Mitochondrial dysfunction in liver disease


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There are many causes of liver disease however the aetiology is often undetermined. It is speculated that mitochondrial dysfunction has a role in liver disease and this presentation explores possible causes and techniques for investigating mitochondrial dysfunction in liver.

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  • Translation – for example by in vivo metabolic 35S-methionine labelling of mitochondrial protein synthesisTranscription – for example by Northern blotting or PCR for mRNA
  • Mitochondrial dysfunction in liver disease

    1. 1. Maria G Lane
    2. 2. ◦ Mitochondria are tiny cellular organelles responsible for energy production via oxidative phosphorylation ◦ Complexes I-IV form the electron transport chain and create a proton gradient ◦ Complex V uses this gradient to make ATP from ADP
    3. 3.  The liver is a vital organ ◦ Has a high energy requirement ◦ Has a role in metabolism of macromolecules, protein biosynthesis and clearing of toxic compounds ◦ Highly dependent on ATP and functioning mitochondria ◦ Mitochondrial disease affects about 1 in 5,000 to 1 in 10,000 of the population and the liver can be affected in as many as 20% of these cases
    4. 4.  Many types of liver disease including alcoholic liver disease, hepatitis, acute liver failure  Symptoms are varied – lactic acidosis, hypoglycaemia, jaundice, anaemia, coagulopathy  Treatment is symptomatic  Transplantation is life saving in severe cases
    5. 5.  Often unknown  Drugs such as valproate  Viruses such as hepatitis B and C  However it is speculated that mitochondrial dysfunction has a role in liver disease  Several studies have associated liver disease with mitochondrial defects
    6. 6.  Defects in assembly of respiratory chain complexes ◦ BCS1L  Missense mutations  Chaperone for Rieske Fe-S subunit of Complex III  Hepatopathy, tubulopathy and encephalopathy ◦ SCO1  Copper insertion into Complex IV  Hepatopathy and ketoacidic coma
    7. 7.  Alpers’ Syndrome ◦ Polymerase gamma mutations ◦ Affects the liver and nervous system  DGUOK deficiency ◦ Deoxyguanosine kinase maintains a stable ribonucleotide pool for mtDNA synthesis ◦ Affects the liver and nervous system
    8. 8.  TSFM ◦ Encodes elongation factor EFTs ◦ A mutation causes a severe hepatopathy with a respiratory chain deficiency  TRMU ◦ Encodes a tRNA modifying enzyme ◦ Required for thiol addition to tRNAs for lysine, glutamate and glutamine ◦ A mutation causes acute liver failure with reduced activity of complexes I, III and IV
    9. 9.  Isolation of mitochondria from frozen liver tissue ◦ Homogenisation of tissue  Cut and washed  Homogenised in a glass Elvehjem potter ◦ Differential centrifugation  Sucrose gradient ◦ Addition of detergents and sample buffer containing aminocaproic acid for BN PAGE only
    10. 10.  Blue native polyacrylamide gel electrophoresis ◦ Native separation of protein complexes on a gradient gel ◦ Uses Coomassie G250  Attaches a negative charge to protein complexes  Doesn’t dissociate complexes into subunits ◦ Digitonin and n-dodecyl B-D-maltoside  Detergents  Break down mitochondrial membrane  Dissociate some supercomplexes ◦ Immunodetection of proteins with antibodies to complexes I-V
    11. 11. ◦ Biochemical assays of the catalytic activity of enzymes ◦ Complex II  reduction of ubiquinone at 595 nm ◦ Complex IV  oxidation of cytochrome c at 550 nm ◦ Citrate Synthase  formation of citrate from oxaloacetate at 405 nm  Indicator of number of functional mitochondria
    12. 12.  Confocal microscopy  Immunocytochemistry  Study mtDNA transcription or translation  Sequencing of genes commonly affected in mitochondrial disease e.g. TRMU, DGUOK, GFM1  Whole exome sequencing
    13. 13.  Chinnery PF, DiMauro S. Mitochondrial hepatopathies. Journal of hepatology. 2005 Aug; 43(2):207-9  Fernandez-Vizarra E, Ferrin G, Perez-Martos A, Fernandez-Silva P, Zeviani M, Enriquez JA. Isolation of mitochondria for biogenetical studies: An update. Mitochondrion. 2010 Apr; 10(3):253-62  Lee WS, Sokol RJ. Liver disease in mitochondrial disorders. Seminars in liver disease. 2007 Aug; 27(3):259-73  Rahman S. Gastrointestinal and hepatic manifestations of mitochondrial disorders. Journal of inherited metabolic disease. Epub 2013/05/16  Schagger H, von Jagow G. Blue native electrophoresis for isolation of membrane protein complexes in enzymatically active form. Analytical biochemistry. 1991 Dec; 199(2):223-31