Doxorubicin-induced mtDNA insertions in nuclear genome of human lymphocytes

Over 750 nuclear DNA sequences of mitochondrial origin have been identified in human reference genome. Recent data indicate that de novo occurrence of mitochondrial DNA (mtDNA) in the nuclear genome may contribute to development of pathological conditions. Moreover, elevation of nuclear mtDNA was observed in tumor cells of patients with different types of malignancies such as breast and colon cancer. It was suggested that DNA double strand breaks (DSBs) may contribute to occurrence of mtDNA in the nuclear genome. Thus we hypothesized that DSB-inducing mutagens can influence the frequency of mtDNA insertions in the human chromosomes. To test our hypothesize whole blood cultures from two male and two female healthy volunteers were treated with DSB-inducing antitumor drug doxorubicin (DOX) at concentrations 0.025, 0.035 and 0.05 µg/mL for 24 h. Insertions of mtDNA in the chromosomes were identified using fluorescence in situ hybridization with application of mtDNA specific probes. Levels of chromosome damage were analyzed by cytokinesis-block micronucleus (MN) assay. DOX significantly elevated the frequency of metaphases with mtDNA insertions. In the control group, six mtDNA insertions were identified. In cells treated with DOX at concentrations of 0.025, 0.035 and 0.05 μg/mL in total 21, 18 and 16 mtDNA insertions were identified, respectively. Positive correlations between mtDNA insertions induced by DOX at concentrations of 0.025 and 0.035 µg/mL and chromosome length (r = 0.631 and r = 0.502, respectively) were revealed indicating the randomness of DOX-induced mtDNA insertions. Positive correlations between mtDNA insertions and levels of MN induced by DOX at concentrations of 0.025 and 0.05 µg/mL were revealed (r = 0.985 and r = 0.956, respectively) indicating the potential influence of chromosome damage on insertion of mtDNA in the nuclear genome. Overall these results indicate that DOX can promote mtDNA translocation in the nuclear genome of human lymphocytes which correlated with the level of DOX-induced chromosome damage determined by MN assay.