In vitro supplementation with dAMP/dGMP leads to partial restoration of mtDNA levels in mitochondrial depletion syndromes

Abstract

Mitochondrial DNA depletion syndrome, a frequent cause of childhood (hepato)encephalomyopathies, is
defined as a reduction of mitochondrial DNA copy number related to nuclear DNA. It was previously
shown that mtDNA depletion can be prevented by dAMP/dGMP supplementation in deoxyguanosine
kinase-deficient fibroblasts. We investigated myotubes of patients diagnosed with mtDNA depletion carrying
pathogenic mutations in DGUOK, POLG1 (Alpers syndrome) and TYMP. Differentiating myotubes of all
patients and controls were supplemented with different doses of dAMP/dGMP or dAMP/dGMP/dCMP in
TYMP deficiency, and analysed for mtDNA/nDNA ratio and for cytochrome c oxidase (COX) activity. Serum
deprivation and myotube formation triggered a decrease in mtDNA copy number in DGUOK or POLG1
deficient myotubes, but not in TYMP deficiency and healthy controls. Supplementation with dAMP/dGMP
leads to a significant and reproducible rescue of mtDNA depletion in DGUOK deficiency. POLG1 deficient
myotubes also showed a mild, not significant increase in mtDNA copy number. MtDNA depletion did not
result in deficient COX staining in DGUOK and POLG1-deficient myotubes. Treatment with ethidium bromide
resulted in very severe depletion and absence of COX staining in all cell types, and no recovery was observed
after supplementation with dAMP/dGMP. We show that supplementation with dAMP/dGMP increases mtDNA
copy number significantly in DGUOK deficient myotubes and, leads to a mild, non-significant improvement of
mtDNA depletion in POLG1 deficiency. No adverse effect on mtDNA copy number was observed on high-dose
supplementation in vitro. Further studies are needed to determine possible therapeutic implications of dAMP/
dGMP supplementation for DGUOK deficiency in vivo.

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Identifiers

ORCID for Joanna D. Stewart: orcid.org/0000-0002-2608-1967

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