS as well as deceased expression of COX IV and Cyt C. Whereas each cytokine led to effects on expression of mitochondrial markers, TNFα exclusively led to mitochondrial fragmentation and decreased the total level of OPA1 while increasing OPA1 cleavage, without expression of levels of mitofusin 2, DRP-1, or mitofilin being affected. In summary, these results indicate that inflammatory cytokines have unique and specialized effects on adipocyte metabolism, but each leads to decreased mitochondrial function and a reprogramming of fat cell biology. PMID:24595304

  11. Simvastatin impairs ADP-stimulated respiration and increases mitochondrial oxidative stress in primary human skeletal myotubes.

    PubMed

    Kwak, Hyo-Bum; Thalacker-Mercer, Anna; Anderson, Ethan J; Lin, Chien-Te; Kane, Daniel A; Lee, Nam-Sihk; Cortright, Ronald N; Bamman, Marcas M; Neufer, P Darrell

    2012-01-01

    Statins, the widely prescribed cholesterol-lowering drugs for the treatment of cardiovascular disease, cause adverse skeletal muscle side effects ranging from fatigue to fatal rhabdomyolysis. The purpose of this study was to determine the effects of simvastatin on mitochondrial respiration, oxidative stress, and cell death in differentiated primary human skeletal muscle cells (i.e., myotubes). Simvastatin induced a dose-dependent decrease in viability of proliferating and differentiating primary human muscle precursor cells, and a similar dose-dependent effect was noted in differentiated myoblasts and myotubes. Additionally, there were decreases in myotube number and size following 48 h of simvastatin treatment (5 μM). In permeabilized myotubes, maximal ADP-stimulated oxygen consumption, supported by palmitoylcarnitine+malate (PCM, complex I and II substrates) and glutamate+malate (GM, complex I substrates), was 32-37% lower (P<0.05) in simvastatin-treated (5 μM) vs control myotubes, providing evidence of impaired respiration at complex I. Mitochondrial superoxide and hydrogen peroxide generation were significantly greater in the simvastatin-treated human skeletal myotube cultures compared to control. In addition, simvastatin markedly increased protein levels of Bax (proapoptotic, +53%) and Bcl-2 (antiapoptotic, +100%, P<0.05), mitochondrial PTP opening (+44%, P<0.05), and TUNEL-positive nuclei in human skeletal myotubes, demonstrating up-regulation of mitochondrial-mediated myonuclear apoptotic mechanisms. These data demonstrate that simvastatin induces myotube atrophy and cell loss associated with impaired ADP-stimulated maximal mitochondrial respiratory capacity, mitochondrial oxidative stress, and apoptosis in primary human skeletal myotubes, suggesting that mitochondrial dysfunction may underlie human statin-induced myopathy.

  12. Analysis of tumor metabolism reveals mitochondrial glucose oxidation in genetically diverse, human glioblastomas in the mouse brain in vivo

    PubMed Central

    Marin-Valencia, Isaac; Yang, Chendong; Mashimo, Tomoyuki; Cho, Steve; Baek, Hyeonman; Yang, Xiao-Li; Rajagopalan, Kartik N.; Maddie, Melissa; Vemireddy, Vamsidhara; Zhao, Zhenze; Cai, Ling; Good, Levi; Tu, Benjamin P.; Hatanpaa, Kimmo J.; Mickey, Bruce E.; Matés, José M.; Pascual, Juan M.; Maher, Elizabeth A.; Malloy, Craig R.; DeBerardinis, Ralph J.; Bachoo, Robert M.

    2012-01-01

    SUMMARY Dysregulated metabolism is a hallmark of cancer cell lines, but little is known about the fate of glucose and other nutrients in tumors growing in their native microenvironment. To study tumor metabolism in vivo, we used an orthotopic mouse model of primary human glioblastoma (GBM). We infused 13C-labeled nutrients into mice bearing three independent GBM lines, each with a distinct set of mutations. All three lines displayed glycolysis, as expected for aggressive tumors. They also displayed unexpected metabolic complexity, oxidizing glucose via pyruvate dehydrogenase and the citric acid cycle, and using glucose to supply anaplerosis and other biosynthetic activities. Comparing the tumors to surrounding brain revealed obvious metabolic differences, notably the accumulation of a large glutamine pool within the tumors. Many of these same activities were conserved in cells cultured ex vivo from the tumors. Thus GBM cells utilize mitochondrial glucose oxidation during aggressive tumor growth in vivo. PMID:22682223

  13. Bridges between mitochondrial oxidative stress, ER stress and mTOR signaling in pancreatic β cells.

    PubMed

    Wang, Jing; Yang, Xin; Zhang, Jingjing

    2016-08-01

    Pancreatic β cell dysfunction, i.e., failure to provide insulin in concentrations sufficient to control blood sugar, is central to the etiology of all types of diabetes. Current evidence implicates mitochondrial oxidative stress and endoplasmic reticulum (ER) stress in pancreatic β cell loss and impaired insulin secretion. Oxidative and ER stress are interconnected so that misfolded proteins induce reactive oxygen species (ROS) production; likewise, oxidative stress disturbs the ER redox state thereby disrupting correct disulfide bond formation and proper protein folding. mTOR signaling regulates many metabolic processes including protein synthesis, cell growth, survival and proliferation. Oxidative stress inhibits mTORC1, which is considered an important suppressor of mitochondrial oxidative stress in β cells, and ultimately, controls cell survival. The interplay between ER stress and mTORC1 is complicated, since the unfolded protein response (UPR) activation can occur upstream or downstream of mTORC1. Persistent activation of mTORC1 initiates protein synthesis and UPR activation, while in the later phase induces ER stress. Chronic activation of ER stress inhibits Akt/mTORC1 pathway, while under particular settings, acute activation of UPR activates Akt-mTOR signaling. Thus, modulating mitochondrial oxidative stress and ER stress via mTOR signaling may be an approach that will effectively suppress obesity- or glucolipotoxicity-induced metabolic disorders such as insulin resistance and type 2 diabetes mellitus (T2DM). In this review, we focus on the regulations between mTOR signaling and mitochondrial oxidative or ER stress in pancreatic β cells.

  14. Formation of a Stabilized Cysteine Sulfinic Acid Is Critical for the Mitochondrial Function of the Parkinsonism Protein DJ-1*

    PubMed Central

    Blackinton, Jeff; Lakshminarasimhan, Mahadevan; Thomas, Kelly J.; Ahmad, Rili; Greggio, Elisa; Raza, Ashraf S.; Cookson, Mark R.; Wilson, Mark A.

    2009-01-01

    The formation of cysteine-sulfinic acid has recently become appreciated as a modification that links protein function to cellular oxidative status. Human DJ-1, a protein associated with inherited parkinsonism, readily forms cysteine-sulfinic acid at a conserved cysteine residue (Cys106 in human DJ-1). Mutation of Cys106 causes the protein to lose its normal protective function in cell culture and model organisms. However, it is unknown whether the loss of DJ-1 protective function in these mutants is due to the absence of Cys106 oxidation or the absence of the cysteine residue itself. To address this question, we designed a series of substitutions at a proximal glutamic acid residue (Glu18) in human DJ-1 that alter the oxidative propensity of Cys106 through changes in hydrogen bonding. We show that two mutations, E18N and E18Q, allow Cys106 to be oxidized to Cys106-sulfinic acid under mild conditions. In contrast, the E18D mutation stabilizes a cysteine-sulfenic acid that is readily reduced to the thiol in solution and in vivo. We show that E18N and E18Q can both partially substitute for wild-type DJ-1 using mitochondrial fission and cell viability assays. In contrast, the oxidatively impaired E18D mutant behaves as an inactive C106A mutant and fails to protect cells. We therefore conclude that formation of Cys106-sulfinic acid is a key modification that regulates the protective function of DJ-1. PMID:19124468

  15. Alterations to mitochondrial fatty-acid use in skeletal muscle after chronic exposure to hypoxia depend on metabolic phenotype.

    PubMed

    Malgoyre, Alexandra; Chabert, Clovis; Tonini, Julia; Koulmann, Nathalie; Bigard, Xavier; Sanchez, Hervé

    2017-03-01

    We investigated the effects of chronic hypoxia on the maximal use of and sensitivity of mitochondria to different substrates in rat slow-oxidative (soleus, SOL) and fast-glycolytic (extensor digitorum longus, EDL) muscles. We studied mitochondrial respiration in situ in permeabilized myofibers, using pyruvate, octanoate, palmitoyl-carnitine (PC), or palmitoyl-coenzyme A (PCoA). The hypophagia induced by hypoxia may also alter metabolism. Therefore, we used a group of pair-fed rats (reproducing the same caloric restriction, as observed in hypoxic animals), in addition to the normoxic control fed ad libitum. The resting respiratory exchange ratio decreased after 21 days of exposure to hypobaric hypoxia (simulated elevation of 5,500 m). The respiration supported by pyruvate and octanoate were unaffected. In contrast, the maximal oxidative respiratory rate for PCoA, the transport of which depends on carnitine palmitoyltransferase 1 (CPT-1), decreased in the rapid-glycolytic EDL and increased in the slow-oxidative SOL, although hypoxia improved affinity for this substrate in both muscle types. PC and PCoA were oxidized similarly in normoxic EDL, whereas chronic hypoxia limited transport at the CPT-1 step in this muscle. The effects of hypoxia were mediated by caloric restriction in the SOL and by hypoxia itself in the EDL. We conclude that improvements in mitochondrial affinity for PCoA, a physiological long-chain fatty acid, would facilitate fatty-acid use at rest after chronic hypoxia independently of quantitative alterations of mitochondria. Conversely, decreasing the maximal oxidation of PCoA in fast-glycolytic muscles would limit fatty-acid use during exercise.NEW & NOTEWORTHY Affinity for low concentrations of long-chain fatty acids (LCFA) in mitochondria skeletal muscles increases after chronic hypoxia. Combined with a lower respiratory exchange ratio, this suggests facility for fatty acid utilization at rest. This fuel preference is related to caloric

  16. Mitochondrial oxidative function in human saponin-skinned muscle fibres: effects of prolonged exercise

    PubMed Central

    Tonkonogi, Michail; Harris, Beorn; Sahlin, Kent

    1998-01-01

    The influence of prolonged exhaustive exercise on mitochondrial oxidative function was investigated in ten men. Muscle biopsies were taken before and after exercise and mitochondrial respiration investigated in fibre bundles made permeable by pretreatment with saponin. After exercise, respiration in the absence of ADP increased by 18 % (P < 0.01), but respiration at suboptimal ADP concentration (0.1 mM) and maximal ADP-stimulated respiration (1 mM ADP) remained unchanged. In the presence of creatine (20 mM), mitochondrial affinity for ADP increased markedly and respiration at suboptimal ADP concentration (0.1 mM) was similar (pre-exercise) or higher (post-exercise; P < 0.05) than with 1 mM ADP alone. The increase in respiratory rate with creatine was correlated to the relative type I fibre area (r = 0.84). Creatine-stimulated respiration increased after prolonged exercise (P < 0.01). The respiratory control index (6.8 ± 0.4, mean ± s.e.m.) and the ratio between respiration at 0.1 and 1 mM ADP (ADP sensitivity index, 0.63 ± 0.03) were not changed after exercise. The sensitivity index was negatively correlated to the relative type I fibre area (r = −0.86). The influence of exercise on muscle oxidative function has for the first time been investigated with the skinned-fibre technique. It is concluded that maximal mitochondrial oxidative power is intact or improved after prolonged exercise, while uncoupled respiration is increased. The latter finding may contribute to the elevated post-exercise oxygen consumption. The finding that the sensitivity of mitochondrial respiration for ADP and creatine are related to fibre-type composition indicates intrinsic differences in the control of mitochondrial respiration between fibres. PMID:9625884

  17. Coenzyme Q10 Inhibits Glutamate Excitotoxicity and Oxidative Stress–Mediated Mitochondrial Alteration in a Mouse Model of Glaucoma

    PubMed Central

    Lee, Dongwook; Shim, Myoung Sup; Kim, Keun-Young; Noh, You Hyun; Kim, Heemin; Kim, Sang Yeop; Weinreb, Robert N.; Ju, Won-Kyu

    2014-01-01

    Purpose. To test whether a diet supplemented with coenzyme Q10 (CoQ10) ameliorates glutamate excitotoxicity and oxidative stress–mediated retinal ganglion cell (RGC) degeneration by preventing mitochondrial alterations in the retina of glaucomatous DBA/2J mice. Methods. Preglaucomatous DBA/2J and age-matched control DBA/2J-Gpnmb+ mice were fed with CoQ10 (1%) or a control diet daily for 6 months. The RGC survival and axon preservation were measured by Brn3a and neurofilament immunohistochemistry and by conventional transmission electron microscopy. Glial fibrillary acidic protein (GFAP), superoxide dismutase-2 (SOD2), heme oxygenase-1 (HO1), N-methyl-d-aspartate receptor (NR) 1 and 2A, and Bax and phosphorylated Bad (pBad) protein expression was measured by Western blot analysis. Apoptotic cell death was assessed by TUNEL staining. Mitochondrial DNA (mtDNA) content and mitochondrial transcription factor A (Tfam)/oxidative phosphorylation (OXPHOS) complex IV protein expression were measured by real-time PCR and Western blot analysis. Results. Coenzyme Q10 promoted RGC survival by approximately 29% and preserved the axons in the optic nerve head (ONH), as well as inhibited astroglial activation by decreasing GFAP expression in the retina and ONH of glaucomatous DBA/2J mice. Intriguingly, CoQ10 significantly blocked the upregulation of NR1 and NR2A, as well as of SOD2 and HO1 protein expression in the retina of glaucomatous DBA/2J mice. In addition, CoQ10 significantly prevented apoptotic cell death by decreasing Bax protein expression or by increasing pBad protein expression. More importantly, CoQ10 preserved mtDNA content and Tfam/OXPHOS complex IV protein expression in the retina of glaucomatous DBA/2J mice. Conclusions. Our findings suggest that CoQ10 may be a promising therapeutic strategy for ameliorating glutamate excitotoxicity and oxidative stress in glaucomatous neurodegeneration. PMID:24458150

  18. Detoxification of Mitochondrial Oxidants and Apoptotic Signaling Are Facilitated by Thioredoxin-2 and Peroxiredoxin-3 during Hyperoxic Injury

    PubMed Central

    Forred, Benjamin J.; Daugaard, Darwin R.; Titus, Brianna K.; Wood, Ryan R.; Floen, Miranda J.; Booze, Michelle L.

    2017-01-01

    Mitochondria play a fundamental role in the regulation of cell death during accumulation of oxidants. High concentrations of atmospheric oxygen (hyperoxia), used clinically to treat tissue hypoxia in premature newborns, is known to elicit oxidative stress and mitochondrial injury to pulmonary epithelial cells. A consequence of oxidative stress in mitochondria is the accumulation of peroxides which are detoxified by the dedicated mitochondrial thioredoxin system. This system is comprised of the oxidoreductase activities of peroxiredoxin-3 (Prx3), thioredoxin-2 (Trx2), and thioredoxin reductase-2 (TrxR2). The goal of this study was to understand the role of the mitochondrial thioredoxin system and mitochondrial injuries during hyperoxic exposure. Flow analysis of the redox-sensitive, mitochondrial-specific fluorophore, MitoSOX, indicated increased levels of mitochondrial oxidant formation in human adenocarcinoma cells cultured in 95% oxygen. Increased expression of Trx2 and TrxR2 in response to hyperoxia were not attributable to changes in mitochondrial mass, suggesting that hyperoxic upregulation of mitochondrial thioredoxins prevents accumulation of oxidized Prx3. Mitochondrial oxidoreductase activities were modulated through pharmacological inhibition of TrxR2 with auranofin and genetically through shRNA knockdown of Trx2 and Prx3. Diminished Trx2 and Prx3 expression was associated with accumulation of mitochondrial superoxide; however, only shRNA knockdown of Trx2 increased susceptibility to hyperoxic cell death and increased phosphorylation of apoptosis signal-regulating kinase-1 (ASK1). In conclusion, the mitochondrial thioredoxin system regulates hyperoxic-mediated death of pulmonary epithelial cells through detoxification of oxidants and regulation of redox-dependent apoptotic signaling. PMID:28045936

  19. Effects of Astragalus Polysaccharides on Dysfunction of Mitochondrial Dynamics Induced by Oxidative Stress

    PubMed Central

    Huang, Yan-Feng; Lu, Lu; Zhu, Da-Jian; Wang, Ming; Yin, Yi; Chen, De-Xiu; Wei, Lian-Bo

    2016-01-01

    This paper studied the chronic fatigue induced by excessive exercise and the restoration effects of Astragalus polysaccharides (APS) on mitochondria. In vivo, we found that excessive exercise could cause oxidative stress statue which led to morphological and functional changes of mitochondria. The changes, including imbalance between mitochondria fusion-fission processes, activation of mitophagy, and decrease of PGC-1α expression, could be restored by APS. We further confirmed in vitro, and what is more, we found that APS may ameliorate mitochondrial dysfunction through Sirt1 pathway. Based on the results, we may figure out part of the molecular mechanism of mitochondrial amelioration by APS. PMID:26881048

  20. Association between heat stress and oxidative stress in poultry; mitochondrial dysfunction and dietary interventions with phytochemicals.

    PubMed

    Akbarian, Abdollah; Michiels, Joris; Degroote, Jeroen; Majdeddin, Maryam; Golian, Abolghasem; De Smet, Stefaan

    2016-01-01

    Heat as a stressor of poultry has been studied extensively for many decades; it affects poultry production on a worldwide basis and has significant impact on well-being and production. More recently, the involvement of heat stress in inducing oxidative stress has received much interest. Oxidative stress is defined as the presence of reactive species in excess of the available antioxidant capacity of animal cells. Reactive species can modify several biologically cellular macromolecules and can interfere with cell signaling pathways. Furthermore, during the last decade, there has been an ever-increasing interest in the use of a wide array of natural feed-delivered phytochemicals that have potential antioxidant properties for poultry. In light of this, the current review aims to (1) summarize the mechanisms through which heat stress triggers excessive superoxide radical production in the mitochondrion and progresses into oxidative stress, (2) illustrate that this pathophysiology is dependent on the intensity and duration of heat stress, (3) present different nutritional strategies for mitigation of mitochondrial dysfunction, with particular focus on antioxidant phytochemicals. Oxidative stress that occurs with heat exposure can be manifest in all parts of the body; however, mitochondrial dysfunction underlies oxidative stress. In the initial phase of acute heat stress, mitochondrial substrate oxidation and electron transport chain activity are increased resulting in excessive superoxide production. During the later stage of acute heat stress, down-regulation of avian uncoupling protein worsens the oxidative stress situation causing mitochondrial dysfunction and tissue damage. Typically, antioxidant enzyme activities are upregulated. Chronic heat stress, however, leads to downsizing of mitochondrial metabolic oxidative capacity, up-regulation of avian uncoupling protein, a clear alteration in the pattern of antioxidant enzyme activities, and depletion of antioxidant

  1. DAPK2 regulates oxidative stress in cancer cells by preserving mitochondrial function

    PubMed Central

    Schlegel, C R; Georgiou, M L; Misterek, M B; Stöcker, S; Chater, E R; Munro, C E; Pardo, O E; Seckl, M J; Costa-Pereira, A P

    2015-01-01

    Death-associated protein kinase (DAPK) 2 is a serine/threonine kinase that belongs to the DAPK family. Although it shows significant structural differences from DAPK1, the founding member of this protein family, DAPK2 is also thought to be a putative tumour suppressor. Like DAPK1, it has been implicated in programmed cell death, the regulation of autophagy and diverse developmental processes. In contrast to DAPK1, however, few mechanistic studies have been carried out on DAPK2 and the majority of these have made use of tagged DAPK2, which almost invariably leads to overexpression of the protein. As a consequence, physiological roles of this kinase are still poorly understood. Using two genetically distinct cancer cell lines as models, we have identified a new role for DAPK2 in the regulation of mitochondrial integrity. RNA interference-mediated depletion of DAPK2 leads to fundamental metabolic changes, including significantly decreased rate of oxidative phosphorylation in combination with overall destabilised mitochondrial membrane potential. This phenotype is further corroborated by an increase in the production of mitochondrial superoxide anions and increased oxidative stress. This then leads to the activation of classical stress-activated kinases such as ERK, JNK and p38, which is observed on DAPK2 genetic ablation. Interestingly, the generation of oxidative stress is further enhanced on overexpression of a kinase-dead DAPK2 mutant indicating that it is the kinase domain of DAPK2 that is important to maintain mitochondrial integrity and, by inference, for cellular metabolism. PMID:25741596

  2. Centella asiatica attenuates β-amyloid-induced oxidative stress and mitochondrial dysfunction

    PubMed Central

    Gray, Nora E.; Sampath, Harini; Zweig, Jonathan A.; Quinn, Joseph F.; Soumyanath, Amala

    2015-01-01

    Background We previously showed that a water extract of the medicinal plant Centella asiatica (CAW) attenuates β-amyloid (Aβ)-induced cognitive deficits in vivo, and prevents Aβ-induced cytotoxicity in vitro. Yet the neuroprotective mechanism of CAW is unknown. Objective The goal of this study was to identify biochemical pathways altered by CAW using in vitro models of Aβ toxicity. Methods The effects of CAW on aberrations in antioxidant response, calcium homeostasis and mitochondrial function induced by Aβ were evaluated in MC65 and SH-SY5Y neuroblastoma cells. Results CAW decreased intracellular ROS and calcium levels elevated in response to Aβ, and induced the expression of antioxidant response genes in both cell lines. In SH-SY5Y cells, CAW increased basal and maximal oxygen consumption without altering spare capacity, and attenuated Aβ-induced decreases in mitochondrial respiration. CAW also prevented Aβ –induced decreases in ATP and induced the expression of mitochondrial genes and proteins in both cell types. Caffeoylquinic acids from CAW were shown to have a similar effect on antioxidant and mitochondrial gene expression in neuroblastoma cells. Primary rat hippocampal neurons treated with CAW also showed an increase in mitochondrial and antioxidant gene expression. Conclusions These data suggest an effect of CAW on mitochondrial biogenesis, which in conjunction with activation of antioxidant response genes and normalizing calcium homeostasis, likely contributes to its neuroprotective action against Aβ toxicity. PMID:25633675

  3. Glutaredoxin 2 catalyzes the reversible oxidation and glutathionylation of mitochondrial membrane thiol proteins: implications for mitochondrial redox regulation and antioxidant DEFENSE.

    PubMed

    Beer, Samantha M; Taylor, Ellen R; Brown, Stephanie E; Dahm, Christina C; Costa, Nikola J; Runswick, Michael J; Murphy, Michael P

    2004-11-12

    The redox poise of the mitochondrial glutathione pool is central in the response of mitochondria to oxidative damage and redox signaling, but the mechanisms are uncertain. One possibility is that the oxidation of glutathione (GSH) to glutathione disulfide (GSSG) and the consequent change in the GSH/GSSG ratio causes protein thiols to change their redox state, enabling protein function to respond reversibly to redox signals and oxidative damage. However, little is known about the interplay between the mitochondrial glutathione pool and protein thiols. Therefore we investigated how physiological GSH/GSSG ratios affected the redox state of mitochondrial membrane protein thiols. Exposure to oxidized GSH/GSSG ratios led to the reversible oxidation of reactive protein thiols by thiol-disulfide exchange, the extent of which was dependent on the GSH/GSSG ratio. There was an initial rapid phase of protein thiol oxidation, followed by gradual oxidation over 30 min. A large number of mitochondrial proteins contain reactive thiols and most of these formed intraprotein disulfides upon oxidation by GSSG; however, a small number formed persistent mixed disulfides with glutathione. Both protein disulfide formation and glutathionylation were catalyzed by the mitochondrial thiol transferase glutaredoxin 2 (Grx2), as were protein deglutathionylation and the reduction of protein disulfides by GSH. Complex I was the most prominent protein that was persistently glutathionylated by GSSG in the presence of Grx2. Maintenance of complex I with an oxidized GSH/GSSG ratio led to a dramatic loss of activity, suggesting that oxidation of the mitochondrial glutathione pool may contribute to the selective complex I inactivation seen in Parkinson's disease. Most significantly, Grx2 catalyzed reversible protein glutathionylation/deglutathionylation over a wide range of GSH/GSSG ratios, from the reduced levels accessible under redox signaling to oxidized ratios only found under severe oxidative

  4. Impact of Antioxidants on Cardiolipin Oxidation in Liposomes: Why Mitochondrial Cardiolipin Serves as an Apoptotic Signal?

    PubMed Central

    Lokhmatikov, Alexey V.; Voskoboynikova, Natalia; Cherepanov, Dmitry A.; Skulachev, Maxim V.; Steinhoff, Heinz-Jürgen; Skulachev, Vladimir P.; Mulkidjanian, Armen Y.

    2016-01-01

    Molecules of mitochondrial cardiolipin (CL) get selectively oxidized upon oxidative stress, which triggers the intrinsic apoptotic pathway. In a chemical model most closely resembling the mitochondrial membrane—liposomes of pure bovine heart CL—we compared ubiquinol-10, ubiquinol-6, and alpha-tocopherol, the most widespread naturally occurring antioxidants, with man-made, quinol-based amphiphilic antioxidants. Lipid peroxidation was induced by addition of an azo initiator in the absence and presence of diverse antioxidants, respectively. The kinetics of CL oxidation was monitored via formation of conjugated dienes at 234 nm. We found that natural ubiquinols and ubiquinol-based amphiphilic antioxidants were equally efficient in protecting CL liposomes from peroxidation; the chromanol-based antioxidants, including alpha-tocopherol, were 2-3 times less efficient. Amphiphilic antioxidants, but not natural ubiquinols and alpha-tocopherol, were able, additionally, to protect the CL bilayer from oxidation by acting from the water phase. We suggest that the previously reported therapeutic efficiency of mitochondrially targeted amphiphilic antioxidants is owing to their ability to protect those CL molecules that are inaccessible to natural hydrophobic antioxidants, being trapped within respiratory supercomplexes. The high susceptibility of such occluded CL molecules to oxidation may have prompted their recruitment as apoptotic signaling molecules by nature. PMID:27313834

  5. Impact of Antioxidants on Cardiolipin Oxidation in Liposomes: Why Mitochondrial Cardiolipin Serves as an Apoptotic Signal?

    PubMed

    Lokhmatikov, Alexey V; Voskoboynikova, Natalia; Cherepanov, Dmitry A; Skulachev, Maxim V; Steinhoff, Heinz-Jürgen; Skulachev, Vladimir P; Mulkidjanian, Armen Y

    2016-01-01

    Molecules of mitochondrial cardiolipin (CL) get selectively oxidized upon oxidative stress, which triggers the intrinsic apoptotic pathway. In a chemical model most closely resembling the mitochondrial membrane-liposomes of pure bovine heart CL-we compared ubiquinol-10, ubiquinol-6, and alpha-tocopherol, the most widespread naturally occurring antioxidants, with man-made, quinol-based amphiphilic antioxidants. Lipid peroxidation was induced by addition of an azo initiator in the absence and presence of diverse antioxidants, respectively. The kinetics of CL oxidation was monitored via formation of conjugated dienes at 234 nm. We found that natural ubiquinols and ubiquinol-based amphiphilic antioxidants were equally efficient in protecting CL liposomes from peroxidation; the chromanol-based antioxidants, including alpha-tocopherol, were 2-3 times less efficient. Amphiphilic antioxidants, but not natural ubiquinols and alpha-tocopherol, were able, additionally, to protect the CL bilayer from oxidation by acting from the water phase. We suggest that the previously reported therapeutic efficiency of mitochondrially targeted amphiphilic antioxidants is owing to their ability to protect those CL molecules that are inaccessible to natural hydrophobic antioxidants, being trapped within respiratory supercomplexes. The high susceptibility of such occluded CL molecules to oxidation may have prompted their recruitment as apoptotic signaling molecules by nature.

