Ethanol drinking, brain mitochondrial DNA, polyunsaturated fatty acids and effects of dietary anthocyanins

Background This study aimed at exploring whether moderate ethanol drinking may have adverse effects on the fatty acids composition and on mitochondrial DNA (mtDNA) of rat brain. A secondary aim was to examine whether dietary antioxidant anthocyanins (ACN) can be protective. Methods One group of rats received ethanol 12% and another water as an exclusive liquid to drink for 8 weeks. In order to test the impact of ACN consumption, two other groups of rats were fed an ACN-rich diet in combination with either ethanol or water. Brain fatty acids were measured by gas chromatography and mtDNA alterations, markers of mitochondrial suffering, were studied through an original real-time qPCR-based protocol. Results Linoleic acid (LA, 18:2n-6) and eicosadienoic acid (20:2n-6) were significantly decreased, by 12% and 31% respectively, in the brains of both ethanol groups. The other brain lipids, including arachidonic acid (20:4n-6) and n-3 polyunsaturated fatty acids, were not modified. These changes were associated with a significant increase in deleted mtDNA (by 28%) in the ethanol group, without total mtDNA depletion. The ACN-rich diet prevented the increase in mtDNA common deletion (mtDNA CD). Conclusion These data demonstrate that moderate ethanol drinking reduces certain brain n-6 and results in mtDNA injury. The antioxidant anthocyanins protect brain mtDNA but do not restore normal n-6 levels. Further studies are required to investigate the consequences of a decrease in n-6 levels in brain

le projet PUNAISES – Pour Une Nutrition et une Alimentation à base d’Insectes : quels enjeux Scienti-fiques, Economiques et Sociaux ?

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Escherichia coli exonuclease III enhances long PCR amplification of damaged DNA templates

International audienceRecent development of the long PCR technology has provided an invaluable tool in many areas of molecular biology. However, long PCR amplification fails whenever the DNA template is imperfectly preserved. We report that Escherichia coli exonuclease III, a major repair enzyme in bacteria, strikingly improves the long PCR amplification of damaged DNA templates. Escherichia coli exonuclease III permitted or improved long PCR amplification with DNA samples submitted to different in vitro treatments known to induce DNA strand breaks and/or apurinic/apyrimidinic (AP) sites, including high temperature (99 degrees C), depurination at low pH and near-UV radiation. Exonuclease III also permitted or improved amplification with DNA samples that had been isolated several years ago by the phenol/chloroform method. Amelioration of long PCR amplification was achieved for PCR products ranging in size from 5 to 15.4 kb and with DNA target sequences located either within mitochondrial DNA or the nuclear genome. Exonuclease III increased the amplification of damaged templates using either rTth DNA polymerase alone or rTth plus Vent DNA polymerases or TAQ: plus PWO: DNA polymerases. However, exonuclease III could not improve PCR amplification from extensively damaged DNA samples. In conclusion, supplementation of long PCR mixes with E.COLI: exonuclease III may represent a major technical advance whenever DNA samples have been partly damaged during isolation or subsequent storage

Acute ethanol administration oxidatively damages and depletes mitochondrial dna in mouse liver, brain, heart, and skeletal muscles: protective effects of antioxidants.

International audienceEthanol metabolism causes oxidative stress and lipid peroxidation not only in liver but also in extra-hepatic tissues. Ethanol administration has been shown to cause oxidative degradation and depletion of hepatic mitochondrial DNA (mtDNA) in rodents, but its in vivo effects on the mtDNA of extra-hepatic tissues have not been assessed. We studied the effects of an acute intragastric ethanol administration (5 g/kg) on brain, heart, skeletal muscle, and liver mtDNA in mice. Ethanol administration caused mtDNA depletion and replacement of its supercoiled form by linearized forms in all tissues examined. Maximal mtDNA depletion was about similar (ca. 50%) in all organs studied. It occurred 2 h after ethanol administration in heart, skeletal muscle, and liver but after 10 h in brain. This mtDNA depletion was followed by increased mtDNA synthesis. A secondary, transient increase in mtDNA levels occurred 24 h after ethanol administration in all organs. In hepatic or extra-hepatic tissues, mtDNA degradation and depletion were prevented by 4-methylpyrazole, an inhibitor of ethanol metabolism, and attenuated by vitamin E, melatonin, or coenzyme Q, three antioxidants. In conclusion, our study shows for the first time that ethanol metabolism also causes oxidative degradation of the mitochondrial genome in brain, heart, and skeletal muscles. These effects could contribute to the development of (cardio)myopathy and brain injury in some alcoholic patients. Antioxidants prevent these effects in mice and could be useful in persevering drinkers

Direct quantification of mitochondrial DNA and its 4.9-kb common deletion without DNA purification

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Effect of stavudine on mitochondrial genome and fatty acid oxidation in lean and obese mice.

International audienceLike other antihuman immunodeficiency virus dideoxynucleosides, stavudine may occasionally induce lactic acidosis and perhaps lipodystrophy in metabolically or genetically susceptible patients. We studied the effects of stavudine on mitochondrial DNA (mtDNA), fatty acid oxidation, and blood metabolites in lean and genetically obese (ob/ob) mice. In lean mice, mtDNA was depleted in liver and skeletal muscle, but not heart and brain, after 6 weeks of stavudine treatment (500 mg/kg/day). With 100 mg/kg/day, mtDNA transiently decreased in liver, but was unchanged at 6 weeks in all organs, including white adipose tissue (WAT). Despite unchanged mtDNA levels, lack of significant oxidative mtDNA lesions (as assessed by long polymerase chain reaction experiments), and normal blood lactate/pyruvate ratios, lean mice treated with stavudine for 6 weeks had increased fasting blood ketone bodies, due to both increased hepatic fatty acid beta-oxidation and decreased peripheral ketolysis. In obese mice, basal WAT mtDNA was low and was further decreased by stavudine. In conclusion, stavudine can decrease hepatic and muscle mtDNA in lean mice and can also cause ketoacidosis during fasting without altering mtDNA. Stavudine depletes WAT mtDNA only in obese mice. Fasting and ketoacidosis could trigger decompensation in patients with incipient lactic acidosis, whereas WAT mtDNA depletion could cause lipodystrophy in genetically susceptible patients