title: Detection of mitochondrial DNA deletions in human skin fibroblasts of patients with Pearson's syndrome by two-color fluorescence in situ hybridization.
abstract: Pearson's marrow/pancreas syndrome is a disease associated with a large mitochondrial DNA (mtDNA) deletion. The various tissues of a patient contain heteroplasmic populations of wild-type (WT) and deleted mtDNA molecules. The clinical phenotype of Pearson's syndrome is variable and is not correlated with the size and position of the deletion. The histo- and cytological distribution of WT and deleted mtDNA molecules may be factors that correlate with the phenotypical expression of the disease. Here we introduce a new application of two-color FISH to visualize WT and deleted mtDNA simultaneously in a cell population of in vitro cultured skin fibroblasts of two patients with Pearson's syndrome. At the third passage of culturing, fibroblasts showed a remarkable heterogeneity of WT and deleted mtDNA: about 90% of the cells contained almost 100% WT mtDNA, and 10% of the cells contained predominantly deleted mtDNA. At the tenth passage of culturing, fibroblasts showed a reduction of intercellular heteroplasmy from 10% to 1%, while intracellular heteroplasmy was maintained. This new approach enables detailed analysis of distribution patterns of WT and deleted mtDNA molecules at the inter- and intracellular levels in clinical samples, and may contribute to a better understanding of genotype-phenotype relationships in patients with mitochondrial diseases.
authors.affiliation: Department of Cytochemistry, Leiden University, The Netherlands.
authors.forename: M P
authors.initials: MP
authors.lastname: van de Corput
authors.forename: J M
authors.initials: JM
authors.lastname: van den Ouweland
authors.forename: R W
authors.initials: RW
authors.lastname: Dirks
authors.forename: L M
authors.initials: LM
authors.lastname: Hart
authors.forename: G J
authors.initials: GJ
authors.lastname: Bruining
authors.forename: J A
authors.initials: JA
authors.lastname: Maassen
authors.forename: A K
authors.initials: AK
authors.lastname: Raap
mesh_terms: D001855:Bone Marrow Diseases / Q000235:genetics*; D002478:Cells, Cultured; D004272:DNA, Mitochondrial / Q000032:analysis / Q000235:genetics*; D005347:Fibroblasts / Q000737:chemistry; D006801:Humans; D017404:In Situ Hybridization, Fluorescence; D008928:Mitochondria / Q000235:genetics; D015345:Oligonucleotide Probes; D010182:Pancreatic Diseases / Q000235:genetics*; D017384:Sequence Deletion; D012867:Skin / Q000737:chemistry*; D013577:Syndrome
chemical_list: D004272:DNA, Mitochondrial; D015345:Oligonucleotide Probes