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. © 2012 Society of Photo-Optical Instrumentation Engineers (SPIE).
Villa, A., Fusi, P., Pastori, V., Amicarelli, G., Pozzi, C., Adlerstein, D., et al. (2012). Ethidium bromide as a marker of mtDNA replication in living cells. JOURNAL OF BIOMEDICAL OPTICS, 17(4), 046001 [10.1117/1.JBO.17.4.046001].
|Citazione:||Villa, A., Fusi, P., Pastori, V., Amicarelli, G., Pozzi, C., Adlerstein, D., et al. (2012). Ethidium bromide as a marker of mtDNA replication in living cells. JOURNAL OF BIOMEDICAL OPTICS, 17(4), 046001 [10.1117/1.JBO.17.4.046001].|
|Tipo:||Articolo in rivista - Articolo scientifico|
|Carattere della pubblicazione:||Scientifica|
|Presenza di un coautore afferente ad Istituzioni straniere:||No|
|Titolo:||Ethidium bromide as a marker of mtDNA replication in living cells|
|Autori:||Villa, A; Fusi, P; Pastori, V; Amicarelli, G; Pozzi, C; Adlerstein, D; Doglia, S|
VILLA, ANNA MARIA (Corresponding)
FUSI, PAOLA ALESSANDRA (Secondo)
DOGLIA, SILVIA MARIA (Ultimo)
|Data di pubblicazione:||2012|
|Rivista:||JOURNAL OF BIOMEDICAL OPTICS|
|Digital Object Identifier (DOI):||http://dx.doi.org/10.1117/1.JBO.17.4.046001|
|Appare nelle tipologie:||01 - Articolo su rivista|