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Fluorescence imaging of mitochondria in cultured skin fibroblasts: a useful method for detection of oxidative phosphorylation defects

(2012) PEDIATRIC RESEARCH. 72(3). p.232-240
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Abstract
BACKGROUND: Protons are pumped from the mitochondrial matrix via oxidative phosphorylation (OXPHOS) into the intermembrane space, creating an electric membrane potential (Delta psi) that is used for adenosine triphosphate (ATP) production. Defects in one or more of the OXPHOS complexes are associated with a variety of clinical symptoms, often making it difficult to pinpoint the causal mutation. METHODS: In this article, a microscopic method for the quantitative evaluation of Delta psi in cultured skin fibroblasts is described. The method using 5,5',6,6'-tetraethylbenzimidazolyl-carbocyanine iodide (JC-1) fluorescence staining was tested in a selection of OXPHOS-deficient cell lines. RESULTS: A significant reduction of Delta psi was found in the cell lines of patients with either an isolated defect in complex I, II, or IV or a combined defect (complex I + complex IV). Delta psi was not reduced in the fibroblasts of two patients with severe complex V deficiency. Addition of the complex I inhibitor rotenone induced a significant reduction of ALP and perinuclear relocalization of the mitochondria. In cells with a heteroplasmic mitochondrial DNA (mtDNA) defect, a more heterogeneous reduction of Delta psi was detected. CONCLUSION: Our data show that imaging of Delta psi in cultured skin fibroblasts is a useful method for the evaluation of OXPHOS functioning in cultured cell lines.
Keywords
COMPLEX I, RESPIRATORY-CHAIN, LEIGH-SYNDROME, MUTATION, DEFICIENCY, GENE, CELLS, DNA

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MLA
De Paepe, Boel, et al. “Fluorescence Imaging of Mitochondria in Cultured Skin Fibroblasts: A Useful Method for Detection of Oxidative Phosphorylation Defects.” PEDIATRIC RESEARCH, vol. 72, no. 3, 2012, pp. 232–40, doi:10.1038/pr.2012.84.
APA
De Paepe, B., Smet, J., Vanlander, A., Seneca, S., Lissens, W., De Meirleir, L., … Van Coster, R. (2012). Fluorescence imaging of mitochondria in cultured skin fibroblasts: a useful method for detection of oxidative phosphorylation defects. PEDIATRIC RESEARCH, 72(3), 232–240. https://doi.org/10.1038/pr.2012.84
Chicago author-date
De Paepe, Boel, Joél Smet, Arnaud Vanlander, Sara Seneca, Willy Lissens, Linda De Meirleir, Mado Vandewoestyne, Dieter Deforce, Richard J Rodenburg, and Rudy Van Coster. 2012. “Fluorescence Imaging of Mitochondria in Cultured Skin Fibroblasts: A Useful Method for Detection of Oxidative Phosphorylation Defects.” PEDIATRIC RESEARCH 72 (3): 232–40. https://doi.org/10.1038/pr.2012.84.
Chicago author-date (all authors)
De Paepe, Boel, Joél Smet, Arnaud Vanlander, Sara Seneca, Willy Lissens, Linda De Meirleir, Mado Vandewoestyne, Dieter Deforce, Richard J Rodenburg, and Rudy Van Coster. 2012. “Fluorescence Imaging of Mitochondria in Cultured Skin Fibroblasts: A Useful Method for Detection of Oxidative Phosphorylation Defects.” PEDIATRIC RESEARCH 72 (3): 232–240. doi:10.1038/pr.2012.84.
Vancouver
1.
De Paepe B, Smet J, Vanlander A, Seneca S, Lissens W, De Meirleir L, et al. Fluorescence imaging of mitochondria in cultured skin fibroblasts: a useful method for detection of oxidative phosphorylation defects. PEDIATRIC RESEARCH. 2012;72(3):232–40.
IEEE
[1]
B. De Paepe et al., “Fluorescence imaging of mitochondria in cultured skin fibroblasts: a useful method for detection of oxidative phosphorylation defects,” PEDIATRIC RESEARCH, vol. 72, no. 3, pp. 232–240, 2012.
@article{3000467,
  abstract     = {{BACKGROUND: Protons are pumped from the mitochondrial matrix via oxidative phosphorylation (OXPHOS) into the intermembrane space, creating an electric membrane potential (Delta psi) that is used for adenosine triphosphate (ATP) production. Defects in one or more of the OXPHOS complexes are associated with a variety of clinical symptoms, often making it difficult to pinpoint the causal mutation. 
METHODS: In this article, a microscopic method for the quantitative evaluation of Delta psi in cultured skin fibroblasts is described. The method using 5,5',6,6'-tetraethylbenzimidazolyl-carbocyanine iodide (JC-1) fluorescence staining was tested in a selection of OXPHOS-deficient cell lines. 
RESULTS: A significant reduction of Delta psi was found in the cell lines of patients with either an isolated defect in complex I, II, or IV or a combined defect (complex I + complex IV). Delta psi was not reduced in the fibroblasts of two patients with severe complex V deficiency. Addition of the complex I inhibitor rotenone induced a significant reduction of ALP and perinuclear relocalization of the mitochondria. In cells with a heteroplasmic mitochondrial DNA (mtDNA) defect, a more heterogeneous reduction of Delta psi was detected. 
CONCLUSION: Our data show that imaging of Delta psi in cultured skin fibroblasts is a useful method for the evaluation of OXPHOS functioning in cultured cell lines.}},
  author       = {{De Paepe, Boel and Smet, Joél and Vanlander, Arnaud and Seneca, Sara and Lissens, Willy and De Meirleir, Linda and Vandewoestyne, Mado and Deforce, Dieter and Rodenburg, Richard J and Van Coster, Rudy}},
  issn         = {{0031-3998}},
  journal      = {{PEDIATRIC RESEARCH}},
  keywords     = {{COMPLEX I,RESPIRATORY-CHAIN,LEIGH-SYNDROME,MUTATION,DEFICIENCY,GENE,CELLS,DNA}},
  language     = {{eng}},
  number       = {{3}},
  pages        = {{232--240}},
  title        = {{Fluorescence imaging of mitochondria in cultured skin fibroblasts: a useful method for detection of oxidative phosphorylation defects}},
  url          = {{http://doi.org/10.1038/pr.2012.84}},
  volume       = {{72}},
  year         = {{2012}},
}

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