Dose-Related Structural and Functional Modifications of Mitochondria Are Induced In Vitro by Low Ozone Concentrations

: In the last decades, ozone (O3)-based medical treatments have become a widely applied complementary therapy for several pathological conditions. O3 is administered at low dosages since the induction of a mild oxidative stress does not cause damage but stimulates the antioxidant cell response through the nuclear factor erythroid 2-related factor 2 (Nrf2). Mitochondria are sensitive to even mild oxidative stress, thus being a responsive target for O3. This study aimed to evaluate the mitochondrial response to low O3 doses used for medical treatments. As the skeletal muscle represents a primary target in local O3 treatments, a murine non-tumoral muscle cell line was selected as an appropriate in vitro model. Transmission electron microscopy, biochemistry, and flow cytometry provided original information on the O3 dose-dependent modifications of mitochondrial structural and molecular features. Low O3 doses promoted an increase in mitochondrial area and in cristae extension, as well as an enhancement of the electron transport chain complexes and of antioxidant catalase and manganese-dependent superoxide dismutase. Nrf2 maintained its association with the outer mitochondrial membrane, thus exerting its protective role. All mitochondrial modifications were observed 24 h after treatment and disappeared after 48 h, demonstrating that cells promptly respond to the O3-driven oxidative stress, effectively restoring homeostasis.