Antioxidant defences and homeostasis of reactive oxygen species in different human mitochondrial DNA-depleted cell lines

Three pairs of parental (rho(+)) and established mitochondrial DNA depleted (rho(0)) cells, derived from bone, lung and muscle were used to verify the influence of the nuclear background and the lack of efficient mitochondrial respiratory chain on antioxidant defences and homeostasis of intracellular reactive oxygen species (ROS). Mitochondrial DNA depletion significantly lowered glutathione reductase activity, glutathione (GSH) content, and consistently altered the GSH(2) : oxidized glutathione ratio in all of the rho(0) cell lines, albeit to differing extents, indicating the most oxidized redox state in bone rho(0) cells. Activity, as well as gene expression and protein content, of superoxide dismutase showed a decrease in bone and muscle rho(0) cell lines but not in lung rho(0) cells. GSH peroxidase activity was four times higher in all three rho(0) cell lines in comparison to the parental rho(+), suggesting that this may be a necessary adaptation for survival without a functional respiratory chain. Taken together, these data suggest that the lack of respiratory chain prompts the cells to reduce their need for antioxidant defences in a tissue-specific manner, exposing them to a major risk of oxidative injury. In fact bone-derived rho(0) cells displayed the highest steady-state level of intracellular ROS (measured directly by 2',7'-dichlorofluorescin, or indirectly by aconitase activity) compared to all the other rho(+) and rho(0) cells, both in the presence or absence of glucose. Analysis of mitochondrial and cytosolic/iron regulatory protein-1 aconitase indicated that most ROS of bone rho(0) cells originate from sources other than mitochondria.