RESUMO
Superoxide dismutase (SOD) enzymes are considered the first line of Defence against reactive oxygen species. Among the intracellular isoforms of SOD, Cu,ZnSOD is widely distributed, and the MnSOD is localised in the mitochondria. The SOD activities have been measured indirectly by inhibiting oxidation reactions. Most techniques assessing the hepatic SOD activity are adaptations of classical methods and differ significantly from each other. This work assessed the hepatic Cu,ZnSOD activity in the supernatants of two different centrifugations, using two isotonic medium and the pyrogallol method. In most studies conducted in rat liver, only the Cu,Zn or total SOD activities were assessed and sometimes the Cu,ZnSOD activity was calculated based on the inhibition by cyanide. But, as demonstrated here, this inhibition is not complete. Besides, the novelty of this work is that we presented a method for the evaluating the Cu,ZnSOD and MnSOD activities separately and from the same liver.
Assuntos
Ensaios Enzimáticos/métodos , Fígado/enzimologia , Superóxido Dismutase/metabolismo , Animais , RatosRESUMO
Female mice fed a cafeteria diet (FCaf) develop higher liver steatosis and oxidative stress than males (MCaf) as a consequence of unresolved ER stress. Here, we investigated whether mitochondria play a role in this sex difference. The isolated mitochondria from FCaf showed more signs of oxidative stress than those of MCaf, correlated with a reduced content of GSH, increased amount of reactive oxygen species (ROS), and lower activities of enzymes involved in ROS neutralisation. Mitochondria from FCaf and MCaf livers exhibited lower rates of succinate-driven state III respiration and reduced ATPase activity in intact coupled mitochondria compared to their controls fed a standard diet (FC and MC), with no differences between the sexes. Fatty acid oxidation in mitochondria and peroxisomes was higher in MCaf and FCaf compared to their respective controls. In the intact perfused liver, there was no difference between sex or diet regarding the fatty acid oxidation rate. These results indicated that cafeteria diet did not affect mitochondrial energy metabolism, even in FCaf livers, which have higher steatosis and cellular oxidative stress. Nevertheless, the increase in mitochondrial ROS generation associated with a decrease in the antioxidant defence capacity, probably contributes to inducing or reinforcing the ER stress in FCaf livers.