  6. Age-related differences in experimental stroke: possible involvement of mitochondrial dysfunction and oxidative damage.

    PubMed

    Li, Nanlin; Kong, Xiangwei; Ye, Ruidong; Yang, Qianzi; Han, Junliang; Xiong, Lize

    2011-06-01

    Age is the single most important risk factor for cerebral stroke. Unfortunately, the effect of age on ischemic brain damage is less clear. In this study, we sought to examine the potential influence of aging on the histologic and functional outcomes after ischemia. Juvenile (4 weeks of age), young adult (4 months of age), mid-aged (11-12 months of age), and aged (18-19 months of age) mice were subjected to transient middle cerebral artery occlusion. There was no remarkable difference of infarct volume on postoperative days 1 and 3. However, on postoperative day 7, aged mice exhibited significantly worsened infarct volume compared with juvenile and young mice. Intriguingly, the increase of infarct volume was most prominent in the striatal area rather than in cortex. Accordingly, aged mice displayed a slower and incomplete functional recovery after stroke. We further evaluated the effects of aging on the oxidative damage and mitochondrial dysfunction following ischemia. Brain tissues were assayed for lipid, DNA, and protein peroxidation products, mitochondrial enzyme activities, mitochondrial membrane potential, production of reactive oxygen species, and antioxidant activities. Aging was associated with declined mitochondrial function and antioxidant detoxification following ischemia, thereby inducing a deteriorated oxidative damage. Regional subanalyses demonstrated that, in accordance with infarct area, the pro-oxidant/antioxidant imbalance occurred more prominently in subcortical areas. Collectively, these findings suggest mitochondria-mediated oxidative damage may be involved in the age-related aggravated injury in subcortical areas. Mitochondrial protection could be a promising target for neuroprotective therapy, especially in the aged population.

  7. Regulation of mitochondrial oxidative stress by β-arrestins in cultured human cardiac fibroblasts

    PubMed Central

    Philip, Jennifer L.; Razzaque, Md. Abdur; Han, Mei; Li, Jinju; Theccanat, Tiju; Xu, Xianyao; Akhter, Shahab A.

    2015-01-01

    ABSTRACT Oxidative stress in cardiac fibroblasts (CFs) promotes transformation to myofibroblasts and collagen synthesis leading to myocardial fibrosis, a precursor to heart failure (HF). NADPH oxidase 4 (Nox4) is a major source of cardiac reactive oxygen species (ROS); however, mechanisms of Nox4 regulation are unclear. β-arrestins are scaffold proteins that signal in G-protein-dependent and -independent pathways; for example, in ERK activation. We hypothesize that β-arrestins regulate oxidative stress in a Nox4-dependent manner and increase fibrosis in HF. CFs were isolated from normal and failing adult human left ventricles. Mitochondrial ROS/superoxide production was quantitated using MitoSox. β-arrestin and Nox4 expressions were manipulated using adenoviral overexpression or short interfering RNA (siRNA)-mediated knockdown. Mitochondrial oxidative stress and Nox4 expression in CFs were significantly increased in HF. Nox4 knockdown resulted in inhibition of mitochondrial superoxide production and decreased basal and TGF-β-stimulated collagen and α-SMA expression. CF β-arrestin expression was upregulated fourfold in HF. β-arrestin knockdown in failing CFs decreased ROS and Nox4 expression by 50%. β-arrestin overexpression in normal CFs increased mitochondrial superoxide production twofold. These effects were prevented by inhibition of either Nox or ERK. Upregulation of Nox4 seemed to be a primary mechanism for increased ROS production in failing CFs, which stimulates collagen deposition. β-arrestin expression was upregulated in HF and plays an important and newly identified role in regulating mitochondrial superoxide production via Nox4. The mechanism for this effect seems to be ERK-mediated. Targeted inhibition of β-arrestins in CFs might decrease oxidative stress as well as pathological cardiac fibrosis. PMID:26449263

  8. Chronically ischemic mouse skeletal muscle exhibits myopathy in association with mitochondrial dysfunction and oxidative damage.

    PubMed

    Pipinos, Iraklis I; Swanson, Stanley A; Zhu, Zhen; Nella, Aikaterini A; Weiss, Dustin J; Gutti, Tanuja L; McComb, Rodney D; Baxter, B Timothy; Lynch, Thomas G; Casale, George P

    2008-07-01

    A myopathy characterized by mitochondrial pathology and oxidative stress is present in patients with peripheral arterial disease (PAD). Patients with PAD differ in disease severity, mode of presentation, and presence of comorbid conditions. In this study, we used a mouse model of hindlimb ischemia to isolate and directly investigate the effects of chronic inflow arterial occlusion on skeletal muscle microanatomy, mitochondrial function and expression, and oxidative stress. Hindlimb ischemia was induced by staged ligation/division of the common femoral and iliac arteries in C57BL/6 mice, and muscles were harvested 12 wk later. Muscle microanatomy was examined by bright-field microscopy, and mitochondrial content was determined as citrate synthase activity in muscle homogenates and ATP synthase expression by fluorescence microscopy. Electron transport chain (ETC) complexes I through IV were analyzed individually by respirometry. Oxidative stress was assessed as total protein carbonyls and 4-hydroxy-2-nonenal (HNE) adducts and altered expression and activity of manganese superoxide dismutase (MnSOD). Ischemic muscle exhibited histological features of myopathy and increased mitochondrial content compared with control muscle. Complex-dependent respiration was significantly reduced for ETC complexes I, III, and IV in ischemic muscle. Protein carbonyls, HNE adducts, and MnSOD expression were significantly increased in ischemic muscle. MnSOD activity was not significantly changed, suggesting MnSOD inactivation. Using a mouse model, we have demonstrated for the first time that inflow arterial occlusion alone, i.e., in the absence of other comorbid conditions, causes myopathy with mitochondrial dysfunction and increased oxidative stress, recapitulating the muscle pathology of PAD patients.

  9. Acrolein cytotoxicity in hepatocytes involves endoplasmic reticulum stress, mitochondrial dysfunction and oxidative stress

    SciTech Connect

    Mohammad, Mohammad K.; Avila, Diana; Zhang, Jingwen; Barve, Shirish; Arteel, Gavin; McClain, Craig; Joshi-Barve, Swati

    2012-11-15

    Acrolein is a common environmental, food and water pollutant and a major component of cigarette smoke. Also, it is produced endogenously via lipid peroxidation and cellular metabolism of certain amino acids and drugs. Acrolein is cytotoxic to many cell types including hepatocytes; however the mechanisms are not fully understood. We examined the molecular mechanisms underlying acrolein hepatotoxicity in primary human hepatocytes and hepatoma cells. Acrolein, at pathophysiological concentrations, caused a dose-dependent loss of viability of hepatocytes. The death was apoptotic at moderate and necrotic at high concentrations of acrolein. Acrolein exposure rapidly and dramatically decreased intracellular glutathione and overall antioxidant capacity, and activated the stress-signaling MAP-kinases JNK, p42/44 and p38. Our data demonstrate for the first time in human hepatocytes, that acrolein triggered endoplasmic reticulum (ER) stress and activated eIF2α, ATF-3 and -4, and Gadd153/CHOP, resulting in cell death. Notably, the protective/adaptive component of ER stress was not activated, and acrolein failed to up-regulate the protective ER-chaperones, GRP78 and GRP94. Additionally, exposure to acrolein disrupted mitochondrial integrity/function, and led to the release of pro-apoptotic proteins and ATP depletion. Acrolein-induced cell death was attenuated by N-acetyl cysteine, phenyl-butyric acid, and caspase and JNK inhibitors. Our data demonstrate that exposure to acrolein induces a variety of stress responses in hepatocytes, including GSH depletion, oxidative stress, mitochondrial dysfunction and ER stress (without ER-protective responses) which together contribute to acrolein toxicity. Our study defines basic mechanisms underlying liver injury caused by reactive aldehyde pollutants such as acrolein. -- Highlights: ► Human primary hepatocytes and cultured cell lines are used. ► Multiple cell death signaling pathways are activated by acrolein. ► Novel finding of

  10. Iron-dependent changes in cellular energy metabolism: influence on citric acid cycle and oxidative phosphorylation.

    PubMed

    Oexle, H; Gnaiger, E; Weiss, G

    1999-11-10

    Iron modulates the expression of the critical citric acid cycle enzyme aconitase via a translational mechanism involving iron regulatory proteins. Thus, the present study was undertaken to investigate the consequences of iron perturbation on citric acid cycle activity, oxidative phosphorylation and mitochondrial respiration in the human cell line K-562. In agreement with previous data iron increases the activity of mitochondrial aconitase while it is reduced upon addition of the iron chelator desferrioxamine (DFO). Interestingly, iron also positively affects three other citric acid cycle enzymes, namely citrate synthase, isocitric dehydrogenase, and succinate dehydrogenase, while DFO decreases the activity of these enzymes. Consequently, iron supplementation results in increased formation of reducing equivalents (NADH) by the citric acid cycle, and thus in increased mitochondrial oxygen consumption and ATP formation via oxidative phosphorylation as shown herein. This in turn leads to downregulation of glucose utilization. In contrast, all these metabolic pathways are reduced upon iron depletion, and thus glycolysis and lactate formation are significantly increased in order to compensate for the decrease in ATP production via oxidative phosphorylation in the presence of DFO. Our results point to a complex interaction between iron homeostasis, oxygen supply and cellular energy metabolism in human cells.

  11. Acid-permanganate oxidation of potassium tetraphenylboron

    SciTech Connect

    Smith, J.R.

    1993-02-01

    Scoping experiments have been performed which show that potassium tetraphenylboron (KTPB) is rapidly oxidized by permanganate in acidic solutions at room temperature. The main Products are CO[sub 2], highly oxidized organic compounds related to tartaric and tartronic acids, boric acid, and potassium phosphate (when phosphoric acid is used as the source of acid). One liter of 0.6M NaMnO[sub 4]/2.5M H[sub 3]PO[sub 4] solution will destroy up to 8 grams of KTPB. The residual benzene concentration has been measured to be less than the RCRA limit of 0.5 ppm. Approximately 30% of the organic material is released as CO[sub 2] (trace CO) and 0.16% as benzene vapor. The reaction is well behaved, no foaming or spattering. Tests were performed from .15M to near 1M permanganate. The phosphoric acid concentration was maintained at a concentration at least three times that of the permanganate since an excess of acid was desired and this is the ratio that these two reagents are consumed in the oxidation.

  12. Acid-permanganate oxidation of potassium tetraphenylboron

    SciTech Connect

    Smith, J.R.

    1993-02-01

    Scoping experiments have been performed which show that potassium tetraphenylboron (KTPB) is rapidly oxidized by permanganate in acidic solutions at room temperature. The main Products are CO{sub 2}, highly oxidized organic compounds related to tartaric and tartronic acids, boric acid, and potassium phosphate (when phosphoric acid is used as the source of acid). One liter of 0.6M NaMnO{sub 4}/2.5M H{sub 3}PO{sub 4} solution will destroy up to 8 grams of KTPB. The residual benzene concentration has been measured to be less than the RCRA limit of 0.5 ppm. Approximately 30% of the organic material is released as CO{sub 2} (trace CO) and 0.16% as benzene vapor. The reaction is well behaved, no foaming or spattering. Tests were performed from .15M to near 1M permanganate. The phosphoric acid concentration was maintained at a concentration at least three times that of the permanganate since an excess of acid was desired and this is the ratio that these two reagents are consumed in the oxidation.

  13. CARDIOSELECTIVE OXIDATION OF MITOCHONDRIAL DNA FOLLOWING SUBCHRONIC ADMINISTRATION OF DOXORUBICIN

    EPA Science Inventory

    This preferential oxidation of cardiac mtDNA is consistent with the bioenergetic failure and the cumulative and irreversible cardiomyopathy that limits the clinical utility of this important antineoplastic drug.

  14. Mitochondrial PKM2 regulates oxidative stress-induced apoptosis by stabilizing Bcl2

    PubMed Central

    Liang, Ji; Cao, Ruixiu; Wang, Xiongjun; Zhang, Yajuan; Wang, Pan; Gao, Hong; Li, Chen; Yang, Fan; Zeng, Rong; Wei, Ping; Li, Dawei; Li, Wenfeng; Yang, Weiwei

    2017-01-01

    Pyruvate kinase M2 isoform (PKM2) catalyzes the last step of glycolysis and plays an important role in tumor cell proliferation. Recent studies have reported that PKM2 also regulates apoptosis. However, the mechanisms underlying such a role of PKM2 remain elusive. Here we show that PKM2 translocates to mitochondria under oxidative stress. In the mitochondria, PKM2 interacts with and phosphorylates Bcl2 at threonine (T) 69. This phosphorylation prevents the binding of Cul3-based E3 ligase to Bcl2 and subsequent degradation of Bcl2. A chaperone protein, HSP90α1, is required for this function of PKM2. HSP90α1's ATPase activity launches a conformational change of PKM2 and facilitates interaction between PKM2 and Bcl2. Replacement of wild-type Bcl2 with phosphorylation-deficient Bcl2 T69A mutant sensitizes glioma cells to oxidative stress-induced apoptosis and impairs brain tumor formation in an orthotopic xenograft model. Notably, a peptide that is composed of the amino acid residues from 389 to 405 of PKM2, through which PKM2 binds to Bcl2, disrupts PKM2-Bcl2 interaction, promotes Bcl2 degradation and impairs brain tumor growth. In addition, levels of Bcl2 T69 phosphorylation, conformation-altered PKM2 and Bcl2 protein correlate with one another in specimens of human glioblastoma patients. Moreover, levels of Bcl2 T69 phosphorylation and conformation-altered PKM2 correlate with both grades and prognosis of glioma malignancy. Our findings uncover a novel mechanism through which mitochondrial PKM2 phosphorylates Bcl2 and inhibits apoptosis directly, highlight the essential role of PKM2 in ROS adaptation of cancer cells, and implicate HSP90-PKM2-Bcl2 axis as a potential target for therapeutic intervention in glioblastoma. PMID:28035139

  15. A Fox2-Dependent Fatty Acid ß-Oxidation Pathway Coexists Both in Peroxisomes and Mitochondria of the Ascomycete Yeast Candida lusitaniae

    PubMed Central

    Bessoule, Jean-Jacques; Salin, Bénédicte; Lucas-Guérin, Marine; Manon, Stephen; Dementhon, Karine; Noël, Thierry

    2014-01-01

    It is generally admitted that the ascomycete yeasts of the subphylum Saccharomycotina possess a single fatty acid ß-oxidation pathway located exclusively in peroxisomes, and that they lost mitochondrial ß-oxidation early during evolution. In this work, we showed that mutants of the opportunistic pathogenic yeast Candida lusitaniae which lack the multifunctional enzyme Fox2p, a key enzyme of the ß-oxidation pathway, were still able to grow on fatty acids as the sole carbon source, suggesting that C. lusitaniae harbored an alternative pathway for fatty acid catabolism. By assaying 14Cα-palmitoyl-CoA consumption, we demonstrated that fatty acid catabolism takes place in both peroxisomal and mitochondrial subcellular fractions. We then observed that a fox2Δ null mutant was unable to catabolize fatty acids in the mitochondrial fraction, thus indicating that the mitochondrial pathway was Fox2p-dependent. This finding was confirmed by the immunodetection of Fox2p in protein extracts obtained from purified peroxisomal and mitochondrial fractions. Finally, immunoelectron microscopy provided evidence that Fox2p was localized in both peroxisomes and mitochondria. This work constitutes the first demonstration of the existence of a Fox2p-dependent mitochondrial β-oxidation pathway in an ascomycetous yeast, C. lusitaniae. It also points to the existence of an alternative fatty acid catabolism pathway, probably located in peroxisomes, and functioning in a Fox2p-independent manner. PMID:25486052

  16. Interrelationships between mitochondrial fusion, energy metabolism and oxidative stress during development in Caenorhabditis elegans.

    PubMed

    Yasuda, Kayo; Hartman, Philip S; Ishii, Takamasa; Suda, Hitoshi; Akatsuka, Akira; Shoyama, Tetsuji; Miyazawa, Masaki; Ishii, Naoaki

    2011-01-21

    Mitochondria are known to be dynamic structures with the energetically and enzymatically mediated processes of fusion and fission responsible for maintaining a constant flux. Mitochondria also play a role of reactive oxygen species production as a byproduct of energy metabolism. In the current study, interrelationships between mitochondrial fusion, energy metabolism and oxidative stress on development were explored using a fzo-1 mutant defective in the fusion process and a mev-1 mutant overproducing superoxide from mitochondrial electron transport complex II of Caenorhabditis elegans. While growth and development of both single mutants was slightly delayed relative to the wild type, the fzo-1;mev-1 double mutant experienced considerable delay. Oxygen sensitivity during larval development, superoxide production and carbonyl protein accumulation of the fzo-1 mutant were similar to wild type. fzo-1 animals had significantly lower metabolism than did N2 and mev-1. These data indicate that mitochondrial fusion can profoundly affect energy metabolism and development.

  17. Fatty acid oxidation and ketogenesis during development.

    PubMed

    Girard, J; Duée, P H; Ferré, P; Pégorier, J P; Escriva, F; Decaux, J F

    1985-01-01

    Fatty acids are the preferred oxidative substrates of the heart, skeletal muscles, kidney cortex and liver in adult mammals. They are supplied to these tissues either as nonesterified fatty acids (NEFA), or as triglycerides after hydrolysis by lipoprotein lipase. During fetal life, tissue capacity to oxidize NEFA is very low, even in species in which the placental transfer of NEFA and carnitine is high. At birth, the ability to oxidize NEFA from endogenous sources or from milk (a high-fat diet) develops rapidly in various tissues and remains very high throughout the suckling period. Ketogenesis appears in the liver by 6 to 12 hrs after birth, and the ketone bodies are used as oxidative fuels by various tissues during the suckling period. At the time of weaning, the transition from a high-fat to a high-carbohydrate diet is attended by a progressive decrease in the ketogenic capacity of the liver, whereas other tissues (skeletal muscle, heart, kidney) maintain a high capacity for NEFA oxidation. The nutritional and hormonal factors involved in changes in fatty acid oxidation during development are discussed.

  18. Malfunction in Mitochondrial β-Oxidation Contributes to Lipid Accumulation in Hepatocyte-Like Cells Derived from Citrin Deficiency-Induced Pluripotent Stem Cells.

    PubMed

    Kim, Yeji; Choi, Jung-Yun; Lee, Sang-Hee; Lee, Beom-Hee; Yoo, Han-Wook; Han, Yong-Mahn

    2016-04-15

    Citrin deficiency (CD) is a recessive genetic disorder caused by mutations in the citrin gene SLC25A13. CD causes various symptoms related to nutrient metabolism such as urea cycle failure, abnormal amino acid levels, and fatty liver. To understand the pathophysiology of CD, the molecular phenotypes were investigated using induced pluripotent stem cells derived from fibroblasts of CD patient (CD-iPSCs). In this study, we demonstrate that aberrant mitochondrial β-oxidation may lead to fatty liver in CD patients. CD-iPSCs normally differentiated into hepatocytes, similar to wild-type iPSCs (WT-iPSCs). However, hepatocytes derived from CD-iPSCs (CD-HLCs) did not exhibit ureogenesis. Cellular triglyceride and lipid granule levels were significantly increased in CD-HLCs compared with WT-HLCs. Peroxisome proliferator-activated receptor-α (PPAR-α) and its target genes which are involved in mitochondrial β-oxidation were downregulated in CD-HLCs, and treatment with a PPAR-α agonist partially reduced the lipid accumulation in CD-HLCs. In addition, the mitochondria in CD-HLCs exhibited abnormal morphologies. Based on these observations, we conclude that the lipid accumulation in CD-HLCs results from dysfunctional mitochondrial β-oxidation and abnormal mitochondrial structure.

  19. Residual mitochondrial transmembrane potential decreases unsaturated fatty acid level in sake yeast during alcoholic fermentation.

    PubMed

    Sawada, Kazutaka; Kitagaki, Hiroshi

    2016-01-01

    Oxygen, a key nutrient in alcoholic fermentation, is rapidly depleted during this process. Several pathways of oxygen utilization have been reported in the yeast Saccharomyces cerevisiae during alcoholic fermentation, namely synthesis of unsaturated fatty acid, sterols and heme, and the mitochondrial electron transport chain. However, the interaction between these pathways has not been investigated. In this study, we showed that the major proportion of unsaturated fatty acids of ester-linked lipids in sake fermentation mash is derived from the sake yeast rather than from rice or koji (rice fermented with Aspergillus). Additionally, during alcoholic fermentation, inhibition of the residual mitochondrial activity of sake yeast increases the levels of unsaturated fatty acids of ester-linked lipids. These findings indicate that the residual activity of the mitochondrial electron transport chain reduces molecular oxygen levels and decreases the synthesis of unsaturated fatty acids, thereby increasing the synthesis of estery flavors by sake yeast. This is the first report of a novel link between residual mitochondrial transmembrane potential and the synthesis of unsaturated fatty acids by the brewery yeast during alcoholic fermentation.

  20. Calcium Uptake via Mitochondrial Uniporter Contributes to Palmitic Acid-induced Apoptosis in Mouse Podocytes.

    PubMed

    Yuan, Zeting; Cao, Aili; Liu, Hua; Guo, Henjiang; Zang, Yingjun; Wang, Yi; Wang, Yunman; Wang, Hao; Yin, Peihao; Peng, Wen

    2017-02-09

    Podocytes are component cells of the glomerular filtration barrier, and their loss by apoptosis is the main cause of proteinuria that leads to diabetic nephropathy (DN). Therefore, insights into podocyte apoptosis mechanism would allow a better understanding of DN pathogenesis and thus help develop adequate therapeutic strategies. Here, we investigated the molecular mechanism of palmitic acid-inhibited cell death in mouse podocytes, and found that palmitic acid increased cell death in a dose- and time-dependent manner. Palmitic acid induces apoptosis in podocytes through up-regulation of cytosolic and mitochondrial Ca(2+) , mitochondrial membrane potential (MMP), cytochrome c release and depletion of endoplasmic reticulum (ER) Ca(2+) , The intracellular calcium chelator, 1,2-bis (2-aminophenoxy) ethane-N,N,N, N'-tetraacetic acid tetrakis acetoxymethyl ester (BAPTA-AM), partially prevented this up-regulation whereas 2-aminoethoxydiphenyl borate (2-APB), an inositol 1,4,5-triphosphate receptor (IP3R) inhibitor; dantrolene, a ryanodine receptor (RyR) inhibitor; and 4,4'-diisothiocyanatostibene-2,2'-disulfonic acid (DIDS), an anion exchange inhibitor, had no effect. Interestingly, ruthenium red and Ru360, both inhibitors of the mitochondrial Ca(2+) uniporter (MCU), blocked palmitic acid-induced mitochondrial Ca(2+) elevation, cytochrome c release from mitochondria to cytosol, and apoptosis. siRNA to MCU markedly reduced curcumin-induced apoptosis. These data indicate that Ca(2+) uptake via mitochondrial uniporter contributes to palmitic acid-induced apoptosis in mouse podocytes. This article is protected by copyright. All rights reserved.

  1. Polyunsaturated fatty acids trigger apoptosis of colon cancer cells through a mitochondrial pathway

    PubMed Central

    Zhang, Chengcheng; Yu, Haining; Shen, Yuzhen; Ni, Xiaofeng; Das, Undurti N.

    2015-01-01

    Introduction Colorectal cancer is common in developed countries. Polyunsaturated fatty acids (PUFAs) have been reported to possess tumoricidal action, but the exact mechanism of their action is not clear. Material and methods In the present study, we studied the effect of various n-6 and n-3 fatty acids on the survival of the colon cancer cells LoVo and RKO and evaluated the possible involvement of a mitochondrial pathway in their ability to induce apoptosis. Results It was observed that n-3 α-linolenic acid, eicosapentaenoic acid and docosahexaenoic acid (ALA, EPA and DHA respectively) and n-6 linoleic acid, gamma-linolenic acid and arachidonic acid (LA, GLA and AA respectively) induced apoptosis of the colon cancer cells LoVo and RKO at concentrations above 120 μM (p < 0.01 compared to control). The semi-differentiated colon cancer cell line RKO was more sensitive to the cytotoxic action of PUFAs compared to the undifferentiated colon cancer cell line LoVo. PUFA-treated cells showed an increased number of lipid droplets in their cytoplasm. PUFA-induced apoptosis of LoVo and RKO cells is mediated through a mitochondria-mediated pathway as evidenced by loss of mitochondrial membrane potential, generation of ROS, accumulation of intracellular Ca2+, activation of caspase-9 and caspase-3, decreased ATP level and increase in the Bax/Bcl2 expression ratio. Conclusions PUFAs induced apoptosis of colon cancer cells through a mitochondrial dependent pathway. PMID:26528354

  2. Chemoproteomic Profiling of Acetanilide Herbicides Reveals Their Role in Inhibiting Fatty Acid Oxidation.

    PubMed

    Counihan, Jessica L; Duckering, Megan; Dalvie, Esha; Ku, Wan-Min; Bateman, Leslie A; Fisher, Karl J; Nomura, Daniel K

    2017-03-17

    Acetanilide herbicides are among the most widely used pesticides in the United States, but their toxicological potential and mechanisms remain poorly understood. Here, we have used chemoproteomic platforms to map proteome-wide cysteine reactivity of acetochlor (AC), the most widely used acetanilide herbicide, in vivo in mice. We show that AC directly reacts with >20 protein targets in vivo in mouse liver, including the catalytic cysteines of several thiolase enzymes involved in mitochondrial and peroxisomal fatty acid oxidation. We show that the fatty acids that are not oxidized, due to impaired fatty acid oxidation, are instead diverted into other lipid pathways, resulting in heightened free fatty acids, triglycerides, cholesteryl esters, and other lipid species in the liver. Our findings show the utility of chemoproteomic approaches for identifying novel mechanisms of toxicity associated with environmental chemicals like acetanilide herbicides.

  3. Demethyleneberberine, a natural mitochondria-targeted antioxidant, inhibits mitochondrial dysfunction, oxidative stress, and steatosis in alcoholic liver disease mouse model.

    PubMed

    Zhang, Pengcheng; Qiang, Xiaoyan; Zhang, Miao; Ma, Dongshen; Zhao, Zheng; Zhou, Cuisong; Liu, Xie; Li, Ruiyan; Chen, Huan; Zhang, Yubin

    2015-01-01

    Excessive alcohol consumption induces oxidative stress and lipid accumulation in the liver. Mitochondria have long been recognized as the key target for alcoholic liver disease (ALD). Recently, the artificial mitochondria-targeted antioxidant MitoQ has been used to treat ALD effectively in mice. Here, we introduce the natural mitochondria-targeted antioxidant demethyleneberberine (DMB), which has been found in Chinese herb Cortex Phellodendri chinensis. The protective effect of DMB on ALD was evaluated with HepG2 cells and acutely/chronically ethanol-fed mice, mimicking two common patterns of drinking in human. The results showed that DMB, which is composed of a potential antioxidant structure, could penetrate the membrane of mitochondria and accumulate in mitochondria either in vitro or in vivo. Consequently, the acute drinking-caused oxidative stress and mitochondrial dysfunction were significantly ameliorated by DMB. Moreover, we also found that DMB suppressed CYP2E1, hypoxia inducible factor α, and inducible nitric oxide synthase, which contributed to oxidative stress and restored sirtuin 1/AMP-activated protein kinase/peroxisome proliferator-activated receptor-γ coactivator-1α pathway-associated fatty acid oxidation in chronic ethanol-fed mice, which in turn ameliorated lipid peroxidation and macrosteatosis in the liver. Taking these findings together, DMB could serve as a novel and potential therapy for ALD in human beings.

