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ARS Home » Pacific West Area » Davis, California » Western Human Nutrition Research Center » Obesity and Metabolism Research » Research » Publications at this Location » Publication #237144
Title: Long-chain fatty acid combustion rate is associated with unique metabolite profiles in skeletal muscle mitochondria

Author
item SEIFERT, ERIN - UNIVERSITY OF OTTAWA
item FIEHN, OLIVER - UCD, GENOME CENTER
item BEZAIRE, VERONIC - UNIVERSITY OF OTTAWA
item BICKEL, DAVID - UNIVERSITY OF OTTAWA
item WOHLGEMUTH, GERT - UCD, GENOME CENTER
item Adams, Sean
item HARPER, MARY-ELLEN - UNIVERSITY OF OTTAWA

Submitted to: PLOS ONE
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 2/17/2010
Publication Date: 3/24/2010
Citation: Seifert, E.L., Fiehn, O., Bezaire, V., Bickel, D.B., Wohlgemuth, G., Adams, S.H., Harper, M. 2010. Long-chain fatty acid combustion rate is associated with unique metabolite profiles in skeletal muscle mitochondria. PLoS One. Volume 5(3) pp. 1-13, 2010.

Interpretive Summary: Incomplete or limited long-chain fatty acid (LCFA) combustion in skeletal muscle has been associated with insulin resistance. Signals that are responsive to shifts in LCFA beta-oxidation rate or degree of intramitochondrial catabolism are hypothesized to regulate second messenger systems downstream of the insulin receptor. Recent evidence supports a causal link between mitochondrial LCFA combustion in skeletal muscle and insulin resistance. We have used unbiased metabolite profiling of mouse muscle mitochondria with the aim of identifying candidate metabolites within or effluxed from mitochondria and that are shifted with LCFA combustion rate. Large-scale unbiased metabolomics analysis was performed using GC/TOF-MS on bufferand mitochondrial matrix fractions obtained prior to and after 20 min of palmitate catabolism (n=7 mice/condition). Three [palmitate] (2, 9 and 19 µM; corresponding to low, intermediate and high oxidation rates) and 9 µM palmitate plus tricarboxylic acid (TCA) cycle and electron transport chain inhibitors were each tested and compared to zero palmitate control incubations. Paired comparisons of the 0 and 20 min samples were made by Student’s t-test. False discovery rate were estimated and Type I error rates assigned. Major metabolite groups were organic acids, amines and amino acids, free fatty acids and sugar phosphates. Palmitate oxidation was associated with unique profiles of metabolites, a subset of which correlated to palmitate oxidation rate. In particular, palmitate oxidation rate was associated with distinct changes in the levels of TCA cycle intermediates within and effluxed from mitochondria. This proof-of-principle study establishes that large-scale metabolomics methods can be applied to organelle-level models to discover metabolite patterns reflective of LCFA combustion, which may lead to identification of molecules linking muscle fat metabolism and insulin signaling. Our results suggest that future studies should focus on the fate of effluxed TCA cycle intermediates and on mechanisms ensuring their replenishment.

Technical Abstract: Incomplete or limited long-chain fatty acid (LCFA) combustion in skeletal muscle has been associated with insulin resistance. Signals that are responsive to shifts in LCFA beta-oxidation rate or degree of intramitochondrial catabolism are hypothesized to regulate second messenger systems downstream of the insulin receptor. Recent evidence supports a causal link between mitochondrial LCFA combustion in skeletal muscle and insulin resistance. We have used unbiased metabolite profiling of mouse muscle mitochondria with the aim of identifying candidate metabolites within or effluxed from mitochondria and that are shifted with LCFA combustion rate. Large-scale unbiased metabolomics analysis was performed using GC/TOF-MS on bufferand mitochondrial matrix fractions obtained prior to and after 20 min of palmitate catabolism (n=7 mice/condition). Three [palmitate] (2, 9 and 19 µM; corresponding to low, intermediate and high oxidation rates) and 9 µM palmitate plus tricarboxylic acid (TCA) cycle and electron transport chain inhibitors were each tested and compared to zero palmitate control incubations. Paired comparisons of the 0 and 20 min samples were made by Student’s t-test. False discovery rate were estimated and Type I error rates assigned. Major metabolite groups were organic acids, amines and amino acids, free fatty acids and sugar phosphates. Palmitate oxidation was associated with unique profiles of metabolites, a subset of which correlated to palmitate oxidation rate. In particular, palmitate oxidation rate was associated with distinct changes in the levels of TCA cycle intermediates within and effluxed from mitochondria. This proof-of-principle study establishes that large-scale metabolomics methods can be applied to organelle-level models to discover metabolite patterns reflective of LCFA combustion, which may lead to identification of molecules linking muscle fat metabolism and insulin signaling. Our results suggest that future studies should focus on the fate of effluxed TCA cycle intermediates and on mechanisms ensuring their replenishment.