Most organisms living on Earth are entirely dependent on the presence of oxygen in the atmosphere. However, the by-products of oxygen metabolism are toxic to living organisms. Reactive oxygen species (ROS) in the cell are produced both during normal cellular metabolism or inflammatory reactions and under the influence of external factors like γ, X and UV radiation, biotransformation of dietary chemicals and some diet components, e.g. transient metal ions (1). Normal cellular metabolism seems to be the primary source of endogenous ROS. An imbalance between the formation of ROS and antioxidant defense leads to increased reactive oxygen species generation and oxidative stress development (2). ROS are radical molecules containing oxygen, for example superoxide (O₂^?–) and hydroxyl radical (^?OH), or non-radical molecules, such as hydrogen peroxide (H₂O₂) and singlet oxygen (¹O₂), which may be converted into radical forms. The most reactive ROS, hydroxyl radicals, are responsible for oxidation and fragmentation of nucleic acids, proteins and lipids. They are produced in the metal-catalysed Haber-Weiss and Fenton reactions mediated by the transition metal ions such as iron and the copper (3). Iron is a cofactor for many biological reactions and is an important component of metabolism in various tissues and organs, including the thyroid. Iron deficiency may affect thyroid hormone synthesis by decreasing the activity of the heme-dependent thyroid peroxidase (TPO). In addition, low iron levels reduce deiodinase activity, i.e. it slows down the conversion of T4 to T3, and also causes a raise in circulating concentrations of thyroid stimulating hormone (TSH) (4). With higher levels of TSH and low free T4 and T3 levels hypothyroidism occurs. Iron overload, on the other hand, may promote the persistence of harmful labile iron, which can catalyze the generation of potentially carcinogenic DNA adducts in the cell (5).

H₂O₂ production was found in vivo in many intracellular structures e.g. mitochondria, endoplasmic reticulum and peroxisomes. A high concentration of hydrogen peroxide was also observed in activated phagocytes, spermatozoids, bacteria and even in exhaled air (6). In the thyroid gland H₂O₂ is produced by one or two NADPH oxidases (Duox1/2) at the apical membrane of thyrocytes and it participates in hormone biosynthesis. To synthesize T3 and T4 hormones, the thyroid takes up iodine and incorporates it into the precursor of the hormones – thyroglobulin. Iodination of thyrosyl residues on thyroglobulin requires high concentrations of H₂O₂ as well as oxidized iodine, which is generated by the thyroid peroxidase (TPO) (7). For the TPO function properly H₂O₂ is necessary. It helps to stabilize the enzyme by autocatalytic covalent heme binding to the TPO molecule, which positively affects TPO activity (8). On the other hand, an excess of H₂O₂ may inhibit TPO activity and consequently inhibit thyroid hormone synthesis (9). Because H₂O₂ and iodine are co-substrates in hormone synthesis, changes of iodine concentrations affect the concentration of H₂O₂. In vitro and in vivo studies demonstrated that iodide inhibits the generation of H₂O₂ in the thyroid (10, 11). Production of H₂O₂ is moreover stimulated through the cAMP cascade by the thyrotropin (TSH), which increases the expression of genes important for hormone synthesis (e.g. TPO) (12).

H₂O₂ has various effects in the cell and it may enhance cell metabolism through diverse mechanisms. Besides that H₂O₂ acts as an oxidant, and also induces oxidative stress and apoptosis (13) working as an intracellular messenger (14). ROS-derived signals regulate growth, proliferation, differentiation and death of the cell (15-17). It has been demonstrated that in thyroid H₂O₂-mediated cytotoxicity appears at low H₂O₂ concentrations and leads to cell apoptosis or less frequently to necrosis (15). Moreover in vivo studies suggest that cytotoxic reaction to oxidative stress may depend on the functional state of the thyroid gland (18).

Despite the fact that hydrogen peroxide does not react directly with components of DNA, it is a precursor to highly reactive hydroxyl radical (^?OH), hypochlorite (ClO^–) and singlet oxygen (¹O₂). Therefore H₂O₂ may facilitate a mutagenic process and DNA modification leading to cancer development (19). A thyrocyte which generates a great amount of H₂O₂ is a long-lived cell and that allows it to accumulate mutations in DNA (20). Consequently, oxidative stress has been suggested to contribute to the pathogenesis of thyroid cancer (21, 22).

An antioxidative defense systems, that protect from the formation and effects of reactive oxygen species, function in all living organisms. In the cell the defense against the destructive effects of ROS works on the three levels.

The first level of the system prevents the formation of excessive quantities of ROS. The main component of this level are proteins that bind transition metal ions which thus inhibits Fenton reactions. Iron ions are bound by ferritin, transferrin and lactoferrin, copper ions by ceruloplasmin. Metallothioneins bind a number of different metal ions, as well as albumin, which non-specifically, is capable of binding many metal ions (23).

The second defense level neutralizes ROS. This system includes antioxidant enzymes such as superoxide dismutase (SOD), glutathione and ascorbate peroxidases (GPX, APX1), and glutathione transferase. The other elements of this protection level are small molecule antioxidants that work as direct or indirect free radical scavengers: glutathione, ascorbic acid, cysteine, tocopherols (vitamin E), retinoids (vitamin A analogs), uric acid, carotenoids, bilirubin, ubiquinol, and even glucose and pyruvate (3, 24). The above antioxidative protectors have been found in thyroid gland, e.g. GPX and TPO and are upregulated during the synthesis of thyroid hormones (25). There is also evidence that GPX3 which affects the H₂O₂ concentration directly interferes with hormone synthesis (26).

The third level of the defense is the elimination of ROS harmful effects on the most important cellular macromolecule – DNA. Oxidative DNA adducts are repaired by enzymes of excision repair systems, which will be described in subsequent chapters.

ROS reactions with DNA cause the most dangerous consequences for multicellular organisms. The ^?OH radical molecule is one of the ROS that is extremely reactive in the oxidation of cellular constituents such as nucleic acids, proteins and lipids. ^?OH interactions with DNA may lead to considerable damage, such as oxidized bases, base and sugar lesions, abasic sites, DNA-DNA intrastrand adducts, single or double strand breaks and DNA-protein cross-links (2, 27-29).