Hypoxia differently modulates the release of cell-free mitochondrial and nuclear DNA

The factors influencing the dynamics of cell-free DNA (cfDNA) release are poorly known. In this study, we investigated the incidence of hypoxia on the release of mitochondrial and nuclear cell-free DNA (McfDNA and NcfDNA, respectively). McfDNA and NcfDNA were measured by an ultra-sensitive qPCR-based assay, in the supernatants of different colorectal cell lines and in the plasma of tumor cell-grafted mice, in normoxic or hypoxic conditions. Our data highlighted the intrinsic stability of cfDNA, what allowed us to demonstrate that cancer cell lines released amounts equivalent to the oligonucleotidic mass of a chromosome in 6 hours, revealing the high dynamics of cfDNA release. Hypoxia induced a huge increase in NcfDNA and McfDNA concentrations within the first 24 hours of culture. After this period, total concentrations remained stable, suggesting a negative regulation of cfDNA release. Noteworthy, daily McfDNA release after 24 hours of culture was highly inhibited. In grafted mice, we observed a significantly higher level of circulating NcfDNA in hypoxia vs normoxia. In contrast, McfDNA release remained stable, consistently with the negative regulation observed in cell model. Our data suggest that the mechanisms underlying NcfDNA release are different from those involved in McfDNA release, opening the way to better understand the variation in circulating DNA concentrations.