  4. Dexmedetomidine attenuates lipopolysaccharide-induced acute lung injury by inhibiting oxidative stress, mitochondrial dysfunction and apoptosis in rats

    PubMed Central

    Fu, Chunlai; Dai, Xingui; Yang, You; Lin, Mengxiang; Cai, Yeping; Cai, Shaoxi

    2016-01-01

    Previous studies have identified that dexmedetomidine (DEX) treatment can ameliorate the acute lung injury (ALI) induced by lipopolysaccharide and ischemia-reperfusion. However, the molecular mechanisms by which DEX ameliorates lung injury remain unclear. The present study investigated whether DEX, which has been reported to exert effects on oxidative stress, mitochondrial permeability transition pores and apoptosis in other disease types, can exert protective effects in lipopolysaccharide (LPS)-induced ALI by inhibiting oxidative stress, mitochondrial dysfunction and mitochondrial-dependent apoptosis. It was revealed that LPS-challenged rats exhibited significant lung injury, characterized by the deterioration of histopathology, vascular hyperpermeability, wet-to-dry weight ratio and oxygenation index (PaO2/FIO2), which was attenuated by DEX treatment. DEX treatment inhibited LPS-induced mitochondrial dysfunction, as evidenced by alleviating the cellular ATP and mitochondrial membrane potential in vitro. In addition, DEX treatment markedly prevented the LPS-induced mitochondrial-dependent apoptotic pathway in vitro (increases of cell apoptotic rate, cytosolic cytochrome c, and caspase 3 activity) and in vivo (increases of |terminal deoxynucleotidyl transferase dUTP nick-end labeling positive cells, cleaved caspase 3, Bax upregulation and Bcl-2 downregulation). Furthermore, DEX treatment markedly attenuated LPS-induced oxidative stress, as evidenced by downregulation of cellular reactive oxygen species in vitro and lipid peroxides in serum. Collectively, the present results demonstrated that DEX ameliorates LPS-induced ALI by reducing oxidative stress, mitochondrial dysfunction and mitochondrial-dependent apoptosis. PMID:27959438

  5. Epigallocatechin-3-gallate prevents oxidative phosphorylation deficit and promotes mitochondrial biogenesis in human cells from subjects with Down's syndrome.

    PubMed

    Valenti, Daniela; De Rasmo, Domenico; Signorile, Anna; Rossi, Leonardo; de Bari, Lidia; Scala, Iris; Granese, Barbara; Papa, Sergio; Vacca, Rosa Anna

    2013-04-01

    A critical role for mitochondrial dysfunction has been proposed in the pathogenesis of Down's syndrome (DS), a human multifactorial disorder caused by trisomy of chromosome 21, associated with mental retardation and early neurodegeneration. Previous studies from our group demonstrated in DS cells a decreased capacity of the mitochondrial ATP production system and overproduction of reactive oxygen species (ROS) in mitochondria. In this study we have tested the potential of epigallocatechin-3-gallate (EGCG) - a natural polyphenol component of green tea - to counteract the mitochondrial energy deficit found in DS cells. We found that EGCG, incubated with cultured lymphoblasts and fibroblasts from DS subjects, rescued mitochondrial complex I and ATP synthase catalytic activities, restored oxidative phosphorylation efficiency and counteracted oxidative stress. These effects were associated with EGCG-induced promotion of PKA activity, related to increased cellular levels of cAMP and PKA-dependent phosphorylation of the NDUFS4 subunit of complex I. In addition, EGCG strongly promoted mitochondrial biogenesis in DS cells, as associated with increase in Sirt1-dependent PGC-1α deacetylation, NRF-1 and T-FAM protein levels and mitochondrial DNA content. In conclusion, this study shows that EGCG is a promoting effector of oxidative phosphorylation and mitochondrial biogenesis in DS cells, acting through modulation of the cAMP/PKA- and sirtuin-dependent pathways. EGCG treatment promises thus to be a therapeutic approach to counteract mitochondrial energy deficit and oxidative stress in DS.

  6. Impaired enzymatic defensive activity, mitochondrial dysfunction and proteasome activation are involved in RTT cell oxidative damage.

    PubMed

    Cervellati, Carlo; Sticozzi, Claudia; Romani, Arianna; Belmonte, Giuseppe; De Rasmo, Domenico; Signorile, Anna; Cervellati, Franco; Milanese, Chiara; Mastroberardino, Pier Giorgio; Pecorelli, Alessandra; Savelli, Vinno; Forman, Henry J; Hayek, Joussef; Valacchi, Giuseppe

    2015-10-01

    A strong correlation between oxidative stress (OS) and Rett syndrome (RTT), a rare neurodevelopmental disorder affecting females in the 95% of the cases, has been well documented although the source of OS and the effect of a redox imbalance in this pathology has not been yet investigated. Using freshly isolated skin fibroblasts from RTT patients and healthy subjects, we have demonstrated in RTT cells high levels of H2O2 and HNE protein adducts. These findings correlated with the constitutive activation of NADPH-oxidase (NOX) and that was prevented by a NOX inhibitor and iron chelator pre-treatment, showing its direct involvement. In parallel, we demonstrated an increase in mitochondrial oxidant production, altered mitochondrial biogenesis and impaired proteasome activity in RTT samples. Further, we found that the key cellular defensive enzymes: glutathione peroxidase, superoxide dismutase and thioredoxin reductases activities were also significantly lower in RTT. Taken all together, our findings suggest that the systemic OS levels in RTT can be a consequence of both: increased endogenous oxidants as well as altered mitochondrial biogenesis with a decreased activity of defensive enzymes that leads to posttranslational oxidant protein modification and a proteasome activity impairment.

  7. Oxidative Stress and Mitochondrial Dysfunction across Broad-Ranging Pathologies: Toward Mitochondria-Targeted Clinical Strategies

    PubMed Central

    d'Ischia, Marco; Gadaleta, Maria Nicola; Pallardó, Federico V.; Petrović, Sandra; Tiano, Luca; Zatterale, Adriana

    2014-01-01

    Beyond the disorders recognized as mitochondrial diseases, abnormalities in function and/or ultrastructure of mitochondria have been reported in several unrelated pathologies. These encompass ageing, malformations, and a number of genetic or acquired diseases, as diabetes and cardiologic, haematologic, organ-specific (e.g., eye or liver), neurologic and psychiatric, autoimmune, and dermatologic disorders. The mechanistic grounds for mitochondrial dysfunction (MDF) along with the occurrence of oxidative stress (OS) have been investigated within the pathogenesis of individual disorders or in groups of interrelated disorders. We attempt to review broad-ranging pathologies that involve mitochondrial-specific deficiencies or rely on cytosol-derived prooxidant states or on autoimmune-induced mitochondrial damage. The established knowledge in these subjects warrants studies aimed at elucidating several open questions that are highlighted in the present review. The relevance of OS and MDF in different pathologies may establish the grounds for chemoprevention trials aimed at compensating OS/MDF by means of antioxidants and mitochondrial nutrients. PMID:24876913

  8. Regulation of mitochondrial function and endoplasmic reticulum stress by nitric oxide in pluripotent stem cells

    PubMed Central

    Caballano-Infantes, Estefania; Terron-Bautista, José; Beltrán-Povea, Amparo; Cahuana, Gladys M; Soria, Bernat; Nabil, Hajji; Bedoya, Francisco J; Tejedo, Juan R

    2017-01-01

    Mitochondrial dysfunction and endoplasmic reticulum stress (ERS) are global processes that are interrelated and regulated by several stress factors. Nitric oxide (NO) is a multifunctional biomolecule with many varieties of physiological and pathological functions, such as the regulation of cytochrome c inhibition and activation of the immune response, ERS and DNA damage; these actions are dose-dependent. It has been reported that in embryonic stem cells, NO has a dual role, controlling differentiation, survival and pluripotency, but the molecular mechanisms by which it modulates these functions are not yet known. Low levels of NO maintain pluripotency and induce mitochondrial biogenesis. It is well established that NO disrupts the mitochondrial respiratory chain and causes changes in mitochondrial Ca2+ flux that induce ERS. Thus, at high concentrations, NO becomes a potential differentiation agent due to the relationship between ERS and the unfolded protein response in many differentiated cell lines. Nevertheless, many studies have demonstrated the need for physiological levels of NO for a proper ERS response. In this review, we stress the importance of the relationships between NO levels, ERS and mitochondrial dysfunction that control stem cell fate as a new approach to possible cell therapy strategies. PMID:28289506

  9. Mitochondrial ATP synthases cluster as discrete domains that reorganize with the cellular demand for oxidative phosphorylation.

    PubMed

    Jimenez, Laure; Laporte, Damien; Duvezin-Caubet, Stephane; Courtout, Fabien; Sagot, Isabelle

    2014-02-15

    Mitochondria are double membrane-bounded organelles that form a dynamic tubular network. Mitochondria energetic functions depend on a complex internal architecture. Cristae, inner membrane invaginations that fold into the matrix space, are proposed to be the site of oxidative phosphorylation, reactions by which ATP synthase produces ATP. ATP synthase is also thought to have a role in crista morphogenesis. To date, the exploration of the processes regulating mitochondrial internal compartmentalization have been mostly limited to electron microscopy. Here, we describe ATP synthase localization in living yeast cells and show that it clusters as discrete inner membrane domains. These domains are dynamic within the mitochondrial network. They are impaired in mutants defective in crista morphology and partially overlap with the crista-associated MICOS-MINOS-MITOS complex. Finally, ATP synthase occupancy increases with the cellular demand for OXPHOS. Overall our data suggest that domains in which ATP synthases are clustered correspond to mitochondrial cristae. Being able to follow mitochondrial sub-compartments in living yeast cells opens new avenues to explore the mechanisms involved in inner membrane remodeling, an architectural feature crucial for mitochondrial activities.

  10. Interrelationships between mitochondrial fusion, energy metabolism and oxidative stress during development in Caenorhabditis elegans

    SciTech Connect

    Yasuda, Kayo; Hartman, Philip S.; Ishii, Takamasa; Suda, Hitoshi; Akatsuka, Akira; Shoyama, Tetsuji; Miyazawa, Masaki; Ishii, Naoaki

    2011-01-21

    Research highlights: {yields} Growth and development of a fzo-1 mutant defective in the fusion process of mitochondria was delayed relative to the wild type of Caenorhabditis elegans. {yields} Oxygen sensitivity during larval development, superoxide production and carbonyl protein accumulation of the fzo-1 mutant were similar to wild type. {yields} fzo-1 animals had significantly lower metabolism than did N2 and mev-1 overproducing superoxide from mitochondrial electron transport complex II. {yields} Mitochondrial fusion can profoundly affect energy metabolism and development. -- Abstract: Mitochondria are known to be dynamic structures with the energetically and enzymatically mediated processes of fusion and fission responsible for maintaining a constant flux. Mitochondria also play a role of reactive oxygen species production as a byproduct of energy metabolism. In the current study, interrelationships between mitochondrial fusion, energy metabolism and oxidative stress on development were explored using a fzo-1 mutant defective in the fusion process and a mev-1 mutant overproducing superoxide from mitochondrial electron transport complex II of Caenorhabditis elegans. While growth and development of both single mutants was slightly delayed relative to the wild type, the fzo-1;mev-1 double mutant experienced considerable delay. Oxygen sensitivity during larval development, superoxide production and carbonyl protein accumulation of the fzo-1 mutant were similar to wild type. fzo-1 animals had significantly lower metabolism than did N2 and mev-1. These data indicate that mitochondrial fusion can profoundly affect energy metabolism and development.

  11. Does the oxidative stress theory of aging explain longevity differences in birds? I. Mitochondrial ROS production.

    PubMed

    Montgomery, Magdalene K; Hulbert, A J; Buttemer, William A

    2012-03-01

    Mitochondrial reactive oxygen species (ROS) production rates are reported to be inversely related to maximum lifespan potential (MLSP) in mammals and also to be higher in short-living mammals compared to short-living birds. The mammal-bird comparison, however, is mainly based on studies of rats and pigeons. To date, there has been no systematic examination of ROS production in birds that differ in MLSP. Here we report a comparison of mitochondrial ROS production in two short-living (quails) and three long-living bird species (parrots) that exhibit, on average, a 5-fold longevity difference. Mitochondrial ROS production was determined both in isolated mitochondria (heart, skeletal muscle and liver) as traditionally done and also in intact erythrocytes. In all four tissues, mitochondrial ROS production was similar in quails and parrots and showed no correspondence with known longevity differences. The lack of a consistent difference between quails and parrots was not due to differences in mitochondrial content as ROS production in relation to oxygen consumption (determined as the free radical leak) showed a similar pattern. These findings cast doubt on the robustness of the oxidative stress theory of aging.

  12. Organization of the human mitochondrial hydrogen sulfide oxidation pathway.

    PubMed

    Libiad, Marouane; Yadav, Pramod Kumar; Vitvitsky, Victor; Martinov, Michael; Banerjee, Ruma

    2014-11-07

    Sulfide oxidation is expected to play an important role in cellular switching between low steady-state intracellular hydrogen sulfide levels and the higher concentrations where the physiological effects are elicited. Yet despite its significance, fundamental questions regarding how the sulfide oxidation pathway is wired remain unanswered, and competing proposals exist that diverge at the very first step catalyzed by sulfide quinone oxidoreductase (SQR). We demonstrate that, in addition to sulfite, glutathione functions as a persulfide acceptor for human SQR and that rhodanese preferentially synthesizes rather than utilizes thiosulfate. The kinetic behavior of these enzymes provides compelling evidence for the flow of sulfide via SQR to glutathione persulfide, which is then partitioned to thiosulfate or sulfite. Kinetic simulations at physiologically relevant metabolite concentrations provide additional support for the organizational logic of the sulfide oxidation pathway in which glutathione persulfide is the first intermediate formed.

  13. Protective effect of S-allylcysteine on 3-nitropropionic acid-induced lipid peroxidation and mitochondrial dysfunction in rat brain synaptosomes.

    PubMed

    Pérez-De La Cruz, Verónica; González-Cortés, Carolina; Pedraza-Chaverrí, José; Maldonado, Perla D; Andrés-Martínez, Leticia; Santamaría, Abel

    2006-01-30

    3-Nitropropionic acid is a neurotoxin that irreversibly inhibits succinate dehydrogenase, a relevant enzyme constituting the complex II of the respiratory chain during mitochondrial electron transport. 3-Nitropropionic acid is known to produce oxidative/nitrosative stress and evokes an experimental model of Huntington's disease. In this work we evaluated the effects of the antioxidant compound and major organosulfur garlic derivative, S-allylcysteine, on lipid peroxidation and mitochondrial dysfunction induced by 3-nitropropionic acid in synaptosomal fractions from rat brain. 3-Nitropropionic acid, at concentrations ranging 0.75-2.5 mM, produced enhanced levels of lipid peroxidation, while increasing concentrations of S-allylcysteine (0.1-2 mM) decreased the peroxidative action of 3-nitropropionic acid (1 mM) in synaptosomal fractions in a concentration-dependent manner. S-Allylcysteine (0.75 mM) also prevented the 3-nitropropionic acid (1mM)-induced mitochondrial dysfunction. These findings suggest that the protective actions that S-allylcysteine exert on the in vitro neurotoxicity induced by 3-nitropropionic acid are mediated by its antioxidant properties.

  14. Uncoupling, metabolic inhibition and induction of mitochondrial permeability transition in rat liver mitochondria caused by the major long-chain hydroxyl monocarboxylic fatty acids accumulating in LCHAD deficiency.

    PubMed

    Hickmann, Fernanda Hermes; Cecatto, Cristiane; Kleemann, Daniele; Monteiro, Wagner Oliveira; Castilho, Roger Frigério; Amaral, Alexandre Umpierrez; Wajner, Moacir

    2015-01-01

    Patients with long-chain 3-hydroxy-acyl-CoA dehydrogenase (LCHAD) deficiency commonly present liver dysfunction whose pathogenesis is unknown. We studied the effects of long-chain 3-hydroxylated fatty acids (LCHFA) that accumulate in LCHAD deficiency on liver bioenergetics using mitochondrial preparations from young rats. We provide strong evidence that 3-hydroxytetradecanoic (3HTA) and 3-hydroxypalmitic (3HPA) acids, the monocarboxylic acids that are found at the highest tissue concentrations in this disorder, act as metabolic inhibitors and uncouplers of oxidative phosphorylation. These conclusions are based on the findings that these fatty acids decreased ADP-stimulated (state 3) and uncoupled respiration, mitochondrial membrane potential and NAD(P)H content, and, in contrast, increased resting (state 4) respiration. We also verified that 3HTA and 3HPA markedly reduced Ca2+ retention capacity and induced swelling in Ca2+-loaded mitochondria. These effects were mediated by mitochondrial permeability transition (MPT) induction since they were totally prevented by the classical MPT inhibitors cyclosporin A and ADP, as well as by ruthenium red, a Ca2+ uptake blocker. Taken together, our data demonstrate that the major monocarboxylic LCHFA accumulating in LCHAD deficiency disrupt energy mitochondrial homeostasis in the liver. It is proposed that this pathomechanism may explain at least in part the hepatic alterations characteristic of the affected patients.

  15. Nitrite-nitric oxide control of mitochondrial respiration at the frontier of anoxia.

    PubMed

    Benamar, Abdelilah; Rolletschek, Hardy; Borisjuk, Ljudmilla; Avelange-Macherel, Marie-Hélène; Curien, Gilles; Mostefai, H Ahmed; Andriantsitohaina, Ramaroson; Macherel, David

    2008-10-01

    Actively respiring animal and plant tissues experience hypoxia because of mitochondrial O(2) consumption. Controlling oxygen balance is a critical issue that involves in mammals hypoxia-inducible factor (HIF) mediated transcriptional regulation, cytochrome oxidase (COX) subunit adjustment and nitric oxide (NO) as a mediator in vasodilatation and oxygen homeostasis. In plants, NO, mainly derived from nitrite, is also an important signalling molecule. We describe here a mechanism by which mitochondrial respiration is adjusted to prevent a tissue to reach anoxia. During pea seed germination, the internal atmosphere was strongly hypoxic due to very active mitochondrial respiration. There was no sign of fermentation, suggesting a down-regulation of O(2) consumption near anoxia. Mitochondria were found to finely regulate their surrounding O(2) level through a nitrite-dependent NO production, which was ascertained using electron paramagnetic resonance (EPR) spin trapping of NO within membranes. At low O(2), nitrite is reduced into NO, likely at complex III, and in turn reversibly inhibits COX, provoking a rise to a higher steady state level of oxygen. Since NO can be re-oxidized into nitrite chemically or by COX, a nitrite-NO pool is maintained, preventing mitochondrial anoxia. Such an evolutionarily conserved mechanism should have an important role for oxygen homeostasis in tissues undergoing hypoxia.

  16. The multiple acyl-coenzyme A dehydrogenation disorders, glutaric aciduria type II and ethylmalonic-adipic aciduria. Mitochondrial fatty acid oxidation, acyl-coenzyme A dehydrogenase, and electron transfer flavoprotein activities in fibroblasts.

    PubMed Central

    Amendt, B A; Rhead, W J

    1986-01-01

    The multiple acyl-coenzyme A (CoA) dehydrogenation disorders (MAD) include severe (S) and mild (M) variants, glutaric aciduria type II (MAD:S) and ethylmalonic-adipic aciduria (MAD:M). Intact MAD:M mitochondria oxidized [1-14C]octanoate, [1-14C]palmityl-CoA, and [1,5-14C]glutarate at 20-46% of control levels; MAD:S mitochondria oxidized these three substrates at 0.4-18% of control levels. In MAD:M mitochondria, acyl-CoA dehydrogenase (ADH) activities were similar to control, whereas MAD:S ADH activities ranged from 38% to 73% of control. Electron transfer flavoprotein (ETF) activities in five MAD:M cell lines ranged from 29 to 51% of control (P less than 0.01); ETF deficiency was the primary enzymatic defect in two MAD:M lines. In four MAD:S patients, ETF activities ranged from 3% to 6% of control (P less than 0.001); flavin adenine dinucleotide addition increased residual ETF activity from 4% to 21% of control in a single MAD:S line (P less than 0.01). Three MAD:S patients had ETF activities ranging from 33 to 53% of control; other investigators found deficient ETF-dehydrogenase activity in these MAD:S and three of our MAD:M cell lines. PMID:3722376

  17. Protection against oxidant-induced apoptosis by mitochondrial thioredoxin in SH-SY5Y neuroblastoma cells

    SciTech Connect

    Chen Yan; Yu Min; Jones, Dean P.; Greenamyre, J. Timothy; Cai Jiyang . E-mail: jiyang.cai@vanderbilt.edu

    2006-10-15

    Mitochondrial oxidative stress plays important roles in aging and age-related degenerative disorders. The newly identified mitochondrial thioredoxin (mtTrx; Trx2) is a key component of the mitochondrial antioxidant system which is responsible for the clearance of reactive intermediates and repairs proteins with oxidative damage. Here, we show that in cultured SH-SY5Y human neuroblastoma 1cells, overexpression of mtTrx inhibited apoptosis and loss of mitochondrial membrane potential induced by a chemical oxidant, tert-butylhydroperoxide (tBH). The effects of calcium ionophore (Br-A23187) were not affected by mtTrx, suggesting the protection was specific against oxidative injury. The mitochondrial glutathione pool was oxidized by tBH, and this oxidation was not inhibited by increased mtTrx. Consequently, the antioxidant function of mtTrx is not redundant, but rather in addition, to that of GSH. Mutations of Cys90 and Cys93 to serines rendered mtTrx ineffective in protection against tBH-induced cytoxicity. These data indicate that mtTrx controls the mitochondrial redox status independently of GSH and is a key component of the defensive mechanism against oxidative stress in cultured neuronal cells.

  18. [Malate oxidation by mitochondrial succinate:ubiquinone-reductase].

    PubMed

    Belikova, Iu O; Kotliar, A B

    1988-04-01

    Succinate:ubiquinone reductase was shown to catalyze the oxidation of L- and D-stereoisomers of malate by artificial electron acceptors and ubiquinone. The rate of malate oxidation by succinate:ubiquinone reductase is by two orders of magnitude lower than that for the natural substrate--succinate. The values of kinetic constants for the oxidation of D- and L-stereoisomers of malate are equal to: V infinity = 0.1 mumol/min/mg protein, Km = 2 mM and V infinity = 0.05 mumol/min/mg protein, Km = 2 mM, respectively. The malate dehydrogenase activity is fully inhibited by the inhibitors of the dicarboxylate-binding site of the enzyme, i.e., N-ethylmaleimide and malonate and is practically insensitive to carboxin, a specific inhibitor of the ubiquinone-binding center. The enol form of oxaloacetate was shown to be the product of malate oxidation by succinate:ubiquinone reductase. The kinetics of inhibition of the enzyme activity by the ketone and enol forms of oxaloacetate was studied. Both forms of oxaloacetate effectively inhibit the succinate:ubiquinone reductase reaction.

  19. Animation Model to Conceptualize ATP Generation: A Mitochondrial Oxidative Phosphorylation

    ERIC Educational Resources Information Center

    Jena, Ananta Kumar

    2015-01-01

    Adenosine triphosphate (ATP) is the molecular unit of intracellular energy and it is the product of oxidative phosphorylation of cellular respiration uses in cellular processes. The study explores the growth of the misconception levels amongst the learners and evaluates the effectiveness of animation model over traditional methods. The data…

  20. Disturbance of mitochondrial functions provoked by the major long-chain 3-hydroxylated fatty acids accumulating in MTP and LCHAD deficiencies in skeletal muscle.

    PubMed

    Cecatto, Cristiane; Godoy, Kálita Dos Santos; da Silva, Janaína Camacho; Amaral, Alexandre Umpierrez; Wajner, Moacir

    2016-10-01

    The pathogenesis of the muscular symptoms and recurrent rhabdomyolysis that are commonly manifested in patients with mitochondrial trifunctional protein (MTP) and long-chain 3-hydroxy-acyl-CoA dehydrogenase (LCHAD) deficiencies is still unknown. In this study we investigated the effects of the major long-chain monocarboxylic 3-hydroxylated fatty acids (LCHFA) accumulating in these disorders, namely 3-hydroxytetradecanoic (3HTA) and 3-hydroxypalmitic (3HPA) acids, on important mitochondrial functions in rat skeletal muscle mitochondria. 3HTA and 3HPA markedly increased resting (state 4) and decreased ADP-stimulated (state 3) and CCCP-stimulated (uncoupled) respiration. 3HPA provoked similar effects in permeabilized skeletal muscle fibers, validating the results obtained in purified mitochondria. Furthermore, 3HTA and 3HPA markedly diminished mitochondrial membrane potential, NAD(P)H content and Ca(2+) retention capacity in Ca(2+)-loaded mitochondria. Mitochondrial permeability transition (mPT) induction probably underlie these effects since they were totally prevented by cyclosporin A and ADP. In contrast, the dicarboxylic analogue of 3HTA did not alter the tested parameters. Our data strongly indicate that 3HTA and 3HPA behave as metabolic inhibitors, uncouplers of oxidative phosphorylation and mPT inducers in skeletal muscle. It is proposed that these pathomechanisms disrupting mitochondrial homeostasis may be involved in the muscle alterations characteristic of MTP and LCHAD deficiencies.

  1. The uncoupling agent 2,4-dinitrophenol improves mitochondrial homeostasis following striatal quinolinic acid injections.

    PubMed

    Korde, Amit S; Sullivan, Patrick G; Maragos, William F

    2005-10-01

    It is now generally accepted that excitotoxic cell death involves bioenergetic failure resulting from the cycling of Ca2+ and the generation of reactive oxygen species (ROS) by mitochondria. Both Ca2+ cycling and ROS formation by mitochondria are dependent on the mitochondrial membrane potential (Deltapsi(m)) that results from the proton gradient that is generated across the inner membrane. Mitochondrial uncoupling refers to a condition in which protons cross the inner membrane back into the matrix while bypassing the ATP synthase. As a consequence of this "short-circuit," there is a reduction in Deltapsi(m). We have previously demonstrated that animals treated with the classic uncoupling agent 2,4-dinitrophenol (DNP) show significant protection against brain damage following striatal injections of the NMDA agonist quinolinic acid (QA). In an effort to elucidate the mechanism of neuroprotection, we have assessed the effects of DNP on several parameters of mitochondrial function caused by QA. The results presented herein demonstrate that treatment with DNP attenuates QA-induced increases in mitochondrial Ca2+ levels and ROS formation and also improves mitochondrial respiration. Our findings indicate that DNP may confer protection against acute brain injury involving excitotoxic pathways by mechanisms that maintain mitochondrial function.

