Ethidium bromide as a marker of mtDNA replication in living cells.
J Biomed Opt
; 17(4): 046001, 2012 Apr.
Article
em En
| MEDLINE
| ID: mdl-22559680
ABSTRACT
Mitochondrial DNA (mtDNA) in tumor cells was found to play an important role in maintaining the malignant phenotype. Using laser scanning confocal fluorescence microscopy (LSCFM) in a recent work, we reported a variable fluorescence intensity of ethidium bromide (EB) in mitochondria nucleoids of living carcinoma cells. Since when EB is bound to nucleic acids its fluorescence is intensified; a higher EB fluorescence intensity could reflect a higher DNA accessibility to EB, suggesting a higher mtDNA replication activity. To prove this hypothesis, in the present work we studied, by LSCFM, the EB fluorescence in mitochondria nucleoids of living neuroblastoma cells, a model system in which differentiation affects the level of mtDNA replication. A drastic decrease of fluorescence was observed after differentiation. To correlate EB fluorescence intensity to the mtDNA replication state, we evaluated the mtDNA nascent strands content by ligation-mediated real-time PCR, and we found a halved amount of replicating mtDNA molecules in differentiating cells. A similar result was obtained by BrdU incorporation. These results indicate that the low EB fluorescence of nucleoids in differentiated cells is correlated to a low content of replicating mtDNA, suggesting that EB may be used as a marker of mtDNA replication in living cells.
Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
DNA Mitocondrial
/
Replicação do DNA
/
Etídio
/
Corantes Fluorescentes
Tipo de estudo:
Prognostic_studies
Limite:
Humans
Idioma:
En
Revista:
J Biomed Opt
Assunto da revista:
ENGENHARIA BIOMEDICA
/
OFTALMOLOGIA
Ano de publicação:
2012
Tipo de documento:
Article
País de afiliação:
Itália
País de publicação:
EEUU
/
ESTADOS UNIDOS
/
ESTADOS UNIDOS DA AMERICA
/
EUA
/
UNITED STATES
/
UNITED STATES OF AMERICA
/
US
/
USA