Osiki 2016 Abstract MitoFit Science Camp 2016
|Suitability of octanoylcarnitine & malate for assessment of beta oxidation capacity by respirometry in aconitase-inhibited samples.|
The capacity of cells, tissues or isolated mitochondria to oxidize fatty acids is often measured by respirometry using acyl carnitines & malate as substrates . Inclusion of malate as a co-substrate is essential to prevent feed forward inhibition of beta oxidation arising from matrix CoA depletion as a result of short-chain acyl-CoA accumulation . Malate is dehydrogenated to oxaloacetate which condenses with the beta-oxidation-derived acetyl-CoA to form citrate and CoA. Therefore, as long as citrate does not accumulate in the mitochondrial matrix and inhibit the above series of reactions, beta oxidation reactions are likely to keep running. However, a number of disease conditions are known to inhibit mitochondrial aconitase, a mitochondrial enzyme that has profound effects on citrate metabolism [3,4].
The purpose of this study was therefore to investigate the validity of beta-oxidation assessment by respirometry using octanoylcarnitine & malate in rat soleus muscle when mitochondrial aconitase is chemically inhibited by 5 mM Oxalomalic acid. Our results indicate that inhibition of aconitase by 25% does not decrease oxygen flux at all respiratory states but increased citrate levels in the mitochondrial media by 40% compared to controls. However, as expected, oxygen fluxes were significantly diminished at OXPHOS and ET-pathway respiratory states by ~50% when medium chain acyl dehydrogenase, a rate limiting beta-oxidation enzyme, was inhibited by 1 mm 2-mercaptoacetate.
Taken together, our data indicate that in respirometric assays, use of octanoylcarnitine + malate allows for a valid assessment of beta-oxidation capacity in skeletal muscle under conditions where mitochondrial aconitase activity is compromised.
Labels: MiParea: Respiration
Organism: Rat Tissue;cell: Skeletal muscle
Regulation: Substrate Coupling state: OXPHOS, ET Pathway: F, N, Other combinations HRR: Oxygraph-2k Event: A1 MitoFit Science Camp 2016
Dept Human Biol, Univ Cape Town, South Africa. - [email protected]
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