  2. Amino Acid Starvation Has Opposite Effects on Mitochondrial and Cytosolic Protein Synthesis

    PubMed Central

    Pearce, Sarah F.; Rorbach, Joanna; He, Jiuya; Brea-Calvo, Gloria; Minczuk, Michal; Reyes, Aurelio; Holt, Ian J.; Spinazzola, Antonella

    2014-01-01

    Amino acids are essential for cell growth and proliferation for they can serve as precursors of protein synthesis, be remodelled for nucleotide and fat biosynthesis, or be burnt as fuel. Mitochondria are energy producing organelles that additionally play a central role in amino acid homeostasis. One might expect mitochondrial metabolism to be geared towards the production and preservation of amino acids when cells are deprived of an exogenous supply. On the contrary, we find that human cells respond to amino acid starvation by upregulating the amino acid-consuming processes of respiration, protein synthesis, and amino acid catabolism in the mitochondria. The increased utilization of these nutrients in the organelle is not driven primarily by energy demand, as it occurs when glucose is plentiful. Instead it is proposed that the changes in the mitochondrial metabolism complement the repression of cytosolic protein synthesis to restrict cell growth and proliferation when amino acids are limiting. Therefore, stimulating mitochondrial function might offer a means of inhibiting nutrient-demanding anabolism that drives cellular proliferation. PMID:24718614

  3. Synergism of antifungal activity between mitochondrial respiration inhibitors and kojic acid

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Co-application of certain types of compounds with conventional antimicrobial drugs results in the enhancement of efficacy of drugs through a mechanism termed chemosensitization. We show that kojic acid (KA), a natural product, is a potent chemosensitizer to complex III inhibitors of mitochondrial re...

  4. Role of mitochondrial permeability transition in human renal tubular epithelial cell death induced by aristolochic acid

    SciTech Connect

    Qi Xinming; Cai Yan; Gong Likun; Liu Linlin; Chen Fangping; Xiao Ying; Wu Xiongfei; Li Yan; Xue Xiang |; Ren Jin . E-mail: cdser_simm@mail.shcnc.ac.cn

    2007-07-01

    Aristolochic acid (AA), a natural nephrotoxin and carcinogen, can induce a progressive tubulointerstitial nephropathy. However, the mechanism by which AA causes renal injury remains largely unknown. Here we reported that the mitochondrial permeability transition (MPT) plays an important role in the renal injury induced by aristolochic acid I (AAI). We found that in the presence of Ca{sup 2+}, AAI caused mitochondrial swelling, leakage of Ca{sup 2+}, membrane depolarization, and release of cytochrome c in isolated kidney mitochondria. These alterations were suppressed by cyclosporin A (CsA), an agent known to inhibit MPT. Culture of HK-2 cell, a human renal tubular epithelial cell line for 24 h with AAI caused a decrease in cellular ATP, mitochondrial membrane depolarization, cytochrome c release, and increase of caspase 3 activity. These toxic effects of AAI were attenuated by CsA and bongkrekic acid (BA), another specific MPT inhibitor. Furthermore, AAI greatly inhibited the activity of mitochondrial adenine nucleotide translocator (ANT) in isolated mitochondria. We suggested that ANT may mediate, at least in part, the AAI-induced MPT. Taken together, these results suggested that MPT plays a critical role in the pathogenesis of HK-2 cell injury induced by AAI and implied that MPT might contribute to human nephrotoxicity of aristolochic acid.

  5. Reference electrode for strong oxidizing acid solutions

    DOEpatents

    Rigdon, Lester P.; Harrar, Jackson E.; Bullock, Sr., Jack C.; McGuire, Raymond R.

    1990-01-01

    A reference electrode for the measurement of the oxidation-reduction potentials of solutions is especially suitable for oxidizing solutions such as highly concentrated and fuming nitric acids, the solutions of nitrogen oxides, N.sub.2 O.sub.4 and N.sub.2 O.sub.5, in nitric acids. The reference electrode is fabricated of entirely inert materials, has a half cell of Pt/Ce(IV)/Ce(III)/70 wt. % HNO.sub.3, and includes a double-junction design with an intermediate solution of 70 wt. % HNO.sub.3. The liquid junctions are made from Corning No. 7930 glass for low resistance and negligible solution leakage.

  6. Excitotoxic mitochondrial depolarisation requires both calcium and nitric oxide in rat hippocampal neurons

    PubMed Central

    Keelan, Julie; Vergun, Olga; Duchen, Michael R

    1999-01-01

    Glutamate neurotoxicity has been attributed to cellular Ca2+ overload. As mitochondrial depolarisation may represent a pivotal step in the progression to cell death, we have used digital imaging techniques to examine the relationship between cytosolic Ca2+ concentration ([Ca2+]c) and mitochondrial potential (ΔΨm) during glutamate toxicity, and to define the mechanisms underlying mitochondrial dysfunction. In cells of > 11 days in vitro (DIV), exposure to 50 mM potassium or 100 μM glutamate had different consequences for ΔΨm. KCl caused a small transient loss of ΔΨm but in response to glutamate there was a profound loss of ΔΨm. In cells of 7–10 DIV, glutamate caused only a modest and reversible drop in ΔΨm. Using fura-2 to measure [Ca2+]c, responses to KCl and glutamate did not appear significantly different. However, use of the low affinity indicator fura-2FF revealed a difference in the [Ca2+]c responses to KCl and glutamate, which clearly correlated with the loss of ΔΨm. Neurons exhibiting a profound mitochondrial depolarisation also showed a large secondary increase in the fura-2FF ratio. The glutamate-induced loss of ΔΨm was dependent on Ca2+ influx. However, inhibition of nitric oxide synthase (NOS) by L-NAME significantly attenuated the loss of ΔΨm. Furthermore, photolysis of caged NO at levels that had no effect alone promoted a profound mitochondrial depolarisation when combined with high [Ca2+]c, either in response to KCl or to glutamate in cultures at 7–10 DIV. In cells that showed only modest mitochondrial responses to glutamate, induction of a mitochondrial depolarisation by the addition of NO was followed by a secondary rise in [Ca2+]c. These data suggest that [Ca2+]c and nitric oxide act synergistically to cause mitochondrial dysfunction and impaired [Ca2+]c homeostasis during glutamate toxicity. PMID:10545145

  7. Exposure to 1800 MHz radiofrequency radiation induces oxidative damage to mitochondrial DNA in primary cultured neurons.

    PubMed

    Xu, Shangcheng; Zhou, Zhou; Zhang, Lei; Yu, Zhengping; Zhang, Wei; Wang, Yuan; Wang, Xubu; Li, Maoquan; Chen, Yang; Chen, Chunhai; He, Mindi; Zhang, Guangbin; Zhong, Min

    2010-01-22

    Increasing evidence indicates that oxidative stress may be involved in the adverse effects of radiofrequency (RF) radiation on the brain. Because mitochondrial DNA (mtDNA) defects are closely associated with various nervous system diseases and mtDNA is particularly susceptible to oxidative stress, the purpose of this study was to determine whether radiofrequency radiation can cause oxidative damage to mtDNA. In this study, we exposed primary cultured cortical neurons to pulsed RF electromagnetic fields at a frequency of 1800 MHz modulated by 217 Hz at an average special absorption rate (SAR) of 2 W/kg. At 24 h after exposure, we found that RF radiation induced a significant increase in the levels of 8-hydroxyguanine (8-OHdG), a common biomarker of DNA oxidative damage, in the mitochondria of neurons. Concomitant with this finding, the copy number of mtDNA and the levels of mitochondrial RNA (mtRNA) transcripts showed an obvious reduction after RF exposure. Each of these mtDNA disturbances could be reversed by pretreatment with melatonin, which is known to be an efficient antioxidant in the brain. Together, these results suggested that 1800 MHz RF radiation could cause oxidative damage to mtDNA in primary cultured neurons. Oxidative damage to mtDNA may account for the neurotoxicity of RF radiation in the brain.

  8. Altered Skeletal Muscle Mitochondrial Proteome As the Basis of Disruption of Mitochondrial Function in Diabetic Mice

    PubMed Central

    Zabielski, Piotr; Lanza, Ian R.; Gopala, Srinivas; Holtz Heppelmann, Carrie J.; Bergen, H. Robert; Dasari, Surendra

    2016-01-01

    Insulin plays pivotal role in cellular fuel metabolism in skeletal muscle. Despite being the primary site of energy metabolism, the underlying mechanism on how insulin deficiency deranges skeletal muscle mitochondrial physiology remains to be fully understood. Here we report an important link between altered skeletal muscle proteome homeostasis and mitochondrial physiology during insulin deficiency. Deprivation of insulin in streptozotocin-induced diabetic mice decreased mitochondrial ATP production, reduced coupling and phosphorylation efficiency, and increased oxidant emission in skeletal muscle. Proteomic survey revealed that the mitochondrial derangements during insulin deficiency were related to increased mitochondrial protein degradation and decreased protein synthesis, resulting in reduced abundance of proteins involved in mitochondrial respiration and β-oxidation. However, a paradoxical upregulation of proteins involved in cellular uptake of fatty acids triggered an accumulation of incomplete fatty acid oxidation products in skeletal muscle. These data implicate a mismatch of β-oxidation and fatty acid uptake as a mechanism leading to increased oxidative stress in diabetes. This notion was supported by elevated oxidative stress in cultured myotubes exposed to palmitate in the presence of a β-oxidation inhibitor. Together, these results indicate that insulin deficiency alters the balance of proteins involved in fatty acid transport and oxidation in skeletal muscle, leading to impaired mitochondrial function and increased oxidative stress. PMID:26718503

  9. The mitochondrial uncoupling agent 2,4-dinitrophenol improves mitochondrial function, attenuates oxidative damage, and increases white matter sparing in the contused spinal cord.

    PubMed

    Jin, Ying; McEwen, Melanie L; Nottingham, Stephanie A; Maragos, William F; Dragicevic, Natasha B; Sullivan, Patrick G; Springer, Joe E

    2004-10-01

    The purpose of this study was to investigate the potential neuroprotective efficacy of the mitochondrial uncoupler 2,4-dinitrophenol (DNP) in rats following a mild to moderate spinal cord contusion injury. Animals received intraperitoneal injections of vehicle (DMSO) or 5 mg/mL of DNP prior to injury. Twenty-four hours following surgery, mitochondrial function was assessed in mitochondria isolated from spinal cord synaptosomes. In addition, synaptosomes were used to measure indicators of reactive oxygen species formation, lipid peroxidation, and protein oxidation. Relative to vehicle-treated animals, pretreatment with DNP maintained mitochondrial bioenergetics and significantly decreased reactive oxygen species levels, lipid peroxidation, and protein carbonyl content following spinal cord injury. Furthermore, pretreatment with DNP significantly increased the amount of remaining white matter at the injury epicenter 6 weeks after injury. These results indicate that treatment with mitochondrial uncoupling agents may provide a novel approach for the treatment of secondary injury following spinal cord contusion.

  10. Mitochondrial proteomics of the acetic acid - induced programmed cell death response in a highly tolerant Zygosaccharomyces bailii - derived hybrid strain

    PubMed Central

    Guerreiro, Joana F.; Sampaio-Marques, Belém; Soares, Renata; Coelho, Ana V.; Leão, Cecília; Ludovico, Paula; Sá-Correia, Isabel

    2016-01-01

    Very high concentrations of acetic acid at low pH induce programmed cell death (PCD) in both the experimental model Saccharomyces cerevisiae and in Zygosaccharomyces bailii, the latter being considered the most problematic acidic food spoilage yeast due to its remarkable intrinsic resistance to this food preservative. However, while the mechanisms underlying S. cerevisiae PCD induced by acetic acid have been previously examined, the corresponding molecular players remain largely unknown in Z. bailii. Also, the reason why acetic acid concentrations known to be necrotic for S. cerevisiae induce PCD with an apoptotic phenotype in Z. bailii remains to be elucidated. In this study, a 2-DE-based expression mitochondrial proteomic analysis was explored to obtain new insights into the mechanisms involved in PCD in the Z. bailii derived hybrid strain ISA1307. This allowed the quantitative assessment of expression of protein species derived from each of the parental strains, with special emphasis on the processes taking place in the mitochondria known to play a key role in acetic acid - induced PCD. A marked decrease in the content of proteins involved in mitochondrial metabolism, in particular, in respiratory metabolism (Cor1, Rip1, Lpd1, Lat1 and Pdb1), with a concomitant increase in the abundance of proteins involved in fermentation (Pdc1, Ald4, Dld3) was registered. Other differentially expressed identified proteins also suggest the involvement of the oxidative stress response, protein translation, amino acid and nucleotide metabolism, among other processes, in the PCD response. Overall, the results strengthen the emerging concept of the importance of metabolic regulation of yeast PCD. PMID:28357336

  11. Mitochondrial oxidative stress-induced hepatocyte apoptosis reflects increased molybdenum intake in caprine.

    PubMed

    Zhuang, Yu; Liu, Ping; Wang, Liqi; Luo, Junrong; Zhang, Caiying; Guo, Xiaoquan; Hu, Guoliang; Cao, Huabin

    2016-03-01

    Molybdenum (Mo) is an essential trace element for animals and humans. However, the high dietary intake of Mo leads to disease conditions in heavy metal pollution areas. To the best of our knowledge, the effect of high levels of Mo on the apoptosis of hepatocyte in goats has not been investigated. Therefore, the aim of the present in vivo study was to investigate the impact of Mo on mitochondrial oxidative stress and apoptosis genes in the liver using real-time quantitative polymerase chain reaction (RT-qPCR) and transmission electron microscopy, respectively. Thirty-six healthy goats were randomly divided into three groups: two groups treated with ammonium molybdate [(NH4)6·Mo7O24·H2O] at 15 and 45 mg Mo kg(-1) BW, respectively, and a control group without treatment. Liver samples were collected from individual goats at different time intervals. The levels of oxidative stress in the mitochondrial membrane and expression of liver-related apoptosis genes, including Bcl-2, Cyt c, caspase-3, and Smac, were examined. The results demonstrated that the levels of superoxide dismutase (SOD) and catalase (CAT) expression were significantly down-regulated in liver cells, whereas malondialdehyde (MDA), nitric oxide (NO), and total nitric oxide synthase (T-NOS) expression was up-regulated (P < 0.01). The expression of Smac, Cyt c, and caspase-3 was significantly up-regulated, whereas Bcl-2 expression was down-regulated in liver cells (P < 0.01). In addition, histopathological examination revealed varying degrees of vacuolization, irregularity, nuclear fission, and mitochondrial swelling and high-density electrons in the cytoplasm of hepatocytes in groups treated with 15 and 45 mg Mo kg(-1) BW. Thus, these results suggested that high molybdenum induced hepatocyte apoptosis and might involve a mitochondrial pathway.

  12. Cisplatin upregulates mitochondrial nitric oxide synthase and peroxynitrite formation to promote renal injury.

    PubMed

    Jung, Michaela; Hotter, Georgina; Viñas, Jose Luis; Sola, Anna

    2009-01-15

    The mitochondria are a critical target for cisplatin-associated nephrotoxicity. Though nitric oxide formation has been implicated in the toxicity of cisplatin, this formation has not so far been related to a possible activation of mitochondrial nitric oxide synthase (mNOS). We show here that the upregulation of oxide mNOS and peroxynitrite formation in cisplatin treatment are key events that influence the development of the harmful parameters described in cisplatin-associated kidney failure. We confirm this by isolating the mitochondrial fraction of the kidney and across different access routes such as the use of a specific inhibitor of neuronal NOS, L-NPA, a peroxynitrite scavenger, FeTMPyP, and a peroxynitrite donor, SIN-1. The in vitro studies corroborated the information obtained in the in vivo experiments. The administration of cisplatin reveals a clear upregulation in the transcription of neuronal NOS and an increase in the levels of nitrites in the mitochondrial fractions of the kidneys. The upregulated transcription directly affects the cytoskeleton structure and the apoptosis. The inhibition of neuronal NOS reduces the levels of nitrites, cell death, and cytoskeleton derangement. Peroxynitrite is involved in the mechanism promoting the NOS transcription. In addition, in controls SIN-1 imitates the effects of cisplatin. In summary, we demonstrate that upregulation of mNOS in cisplatin treatment is a key component in both the initiation and the spread of cisplatin-associated damage in the kidney. Furthermore, peroxynitrite formation is directly involved in this process.

  13. Cisplatin upregulates mitochondrial nitric oxide synthase and peroxynitrite formation to promote renal injury

    SciTech Connect

    Jung, Michaela; Sola, Anna

    2009-01-15

    The mitochondria are a critical target for cisplatin-associated nephrotoxicity. Though nitric oxide formation has been implicated in the toxicity of cisplatin, this formation has not so far been related to a possible activation of mitochondrial nitric oxide synthase (mNOS). We show here that the upregulation of oxide mNOS and peroxynitrite formation in cisplatin treatment are key events that influence the development of the harmful parameters described in cisplatin-associated kidney failure. We confirm this by isolating the mitochondrial fraction of the kidney and across different access routes such as the use of a specific inhibitor of neuronal NOS, L-NPA, a peroxynitrite scavenger, FeTMPyP, and a peroxynitrite donor, SIN-1. The in vitro studies corroborated the information obtained in the in vivo experiments. The administration of cisplatin reveals a clear upregulation in the transcription of neuronal NOS and an increase in the levels of nitrites in the mitochondrial fractions of the kidneys. The upregulated transcription directly affects the cytoskeleton structure and the apoptosis. The inhibition of neuronal NOS reduces the levels of nitrites, cell death, and cytoskeleton derangement. Peroxynitrite is involved in the mechanism promoting the NOS transcription. In addition, in controls SIN-1 imitates the effects of cisplatin. In summary, we demonstrate that upregulation of mNOS in cisplatin treatment is a key component in both the initiation and the spread of cisplatin-associated damage in the kidney. Furthermore, peroxynitrite formation is directly involved in this process.

  14. Infertility and recurrent miscarriage with complex II deficiency-dependent mitochondrial oxidative stress in animal models.

    PubMed

    Ishii, Takamasa; Yasuda, Kayo; Miyazawa, Masaki; Mitsushita, Junji; Johnson, Thomas E; Hartman, Phil S; Ishii, Naoaki

    2016-04-01

    Oxidative stress is associated with some forms of both male and female infertility. However, there is insufficient knowledge of the influence of oxidative stress on the maintenance of a viable pregnancy, including pregnancy complications and fetal development. There are a number of animal models for understanding age-dependent decrease of reproductive ability and diabetic embryopathy, especially abnormal spermatogenesis, oogenesis and embryogenesis with mitochondrial dysfunctions. Several important processes occur in mitochondria, including ATP synthesis, calcium ion storage, induction of apoptosis and production of reactive oxygen species (ROS). These events have different effects on the several aspects of reproductive function. Tet-mev-1 conditional transgenic mice, developed after studies with the mev-1 mutant of the nematode C. elegans, offer the ability to carefully regulate expression of doxycycline-induced mutated SDHC(V69E) levels and hence modulate endogenous oxidative stress. The mev-1 models have served to illuminate the effects of complex II deficiency-dependent mitochondrial ROS production, although interestingly they maintain normal mitochondrial and intracellular ATP levels. In this review, the reproductive dysfunctions are presented focusing on fertility potentials in each gamete, early embryogenesis, maternal conditions with placental function and neonatal development.

  15. Modulation of mitochondrial gene expression in pulmonary epithelial cells exposed to oxidants.

    PubMed Central

    Janssen, Y M; Driscoll, K E; Timblin, C R; Hassenbein, D; Mossman, B T

    1998-01-01

    Oxidants are important in the regulation of signal transduction and gene expression. Multiple classes of genes are transcriptionally activated by oxidants and are implicated in different phenotypic responses. In the present study, we performed differential mRNA display to elucidate genes that are induced or repressed after exposure of rat lung epithelial (RLE) cells to H2O2 or crocidolite asbestos, a pathogenic mineral that generates oxidants. After 8 or 24 hr of exposure, RNA was extracted, reverse transcribed, and amplified by polymerase chain reaction with degenerate primers to visualize alterations in gene expression. The seven clones obtained were sequenced and encoded the mitochondrial genes, NADH dehydrogenase subunits ND5 and ND6, and 16S ribosomal RNA. Evaluation of their expression by Northern blot analysis revealed increased expression of 16S rRNA after 1 or 2 hr of exposure to H2O2. At later time periods (4 and 24 hr), mRNA levels of 16S rRNA and NADH dehydrogenase were decreased in H2O2-treated RLE cells when compared to sham controls. Crocidolite asbestos caused increases in 16S rRNA levels after 8 hr of exposure, whereas after 24 hr of exposure to asbestos, 16S rRNA levels were decreased in comparison to sham controls. In addition to these oxidants, the nitric oxide generator spermine NONOate caused similar decreases in NADH dehydrogenase mRNA levels after 4 hr of exposure. The present data and previous studies demonstrated that all oxidants examined resulted in apoptosis in RLE cells during the time frame where alterations of mitochondrial gene expression were observed. As the mitochondrion is a major organelle that controls apoptosis, alterations in expression of mitochondrial genes may be involved in the regulation of apoptosis. Images Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 PMID:9788897

  16. Prediction of mitochondrial proteins of malaria parasite using split amino acid composition and PSSM profile.

    PubMed

    Verma, Ruchi; Varshney, Grish C; Raghava, G P S

    2010-06-01

    The rate of human death due to malaria is increasing day-by-day. Thus the malaria causing parasite Plasmodium falciparum (PF) remains the cause of concern. With the wealth of data now available, it is imperative to understand protein localization in order to gain deeper insight into their functional roles. In this manuscript, an attempt has been made to develop prediction method for the localization of mitochondrial proteins. In this study, we describe a method for predicting mitochondrial proteins of malaria parasite using machine-learning technique. All models were trained and tested on 175 proteins (40 mitochondrial and 135 non-mitochondrial proteins) and evaluated using five-fold cross validation. We developed a Support Vector Machine (SVM) model for predicting mitochondrial proteins of P. falciparum, using amino acids and dipeptides composition and achieved maximum MCC 0.38 and 0.51, respectively. In this study, split amino acid composition (SAAC) is used where composition of N-termini, C-termini, and rest of protein is computed separately. The performance of SVM model improved significantly from MCC 0.38 to 0.73 when SAAC instead of simple amino acid composition was used as input. In addition, SVM model has been developed using composition of PSSM profile with MCC 0.75 and accuracy 91.38%. We achieved maximum MCC 0.81 with accuracy 92% using a hybrid model, which combines PSSM profile and SAAC. When evaluated on an independent dataset our method performs better than existing methods. A web server PFMpred has been developed for predicting mitochondrial proteins of malaria parasites ( http://www.imtech.res.in/raghava/pfmpred/).

  17. Arachnid relationships based on mitochondrial genomes: asymmetric nucleotide and amino acid bias affects phylogenetic analyses.

    PubMed

    Masta, Susan E; Longhorn, Stuart J; Boore, Jeffrey L

    2009-01-01

    Phylogenetic analyses based on mitochondrial DNA have yielded widely differing relationships among members of the arthropod lineage Arachnida, depending on the nucleotide coding schemes and models of evolution used. We enhanced taxonomic coverage within the Arachnida greatly by sequencing seven new arachnid mitochondrial genomes from five orders. We then used all 13 mitochondrial protein-coding genes from these genomes to evaluate patterns of nucleotide and amino acid biases. Our data show that two of the six orders of arachnids (spiders and scorpions) have experienced shifts in both nucleotide and amino acid usage in all their protein-coding genes, and that these biases mislead phylogeny reconstruction. These biases are most striking for the hydrophobic amino acids isoleucine and valine, which appear to have evolved asymmetrical exchanges in response to shifts in nucleotide composition. To improve phylogenetic accuracy based on amino acid differences, we tested two recoding methods: (1) removing all isoleucine and valine sites and (2) recoding amino acids based on their physiochemical properties. We find that these methods yield phylogenetic trees that are consistent in their support of ancient intraordinal divergences within the major arachnid lineages. Further refinement of amino acid recoding methods may help us better delineate interordinal relationships among these diverse organisms.

  18. Effect of nitric oxide on mitochondrial respiratory activity of human articular chondrocytes

    PubMed Central

    Maneiro, E; Lopez-Armada, M; de Andres, M C; Carames, B; Martin, M; Bonilla, A; del Hoyo, P; Galdo, F; Arenas, J; Blanco, F

    2005-01-01

    Objective: To investigate the effect of nitric oxide (NO) on mitochondrial activity and its relation with the apoptosis of human articular chondrocytes. Materials and methods: Mitochondrial function was evaluated by analysing respiratory chain enzyme complexes, citrate synthase (CS) activities, and mitochondrial membrane potential (Δψm). The activities of the mitochondrial respiratory chain (MRC) complexes (complex I: NADH CoQ1 reductase, complex II: succinate dehydrogenase, complex III: ubiquinol cytochrome c reductase, complex IV: cytochrome c oxidase) and CS were measured in human articular chondrocytes isolated from normal cartilage. The Δψm was measured by 5,5',6,6'-tetracholoro-1,1',3,3'-tetraethylbenzimidazole carbocyanide iodide (JC-1) using flow cytometry. Apoptosis was analysed by flow cytometry. The mRNA expression of caspases was analysed by ribonuclease protection analysis and the detection of protein synthesis by western blotting. Sodium nitroprusside (SNP) was used as an NO compound donor. Results: SNP at concentrations higher than 0.5 mmol/l for 24 hours induced cellular changes characteristic of apoptosis. SNP elicited mRNA expression of caspase-3 and caspase-7 and down regulated bcl-2 synthesis in a dose and time dependent manner. Furthermore, 0.5 mM SNP induced depolarisation of the mitochondrial membrane at 5, 12, and 24 hours. Analysis of the MRC showed that at 5 hours, 0.5 mM SNP reduced the activity of complex IV by 33%. The individual inhibition of mitochondrial complex IV with azide modified the Δψm and induced apoptosis. Conclusions: This study suggests that the effect of NO on chondrocyte survival is mediated by its effect on complex IV of the MRC. PMID:15708893

  19. Prevention of oxidative stress, inflammation and mitochondrial dysfunction in the intestine by different cranberry phenolic fractions.

    PubMed

    Denis, Marie-Claude; Desjardins, Yves; Furtos, Alexandra; Marcil, Valérie; Dudonné, Stéphanie; Montoudis, Alain; Garofalo, Carole; Delvin, Edgard; Marette, André; Levy, Emile

    2015-02-01

    Cranberry fruit has been reported to have high antioxidant effectiveness that is potentially linked to its richness in diversified polyphenolic content. The aim of the present study was to determine the role of cranberry polyphenolic fractions in oxidative stress (OxS), inflammation and mitochondrial functions using intestinal Caco-2/15 cells. The combination of HPLC and UltraPerformance LC®-tandem quadrupole (UPLC-TQD) techniques allowed us to characterize the profile of low, medium and high molecular mass polyphenolic compounds in cranberry extracts. The medium molecular mass fraction was enriched with flavonoids and procyanidin dimers whereas procyanidin oligomers (DP > 4) were the dominant class of polyphenols in the high molecular mass fraction. Pre-incubation of Caco-2/15 cells with these cranberry extracts prevented iron/ascorbate-mediated lipid peroxidation and counteracted lipopolysaccharide-mediated inflammation as evidenced by the decrease in pro-inflammatory cytokines (TNF-α and interleukin-6), cyclo-oxygenase-2 and prostaglandin E2. Cranberry polyphenols (CP) fractions limited both nuclear factor κB activation and Nrf2 down-regulation. Consistently, cranberry procyanidins alleviated OxS-dependent mitochondrial dysfunctions as shown by the rise in ATP production and the up-regulation of Bcl-2, as well as the decline of protein expression of cytochrome c and apoptotic-inducing factor. These mitochondrial effects were associated with a significant stimulation of peroxisome-proliferator-activated receptor γ co-activator-1-α, a central inducing factor of mitochondrial biogenesis and transcriptional co-activator of numerous downstream mediators. Finally, cranberry procyanidins forestalled the effect of iron/ascorbate on the protein expression of mitochondrial transcription factors (mtTFA, mtTFB1, mtTFB2). Our findings provide evidence for the capacity of CP to reduce intestinal OxS and inflammation while improving mitochondrial dysfunction.

