RESUMO

The effect of exogenous cytochrome c on reactive oxygen species (ROS) formation and its dependence on mitochondrial permeability transition pore (MPTP) opening were studied in rat liver mitochondria. ROS generation was evaluated by changes in dichlorofluorescein (DCF) fluorescence. It has been shown that MPTP activation by increasing Ca²⁺ content in the medium resulted in an enhancement of the mitochondrial ROS production and oxygen consumption, despite a decrease in matrix calcium retention, dependent on the amount of added Ca²⁺. Cytochrome c in the incubation medium did not influence ROS formation significantly when MPTP opening was blocked by cyclosporine A. However, in the presence of cytochrome c, MPTP opening was accompanied by dramatic increase in ROS production. Steep rise in DCF fluorescence because of matrix ROS formation was sensitive to MPTP opening, and it did not result from the direct interaction between the probe and cytochrome c outside the mitochondria. To explain obtained data, the hypothesis has been put forward that MPTP could serve for ROS exchange between the matrix and the medium where heme iron of cytochrome c would act as a catalytic center to enhance ROS production. We suppose that apart of its conventional function, cytochrome c which is not involved in electron transport could serve as an amplifier of ROS production, which in turn would provide a background for the development of apoptosis due to MPTP opening.