Also, F-inhibits mitochondrial VO2 in both state 4 and state 3 and increases superoxide production. According with these results, the oxygen uptake rate also decreased in chronic treatments with F-. The increase in superoxide radicals as soon as F-reaches the mitochondrion could explain the increase in GSSG/TGSH ratio due to an augmented amount of peroxides reacting with reduced glutathione and producing higher amounts of oxidized glutathione. The antioxidant enzymes activities were not modified after three pulses of F- and the decreased in GSSG/TGSH could explain the increase in lipid peroxidation after three pulses of F-. Recently it has been reported that the constant exposure to 
Ffor 72 h is able to induce apoptosis in osteoblasts through Ursolic-acid increasing oxidative stress. In that work F- was administered to osteoblasts in constant concentration for 72 h. In the experiments described in our paper, osteoblasts are exposed for a few minutes to a high concentration of F-, as it occurs in vivo. Although F- levels returned to basal levels 24 h after each pulse of F-, an inhibition of oxygen consumption and respiratory complexes activities and an increase in oxidative stress status were observed. The results shown in this paper also demonstrate that chronic administration of F- produces a significant decrease in mitochondrial respiratory chain activity. F- treatment significantly inhibited complex IV and partially decreasedcomplex I-II and complex I-III activities. As complex I and III are the main sites of superoxide radical synthesis, F- could enhance its production by inhibiting the mitochondrial activity at respiratory chain level. F- inhibition at complex IV level could enhance the formation of ubiquinone radical which in turns can react with molecular oxygen increasing superoxide radical production. When the respiratory chain is inhibited, the electron supply reduces the ubiquinonepool and in the presence of large proton motive force, the electrons are forced back from reduced Loganin ubiquinoneinto complex I, which has two possible sites of superoxide production: the flavin in the NADH-oxidizing site and the ubiquinone-reducing site. Recently it has been demonstrated that when succinate is used as electron donor, most superoxide is produced at the ubiquinone reduction site. Taking all these results into account, we are able to describe the effect of F- on mitochondrial ROS production and its relationship with oxidative stress and inflammation. After an oral dose, F- may inhibit the respiratory chain, increasing the production of superoxide radicaland thereby of hydroxide peroxide and peroxynitrite. Antioxidant enzymes activities cannot prevent increased free radical formation. Therefore, there is an increase in ROS that finally produce oxidations in membranes and damage the cell macromoleculesand may be the cause of the inflammatory foci observed in the bone. It has already been demonstrated that ROS production induced inflammatory gene expression in alveolar macrophages, fibroblastsand kidney. Therefore, bone inflammatory foci could be enhanced via ROS-dependent activation of pro-inflammatory genes. Several methods exist for SNP detection. In the present study, we adapted the snapshot method for SNP detection. As compared to the direct sequencing method, the snapshot method is both time- and cost-efficient, and is therefore, suitable for examining large samples and multilocus genotyping.