  20. Mulberry leaf phenolics ameliorate hyperglycemia-induced oxidative stress and stabilize mitochondrial membrane potential in HepG2 cells.

    PubMed

    Zou, Yu-Xiao; Shen, Wei-Zhi; Liao, Sen-Tai; Liu, Fan; Zheng, Shan-Qing; Blumberg, Jeffrey B; Chen, C-Y Oliver

    2014-12-01

    To investigate the effect of phenolics in mulberry leaves (mulberry leaf phenolics; MLP) on hyperglycemia-induced oxidative stress and mitochondrial membrane potential (ΔΨm) in HepG2 cells; we treated HepG2 with glucose [5.5 (N-Glc) or 50 mmol/L (Hi-Glc)] with or without MLP at 10 or 100 µmol/L gallic acid equivalents and assessed level of reactive oxidant species (ROS), ΔΨm, malondialdehyde (MDA) and nuclear factor-kappaB (NF-κB) activation. Hi-Glc-induced oxidative damage was demonstrated by a series of increase in superoxides (560%, 0.5 h), MDA (400%, 24 h), NF-κB activation (474%, 4 h) and a wild fluctuation of ΔΨm relative to the control cells (p ≤ 0.05). MLP treatments ameliorate Hi-Glc-induced negative effects by a 40% reduction in ROS production, 34-44% reduction in MDA production, over 35% inhibition of NF-κB activation, as well as exert protective effect on HepG2 cells from change in ΔΨm. Our data show that MLP in vitro can protect hepatoctyes from hyperglycemia-induced oxidative damages.

  1. Hyperactivation of oxidative mitochondrial metabolism in epithelial cancer cells in situ

    PubMed Central

    Whitaker-Menezes, Diana; Martinez-Outschoorn, Ubaldo E; Flomenberg, Neal; Birbe, Ruth C; Witkiewicz, Agnieszka K; Howell, Anthony; Pavlides, Stephanos; Tsirigos, Aristotelis; Ertel, Adam; Pestell, Richard G; Broda, Paolo; Minetti, Carlo

    2011-01-01

    We have recently proposed a new mechanism for explaining energy transfer in cancer metabolism. In this scenario, cancer cells behave as metabolic parasites, by extracting nutrients from normal host cells, such as fibroblasts, via the secretion of hydrogen peroxide as the initial trigger. Oxidative stress in the tumor microenvironment then leads to autophagy-driven catabolism, mitochondrial dys-function and aerobic glycolysis. This, in turn, produces high-energy nutrients (such as L-lactate, ketones and glutamine) that drive the anabolic growth of tumor cells, via oxidative mitochondrial metabolism. A logical prediction of this new “parasitic” cancer model is that tumor-associated fibroblasts should show evidence of mitochondrial dys-function (mitophagy and aerobic glycolysis). In contrast, epithelial cancer cells should increase their oxidative mitochondrial capacity. To further test this hypothesis, here we subjected frozen sections from human breast tumors to a staining procedure that only detects functional mitochondria. This method detects the in situ enzymatic activity of cytochrome C oxidase (COX), also known as Complex IV. Remarkably, cancer cells show an over-abundance of COX activity, while adjacent stromal cells remain essentially negative. Adjacent normal ductal epithelial cells also show little or no COX activity, relative to epithelial cancer cells. Thus, oxidative mitochondrial activity is selectively amplified in cancer cells. Although COX activity staining has never been applied to cancer tissues, it could now be used routinely to distinguish cancer cells from normal cells, and to establish negative margins during cancer surgery. Similar results were obtained with NADH activity staining, which measures Complex I activity, and succinate dehydrogenase (SDH) activity staining, which measures Complex II activity. COX and NADH activities were blocked by electron transport inhibitors, such as Metformin. This has mechanistic and clinical implications

  2. Salicylic acid induces mitochondrial injury by inhibiting ferrochelatase heme biosynthesis activity.

    PubMed

    Gupta, Vipul; Liu, Shujie; Ando, Hideki; Ishii, Ryohei; Tateno, Shumpei; Kaneko, Yuki; Yugami, Masato; Sakamoto, Satoshi; Yamaguchi, Yuki; Nureki, Osamu; Handa, Hiroshi

    2013-12-01

    Salicylic acid is a classic nonsteroidal anti-inflammatory drug. Although salicylic acid also induces mitochondrial injury, the mechanism of its antimitochondrial activity is not well understood. In this study, by using a one-step affinity purification scheme with salicylic acid-immobilized beads, ferrochelatase (FECH), a homodimeric enzyme involved in heme biosynthesis in mitochondria, was identified as a new molecular target of salicylic acid. Moreover, the cocrystal structure of the FECH-salicylic acid complex was determined. Structural and biochemical studies showed that salicylic acid binds to the dimer interface of FECH in two possible orientations and inhibits its enzymatic activity. Mutational analysis confirmed that Trp301 and Leu311, hydrophobic amino acid residues located at the dimer interface, are directly involved in salicylic acid binding. On a gel filtration column, salicylic acid caused a shift in the elution profile of FECH, indicating that its conformational change is induced by salicylic acid binding. In cultured human cells, salicylic acid treatment or FECH knockdown inhibited heme synthesis, whereas salicylic acid did not exert its inhibitory effect in FECH knockdown cells. Concordantly, salicylic acid treatment or FECH knockdown inhibited heme synthesis in zebrafish embryos. Strikingly, the salicylic acid-induced effect in zebrafish was partially rescued by FECH overexpression. Taken together, these findings illustrate that FECH is responsible for salicylic acid-induced inhibition of heme synthesis, which may contribute to its antimitochondrial and anti-inflammatory function. This study establishes a novel aspect of the complex pharmacological effects of salicylic acid.

  3. Mitochondrial dysfunction and oxidative stress in patients with chronic kidney disease.

    PubMed

    Gamboa, Jorge L; Billings, Frederic T; Bojanowski, Matthew T; Gilliam, Laura A; Yu, Chang; Roshanravan, Baback; Roberts, L Jackson; Himmelfarb, Jonathan; Ikizler, T Alp; Brown, Nancy J

    2016-05-01

    Mitochondria abnormalities in skeletal muscle may contribute to frailty and sarcopenia, commonly present in patients with chronic kidney disease (CKD). Dysfunctional mitochondria are also a major source of oxidative stress and may contribute to cardiovascular disease in CKD We tested the hypothesis that mitochondrial structure and function worsens with the severity of CKD Mitochondrial volume density, mitochondrial DNA (mtDNA) copy number, BNIP3, and PGC1α protein expression were evaluated in skeletal muscle biopsies obtained from 27 subjects (17 controls and 10 with CKD stage 5 on hemodialysis). We also measured mtDNA copy number in peripheral blood mononuclear cells (PBMCs), plasma isofurans, and plasma F2-isoprostanes in 208 subjects divided into three groups: non-CKD (eGFR>60 mL/min), CKD stage 3-4 (eGFR 60-15 mL/min), and CKD stage 5 (on hemodialysis). Muscle biopsies from patients with CKD stage 5 revealed lower mitochondrial volume density, lower mtDNA copy number, and higher BNIP3 content than controls. mtDNA copy number in PBMCs was decreased with increasing severity of CKD: non-CKD (6.48, 95% CI 4.49-8.46), CKD stage 3-4 (3.30, 95% CI 0.85-5.75, P = 0.048 vs. non-CKD), and CKD stage 5 (1.93, 95% CI 0.27-3.59, P = 0.001 vs. non-CKD). Isofurans were higher in patients with CKD stage 5 (median 59.21 pg/mL, IQR 41.76-95.36) compared to patients with non-CKD (median 49.95 pg/mL, IQR 27.88-83.46, P = 0.001), whereas F2-isoprostanes did not differ among groups. Severity of CKD is associated with mitochondrial dysfunction and markers of oxidative stress. Mitochondrial abnormalities, which are common in skeletal muscle from patients with CKD stage 5, may explain the muscle dysfunction associated with frailty and sarcopenia in CKD Further studies are required to evaluate mitochondrial function in vivo in patients with different CKD stages.

  4. Methoxychlor causes mitochondrial dysfunction and oxidative damage in the mouse ovary

    SciTech Connect

    Gupta, R.K.; Schuh, R.A.; Fiskum, G.; Flaws, J.A. . E-mail: jflaws@epi.umaryland.edu

    2006-11-01

    Methoxychlor (MXC) is an organochlorine pesticide that reduces fertility in female rodents by causing ovarian atrophy, persistent estrous cyclicity, and antral follicle atresia (apoptotic cell death). Oxidative damage resulting from reactive oxygen species (ROS) generation has been demonstrated to lead to toxicant-induced cell death. Thus, this work tested the hypothesis that MXC causes oxidative damage to the mouse ovary and affects mitochondrial respiration in a manner that stimulates ROS production. For the in vitro experiments, mitochondria were collected from adult cycling mouse ovaries, treated with vehicle (dimethyl sulfoxide; DMSO) or MXC, and subjected to polarographic measurements of respiration. For the in vivo experiments, adult cycling CD-1 mice were dosed with either vehicle (sesame oil) or MXC for 20 days. After treatment, ovarian mitochondria were isolated and subjected to measurements of respiration and fluorimetric measurements of H{sub 2}O{sub 2} production. Some ovaries were also fixed and processed for immunohistochemistry using antibodies for ROS production markers: nitrotyrosine and 8-hydroxy-2'-deoxyguanosine (8-OHG). Ovaries from in vivo experiments were also used to measure the mRNA expression and activity of antioxidants such as Cu/Zn superoxide dismutase (SOD1), glutathione peroxidase (GPX), and catalase (CAT). The results indicate that MXC significantly impairs mitochondrial respiration, increases production of H{sub 2}O{sub 2}, causes more staining for nitrotyrosine and 8-OHG in antral follicles, and decreases the expression and activity of SOD1, GPX, and CAT as compared to controls. Collectively, these data indicate that MXC inhibits mitochondrial respiration, causes ROS production, and decreases antioxidant expression and activity in the ovary, specifically in the antral follicles. Therefore, it is possible that MXC causes atresia of ovarian antral follicles by inducing oxidative stress through mitochondrial production of ROS.

  5. Application of Markov chain to the pattern of mitochondrial deoxyribonucleic acid mutations

    NASA Astrophysics Data System (ADS)

    Vantika, Sandy; Pasaribu, Udjianna S.

    2014-03-01

    This research explains how Markov chain used to model the pattern of deoxyribonucleic acid mutations in mitochondrial (mitochondrial DNA). First, sign test was used to see a pattern of nucleotide bases that will appear at one position after the position of mutated nucleotide base. Results obtained from the sign test showed that for most cases, there exist a pattern of mutation except in the mutation cases of adenine to cytosine, adenine to thymine, and cytosine to guanine. Markov chain analysis results on data of mutations that occur in mitochondrial DNA indicate that one and two positions after the position of mutated nucleotide bases tend to be occupied by particular nucleotide bases. From this analysis, it can be said that the adenine, cytosine, guanine and thymine will mutate if the nucelotide base at one and/or two positions after them is cytosine.

  6. Ethanol-induced oxidative stress precedes mitochondrially mediated apoptotic death of cultured fetal cortical neurons.

    PubMed

    Ramachandran, Vinitha; Watts, Lora Talley; Maffi, Shivani Kaushal; Chen, Juanjuan; Schenker, Steven; Henderson, George

    2003-11-15

    In utero ethanol exposure elicits apoptotic cell death in the fetal brain, and this may be mediated by oxidative stress. Our studies utilize cultured fetal rat cortical neurons and illustrate that ethanol elicits a rapid onset of oxidative stress, which culminates in mitochondrially mediated apoptotic cell death. Cells exposed to ethanol (2.5 mg/ml) remained attached to their polylysine matrix during a 24-hr exposure, but they exhibited distinct signs of oxidative stress, decreased viability, and apoptosis. Confocal microscopy of live cortical neurons pretreated with dichlorodihydrofluorescein diacetate demonstrated an increase in reactive oxygen species (ROS) within 5 min of ethanol exposure. The levels of ROS further increased by 58% within 1 hr (P <.05) and by 82% within 2 hr (P <.05), accompanied by increases of mitochondrial 4-hydroxynonenal (HNE). These early events were followed by decreased trypan blue exclusion of 10% to 32% (P <.05) at the 6- to 24-hr time points, respectively. This culminates in apoptotic death, with increases of Annexin V binding of 43%, 89%, 123%, and 238%, at 2, 6, 12, and 24 hr of ethanol treatment, respectively, as well as DNA fragmentation increases of 50% and 65% by 12 and 24 hr, respectively. Release of cytochrome c by mitochondria increased by 53% at 6 hr of exposure (P <.05), concomitant with activation of caspase 3 (52% at 12 hr, P <.05). Pretreatment with N-acetylcysteine increased cellular glutathione and prevented apoptosis. These studies provide a time line illustrating that oxidative stress and formation of a proapoptotic lipid peroxidation product, HNE, precede a cascade of mitochondrially mediated events in cultured fetal cortical neurons, culminating in apoptotic death. The prevention of apoptosis by augmentation of glutathione stores also strongly supports a role for oxidative stress in ethanol-mediated apoptotic death of fetal cortical neurons.

  7. The development of structure-activity relationships for mitochondrial dysfunction: uncoupling of oxidative phosphorylation.

    PubMed

    Naven, Russell T; Swiss, Rachel; Klug-McLeod, Jacquelyn; Will, Yvonne; Greene, Nigel

    2013-01-01

    Mitochondrial dysfunction has been implicated as an important factor in the development of idiosyncratic organ toxicity. An ability to predict mitochondrial dysfunction early in the drug development process enables the deselection of those drug candidates with potential safety liabilities, allowing resources to be focused on those compounds with the highest chance of success to the market. A database of greater than 2000 compounds was analyzed to identify structural and physicochemical features associated with the uncoupling of oxidative phosphorylation (herein defined as an increase in basal respiration). Many toxicophores associated with potent uncoupling activity were identified, and these could be divided into two main mechanistic classes, protonophores and redox cyclers. For the protonophores, potent uncoupling activity was often promoted by high lipophilicity and apparent stabilization of the anionic charge resulting from deprotonation of the protonophore. The potency of redox cyclers did not appear to be prone to variations in lipophilicity. Only 11 toxicophores were of sufficient predictive performance that they could be incorporated into a structural-alert model. Each alert was associated with one of three confidence levels (high, medium, and low) depending upon the lipophilicity-activity profile of the structural class. The final model identified over 68% of those compounds with potent uncoupling activity and with a value for specificity above 99%. We discuss the advantages and limitations of this approach and conclude that although structural alert methodology is useful for identifying toxicophores associated with mitochondrial dysfunction, they are not a replacement for the mitochondrial dysfunction assays in early screening paradigms.

  8. Mitochondrial dependent oxidative stress in cell culture induced by laser radiation at 1265 nm.

    PubMed

    Saenko, Yury V; Glushchenko, Eugenia S; Zolotovskii, Igor O; Sholokhov, Evgeny; Kurkov, Andrey

    2016-04-01

    Photodynamic therapy is the main technique applied for surface carcinoma treatment. This technique employs singlet oxygen generated via a laser excited photosensitizer as a main damaging agent. However, prolonged sensitivity to intensive light, relatively low tissue penetration by activating light the cost of photosensitizer (PS) administration can limit photodynamic therapy applications. Early was reported singlet oxygen generation without photosensitizer induced by a laser irradiation at the wavelength of 1250-1270 nm. Here, we study the dynamics of oxidative stress, DNA damage, changes of mitochondrial potential, and mitochondrial mass induced by a laser at 1265 nm have been studied in HCT-116 and CHO-K cells. Laser irradiation of HCT-116 and CHO-K cells has induced a dose-dependent cell death via increasing intracellular reactive oxygen species (ROS) concentration, increase of DNA damage, decrease of mitochondrial potential, and reduced glutathione. It has been shown that, along with singlet oxygen generation, the increase of the intracellular ROS concentration induced by mitochondrial damage contributes to the damaging effect of the laser irradiation at 1265 nm.

  9. Integrative Approaches for Studying Mitochondrial and Nuclear Genome Co-evolution in Oxidative Phosphorylation

    PubMed Central

    Sunnucks, Paul; Morales, Hernán E.; Lamb, Annika M.; Pavlova, Alexandra; Greening, Chris

    2017-01-01

    In animals, interactions among gene products of mitochondrial and nuclear genomes (mitonuclear interactions) are of profound fitness, evolutionary, and ecological significance. Most fundamentally, the oxidative phosphorylation (OXPHOS) complexes responsible for cellular bioenergetics are formed by the direct interactions of 13 mitochondrial-encoded and ∼80 nuclear-encoded protein subunits in most animals. It is expected that organisms will develop genomic architecture that facilitates co-adaptation of these mitonuclear interactions and enhances biochemical efficiency of OXPHOS complexes. In this perspective, we present principles and approaches to understanding the co-evolution of these interactions, with a novel focus on how genomic architecture might facilitate it. We advocate that recent interdisciplinary advances assist in the consolidation of links between genotype and phenotype. For example, advances in genomics allow us to unravel signatures of selection in mitochondrial and nuclear OXPHOS genes at population-relevant scales, while newly published complete atomic-resolution structures of the OXPHOS machinery enable more robust predictions of how these genes interact epistatically and co-evolutionarily. We use three case studies to show how integrative approaches have improved the understanding of mitonuclear interactions in OXPHOS, namely those driving high-altitude adaptation in bar-headed geese, allopatric population divergence in Tigriopus californicus copepods, and the genome architecture of nuclear genes coding for mitochondrial functions in the eastern yellow robin. PMID:28316610

  10. Stat3 promotes mitochondrial transcription and oxidative respiration during maintenance and induction of naive pluripotency.

    PubMed

    Carbognin, Elena; Betto, Riccardo M; Soriano, Maria E; Smith, Austin G; Martello, Graziano

    2016-03-15

    Transcription factor Stat3 directs self-renewal of pluripotent mouse embryonic stem (ES) cells downstream of the cytokine leukemia inhibitory factor (LIF). Stat3 upregulates pivotal transcription factors in the ES cell gene regulatory network to sustain naïve identity. Stat3 also contributes to the rapid proliferation of ES cells. Here, we show that Stat3 increases the expression of mitochondrial-encoded transcripts and enhances oxidative metabolism. Chromatin immunoprecipitation reveals that Stat3 binds to the mitochondrial genome, consistent with direct transcriptional regulation. An engineered form of Stat3 that localizes predominantly to mitochondria is sufficient to support enhanced proliferation of ES cells, but not to maintain their undifferentiated phenotype. Furthermore, during reprogramming from primed to naïve states of pluripotency, Stat3 similarly upregulates mitochondrial transcripts and facilitates metabolic resetting. These findings suggest that the potent stimulation of naïve pluripotency by LIF/Stat3 is attributable to parallel and synergistic induction of both mitochondrial respiration and nuclear transcription factors.

  11. Prolonged exposure to insulin induces mitochondrion-derived oxidative stress through increasing mitochondrial cholesterol content in hepatocytes.

    PubMed

    Mei, Shuang; Gu, Haihua; Yang, Xuefeng; Guo, Huailan; Liu, Zhenqi; Cao, Wenhong

    2012-05-01

    We addressed the link between excessive exposure to insulin and mitochondrion-derived oxidative stress in this study and found that prolonged exposure to insulin increased mitochondrial cholesterol in cultured hepatocytes and in mice and stimulated production of reactive oxygen species (ROS) and decreased the reduced glutathione to glutathione disulfide ratio in cultured hepatocytes. Exposure of isolated hepatic mitochondria to cholesterol alone promoted ROS emission. The oxidative stress induced by the prolonged exposure to insulin was prevented by inhibition of cholesterol synthesis with simvastatin. We further found that prolonged exposure to insulin decreased mitochondrial membrane potential and the increased ROS production came from mitochondrial respiration complex I. Finally, we observed that prolonged exposure to insulin decreased mitochondrial membrane fluidity in a cholesterol synthesis-dependent manner. Together our results demonstrate that excess exposure to insulin causes mitochondrion-derived oxidative stress through cholesterol synthesis in hepatocytes.

  12. Citrus Flavanones Affect Hepatic Fatty Acid Oxidation in Rats by Acting as Prooxidant Agents

    PubMed Central

    Constantin, Rodrigo Polimeni; do Nascimento, Gilson Soares; Constantin, Renato Polimeni; Salgueiro, Clairce Luzia; Bracht, Adelar; Ishii-Iwamoto, Emy Luiza; Yamamoto, Nair Seiko

    2013-01-01

    Citrus flavonoids have a wide range of biological activities and positive health effects on mammalian cells because of their antioxidant properties. However, they also act as prooxidants and thus may interfere with metabolic pathways. The purpose of this work was to evaluate the effects of three citrus flavanones, hesperidin, hesperetin, and naringenin, on several parameters linked to fatty acid oxidation in mitochondria, peroxisomes, and perfused livers of rats. When exogenous octanoate was used as substrate, hesperetin and naringenin reduced the mitochondrial NADH/NAD+ ratio and stimulated the citric acid cycle without significant changes on oxygen uptake or ketogenesis. When fatty acid oxidation from endogenous sources was evaluated, hesperetin and naringenin strongly reduced the mitochondrial NADH/NAD+ ratio. They also inhibited both oxygen uptake and ketogenesis and stimulated the citric acid cycle. Hesperidin, on the other hand, had little to no effect on these parameters. These results confirm the hypothesis that citrus flavanones are able to induce a more oxidised state in liver cells, altering parameters related to hepatic fatty acid oxidation. The prooxidant effect is most likely a consequence of the ability of these substances to oxidise NADH upon production of phenoxyl radicals in the presence of peroxidases and hydrogen peroxide. PMID:24288675

  13. Induction of mitochondrial fusion by cysteine-alkylators ethacrynic acid and N-ethylmaleimide.

    PubMed

    Bowes, Timothy J; Gupta, Radhey S

    2005-03-01

    Mitochondrial fusion and fission are important aspects of eukaryotic cell function that permit the adoption of varied mitochondrial morphologies depending upon cellular physiology. We previously observed that ethacrynic acid (EA) induced mitochondrial fusion in cultured BSC-1 and CHO/wt cells. However, the mechanism responsible for it was not clear since EA has a number of known cellular effects including glutathione (GSH) depletion and alkylation of cysteine residues. To gain insight, we have tested the effects of a variety of compounds on EA induced cellular toxicity and mitochondrial fusion. N-acetyl cysteine (NAC), a GSH precursor, was found to abrogate both the toxic and fusion-inductive effects, whereas diethylmaleate (dEM), a GSH depletor, potentiated both these effects in a dose-dependent manner. However, treatment with dEM alone, which depleted GSH to the same degree as EA, did not induce mitochondrial fusion. These results indicate that although detoxification of EA via formation of GSH conjugates is dependant upon GSH levels, the depletion of GSH by EA is not responsible for its effect on mitochondrial fusion. Dihydro-EA (DH-EA), a saturated EA analogue, lacked EA's toxicity and effect on fusion, indicating that the alpha,beta-unsaturated ketone is central to its observed effects. N-ethylmaleimide (NEM), another well-known cysteine-alkylator, also induced mitochondrial fusion at near toxic concentrations. These data suggests that cysteine-alkylation is the causative factor for fusion and toxicity. In live BSC-1 cells, EA induced fusion of mitochondria occurred very rapidly (<20 min), which suggests that it is inducing fusion by modifying certain critical cysteine residue(s) in proteins involved in the process.

  14. Biochemical properties of rat liver mitochondrial aldehyde dehydrogenase with respect to oxidation of formaldehyde.

    PubMed

    Cinti, D L; Keyes, S R; Lemelin, M A; Denk, H; Schenkman, J B

    1976-03-25

    The oxidation of formaldehyde by rat liver mitochondria in the presence of 50 mM phosphate was enhanced 2-fold by exogenous NAD+. Absolute requirement of NAD+ for formaldehyde oxidation was demonstrated by depleting the mitochondria of their NAD+ content (4.6 nmol/mg of protein), followed by reincorporation of the NAD+ into the depleted mitochondria. Aldehyde (formaldehyde) dehydrogenase activity was completely abolished in the depleted mitochondria, but the enzyme activity was restored to control levels following reincorporation of the pyridine nucleotide. Phosphate stimulation of formaldehyde oxidation could not be explained fully by the phosphate-induced swelling which enhances membrane permeability to NAD+, since stimulation of the enzyme activity by increased phosphate concentrations was still observed in the absence of exogenous NAD+. The Km for formaldehyde oxidation by the mitochondria was found to be 0.38 nM, a value similar to that obtained with varying concentrations of NAD+; both Vmax values were very similar, giving a value of 70 to 80 nmol/min/mg of protein. The pH optimum for the mitochondrial enzyme was 8.0. Inhibition of the enzyme activity by anaerobiosis was apparently due to the inability of the respiratory chain to oxidize the generated NADH. The inhibition of mitochondrial formaldehyde oxidation by succinate was found to be due to a lowering of the NAD+ level in the mitochondria. Succinate also inhibited acetaldehyde oxidation by the mitochondria. Malonate, a competitive inhibitor of succinic dehydrogenase, blocked the inhibitory effect of succinate. The respiratory chain inhibitors, rotenone, and antimycin A plus succinate, strongly inhibited formaldehyde oxidation by apparently the same mechanism, although the crude enzyme preparation (freed from the membrane) was slightly sensitive to rotenone. The mitochondria were subfractionated, and 85% of the enzyme activity was found in the inner membrane fraction (mitoplast). Furthermore, separation

  15. Long-Term Exposure to AZT, but not d4T, Increases Endothelial Cell Oxidative Stress and Mitochondrial Dysfunction

    PubMed Central

    Kline, Erik R.; Bassit, Leda; Hernandez-Santiago, Brenda I.; Detorio, Mervi A.; Liang, Bill; Kleinhenz, Dean J.; Walp, Erik R.; Dikalov, Sergey; Jones, Dean P.; Schinazi, Raymond F.

