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

Boel De Paepe UGent, Joél Smet UGent, Arnaud Vanlander UGent, Sara Seneca, Willy Lissens, Linda De Meirleir, Mado Vandewoestyne UGent, Dieter Deforce UGent, Richard J Rodenburg and Rudy Van Coster UGent (2012) PEDIATRIC RESEARCH. 72(3). p.232-240
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.
Please use this url to cite or link to this publication:
author
organization
year
type
journalArticle (original)
publication status
published
subject
keyword
COMPLEX I, RESPIRATORY-CHAIN, LEIGH-SYNDROME, MUTATION, DEFICIENCY, GENE, CELLS, DNA
journal title
PEDIATRIC RESEARCH
Pediatr. Res.
volume
72
issue
3
pages
232 - 240
Web of Science type
Article
Web of Science id
000308280500002
JCR category
PEDIATRICS
JCR impact factor
2.673 (2012)
JCR rank
18/119 (2012)
JCR quartile
1 (2012)
ISSN
0031-3998
DOI
10.1038/pr.2012.84
language
English
UGent publication?
yes
classification
A1
copyright statement
I have transferred the copyright for this publication to the publisher
id
3000467
handle
http://hdl.handle.net/1854/LU-3000467
date created
2012-09-27 15:36:50
date last changed
2013-07-08 11:31:25
@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, Jo{\'e}l 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},
  keyword      = {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://dx.doi.org/10.1038/pr.2012.84},
  volume       = {72},
  year         = {2012},
}

Chicago
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.
APA
De Paepe, B., Smet, J., Vanlander, A., Seneca, S., Lissens, W., De Meirleir, L., Vandewoestyne, M., et al. (2012). Fluorescence imaging of mitochondria in cultured skin fibroblasts: a useful method for detection of oxidative phosphorylation defects. PEDIATRIC RESEARCH, 72(3), 232–240.
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.
MLA
De Paepe, Boel, Joél Smet, Arnaud Vanlander, et al. “Fluorescence Imaging of Mitochondria in Cultured Skin Fibroblasts: a Useful Method for Detection of Oxidative Phosphorylation Defects.” PEDIATRIC RESEARCH 72.3 (2012): 232–240. Print.