The effect of ouabain on mitochondrial DNA damage in HepG2 cell lines

Our purpose in this study is to analyze mitochondrial DNA (mtDNA) lesion frequencies and mtDNA⁴⁹⁷⁷ deletion in HepG2 cells to examine the effects of ouabain on mtDNA. HepG2 cells were treated with 0.75, 7.5, 75, and 750 nM of ouabain for 24 h in the presence and absence of 10 mM 2-deoxyglucose (2-DG). The frequency of mtDNA⁴⁹⁷⁷ deletions and mitochondrial lesions were determined by real-time polymerase chain reaction. A ≥1.2-fold change or greater was considered significant. Ouabain doses of 750, 75, and 7.5 nM alone increased the frequency of mtDNA⁴⁹⁷⁷ deletions 1.39, 1.92, and 1.44 times, respectively. The 750 and 75 nM ouabain doses combined with 2-DG increased the mtDNA⁴⁹⁷⁷ deletion frequency 4.94 and 1.57 times, respectively. The 750 and 75 nM ouabain doses alone increased the mtDNA lesion frequency 2.5 and 1.5 times, respectively. The 750 nM ouabain dose combined with 2-DG increased the mtDNA lesion frequency 2.28 times. The 7.5 nM ouabain dose alone and combined with 2-DG decreased the mtDNA lesion frequency 0.67 and 0.45 times, respectively. Ouabain alone and when combined with 2-DG increases mtDNA lesion and mtDNA⁴⁹⁷⁷ deletion frequencies. This supports the thesis that ouabain creates oxidative stress and induces DNA damage and apoptosis.