    2009-01-01

    Nucleoside reverse transcriptase inhibitors (NRTIs), such as zidovudine (AZT) and stavudine (d4T), cause toxicities to numerous tissues, including the liver and vasculature. While much is known about hepatic NRTI toxicity, the mechanism of toxicity in endothelial cells is incompletely understood. Human aortic endothelial and HepG2 liver cells were exposed to 1 μM AZT or d4T for up to 5 weeks. Markers of oxidative stress, mitochondrial function, NRTI phosphorylation, mitochondrial DNA (mtDNA) levels, and cytotoxicity were monitored over time. In endothelial cells, AZT significantly oxidized glutathione redox potential, increased total cellular and mitochondrial-specific superoxide, decreased mitochondrial membrane potential, increased lactate release, and caused cell death from weeks 3 through 5. Toxicity occurred in the absence of di- and tri-phosphorylated AZT and mtDNA depletion. These data show that oxidative stress and mitochondrial dysfunction in endothelial cells occur with a physiologically relevant concentration of AZT, and require long-term exposure to develop. In contrast, d4T did not induce endothelial oxidative stress, mitochondrial dysfunction, or cytotoxicity despite the presence of d4T-triphosphate. Both drugs depleted mtDNA in HepG2 cells without causing cell death. Endothelial cells are more susceptible to AZT-induced toxicity than HepG2 cells, and AZT caused greater endothelial dysfunction than d4T because of its pro-oxidative effects. PMID:19067249

  16. Oxidants, antioxidants and mitochondrial function in non-proliferative diabetic retinopathy

    PubMed Central

    Rodríguez-Carrizalez, Adolfo Daniel; Castellanos-González, José Alberto; Martínez-Romero, Esaú César; Miller-Arrevillaga, Guillermo; Villa-Hernández, David; Hernández-Godínez, Pedro Pablo; Ortiz, Genaro Gabriel; Pacheco-Moisés, Fermín Paul; Cardona-Muñoz, Ernesto Germán; Miranda-Díaz, Alejandra Guillermina

    2014-01-01

    Background Diabetic retinopathy (DR) is a preventable cause of visual disability. The aims of the present study were to investigate levels and behavior oxidative stress markers and mitochondrial function in non-proliferative DR (NPDR) and to establish the correlation between the severity of NPDR and markers of oxidative stress and mitochondrial function. Methods In a transverse analysis, type 2 diabetes mellitus (T2DM) patients with mild, moderate and severe non-proliferative DR (NPDR) were evaluated for markers of oxidative stress (i.e. products of lipid peroxidation (LPO) and nitric oxide (NO) catabolites) and antioxidant activity (i.e. total antioxidant capacity (TAC), catalase, and glutathione peroxidase (GPx) activity of erythrocytes). Mitochondrial function was also determined as the fluidity of the submitochondrial particles of platelets and the hydrolytic activity of F0/F1-ATPase. Results Levels of LPO and NO were significantly increased in T2DM patients with severe NPDR (3.19 ± 0.05 μmol/mL and 45.62 ± 1.27 pmol/mL, respectively; P < 0.007 and P < 0.0001 vs levels in health volunteers, respectively), suggesting the presence of oxidative stress. TAC had significant decrease levels with minimum peak in severe retinopathy with 7.98 ± 0.48 mEq/mL (P < 0.0001). In contrast with TAC, erythrocyte catalase and GPx activity was increased in patients with severe NPDR (139.4 ± 4.4 and 117.13 ± 14.84 U/mg, respectively; P < 0.0001 vs healthy volunteers for both), suggesting an imbalance between oxidants and antioxidants. The fluidity of membrane submitochondrial particles decreased significantly in T2DM patients with mild, moderate, or severe NPDR compared with that in healthy volunteers (P < 0.0001 for all). Furthermore, there was a significant increase in the hydrolytic activity of the F0/F1-ATPase in T2DM patients with mild NPDR (265.07 ± 29.55 nmol/PO4; P < 0.0001 vs healthy volunteers), suggesting

  17. Mechanism underlying mitochondrial protection of asiatic acid against hepatotoxicity in mice.

    PubMed

    Gao, Jing; Chen, Jin; Tang, Xinhui; Pan, Liya; Fang, Feng; Xu, Lizhi; Zhao, Xiaoning; Xu, Qiang

    2006-02-01

    Asiatic acid (AA) is one of the triterpenoid components of Terminalia catappa L., which has antioxidative, anti-inflammatory and hepatoprotective activity. This research focused on the mitochondrial protection of AA against acute liver injury induced by lipopolysaccharide (LPS) and D-galactosamine (D-GalN) in mice. It was found that pretreatment with 25, 50 or 100 mg kg(-1) AA significantly blocked the LPS + D-GalN-induced increase in both serum aspartate aminotransferase (sAST) and serum alanine aminotransferase (sALT) levels, which was confirmed by ultrastructural observation under an electron microscope, showing improved nuclear condensation, ameliorated mitochondrion proliferation and less lipid deposition. Meanwhile, different doses of AA could decrease both the transcription and the translation level of voltage-dependent anion channels (VDACs), the most important mitochondrial PTP component protein, and block the translocation of cytochrome c from mitochondria to cytosol. On the other hand, pre-incubation with 25, 50 and 100 microg mL(-1) AA inhibited the Ca(2+)-induced mitochondrial permeability transition (MPT), including mitochondrial swelling, membrane potential dissipation and releasing of matrix Ca(2+) in liver mitochondria separated from normal mice, indicating the direct role of AA on mitochondria. Collectively, the above data suggest that AA could protect liver from damage and the mechanism might be related to up-regulating mitochondrial VDACs and inhibiting the process of MPT.

  18. Methane activation and oxidation in sulfuric acid.

    PubMed

    Goeppert, Alain; Dinér, Peter; Ahlberg, Per; Sommer, Jean

    2002-07-15

    The H/D exchange observed when methane is contacted with D(2)SO(4) at 270-330 degrees C shows that the alkane behaves as a sigma base and undergoes rapid and reversible protonation at this temperature. DFT studies of the hydrogen exchange between a monomer and a dimer of sulfuric acid and methane show that the transition states involved in the exchange are bifunctional, that is one hydrogen atom is transferred from a hydroxy group in sulfuric acid to methane, while one hydrogen atom is abstracted from methane by a non-hydroxy oxygen atom in sulfuric acid. All the transition states include a CH(5) moiety, which shows similarities to the methanium ion CH(5) (+). The calculated potential activation energy of the hydrogen exchange for the monomer is 174 kJ mol(-1), which is close to the experimental value (176 kJ mol(-1)). Solvation of the monomer and the transition state of the monomer with an extra sulfuric acid molecule, decrease the potential activation energy by 6 kJ mol(-1). The acid-base process is in competition, however, with an oxidative process involving methane and sulfuric acid which leads to CO(2), SO(2), and water, and thus to a decrease of acidity and loss of reactivity of the medium.

  19. Nickel(II)-induced nasal epithelial toxicity and oxidative mitochondrial damage.

    PubMed

    Lee, Yoon-Jin; Lim, Soo-Sung; Baek, Byoung Joon; An, Je-Min; Nam, Hae-Seon; Woo, Kee-Min; Cho, Moon-Kyun; Kim, Sung-Ho; Lee, Sang-Han

    2016-03-01

    In probing the underlying mechanisms of nickel(II)-induced cytotoxicity on nasal epithelium, we investigated the effects of nickel(II) acetate on nasal epithelial RPMI-2650 cells. Nickel(II) elicited apoptosis, as signified by pyknotic and fragmented nuclei, increased caspase-3/7 activity, and an increase in annexin V binding, hypodiploid DNA, and Bax/Bcl-2 protein ratio. Nickel(II)-induced G2/M arrest was associated with up-regulation of p21(WAF1/CIP1) expression, decrease in phosphorylation at Thr(161) of Cdc2, and down-regulation of cyclin B1. Associated with these responses, ROS generation and mitochondrial depolarization increased in a nickel(II) concentration-dependent fashion. Pretreatment with N-acetylcysteine (NAC) attenuated these changes. p53 reporter gene assay and analyses of p53, Puma, Bax, and Bcl-2 protein levels indicated that NAC inhibited nickel(II)-induced activation of p53-mediated mitochondrial apoptotic pathway. Collectively, our study provides evidences that nickel(II) may induce oxidative damage on nasal epithelium in which antioxidant NAC protects cells against nickel(II)-induced apoptosis through the prevention of oxidative stress-mediated mitochondrial damage.

  20. Oxidative stress generated during monensin treatment contributes to altered Toxoplasma gondii mitochondrial function

    PubMed Central

    Charvat, Robert A.; Arrizabalaga, Gustavo

    2016-01-01

    The ionophore monensin displays potent activities against several coccidian parasites of veterinary and medical importance including the opportunistic pathogen of humans, Toxoplasma gondii. While monensin is used widely in animals, toxicity impedes its use in humans. Nonetheless, given its potency, understanding its mode of action would reveal vulnerable aspects of the parasite that can be exploited for drug development. We previously established that monensin induces Toxoplasma to undergo cell cycle arrest and an autophagy-like cell death. Interestingly, these effects are dependent on the mitochondrion-localized TgMSH-1 protein, suggesting that monensin disrupts mitochondrial function. We demonstrate that monensin treatment results in decreased mitochondrial membrane potential and altered morphology. These effects are mitigated by the antioxidant compound N-acetyl-cysteine suggesting that monensin causes an oxidative stress, which was indeed the case based on direct detection of reactive oxygen species. Moreover, over-expression of the antioxidant proteins glutaredoxin and peroxiredoxin 2 protect Toxoplasma from the deleterious effects of monensin. Thus, our studies show that the effects of monensin on Toxoplasma are due to a disruption of mitochondrial function caused by the induction of an oxidative stress and implicate parasite redox biology as a viable target for the development of drugs against Toxoplasma and related pathogenic parasites. PMID:26976749

  1. Mitochondrial Dysfunction and Oxidative Stress in Asthma: Implications for Mitochondria-Targeted Antioxidant Therapeutics

    PubMed Central

    Reddy, P. Hemachandra

    2011-01-01

    Asthma is a complex, inflammatory disorder characterized by airflow obstruction of variable degrees, bronchial hyper-responsiveness, and airway inflammation. Asthma is caused by environmental factors and a combination of genetic and environmental stimuli. Genetic studies have revealed that multiple loci are involved in the etiology of asthma. Recent cellular, molecular, and animal-model studies have revealed several cellular events that are involved in the progression of asthma, including: increased Th2 cytokines leading to the recruitment of inflammatory cells to the airway, and an increase in the production of reactive oxygen species and mitochondrial dysfunction in the activated inflammatory cells, leading to tissue injury in the bronchial epithelium. Further, aging and animal model studies have revealed that mitochondrial dysfunction and oxidative stress are involved and play a large role in asthma. Recent studies using experimental allergic asthmatic mouse models and peripheral cells and tissues from asthmatic humans have revealed antioxidants as promising treatments for people with asthma. This article summarizes the latest research findings on the involvement of inflammatory changes, and mitochondrial dysfunction/oxidative stress in the development and progression of asthma. This article also addresses the relationship between aging and age-related immunity in triggering asthma, the antioxidant therapeutic strategies in treating people with asthma. PMID:21461182

  2. Oxidative stress generated during monensin treatment contributes to altered Toxoplasma gondii mitochondrial function.

    PubMed

    Charvat, Robert A; Arrizabalaga, Gustavo

    2016-03-15

    The ionophore monensin displays potent activities against several coccidian parasites of veterinary and medical importance including the opportunistic pathogen of humans, Toxoplasma gondii. While monensin is used widely in animals, toxicity impedes its use in humans. Nonetheless, given its potency, understanding its mode of action would reveal vulnerable aspects of the parasite that can be exploited for drug development. We previously established that monensin induces Toxoplasma to undergo cell cycle arrest and an autophagy-like cell death. Interestingly, these effects are dependent on the mitochondrion-localized TgMSH-1 protein, suggesting that monensin disrupts mitochondrial function. We demonstrate that monensin treatment results in decreased mitochondrial membrane potential and altered morphology. These effects are mitigated by the antioxidant compound N-acetyl-cysteine suggesting that monensin causes an oxidative stress, which was indeed the case based on direct detection of reactive oxygen species. Moreover, over-expression of the antioxidant proteins glutaredoxin and peroxiredoxin 2 protect Toxoplasma from the deleterious effects of monensin. Thus, our studies show that the effects of monensin on Toxoplasma are due to a disruption of mitochondrial function caused by the induction of an oxidative stress and implicate parasite redox biology as a viable target for the development of drugs against Toxoplasma and related pathogenic parasites.

  3. Salvianolate Protects Hepatocytes from Oxidative Stress by Attenuating Mitochondrial Injury

    PubMed Central

    Zhao, Qiang; Peng, Yuan; Huang, Kai; Lei, Yang; Liu, Hong-Liang; Tao, Yan-Yan

    2016-01-01

    Salvianolate is widely used to treat angiocardiopathy in clinic in China, but its application in liver diseases remains unclear. Our study aims to investigate the effect of Salvianolate on rat hepatic injury by protecting hepatocyte mitochondria. To evaluate the effects of Salvianolate on injured hepatocytes, alpha mouse liver 12 (AML-12) cells were induced with hydrogen peroxide (H2O2) and treated with Salvianolate. Cell viability and MitoTracker Green for mitochondria and 5,5′,6,6′-tetrachloro-1,1′,3,3′-tetraethylbenzimidazole-carbocyanide iodine (JC-1) levels and cytochrome C (Cyto-C) expressions were detected in vitro. To identify the effect of Salvianolate on protecting against mitochondria injury, male Wistar rats were injected with carbon tetrachloride (CCl4) and treated with Salvianolate (40 mg·kg−1). Serum liver function, parameters for peroxidative damage, hematoxylin and eosin (H&E) staining, and transmission electron microscope (TEM) of hepatocyte mitochondria were assayed. Our results showed that Salvianolate effectively protected hepatocytes, increased mitochondria vitality, and decreased Cyto-C expressions in vitro. Besides, Salvianolate alleviated the liver function, attenuated the indicators of peroxidation, and relieved the mitochondria injury in vivo. In conclusion, Salvianolate is effective in protecting hepatocytes from injury in vitro and in vivo, and the mechanism might be related to its protective effect on hepatocyte mitochondria against oxidative stress. PMID:27340417

  4. Quercetin protects against aluminium induced oxidative stress and promotes mitochondrial biogenesis via activation of the PGC-1α signaling pathway.

    PubMed

    Sharma, Deep Raj; Sunkaria, Aditya; Wani, Willayat Yousuf; Sharma, Reeta Kumari; Verma, Deepika; Priyanka, Kumari; Bal, Amanjit; Gill, Kiran Dip

    2015-12-01

    The present investigation was carried out to elucidate a possible molecular mechanism related to the protective effect of quercetin administration against aluminium-induced oxidative stress on various mitochondrial respiratory complex subunits with special emphasis on the role of PGC-1α and its downstream targets, i.e. NRF-1, NRF-2 and Tfam in mitochondrial biogenesis. Aluminium lactate (10mg/kg b.wt./day) was administered intragastrically to rats, which were pre-treated with quercetin 6h before aluminium (10mg/kg b.wt./day, intragastrically) for 12 weeks. We found a decrease in ROS levels, mitochondrial DNA oxidation and citrate synthase activity in the hippocampus (HC) and corpus striatum (CS) regions of rat brain treated with quercetin. Besides this an increase in the mRNA levels of the mitochondrial encoded subunits - ND1, ND2, ND3, Cyt b, COX1, COX3 and ATPase6 along with increased expression of nuclear encoded subunits COX4, COX5A and COX5B of electron transport chain (ETC). In quercetin treated group an increase in the mitochondrial DNA copy number and mitochondrial content in both the regions of rat brain was observed. The PGC-1α was up regulated in quercetin treated rats along with NRF-1, NRF-2 and Tfam, which act downstream from PGC-1α. Electron microscopy results revealed a significant decrease in the mitochondrial cross-section area, mitochondrial perimeter length and increase in mitochondrial number in case of quercetin treated rats as compared to aluminium treated ones. Therefore it seems quercetin increases mitochondrial biogenesis and makes it an almost ideal flavanoid to control or limit the damage that has been associated with the defective mitochondrial function seen in many neurodegenerative diseases.

  5. Melatonin prevents the dynamin-related protein 1-dependent mitochondrial fission and oxidative insult in the cortical neurons after 1-methyl-4-phenylpyridinium treatment.

    PubMed

    Chuang, Jih-Ing; Pan, I-Ling; Hsieh, Chia-Yun; Huang, Chiu-Ying; Chen, Pei-Chun; Shin, Jyh Wei

    2016-09-01

    Mitochondrial dysfunction and oxidative stress are involved in the pathogenesis of Parkinson's disease (PD). Mitochondrial morphology is dynamic and precisely regulated by the mitochondrial fission and fusion machinery. Aberrant mitochondrial fragmentation controlled by the mitochondrial fission protein, dynamin-related protein 1 (Drp1), may result in cell death. Our previous results showed that melatonin protected neurons by inhibiting oxidative stress in a 1-methyl-4-phenylpyridinium (MPP(+) )-induced PD model. However, the effect of melatonin on mitochondrial dynamics remains uncharacterized. Herein, we investigated the effect of melatonin and the role of Drp1 on MPP(+) -induced mitochondrial fission in rat primary cortical neurons. We found that MPP(+) induced a rapid increase in the ratio of GSSG:total glutathione (a marker of oxidative stress) and mitochondrial fragmentation, Drp1 upregulation within 4 hours, and finally resulted in neuron loss 48 hours after the treatment. Neurons overexpressing wild-type Drp1 promoted mitochondrial and nuclear fragmentation; however, neurons overexpressing dominant-negative Drp1(K38A) or cotreated with melatonin exhibited significantly reduced MPP(+) -induced mitochondrial fragmentation and neuron death. Moreover, melatonin cotreatment prevented an MPP(+) -induced high ratio of GSSG and mitochondrial Drp1 upregulation. The prevention of mitochondrial fission by melatonin was not found in neurons transfected with wild-type Drp1. These results provide a new insight that the neuroprotective effect of melatonin against MPP(+) toxicity is mediated by inhibiting the oxidative stress and Drp1-mediated mitochondrial fragmentation.

  6. Systemic induction and role of mitochondrial alternative oxidase and nitric oxide in a compatible tomato-Tobacco mosaic virus interaction.

    PubMed

    Fu, Li-Jun; Shi, Kai; Gu, Min; Zhou, Yan-Hong; Dong, De-Kun; Liang, Wu-Sheng; Song, Feng-Ming; Yu, Jing-Quan

    2010-01-01

    The role of mitochondrial alternative oxidase (AOX) and the relationship between AOX and nitric oxide (NO) in virus-induced systemic defense to Tobacco mosaic virus (TMV) were investigated in susceptible tomato (Solanum lycopersicum) plants. TMV inoculation to the lower leaves induced a rapid NO synthesis and AOX activation in upper uninoculated leaves as early as 0.5 day postinoculation. Application of exogenous potassium cyanide (KCN, a cytochrome pathway inhibitor) at nonlethal concentrations and NO donor diethylamine NONOate (DEA/NO) to the upper uninoculated leaves greatly induced accumulation of AOX transcript, reduced TMV viral RNA accumulation, and increased the leaf photochemical quantum yield at photosystem II. Pretreatment with NO scavenger almost completely blocked TMV-induced AOX induction and substantially increased TMV susceptibility. Salicylhydroxamic acid (SHAM, an AOX inhibitor) pretreatment reduced the DEA/NO-induced cyanide-resistant respiration and partially compromised induced resistance to TMV. Conversely, KCN and SHAM pretreatment had very little effect on generation of NO, and pretreatment with NO scavenger did not affect KCN-induced AOX induction and TMV resistance. These results suggest that TMV-induced NO generation acts upstream and mediates AOX induction which, in turn, induces mitochondrial alternative electron transport and triggers systemic basal defense against the viral pathogen.

  7. Hypochlorous and peracetic acid induced oxidation of dairy proteins.

    PubMed

    Kerkaert, Barbara; Mestdagh, Frédéric; Cucu, Tatiana; Aedo, Philip Roger; Ling, Shen Yan; De Meulenaer, Bruno

    2011-02-09

    Hypochlorous and peracetic acids, both known disinfectants in the food industry, were compared for their oxidative capacity toward dairy proteins. Whey proteins and caseins were oxidized under well controlled conditions at pH 8 as a function of the sanitizing concentration. Different markers for protein oxidation were monitored. The results established that the protein carbonyl content was a rather unspecific marker for protein oxidation, which did not allow one to differentiate the oxidant used especially at the lower concentrations. Cysteine, tryptophan, and methionine were proven to be the most vulnerable amino acids for degradation upon hypochlorous and peracetic acid treatment, while tyrosine was only prone to degradation in the presence of hypochlorous acid. Hypochlorous acid induced oxidation gave rise to protein aggregation, while during peracetic acid induced oxidation, no high molecular weight aggregates were observed. Protein aggregation upon hypochlorous acid oxidation could primarily be linked to tryptophan and tyrosine degradation.

  8. Spongionella Secondary Metabolites Protect Mitochondrial Function in Cortical Neurons against Oxidative Stress

    PubMed Central

    Leirós, Marta; Sánchez, Jon A.; Alonso, Eva; Rateb, Mostafa E.; Houssen, Wael E.; Ebel, Rainer; Jaspars, Marcel; Alfonso, Amparo; Botana, Luis M.

    2014-01-01

    The marine habitat provides a large number of structurally-diverse bioactive compounds for drug development. Marine sponges have been studied over many years and are found to be a rich source of these bioactive chemicals. This study is focused on the evaluation of the activity of six diterpene derivatives isolated from Spongionella sp. on mitochondrial function using an oxidative in vitro stress model. The test compounds include the Gracilins (A, H, K, J and L) and tetrahydroaplysulphurin-1. Compounds were co-incubated with hydrogen peroxide for 12 hours to determine their protective capacities and their effect on markers of apoptosis and Nrf2/ARE pathways was evaluated. Results conclude that Gracilins preserve neurons against oxidative damage, and that in particular, tetrahydroaplysulphurin-1 shows a complete neuroprotective activity. Oxidative stress is linked to mitochondrial dysfunction and consequently to neurodegenerative disorders like Parkinson and Alzheimer diseases, Friedreich ataxia or Amyotrophic lateral sclerosis. This neuroprotection against oxidation conditions suggest that these metabolites could be interesting lead candidates in drug development for neurodegenerative diseases. PMID:24473170

  9. Therapeutic efficacy of chlorogenic acid on cadmium-induced oxidative neuropathy in a murine model

    PubMed Central

    HAO, MAO-LIN; PAN, NING; ZHANG, QING-HUA; WANG, XIAO-HONG

    2015-01-01

    The aim of the present study was to determine whether chlorogenic acid (CA) is able to modulate cadmium (Cd)-induced oxidative brain damage. Cd-treated rats displayed numerous pathological effects, including the inhibition of acetylcholinesterase, elevated lipid peroxidation, the depletion of enzymatic and non-enzymatic antioxidants, the reduction of membrane-bound ATPase activity, mitochondrial dysfunction and DNA fragmentation. Pretreatment of the rats with CA significantly attenuated these effects. These results lead to the hypothesis that the mechanisms by which CA attenuates the effects of Cd-induced oxidative brain damage include the maintenance of antioxidant homeostasis, inhibition of the membrane effects and the perpetuation of mitochondrial dysfunction. These data support the potential of CA as a beneficial intervention in the prevention of heavy metal poisoning due to Cd exposure. PMID:26136910

  10. Conservative and compensatory evolution in oxidative phosphorylation complexes of angiosperms with highly divergent rates of mitochondrial genome evolution.

    PubMed

    Havird, Justin C; Whitehill, Nicholas S; Snow, Christopher D; Sloan, Daniel B

    2015-12-01

    Interactions between nuclear and mitochondrial gene products are critical for eukaryotic cell function. Nuclear genes encoding mitochondrial-targeted proteins (N-mt genes) experience elevated rates of evolution, which has often been interpreted as evidence of nuclear compensation in response to elevated mitochondrial mutation rates. However, N-mt genes may be under relaxed functional constraints, which could also explain observed increases in their evolutionary rate. To disentangle these hypotheses, we examined patterns of sequence and structural evolution in nuclear- and mitochondrial-encoded oxidative phosphorylation proteins from species in the angiosperm genus Silene with vastly different mitochondrial mutation rates. We found correlated increases in N-mt gene evolution in species with fast-evolving mitochondrial DNA. Structural modeling revealed an overrepresentation of N-mt substitutions at positions that directly contact mutated residues in mitochondrial-encoded proteins, despite overall patterns of conservative structural evolution. These findings support the hypothesis that selection for compensatory changes in response to mitochondrial mutations contributes to the elevated rate of evolution in N-mt genes. We discuss these results in light of theories implicating mitochondrial mutation rates and mitonuclear coevolution as drivers of speciation and suggest comparative and experimental approaches that could take advantage of heterogeneity in rates of mtDNA evolution across eukaryotes to evaluate such theories.

  11. Time courses of post-injury mitochondrial oxidative damage and respiratory dysfunction and neuronal cytoskeletal degradation in a rat model of focal traumatic brain injury.

    PubMed

    Hill, Rachel L; Singh, Indrapal N; Wang, Juan A; Hall, Edward D

    2017-03-23

    Traumatic brain injury (TBI) results in rapid reactive oxygen species (ROS) production and oxidative damage to essential brain cellular components leading to neuronal dysfunction and cell death. It is increasingly appreciated that a major player in TBI-induced oxidative damage is the reactive nitrogen species (RNS) peroxynitrite (PN) which is produced in large part in injured brain mitochondria. Once formed, PN decomposes into highly reactive free radicals that trigger membrane lipid peroxidation (LP) of polyunsaturated fatty acids (e.g. arachidonic acid) and protein nitration (3-nitrotyrosine, 3-NT) in mitochondria and other cellular membranes causing various functional impairments to mitochondrial oxidative phosphorylation and calcium (Ca(2+)) buffering capacity. The LP also results in the formation of neurotoxic reactive aldehyde byproducts including 4-hydroxynonenal (4-HNE) and propenal (acrolein) which exacerbates ROS/RNS production and oxidative protein damage in the injured brain. Ultimately, this results in intracellular Ca(2+) overload that activates proteolytic degradation of α-spectrin, a neuronal cytoskeletal protein. Therefore, the aim of this study was to establish the temporal evolution of mitochondrial dysfunction, oxidative damage and cytoskeletal degradation in the brain following a severe controlled cortical impact (CCI) TBI in young male adult rats. In mitochondria isolated from an 8 mm diameter cortical punch including the 5 mm wide impact site and their respiratory function studied ex vivo, we observed an initial decrease in complex I and II mitochondrial bioenergetics within 3 h (h). For complex I bioenergetics, this partially recovered by 12-16 h, whereas for complex II respiration the recovery was complete by 12 h. During the first 24 h, there was no evidence of an injury-induced increase in LP or protein nitration in mitochondrial or cellular homogenates. However, beginning at 24 h, there was a gradual secondary decline in complex

  12. Behavioral deficit, oxidative stress, and mitochondrial dysfunction precede tau pathology in P301S transgenic mice

    PubMed Central

    Dumont, Magali; Stack, Cliona; Elipenahli, Ceyhan; Jainuddin, Shari; Gerges, Meri; Starkova, Natalia N.; Yang, Lichuan; Starkov, Anatoly A.; Beal, Flint

    2011-01-01

    Abnormal tau accumulation can lead to the development of neurodegenerative diseases. P301S mice overexpress the human tau mutated gene, resulting in tau hyperphosphorylation and tangle formation. Mice also develop synaptic deficits and microglial activation prior to any neurodegeneration and tangles. Oxidative stress can also affect tauopathy. We studied the role of oxidative stress in relationship to behavioral abnormalities and disease progression in P301S mice at 2, 7, and 10 mo of age. At 7 mo of age, P301S mice had behavioral abnormalities, such as hyperactivity and disinhibition. At the same age, we observed increased carbonyls in P301S mitochondria (∼215 and 55% increase, males/females), and deregulation in the activity and content of mitochondrial enzymes involved in reactive oxygen species formation and energy metabolism, such as citrate synthase (∼19 and ∼5% decrease, males/females), MnSOD (∼16% decrease, males only), cytochrome C (∼19% decrease, females only), and cytochrome C oxidase (∼20% increase, females only). These changes in mitochondria proteome appeared before tau hyperphosphorylation and tangle formation, which were observed at 10 mo and were associated with GSK3β activation. At that age, mitochondria proteome deregulation became more apparent in male P301S mitochondria. The data strongly suggest that oxidative stress and mitochondrial abnormalities appear prior to tau pathology.—Dumont, M., Stack, C., Elipenahli, C., Jainuddin, S., Gerges, M., Starkova, M. N., Yang, L., Starkov, A. A., Beal, F. Behavioral deficit, oxidative stress, and mitochondrial dysfunction precede tau pathology in P301S transgenic mice. PMID:21825035

  13. The Mitochondrial Sulfur Dioxygenase ETHYLMALONIC ENCEPHALOPATHY PROTEIN1 Is Required for Amino Acid Catabolism during Carbohydrate Starvation and Embryo Development in Arabidopsis1[C][W

    PubMed Central

    Krüßel, Lena; Junemann, Johannes; Wirtz, Markus; Birke, Hannah; Thornton, Jeremy D.; Browning, Luke W.; Poschet, Gernot; Hell, Rüdiger; Balk, Janneke; Braun, Hans-Peter; Hildebrandt, Tatjana M.

