Ethidium bromide as a vital probe of mitochondrial DNA in carcinoma cells

The interaction of ethidium bromide (EB) with mitochondria in human breast and lung carcinoma cells was investigated under living conditions, employing a laser scanning confocal fluorescence microscopy (LSCFM) with a photon counting detection to reduce drastically the laser power excitation and the fluorescent probe concentration. In sensitive and multidrug-resistant MCF-7 cell lines, which are important model systems for the study of mitochondria in tumour cells, two distinct populations of mitochondria were observed, each characterised by a different EB fluorescence, membrane potential, cellular localisation and morphology. By image analysis, these peripheral mitochondria showed a peculiar morphology, consisting of punctuate organelles (0.8 μm in size) organised in rosette-like assemblies. Unexpectedly, an intense EB fluorescence was observed in these mitochondria, indicating a high accessibility to EB of their mtDNA, which is likely to be in an active replicative or transcriptional state. These results might, therefore, suggest an active biogenesis and metabolism of the peripheral mitochondria that could be a consequence of the increased energetic needs of the cells, after their tumour transformation. Indeed, the pool of peripheral mitochondria, as well as their peculiar morphology and spatial organisation, was found to be a characteristic feature of all the carcinoma cells examined here, but not of their non-transformed parental MCF10A cells.