Abstract
Introduction
A decline in mitochondrial function represents a key factor of a large number of inborn errors of metabolism, which lead to an extremely heterogeneous group of disorders.
Objectives
To gain insight into the biochemical consequences of mitochondrial dysfunction, we performed a metabolic profiling study in human skin fibroblasts using galactose stress medium, which forces cells to rely on mitochondrial metabolism.
Methods
Fibroblasts from controls, complex I and pyruvate dehydrogenase (PDH) deficient patients were grown under glucose or galactose culture condition. We investigated extracellular flux using Seahorse XF24 cell analyzer and assessed metabolome fingerprints using NMR spectroscopy.
Results
Incubation of fibroblasts in galactose leads to an increase in oxygen consumption and decrease in extracellular acidification rate, confirming adaptation to a more aerobic metabolism. NMR allowed rapid profiling of 41 intracellular metabolites and revealed clear separation of mitochondrial defects from controls under galactose using partial least squares discriminant analysis. We found changes in classical markers of mitochondrial metabolic dysfunction, as well as unexpected markers of amino acid and choline metabolism. PDH deficient cell lines showed distinct upregulation of glutaminolytic metabolism and accumulation of branched-chain amino acids, while complex I deficient cell lines were characterized by increased levels in choline metabolites under galactose.
Conclusion
Our results show the relevance of selective culture methods in discriminating normal from metabolic deficient cells. The study indicates that untargeted fingerprinting NMR profiles provide physiological insight on metabolic adaptations and can be used to distinguish cellular metabolic adaptations in PDH and complex I deficient fibroblasts.
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Change history
19 March 2019
The original version of this article contained an error in Table 2. The text in the second header line should read “GAL supernatant” and “GAL Medium” instead of “GLC supernatant” and “GLC Medium”. The corrected Table 2 is given below. The original article has been corrected.
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Acknowledgements
We thank André Schaller (Division of Human Genetics and Department of Paediatrics, Inselspital, Bern) for providing the genetic characteristics of the patient fibroblasts.
Funding
This work was supported by a grant from the Batzebär foundation of the children’s university hospitals Bern to JMN.
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DH, AF, GD, SK, PV, and JMN conceived the work and designed the experiments; DH, AF, and SK cultured cells, performed metabolic flux experiments and analyzed the data. DH and GD performed NMR analysis of cells or supernatant and analyzed the data. PV and JMN provided experimental advice and overall guidance. DH, AF, GD, SK, PV and JMN wrote the manuscript or revised it critically for important intellectual content. All authors approved the final manuscript.
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All procedures performed in studies involving human participants were in accordance with the 1964 Helsinki declaration and its later amendments and approved by the Ethics Committee of the University Hospital of Bern.
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The original version of this article was revised: The errors in the Table 2 have been corrected.
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Hertig, D., Felser, A., Diserens, G. et al. Selective galactose culture condition reveals distinct metabolic signatures in pyruvate dehydrogenase and complex I deficient human skin fibroblasts. Metabolomics 15, 32 (2019). https://doi.org/10.1007/s11306-019-1497-2
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DOI: https://doi.org/10.1007/s11306-019-1497-2