    2014-01-01

    The sulfur dioxygenase ETHYLMALONIC ENCEPHALOPATHY PROTEIN1 (ETHE1) catalyzes the oxidation of persulfides in the mitochondrial matrix and is essential for early embryo development in Arabidopsis (Arabidopsis thaliana). We investigated the biochemical and physiological functions of ETHE1 in plant metabolism using recombinant Arabidopsis ETHE1 and three transfer DNA insertion lines with 50% to 99% decreased sulfur dioxygenase activity. Our results identified a new mitochondrial pathway catalyzing the detoxification of reduced sulfur species derived from cysteine catabolism by oxidation to thiosulfate. Knockdown of the sulfur dioxygenase impaired embryo development and produced phenotypes of starvation-induced chlorosis during short-day growth conditions and extended darkness, indicating that ETHE1 has a key function in situations of high protein turnover, such as seed production and the use of amino acids as alternative respiratory substrates during carbohydrate starvation. The amino acid profile of mutant plants was similar to that caused by defects in the electron-transfer flavoprotein/electron-transfer flavoprotein:ubiquinone oxidoreductase complex and associated dehydrogenases. Thus, in addition to sulfur amino acid catabolism, ETHE1 also affects the oxidation of branched-chain amino acids and lysine. PMID:24692429

  14. Oxalic acid mineralization by electrochemical oxidation processes.

    PubMed

    Huang, Yao-Hui; Shih, Yu-Jen; Liu, Cheng-Hong

    2011-04-15

    In this study, two electrochemical oxidation processes were utilized to mineralize oxalic acid which was a major intermediate compound in the oxidation of phenols and other aromatic compounds. The anode rod and cathode net were made of a titanium coated with RuO(2)/IrO(2) (Ti-DSA) and stainless steel (S.S. net, SUS304), respectively. First, the Fered-Fenton process, which used H(2)O(2) and Fe(2+) as additive reagents, achieved 85% of TOC removal. It proceeded with ligand-to-metal charge-transfer (LMCT), which was evidenced by the accumulation of metallic foil on the selected cathode. However, in the absence of H(2)O(2)/Fe(2+), it showed a higher TOC removal efficiency while using Cl(-) only as an additive reagent due to the formation of hypochlorite on the anode. It was also found that the mineralization of oxalic acid by electrolysis generated hypochlorite better than the dosage of commercial hypochlorite without electricity. Also, pH value was a major factor that affected the mineralization efficiency of the oxalic acid due to the chlorine chemistry. 99% TOC removal could be obtained by Cl(-) electrolysis in an acidic environment.

  15. Human Cytomegalovirus Infection Upregulates the Mitochondrial Transcription and Translation Machineries

    PubMed Central

    Weekes, M. P.; Antrobus, R.; Rorbach, J.; van Haute, L.; Umrania, Y.; Smith, D. L.; Minczuk, M.; Lehner, P. J.; Sinclair, J. H.

    2016-01-01

    ABSTRACT Infection with human cytomegalovirus (HCMV) profoundly affects cellular metabolism. Like in tumor cells, HCMV infection increases glycolysis, and glucose carbon is shifted from the mitochondrial tricarboxylic acid cycle to the biosynthesis of fatty acids. However, unlike in many tumor cells, where aerobic glycolysis is accompanied by suppression of mitochondrial oxidative phosphorylation, HCMV induces mitochondrial biogenesis and respiration. Here, we affinity purified mitochondria and used quantitative mass spectrometry to determine how the mitochondrial proteome changes upon HCMV infection. We found that the mitochondrial transcription and translation systems are induced early during the viral replication cycle. Specifically, proteins involved in biogenesis of the mitochondrial ribosome were highly upregulated by HCMV infection. Inhibition of mitochondrial translation with chloramphenicol or knockdown of HCMV-induced ribosome biogenesis factor MRM3 abolished the HCMV-mediated increase in mitochondrially encoded proteins and significantly impaired viral growth under bioenergetically restricting conditions. Our findings demonstrate how HCMV manipulates mitochondrial biogenesis to support its replication. PMID:27025248

  16. Modulation of mitochondrial capacity and angiogenesis by red wine polyphenols via estrogen receptor, NADPH oxidase and nitric oxide synthase pathways.

    PubMed

    Duluc, Lucie; Jacques, Caroline; Soleti, Raffaella; Iacobazzi, Francesco; Simard, Gilles; Andriantsitohaina, Ramaroson

    2013-04-01

    Red wine polyphenolic compounds (RWPC) are reported to exert vasculoprotective properties on endothelial cells, involving nitric oxide (NO) release via a redox-sensitive pathway. This NO release involves the activation of the estrogen receptor-alpha (ERα). Paradoxical effects of a RWPC treatment occur in a rat model of post-ischemic neovascularization, where a low-dose is pro-angiogenic while a higher dose is anti-angiogenic. NO and ERα are key regulators of mitochondrial capacity, and angiogenesis is a highly energetic process associated with mitochondrial biogenesis. However, whether RWPC induces changes in mitochondrial capacity has never been addressed. We investigated the effects of RWPC at low (10(-4)g/l, LCP) and high concentration (10(-2)g/l, HCP) in human endothelial cells. Mitochondrial respiration, expression of mitochondrial biogenesis factors and mitochondrial DNA content were assessed using oxygraphy and quantitative PCR respectively. In vitro capillary formation using ECM gel(®) was also performed. Treatment with LCP increased mitochondrial respiration, with a maximal effect achieved at 48h. LCP also increased expression of several mitochondrial biogenesis factors and mitochondrial DNA content. In contrast, HCP did not affect these parameters. Furthermore, LCP modulated both mitochondrial capacity and angiogenesis through mechanisms sensitive to ER, NADPH oxidase and NO-synthase inhibitors. Finally, the inhibition of mitochondrial protein synthesis abolished the pro-angiogenic capacity of LCP. These results suggest a possible association between the modulation of mitochondrial capacity by LCP and its pro-angiogenic activity. These data provide evidence for a role of mitochondria in the regulation of angiogenesis by RWPC.

  17. The novel mitochondrial iron chelator 5-((methylamino)methyl)-8-hydroxyquinoline protects against mitochondrial-induced oxidative damage and neuronal death.

    PubMed

    Mena, Natalia P; García-Beltrán, Olimpo; Lourido, Fernanda; Urrutia, Pamela J; Mena, Raúl; Castro-Castillo, Vicente; Cassels, Bruce K; Núñez, Marco T

    2015-08-07

    Abundant evidence indicates that iron accumulation, oxidative damage and mitochondrial dysfunction are common features of Huntington's disease, Parkinson's disease, Friedreich's ataxia and a group of disorders known as Neurodegeneration with Brain Iron Accumulation. In this study, we evaluated the effectiveness of two novel 8-OH-quinoline-based iron chelators, Q1 and Q4, to decrease mitochondrial iron accumulation and oxidative damage in cellular and animal models of PD. We found that at sub-micromolar concentrations, Q1 selectively decreased the mitochondrial iron pool and was extremely effective in protecting against rotenone-induced oxidative damage and death. Q4, in turn, preferentially chelated the cytoplasmic iron pool and presented a decreased capacity to protect against rotenone-induced oxidative damage and death. Oral administration of Q1 to mice protected substantia nigra pars compacta neurons against oxidative damage and MPTP-induced death. Taken together, our results support the concept that oral administration of Q1 is a promising therapeutic strategy for the treatment of NBIA.

  18. Mitochondrial cytochrome c oxidase: mechanism of action and role in regulating oxidative phosphorylation.

    PubMed

    Wilson, David F; Vinogradov, Sergei A

    2014-12-15

    Mitochondrial oxidative phosphorylation has a central role in eukaryotic metabolism, providing the energy (ATP) required for survival. Regulation of this important pathway is, however, still not understood, largely due to limitations in the ability to measure the essential metabolites, including oxygen (pO2, oxygen pressure), ADP, and AMP. In addition, neither the mechanism of oxygen reduction by mitochondrial cytochrome c oxidase nor how its rate is controlled is understood, although this enzyme determines the rate of oxygen consumption and thereby the rate of ATP synthesis. Cytochrome c oxidase is responsible for reduction of molecular oxygen to water using reducing equivalents donated by cytochrome c and for site 3 energy coupling in oxidative phosphorylation. A mechanism-based model of the cytochrome c oxidase reaction is presented in which transfer of reducing equivalents from the lower- to the higher-potential region of the coupling site occurs against an opposing energy barrier, Q. The steady-state rate equation is fitted to data for the dependence of mitochondrial respiratory rate on cytochrome c reduction, oxygen pressure (pO2), and [ATP]/[ADP][Pi] at pH 6.5 to 8.35 (where Pi is inorganic phosphate). The fit of the rate expression to the experimental data is very good for all experimental conditions. Levels of the intermediates in oxygen reduction in the oxidase reaction site have been calculated. An intermediate in the reaction, tentatively identified as peroxide, bridged between the iron and copper atoms of the reaction site has a central role in coupling mitochondrial respiration to the [ATP]/[ADP][Pi].

  19. Isotope-reinforced polyunsaturated fatty acids protect mitochondria from oxidative stress.

    PubMed

    Andreyev, Alexander Y; Tsui, Hui S; Milne, Ginger L; Shmanai, Vadim V; Bekish, Andrei V; Fomich, Maksim A; Pham, Minhhan N; Nong, Yvonne; Murphy, Anne N; Clarke, Catherine F; Shchepinov, Mikhail S

    2015-05-01

    Polyunsaturated fatty acid (PUFA) peroxidation is initiated by hydrogen atom abstraction at bis-allylic sites and sets in motion a chain reaction that generates multiple toxic products associated with numerous disorders. Replacement of bis-allylic hydrogens of PUFAs with deuterium atoms (D-PUFAs), termed site-specific isotope reinforcement, inhibits PUFA peroxidation and confers cell protection against oxidative stress. We demonstrate that structurally diverse deuterated PUFAs similarly protect against oxidative stress-induced injury in both yeast and mammalian (myoblast H9C2) cells. Cell protection occurs specifically at the lipid peroxidation step, as the formation of isoprostanes, immediate products of lipid peroxidation, is drastically suppressed by D-PUFAs. Mitochondrial bioenergetics function is a likely downstream target of oxidative stress and a subject of protection by D-PUFAs. Pretreatment of cells with D-PUFAs is shown to prevent inhibition of maximal uncoupler-stimulated respiration as well as increased mitochondrial uncoupling, in response to oxidative stress induced by agents with diverse mechanisms of action, including t-butylhydroperoxide, ethacrynic acid, or ferrous iron. Analysis of structure-activity relationships of PUFAs harboring deuterium at distinct sites suggests that there may be a mechanism supplementary to the kinetic isotope effect of deuterium abstraction off the bis-allylic sites that accounts for the protection rendered by deuteration of PUFAs. Paradoxically, PUFAs with partially deuterated bis-allylic positions that retain vulnerable hydrogen atoms (e.g., monodeuterated 11-D1-Lin) protect in a manner similar to that of PUFAs with completely deuterated bis-allylic positions (e.g., 11,11-D2-Lin). Moreover, inclusion of just a fraction of deuterated PUFAs (20-50%) in the total pool of PUFAs preserves mitochondrial respiratory function and confers cell protection. The results indicate that the therapeutic potential of D-PUFAs may derive

  20. Dexmedetomidine attenuates lipopolysaccharide-induced acute lung injury by inhibiting oxidative stress, mitochondrial dysfunction and apoptosis in rats.

    PubMed

    Fu, Chunlai; Dai, Xingui; Yang, You; Lin, Mengxiang; Cai, Yeping; Cai, Shaoxi

    2017-01-01

    Previous studies have identified that dexmedetomidine (DEX) treatment can ameliorate the acute lung injury (ALI) induced by lipopolysaccharide and ischemia-reperfusion. However, the molecular mechanisms by which DEX ameliorates lung injury remain unclear. The present study investigated whether DEX, which has been reported to exert effects on oxidative stress, mitochondrial permeability transition pores and apoptosis in other disease types, can exert protective effects in lipopolysaccharide (LPS)‑induced ALI by inhibiting oxidative stress, mitochondrial dysfunction and mitochondrial‑dependent apoptosis. It was revealed that LPS‑challenged rats exhibited significant lung injury, characterized by the deterioration of histopathology, vascular hyperpermeability, wet‑to‑dry weight ratio and oxygenation index (PaO2/FIO2), which was attenuated by DEX treatment. DEX treatment inhibited LPS‑induced mitochondrial dysfunction, as evidenced by alleviating the cellular ATP and mitochondrial membrane potential in vitro. In addition, DEX treatment markedly prevented the LPS‑induced mitochondrial‑dependent apoptotic pathway in vitro (increases of cell apoptotic rate, cytosolic cytochrome c, and caspase 3 activity) and in vivo (increases of |terminal deoxynucleotidyl transferase dUTP nick‑end labeling positive cells, cleaved caspase 3, Bax upregulation and Bcl‑2 downregulation). Furthermore, DEX treatment markedly attenuated LPS‑induced oxidative stress, as evidenced by downregulation of cellular reactive oxygen species in vitro and lipid peroxides in serum. Collectively, the present results demonstrated that DEX ameliorates LPS‑induced ALI by reducing oxidative stress, mitochondrial dysfunction and mitochondrial-dependent apoptosis.

  1. Import of desired nucleic acid sequences using addressing motif of mitochondrial ribosomal 5S-rRNA for fluorescent in vivo hybridization of mitochondrial DNA and RNA.

    PubMed

    Zelenka, Jaroslav; Alán, Lukáš; Jabůrek, Martin; Ježek, Petr

    2014-04-01

    Based on the matrix-addressing sequence of mitochondrial ribosomal 5S-rRNA (termed MAM), which is naturally imported into mitochondria, we have constructed an import system for in vivo targeting of mitochondrial DNA (mtDNA) or mt-mRNA, in order to provide fluorescence hybridization of the desired sequences. Thus DNA oligonucleotides were constructed, containing the 5'-flanked T7 RNA polymerase promoter. After in vitro transcription and fluorescent labeling with Alexa Fluor(®) 488 or 647 dye, we obtained the fluorescent "L-ND5 probe" containing MAM and exemplar cargo, i.e., annealing sequence to a short portion of ND5 mRNA and to the light-strand mtDNA complementary to the heavy strand nd5 mt gene (5'-end 21 base pair sequence). For mitochondrial in vivo fluorescent hybridization, HepG2 cells were treated with dequalinium micelles, containing the fluorescent probes, bringing the probes proximally to the mitochondrial outer membrane and to the natural import system. A verification of import into the mitochondrial matrix of cultured HepG2 cells was provided by confocal microscopy colocalizations. Transfections using lipofectamine or probes without 5S-rRNA addressing MAM sequence or with MAM only were ineffective. Alternatively, the same DNA oligonucleotides with 5'-CACC overhang (substituting T7 promoter) were transcribed from the tetracycline-inducible pENTRH1/TO vector in human embryonic kidney T-REx®-293 cells, while mitochondrial matrix localization after import of the resulting unlabeled RNA was detected by PCR. The MAM-containing probe was then enriched by three-order of magnitude over the natural ND5 mRNA in the mitochondrial matrix. In conclusion, we present a proof-of-principle for mitochondrial in vivo hybridization and mitochondrial nucleic acid import.

  2. Mitochondrial-Derived Oxidants and Cellular Responses to Low Dose/Low LET Ionizing Radiation

    SciTech Connect

    Spitz, Douglas R.

    2009-11-09

    Exposure to ionizing radiation results in the immediate formation of free radicals and other reactive oxygen species (ROS). It has been assumed that the subsequent injury processes leading to genomic instability and carcinogenesis following radiation, derive from the initial oxidative damage caused by these free radicals and ROS. It is now becoming increasingly obvious that metabolic oxidation/reduction (redox) reactions can be altered by irradiation leading to persistent increases in steady-state levels of intracellular free radicals and ROS that contribute to the long term biological effects of radiation exposure by causing chronic oxidative stress. The objective during the last period of support (DE-FG02-05ER64050; 5/15/05-12/31/09) was to determine the involvement of mitochondrial genetic defects in metabolic oxidative stress and the biological effects of low dose/low LET radiation. Aim 1 was to determine if cells with mutations in succinate dehydrogenase (SDH) subunits C and D (SDHC and SDHD in mitochondrial complex II) demonstrated increases in steady-state levels of reactive oxygen species (ROS; O2•- and H2O2) as well as demonstrating increased sensitivity to low dose/low LET radiation (10 cGy) in cultured mammalian cells. Aim #2 was to determine if mitochondrially-derived ROS contributed to increased sensitivity to low dose/low LET radiation in mammalian cells containing mutations in SDH subunits. Aim #3 was to determine if a causal relationship existed between increases in mitochondrial ROS production, alterations in electron transport chain proteins, and genomic instability in the progeny of irradiated cells. Evidence gathered in the 2005-2009 period of support demonstrated that mutations in genes coding for mitochondrial electron transport chain proteins (ETC); either Succinate Dehydrogenase (SDH) subunit C (SDHC) or subunit D (SDHD); caused increased ROS production, increased genomic instability, and increased sensitivity to low dose/low LET radiation

  3. PPARα augments heart function and cardiac fatty acid oxidation in early experimental polymicrobial sepsis.

    PubMed

    Standage, Stephen W; Bennion, Brock G; Knowles, Taft O; Ledee, Dolena R; Portman, Michael A; McGuire, John K; Liles, W Conrad; Olson, Aaron K

    2017-02-01

    Children with sepsis and multisystem organ failure have downregulated leukocyte gene expression of peroxisome proliferator-activated receptor-α (PPARα), a nuclear hormone receptor transcription factor that regulates inflammation and lipid metabolism. Mouse models of sepsis have likewise demonstrated that the absence of PPARα is associated with decreased survival and organ injury, specifically of the heart. Using a clinically relevant mouse model of early sepsis, we found that heart function increases in wild-type (WT) mice over the first 24 h of sepsis, but that mice lacking PPARα (Ppara(-/-)) cannot sustain the elevated heart function necessary to compensate for sepsis pathophysiology. Left ventricular shortening fraction, measured 24 h after initiation of sepsis by echocardiography, was higher in WT mice than in Ppara(-/-) mice. Ex vivo working heart studies demonstrated greater developed pressure, contractility, and aortic outflow in WT compared with Ppara(-/-) mice. Furthermore, cardiac fatty acid oxidation was increased in WT but not in Ppara(-/-) mice. Regulatory pathways controlling pyruvate incorporation into the citric acid cycle were inhibited by sepsis in both genotypes, but the regulatory state of enzymes controlling fatty acid oxidation appeared to be permissive in WT mice only. Mitochondrial ultrastructure was not altered in either genotype indicating that severe mitochondrial dysfunction is unlikely at this stage of sepsis. These data suggest that PPARα expression supports the hyperdynamic cardiac response early in the course of sepsis and that increased fatty acid oxidation may prevent morbidity and mortality.

  4. NOX4 NADPH Oxidase-Dependent Mitochondrial Oxidative Stress in Aging-Associated Cardiovascular Disease

    PubMed Central

    Vendrov, Aleksandr E.; Vendrov, Kimberly C.; Smith, Alberto; Yuan, Jinling; Sumida, Arihiro; Robidoux, Jacques; Madamanchi, Nageswara R.

    2015-01-01

    Abstract Aims: Increased oxidative stress and vascular inflammation are implicated in increased cardiovascular disease (CVD) incidence with age. We and others demonstrated that NOX1/2 NADPH oxidase inhibition, by genetic deletion of p47phox, in Apoe−/− mice decreases vascular reactive oxygen species (ROS) generation and atherosclerosis in young age. The present study examined whether NOX1/2 NADPH oxidases are also pivotal to aging-associated CVD. Results: Both aged (16 months) Apoe−/− and Apoe−/−/p47phox−/− mice had increased atherosclerotic lesion area, aortic stiffness, and systolic dysfunction compared with young (4 months) cohorts. Cellular and mitochondrial ROS (mtROS) levels were significantly higher in aortic wall and vascular smooth muscle cells (VSMCs) from aged wild-type and p47phox−/− mice. VSMCs from aged mice had increased mitochondrial protein oxidation and dysfunction and increased vascular cell adhesion molecule 1 expression, which was abrogated with (2-(2,2,6,6-Tetramethylpiperidin-1-oxyl-4-ylamino)-2-oxoethyl)triphenylphosphonium chloride (MitoTEMPO) treatment. NOX4 expression was increased in the vasculature and mitochondria of aged mice and its suppression with shRNA in VSMCs from aged mice decreased mtROS levels and improved function. Increased mtROS levels were associated with enhanced mitochondrial NOX4 expression in aortic VSMCs from aged subjects, and NOX4 expression levels in arterial wall correlated with age and atherosclerotic severity. Aged Apoe−/− mice treated with MitoTEMPO and 2-(2-chlorophenyl)-4-methyl-5-(pyridin-2-ylmethyl)-1H-pyrazolo[4,3-c]pyridine-3,6(2H,5H)-dione had decreased vascular ROS levels and atherosclerosis and preserved vascular and cardiac function. Innovation and Conclusion: These data suggest that NOX4, but not NOX1/2, and mitochondrial oxidative stress are mediators of CVD in aging under hyperlipidemic conditions. Regulating NOX4 activity/expression and using mitochondrial antioxidants are

  5. Impaired translocation and activation of mitochondrial Akt1 mitigated mitochondrial oxidative phosphorylation Complex V activity in diabetic myocardium.

    PubMed

    Yang, Jia-Ying; Deng, Wu; Chen, Yumay; Fan, Weiwei; Baldwin, Kenneth M; Jope, Richard S; Wallace, Douglas C; Wang, Ping H

    2013-06-01

    Insulin can translocate Akt to mitochondria in cardiac muscle. The goals of this study were to define sub-mitochondrial localization of the translocated Akt, to dissect the effects of insulin on Akt isoform translocation, and to determine the direct effect of mitochondrial Akt activation on Complex V activity in normal and diabetic myocardium. The translocated Akt sequentially localized to the mitochondrial intermembrane space, inner membrane, and matrix. To confirm Akt translocation, in vitro import assay showed rapid entry of Akt into mitochondria. Akt isoforms were differentially regulated by insulin stimulation, only Akt1 translocated into mitochondria. In the insulin-resistant Type 2 diabetes model, Akt1 translocation was blunted. Mitochondrial activation of Akt1 increased Complex V activity by 24% in normal myocardium in vivo and restored Complex V activity in diabetic myocardium. Basal mitochondrial Complex V activity was lower by 22% in the Akt1(-/-) myocardium. Insulin-stimulated Complex V activity was not impaired in the Akt1(-/-) myocardium, due to compensatory translocation of Akt2 to mitochondria. Akt1 is the primary isoform that relayed insulin signaling to mitochondria and modulated mitochondrial Complex V activity. Activation of mitochondrial Akt1 enhanced ATP production and increased phosphocreatine in cardiac muscle cells. Dysregulation of this signal pathway might impair mitochondrial bioenergetics in diabetic myocardium.

  6. Iontophoresis with gold nanoparticles improves mitochondrial activity and oxidative stress markers of burn wounds.

    PubMed

    Silveira, Paulo C L; Venâncio, Mirelli; Souza, Priscila S; Victor, Eduardo G; de Souza Notoya, Frederico; Paganini, Carla S; Streck, Emilio L; da Silva, Luciano; Pinho, Ricardo A; Paula, Marcos M S

    2014-11-01

    The aim of this study was to analyse the effects of microcurrent and gold nanoparticles on oxidative stress parameters and the mitochondrial respiratory chain in the healing of skin wounds. Thirty 60-day old male Wistar rats (250-300 g) were divided into five groups (N=6): Control; Burn wounds; Microcurrent (MIC); Gold nanoparticle gel (GNP gel) and Microcurrent+Gold nanoparticle gel (MIC+GNP gel). The microcurrent treatment was applied for five consecutive days at a dose of 300 μA. The results demonstrate a significant decrease in the activity of complexes I, II-III and IV in the Burn Wounds group compared to the control, and the MIC+GNP gel group was able to reverse this inhibition in complexes I, III and IV. Furthermore, a significant reduction in oxidative damage parameters and a significant increase in the levels of antioxidant defence enzymes were induced in the MIC+GNP gel group compared to the Burn Wounds group. The data strongly indicate that the group receiving treatment with MIC+GNP gel had improved mitochondrial functioning and oxidative stress parameters, which contributed to tissue repair.

  7. Cerium oxide nanoparticles prevent apoptosis in primary cortical culture by stabilizing mitochondrial membrane potential.

    PubMed

    Arya, A; Sethy, N K; Das, M; Singh, S K; Das, A; Ujjain, S K; Sharma, R K; Sharma, M; Bhargava, K

    2014-07-01

    Cerium oxide nanoparticles (CNPs) of spherical shape have unique antioxidant capacity primarily due to alternating + 3 and + 4 oxidation states and crystal defects. Several studies revealed the protective efficacies of CNPs in cells and tissues against the oxidative damage. However, its effect on mitochondrial functioning, downstream effectors of radical burst and apoptosis remains unknown. In this study, we investigated whether CNPs treatment could protect the primary cortical cells from loss of mitochondrial membrane potential (Δψm) and Δψm-dependent cell death. CNPs with spherical morphology and size range 7-10 nm were synthesized and utilized at a concentration of 25 nM on primary neuronal culture challenged with 50 μM of hydrogen peroxide (H2O2). We showed that optimal dose of CNPs minimized ROS content of the cells and also curbed related surge in cellular calcium flux. Importantly, CNPs treatment prevented apoptotic loss of cell viability. Reduction in the apoptosis could be successfully attributed to the maintenance of Δψm and restoration of major redox equivalents NADH/NAD(+) ratio and cellular ATP. These findings, therefore, suggest possible route of CNPs protective efficacies in primary cortical culture.

  8. Sources of superoxide/H2O2 during mitochondrial proline oxidation.

    PubMed

    Goncalves, Renata L S; Rothschild, Daniel E; Quinlan, Casey L; Scott, Gary K; Benz, Christopher C; Brand, Martin D

    2014-01-01

    p53 Inducible gene 6 (PIG6) encodes mitochondrial proline dehydrogenase (PRODH) and is up-regulated several fold upon p53 activation. Proline dehydrogenase is proposed to generate radicals that contribute to cancer cell apoptosis. However, there are at least 10 mitochondrial sites that can produce superoxide and/or H2O2, and it is unclear whether proline dehydrogenase generates these species directly, or instead drives production by other sites. Amongst six cancer cell lines, ZR75-30 human breast cancer cells had the highest basal proline dehydrogenase levels, and mitochondria isolated from ZR75-30 cells consumed oxygen and produced H2O2 with proline as sole substrate. Insects use proline oxidation to fuel flight, and mitochondria isolated from Drosophila melanogaster were even more active with proline as sole substrate than ZR75-30 mitochondria. Using mitochondria from these two models we identified the sites involved in formation of superoxide/H2O2 during proline oxidation. In mitochondria from Drosophila the main sites were respiratory complexes I and II. In mitochondria from ZR75-30 breast cancer cells the main sites were complex I and the oxoglutarate dehydrogenase complex. Even with combinations of substrates and respiratory chain inhibitors designed to minimize the contributions of other sites and maximize any superoxide/H2O2 production from proline dehydrogenase itself, there was no significant direct contribution of proline dehydrogenase to the observed H2O2 production. Thus proline oxidation by proline dehydrogenase drives superoxide/H2O2 production, but it does so mainly or exclusively by providing anaplerotic carbon for other mitochondrial dehydrogenases and not by producing superoxide/H2O2 directly.

  9. Hydroxytyrosol ameliorates oxidative stress and mitochondrial dysfunction in doxorubicin-induced cardiotoxicity in rats with breast cancer.

    PubMed

    Granados-Principal, Sergio; El-Azem, Nuri; Pamplona, Reinald; Ramirez-Tortosa, Cesar; Pulido-Moran, Mario; Vera-Ramirez, Laura; Quiles, Jose L; Sanchez-Rovira, Pedro; Naudí, Alba; Portero-Otin, Manuel; Perez-Lopez, Patricia; Ramirez-Tortosa, Mcarmen

    2014-07-01

    Oxidative stress is involved in several processes including cancer, aging and cardiovascular disease, and has been shown to potentiate the therapeutic effect of drugs such as doxorubicin. Doxorubicin causes significant cardiotoxicity characterized by marked increases in oxidative stress and mitochondrial dysfunction. Herein, we investigate whether doxorubicin-associated chronic cardiac toxicity can be ameliorated with the antioxidant hydroxytyrosol in rats with breast cancer. Thirty-six rats bearing breast tumors induced chemically were divided into 4 groups: control, hydroxytyrosol (0.5mg/kg, 5days/week), doxorubicin (1mg/kg/week), and doxorubicin plus hydroxytyrosol. Cardiac disturbances at the cellular and mitochondrial level, mitochondrial electron transport chain complexes I-IV and apoptosis-inducing factor, and oxidative stress markers have been analyzed. Hydroxytyrosol improved the cardiac disturbances enhanced by doxorubicin by significantly reducing the percentage of altered mitochondria and oxidative damage. These results suggest that hydroxytyrosol improve the mitochondrial electron transport chain. This study demonstrates that hydroxytyrosol protect rat heart damage provoked by doxorubicin decreasing oxidative damage and mitochondrial alterations.

  10. Enhanced oxidative stress and aberrant mitochondrial biogenesis in human neuroblastoma SH-SY5Y cells during methamphetamine induced apoptosis

    SciTech Connect

    Wu, C.-W.; Ping, Y.-H.; Yen, J.-C.; Chang, C.-Y.; Wang, S.-F.; Yeh, C.-L.; Chi, C.-W.; Lee, H.-C. . E-mail: hclee2@ym.edu.tw

    2007-05-01

    Methamphetamine (METH) is an abused drug that may cause psychiatric and neurotoxic damage, including degeneration of monoaminergic terminals and apoptosis of non-monoaminergic cells in Brain. The cellular and molecular mechanisms underlying these METH-induced neurotoxic effects remain to be clarified. In this study, we performed a time course assessment to investigate the effects of METH on intracellular oxidative stress and mitochondrial alterations in a human dopaminergic neuroblastoma SH-SY5Y cell line. We characterized that METH induces a temporal sequence of several cellular events including, firstly, a decrease in mitochondrial membrane potential within 1 h of the METH treatment, secondly, an extensive decline in mitochondrial membrane potential and increase in the level of reactive oxygen species (ROS) after 8 h of the treatment, thirdly, an increase in mitochondrial mass after the drug treatment for 24 h, and finally, a decrease in mtDNA copy number and mitochondrial proteins per mitochondrion as well as the occurrence of apoptosis after 48 h of the treatment. Importantly, vitamin E attenuated the METH-induced increases in intracellular ROS level and mitochondrial mass, and prevented METH-induced cell death. Our observations suggest that enhanced oxidative stress and aberrant mitochondrial biogenesis may play critical roles in METH-induced neurotoxic effects.

  11. Expression of mitochondrial branched-chain aminotransferase and α-keto-acid dehydrogenase in rat brain: implications for neurotransmitter metabolism

    PubMed Central

    Cole, Jeffrey T.; Sweatt, Andrew J.; Hutson, Susan M.

    2012-01-01

    In the brain, metabolism of the essential branched chain amino acids (BCAAs) leucine, isoleucine, and valine, is regulated in part by protein synthesis requirements. Excess BCAAs are catabolized or excreted. The first step in BCAA catabolism is catalyzed by the branched chain aminotransferase (BCAT) isozymes, mitochondrial BCATm and cytosolic BCATc. A product of this reaction, glutamate, is the major excitatory neurotransmitter and precursor of the major inhibitory neurotransmitter γ-aminobutyric acid (GABA). The BCATs are thought to participate in a α-keto-acid nitrogen shuttle that provides nitrogen for synthesis of glutamate from α-ketoglutarate. The branched-chain α-keto acid dehydrogenase enzyme complex (BCKDC) catalyzes the second, irreversible step in BCAA metabolism, which is oxidative decarboxylation of the branched-chain α-keto acid (BCKA) products of the BCAT reaction. Maple Syrup Urine Disease (MSUD) results from genetic defects in BCKDC, which leads to accumulation of toxic levels of BCAAs and BCKAs that result in brain swelling. Immunolocalization of BCATm and BCKDC in rats revealed that BCATm is present in astrocytes in white matter and in neuropil, while BCKDC is expressed only in neurons. BCATm appears uniformly distributed in astrocyte cell bodies throughout the brain. The segregation of BCATm to astrocytes and BCKDC to neurons provides further support for the existence of a BCAA-dependent glial-neuronal nitrogen shuttle since the data show that BCKAs produced by glial BCATm must be exported to neurons. Additionally, the neuronal localization of BCKDC suggests that MSUD is a neuronal defect involving insufficient oxidation of BCKAs, with secondary effects extending beyond the neuron. PMID:22654736

  12. Reduction of Brain Mitochondrial β-Oxidation Impairs Complex I and V in Chronic Alcohol Intake: The Underlying Mechanism for Neurodegeneration

    PubMed Central

    Haorah, James; Rump, Travis J.; Xiong, Huangui

    2013-01-01

    Neuropathy and neurocognitive deficits are common among chronic alcohol users, which are believed to be associated with mitochondrial dysfunction in the brain. The specific type of brain mitochondrial respiratory chain complexes (mRCC) that are adversely affected by alcohol abuse has not been studied. Thus, we examined the alterations of mRCC in freshly isolated mitochondria from mice brain that were pair-fed the ethanol (4% v/v) and control liquid diets for 7–8 weeks. We observed that alcohol intake severely reduced the levels of complex I and V. A reduction in complex I was associated with a decrease in carnitine palmitoyltransferase 1 (cPT1) and cPT2 levels. The mitochondrial outer (cPT1) and inner (cPT2) membrane transporter enzymes are specialized in acylation of fatty acid from outer to inner membrane of mitochondria for ATP production. Thus, our results showed that alterations of cPT1 and cPT2 paralleled a decrease β-oxidation of palmitate and ATP production, suggesting that impairment of substrate entry step (complex I function) can cause a negative impact on ATP production (complex V function). Disruption of cPT1/cPT2 was accompanied by an increase in cytochrome C leakage, while reduction of complex I and V paralleled a decrease in depolarization of mitochondrial membrane potential (ΔΨ, monitored by JC-1 fluorescence) and ATP production in alcohol intake. We noted that acetyl-L-carnitine (ALC, a cofactor of cPT1 and cPT2) prevented the adverse effects of alcohol while coenzyme Q10 (CoQ10) was not very effective against alcohol insults. These results suggest that understanding the molecular, biochemical, and signaling mechanisms of the CNS mitochondrial β-oxidation such as ALC can mitigate alcohol related neurological disorders. PMID:23967116

  13. Phosphocreatine protects against LPS-induced human umbilical vein endothelial cell apoptosis by regulating mitochondrial oxidative phosphorylation.

    PubMed

    Sun, Zhengwu; Lan, Xiaoyan; Ahsan, Anil; Xi, Yalin; Liu, Shumin; Zhang, Zonghui; Chu, Peng; Song, Yushu; Piao, Fengyuan; Peng, Jinyong; Lin, Yuan; Han, Guozhu; Tang, Zeyao

    2016-03-01

    Phosphocreatine (PCr) is an exogenous energy substance, which provides phosphate groups for adenosine triphosphate (ATP) cycle and promotes energy metabolism in cells. However, it is still unclear whether PCr has influenced on mitochondrial energy metabolism as well as oxidative phosphorylation (OXPHO) in previous studies. Therefore, the aim of the present study was to investigate the regulation of PCr on lipopolsaccharide (LPS)-induced human umbilical vein endothelial cells (HUVECs) and mitochondrial OXPHO pathway. PCr protected HUVECs against LPS-induced apoptosis by suppressing the mitochondrial permeability transition, cytosolic release of cytochrome c (Cyt C), Ca(2+), reactive oxygen species and subsequent activation of caspases, and increasing Bcl2 expression, while suppressing Bax expression. More importantly, PCr significantly improved mitochondrial swelling and membrane potential, enhanced the activities of ATP synthase and mitochondrial creatine kinase (CKmt) in creatine shuttle, influenced on respiratory chain enzymes, respiratory control ratio, phosphorus/oxygen ratio and ATP production of OXPHO. Above PCr-mediated mitochondrial events were effectively more favorable to reduced form of flavin adenine dinucleotide (FADH2) pathway than reduced form of nicotinamide-adenine dinucleotid pathway in the mitochondrial respiratory chain. Our results revealed that PCr protects against LPS-induced HUVECs apoptosis, which probably related to stabilization of intracellular energy metabolism, especially for FADH2 pathway in mitochondrial respiratory chain, ATP synthase and CKmt. Our findings suggest that PCr may play a certain role in the treatment of atherosclerosis via protecting endothelial cell function.

  14. A Computational Screen for Regulators of Oxidative Phosphorylation Implicates SLIRP in Mitochondrial RNA Homeostasis

    PubMed Central

    Baughman, Joshua M.; Nilsson, Roland; Gohil, Vishal M.; Arlow, Daniel H.; Gauhar, Zareen; Mootha, Vamsi K.

    2009-01-01

    The human oxidative phosphorylation (OxPhos) system consists of approximately 90 proteins encoded by nuclear and mitochondrial genomes and serves as the primary cellular pathway for ATP biosynthesis. While the core protein machinery for OxPhos is well characterized, many of its assembly, maturation, and regulatory factors remain unknown. We exploited the tight transcriptional control of the genes encoding the core OxPhos machinery to identify novel regulators. We developed a computational procedure, which we call expression screening, which integrates information from thousands of microarray data sets in a principled manner to identify genes that are consistently co-expressed with a target pathway across biological contexts. We applied expression screening to predict dozens of novel regulators of OxPhos. For two candidate genes, CHCHD2 and SLIRP, we show that silencing with RNAi results in destabilization of OxPhos complexes and a marked loss of OxPhos enzymatic activity. Moreover, we show that SLIRP plays an essential role in maintaining mitochondrial-localized mRNA transcripts that encode OxPhos protein subunits. Our findings provide a catalogue of potential novel OxPhos regulators that advance our understanding of the coordination between nuclear and mitochondrial genomes for the regulation of cellular energy metabolism. PMID:19680543

  15. The transcriptional coregulator PGC-1β controls mitochondrial function and anti-oxidant defence in skeletal muscles

    PubMed Central

    Gali Ramamoorthy, Thanuja; Laverny, Gilles; Schlagowski, Anna-Isabel; Zoll, Joffrey; Messaddeq, Nadia; Bornert, Jean-Marc; Panza, Salvatore; Ferry, Arnaud; Geny, Bernard; Metzger, Daniel

    2015-01-01

    The transcriptional coregulators PGC-1α and PGC-1β modulate the expression of numerous partially overlapping genes involved in mitochondrial biogenesis and energetic metabolism. The physiological role of PGC-1β is poorly understood in skeletal muscle, a tissue of high mitochondrial content to produce ATP levels required for sustained contractions. Here we determine the physiological role of PGC-1β in skeletal muscle using mice, in which PGC-1β is selectively ablated in skeletal myofibres at adulthood (PGC-1β(i)skm−/− mice). We show that myofibre myosin heavy chain composition and mitochondrial number, muscle strength and glucose homeostasis are unaffected in PGC-1β(i)skm−/− mice. However, decreased expression of genes controlling mitochondrial protein import, translational machinery and energy metabolism in PGC-1β(i)skm−/− muscles leads to mitochondrial structural and functional abnormalities, impaired muscle oxidative capacity and reduced exercise performance. Moreover, enhanced free-radical leak and reduced expression of the mitochondrial anti-oxidant enzyme Sod2 increase muscle oxidative stress. PGC-1β is therefore instrumental for skeletal muscles to cope with high energetic demands. PMID:26674215

  16. Mitochondrial oxidative stress caused by Sod2 deficiency promotes cellular senescence and aging phenotypes in the skin.

    PubMed

    Velarde, Michael C; Flynn, James M; Day, Nicholas U; Melov, Simon; Campisi, Judith

    2012-01-01

    Cellular senescence arrests the proliferation of mammalian cells at risk for neoplastic transformation, and is also associated with aging. However, the factors that cause cellular senescence during aging are unclear. Excessive reactive oxygen species (ROS) have been shown to cause cellular senescence in culture, and accumulated molecular damage due to mitochondrial ROS has long been thought to drive aging phenotypesin vivo. Here, we test the hypothesis that mitochondrial oxidative stress can promote cellular senescence in vivo and contribute to aging phenotypes in vivo, specifically in the skin. We show that the number of senescent cells, as well as impaired mitochondrial (complex II) activity increase in naturally aged mouse skin. Using a mouse model of genetic Sod2 deficiency, we show that failure to express this important mitochondrial anti-oxidant enzyme also impairs mitochondrial complex II activity, causes nuclear DNA damage, and induces cellular senescence but not apoptosis in the epidermis. Sod2 deficiency also reduced the number of cells and thickness of the epidermis, while increasing terminal differentiation. Our results support the idea that mitochondrial oxidative stress and cellular senescence contribute to aging skin phenotypes in vivo.

  17. The transcriptional coregulator PGC-1β controls mitochondrial function and anti-oxidant defence in skeletal muscles.

    PubMed

    Gali Ramamoorthy, Thanuja; Laverny, Gilles; Schlagowski, Anna-Isabel; Zoll, Joffrey; Messaddeq, Nadia; Bornert, Jean-Marc; Panza, Salvatore; Ferry, Arnaud; Geny, Bernard; Metzger, Daniel

    2015-12-17

    The transcriptional coregulators PGC-1α and PGC-1β modulate the expression of numerous partially overlapping genes involved in mitochondrial biogenesis and energetic metabolism. The physiological role of PGC-1β is poorly understood in skeletal muscle, a tissue of high mitochondrial content to produce ATP levels required for sustained contractions. Here we determine the physiological role of PGC-1β in skeletal muscle using mice, in which PGC-1β is selectively ablated in skeletal myofibres at adulthood (PGC-1β((i)skm-/-) mice). We show that myofibre myosin heavy chain composition and mitochondrial number, muscle strength and glucose homeostasis are unaffected in PGC-1β((i)skm-/-) mice. However, decreased expression of genes controlling mitochondrial protein import, translational machinery and energy metabolism in PGC-1β((i)skm-/-) muscles leads to mitochondrial structural and functional abnormalities, impaired muscle oxidative capacity and reduced exercise performance. Moreover, enhanced free-radical leak and reduced expression of the mitochondrial anti-oxidant enzyme Sod2 increase muscle oxidative stress. PGC-1β is therefore instrumental for skeletal muscles to cope with high energetic demands.

  18. Oxidative stress is not a major contributor to somatic mitochondrial DNA mutations.

    PubMed

    Itsara, Leslie S; Kennedy, Scott R; Fox, Edward J; Yu, Selina; Hewitt, Joshua J; Sanchez-Contreras, Monica; Cardozo-Pelaez, Fernando; Pallanck, Leo J

    2014-02-01

    The accumulation of somatic mitochondrial DNA (mtDNA) mutations is implicated in aging and common diseases of the elderly, including cancer and neurodegenerative disease. However, the mechanisms that influence the frequency of somatic mtDNA mutations are poorly understood. To develop a simple invertebrate model system to address this matter, we used the Random Mutation Capture (RMC) assay to characterize the age-dependent frequency and distribution of mtDNA mutations in the fruit fly Drosophila melanogaster. Because oxidative stress is a major suspect in the age-dependent accumulation of somatic mtDNA mutations, we also used the RMC assay to explore the influence of oxidative stress on the somatic mtDNA mutation frequency. We found that many of the features associated with mtDNA mutations in vertebrates are conserved in Drosophila, including a comparable somatic mtDNA mutation frequency (∼10(-5)), an increased frequency of mtDNA mutations with age, and a prevalence of transition mutations. Only a small fraction of the mtDNA mutations detected in young or old animals were G∶C to T∶A transversions, a signature of oxidative damage, and loss-of-function mutations in the mitochondrial superoxide dismutase, Sod2, had no detectable influence on the somatic mtDNA mutation frequency. Moreover, a loss-of-function mutation in Ogg1, which encodes a DNA repair enzyme that removes oxidatively damaged deoxyguanosine residues (8-hydroxy-2'-deoxyguanosine), did not significantly influence the somatic mtDNA mutation frequency of Sod2 mutants. Together, these findings indicate that oxidative stress is not a major cause of somatic mtDNA mutations. Our data instead suggests that somatic mtDNA mutations arise primarily from errors that occur during mtDNA replication. Further studies using Drosophila should aid in the identification of factors that influence the frequency of somatic mtDNA mutations.

  19. CPT1{alpha} over-expression increases long-chain fatty acid oxidation and reduces cell viability with incremental palmitic acid concentration in 293T cells

    SciTech Connect

    Jambor de Sousa, Ulrike L.; Koss, Michael D.; Fillies, Marion; Gahl, Anja; Scheeder, Martin R.L.; Cardoso, M. Cristina; Leonhardt, Heinrich; Geary, Nori; Langhans, Wolfgang; Leonhardt, Monika . E-mail: monika.leonhardt@inw.agrl.ethz.ch

    2005-12-16

    To test the cellular response to an increased fatty acid oxidation, we generated a vector for an inducible expression of the rate-limiting enzyme carnitine palmitoyl-transferase 1{alpha} (CPT1{alpha}). Human embryonic 293T kidney cells were transiently transfected and expression of the CPT1{alpha} transgene in the tet-on vector was activated with doxycycline. Fatty acid oxidation was measured by determining the conversion of supplemented, synthetic cis-10-heptadecenoic acid (C17:1n-7) to C15:ln-7. CPT1{alpha} over-expression increased mitochondrial long-chain fatty acid oxidation about 6-fold. Addition of palmitic acid (PA) decreased viability of CPT1{alpha} over-expressing cells in a concentration-dependent manner. Both, PA and CPT1{alpha} over-expression increased cell death. Interestingly, PA reduced total cell number only in cells over-expressing CPT1{alpha}, suggesting an effect on cell proliferation that requires PA translocation across the mitochondrial inner membrane. This inducible expression system should be well suited to study the roles of CPT1 and fatty acid oxidation in lipotoxicity and metabolism in vivo.

  20. On the origin of 3-methylglutaconic acid in disorders of mitochondrial energy metabolism.

    PubMed

    Ikon, Nikita; Ryan, Robert O

    2016-09-01

    3-methylglutaconic acid (3MGA)-uria occurs in numerous inborn errors of metabolism (IEM) associated with compromised mitochondrial energy metabolism. This organic acid arises from thioester cleavage of 3-methylglutaconyl CoA (3MG CoA), an intermediate in leucine catabolism. In individuals harboring mutations in 3MG CoA hydratase (i.e., primary 3MGA-uria), dietary leucine is the source of 3MGA. In secondary 3MGA-uria, however, no leucine metabolism defects have been reported. While others have suggested 3MGA arises from aberrant isoprenoid shunting from cytosol to mitochondria, an alternative route posits that 3MG CoA arises in three steps from mitochondrial acetyl CoA. Support for this biosynthetic route in IEMs is seen by its regulated occurrence in microorganisms. The fungus, Ustilago maydis, the myxobacterium, Myxococcus xanthus and the marine cyanobacterium, Lyngbya majuscule, generate 3MG CoA (or acyl carrier protein derivative) in the biosynthesis of iron chelating siderophores, iso-odd chain fatty acids and polyketide/nonribosomal peptide products, respectively. The existence of this biosynthetic machinery in these organisms supports a model wherein, under conditions of mitochondrial dysfunction, accumulation of acetyl CoA in the inner mitochondrial space as a result of inefficient fuel utilization drives de novo synthesis of 3MG CoA. Since humans lack the downstream biosynthetic capability of the organisms mentioned above, as 3MG CoA levels rise, thioester hydrolysis yields 3MGA, which is excreted in urine as unspent fuel. Understanding the metabolic origins of 3MGA may increase its utility as a biomarker.

  1. Hyperglycemic switch from mitochondrial nitric oxide to superoxide production in endothelial cells.

    PubMed

    Brodsky, Sergey V; Gao, Shujuan; Li, Hong; Goligorsky, Michael S

    2002-11-01

    The accumulated ultrastructural and biochemical evidence is highly suggestive of the existence of mitochondrial nitric oxide (NO) synthase (mtNOS), where local production of NO regulates the electron transport along the respiratory chain. Here, the functional competence of mtNOS in situ in a living cell was examined using an intravital fluorescent NO indicator, 4,5-diaminofluorescein, employing a new procedure for loading it into the mitochondria to demonstrate local NO generation in undisrupted endothelial cells and in isolated mitochondria as well as in human embryonic kidney cells stably expressing endothelial NOS. With the use of this approach, we showed that endothelial cells incubated in the presence of high concentration of D-glucose (but not L-glucose) are characterized by the reduced NO synthetic function of mitochondria despite the unaltered abundance of the enzyme. In parallel, mitochondrial generation of superoxide was augmented in endothelial cells incubated in the presence of a high concentration of D-glucose. Both the NO generation and superoxide production in hyperglycemic environment could be restored to control levels by treating cells with a cell-permeable superoxide dismutase mimetic. In addition, enhanced mitochondrial superoxide production could be suppressed with an inhibitor of NOS in stimulated endothelial cells. In conclusion, the data 1) provide direct evidence of mitochondrial NO production in endothelial cells, 2) demonstrate its suppression and enhanced superoxide generation in hyperglycemic environment, and 3) provide evidence that "uncoupled" mtNOS represents an important source of superoxide anions in endothelial cells incubated in high glucose-containing medium.

  2. Pharmacologic inhibition of fatty acid oxidation sensitizes human leukemia cells to apoptosis induction

    PubMed Central

    Samudio, Ismael; Harmancey, Romain; Fiegl, Michael; Kantarjian, Hagop; Konopleva, Marina; Korchin, Borys; Kaluarachchi, Kumar; Bornmann, William; Duvvuri, Seshagiri; Taegtmeyer, Heinrich; Andreeff, Michael

    2009-01-01

    The traditional view is that cancer cells predominately produce ATP by glycolysis, rather than by oxidation of energy-providing substrates. Mitochondrial uncoupling — the continuing reduction of oxygen without ATP synthesis — has recently been shown in leukemia cells to circumvent the ability of oxygen to inhibit glycolysis, and may promote the metabolic preference for glycolysis by shifting from pyruvate oxidation to fatty acid oxidation (FAO). Here we have demonstrated that pharmacologic inhibition of FAO with etomoxir or ranolazine inhibited proliferation and sensitized human leukemia cells — cultured alone or on bone marrow stromal cells — to apoptosis induction by ABT-737, a molecule that releases proapoptotic Bcl-2 proteins such as Bak from antiapoptotic family members. Likewise, treatment with the fatty acid synthase/lipolysis inhibitor orlistat also sensitized leukemia cells to ABT-737, which supports the notion that fatty acids promote cell survival. Mechanistically, we generated evidence suggesting that FAO regulates the activity of Bak-dependent mitochondrial permeability transition. Importantly, etomoxir decreased the number of quiescent leukemia progenitor cells in approximately 50% of primary human acute myeloid leukemia samples and, when combined with either ABT-737 or cytosine arabinoside, provided substantial therapeutic benefit in a murine model of leukemia. The results support the concept of FAO inhibitors as a therapeutic strategy in hematological malignancies. PMID:20038799

  3. Dietary whey protein stimulates mitochondrial activity and decreases oxidative stress in mouse female brain.

    PubMed

    Shertzer, Howard G; Krishan, Mansi; Genter, Mary Beth

    2013-08-26

    In humans and experimental animals, protein-enriched diets are beneficial for weight management, muscle development, managing early stage insulin resistance and overall health. Previous studies have shown that in mice consuming a high fat diet, whey protein isolate (WPI) reduced hepatosteatosis and insulin resistance due in part to an increase in basal metabolic rate. In the current study, we examined the ability of WPI to increase energy metabolism in mouse brain. Female C57BL/6J mice were fed a normal AIN-93M diet for 12 weeks, with (WPI group) or without (Control group) 100g WPI/L drinking water. In WPI mice compared to controls, the oxidative stress biomarkers malondialdehyde and 4-hydroxyalkenals were 40% lower in brain homogenates, and the production of hydrogen peroxide and superoxide were 25-35% less in brain mitochondria. Brain mitochondria from WPI mice remained coupled, and exhibited higher rates of respiration with proportionately greater levels of cytochromes a+a3 and c+c1. These results suggested that WPI treatment increased the number or improved the function of brain mitochondria. qRT-PCR revealed that the gene encoding a master regulator of mitochondrial activity and biogenesis, Pgc-1alpha (peroxisome proliferator-activated receptor-gamma coactivator-1alpha) was elevated 2.2-fold, as were the PGC-1alpha downstream genes, Tfam (mitochondrial transcription factor A), Gabpa/Nrf-2a (GA-binding protein alpha/nuclear respiratory factor-2a), and Cox-6a1 (cytochrome oxidase-6a1). Each of these genes had twice the levels of transcript in brain tissue from WPI mice, relative to controls. There was no change in the expression of the housekeeping gene B2mg (beta-2 microglobulin). We conclude that dietary whey protein decreases oxidative stress and increases mitochondrial activity in mouse brain. Dietary supplementation with WPI may be a useful clinical intervention to treat conditions associated with oxidative stress or diminished mitochondrial activity in the

  4. Tetrahydrocannabinol induces brain mitochondrial respiratory chain dysfunction and increases oxidative stress: a potential mechanism involved in cannabis-related stroke.

    PubMed

    Wolff, Valérie; Schlagowski, Anna-Isabel; Rouyer, Olivier; Charles, Anne-Laure; Singh, François; Auger, Cyril; Schini-Kerth, Valérie; Marescaux, Christian; Raul, Jean-Sébastien; Zoll, Joffrey; Geny, Bernard

    2015-01-01

    Cannabis has potential therapeutic use but tetrahydrocannabinol (THC), its main psychoactive component, appears as a risk factor for ischemic stroke in young adults. We therefore evaluate the effects of THC on brain mitochondrial function and oxidative stress, key factors involved in stroke. Maximal oxidative capacities V max (complexes I, III, and IV activities), V succ (complexes II, III, and IV activities), V tmpd (complex IV activity), together with mitochondrial coupling (V max/V 0), were determined in control conditions and after exposure to THC in isolated mitochondria extracted from rat brain, using differential centrifugations. Oxidative stress was also assessed through hydrogen peroxide (H2O2) production, measured with Amplex Red. THC significantly decreased V max (-71%; P < 0.0001), V succ (-65%; P < 0.0001), and V tmpd (-3.5%; P < 0.001). Mitochondrial coupling (V max/V 0) was also significantly decreased after THC exposure (1.8±0.2 versus 6.3±0.7; P < 0.001). Furthermore, THC significantly enhanced H2O2 production by cerebral mitochondria (+171%; P < 0.05) and mitochondrial free radical leak was increased fr