Francisco et al. Increase of reactive oxygen species by desferrioxamine during experimental Chagas’ disease Introduction Chagas’ Disease continues to pose a serious threat to human health in much of Latin America, and has become the most important emerging parasitic disease in developed countries. There is growing evidence to suggest that during the course of infection by Trypanosoma cruzi, the myocardium is exposed to injuries induced by sustained oxidative stress, and that these may contribute to disease progression. The principal causes of oxidative stress are reactive oxygen species (ROS), which may be broadly defined as derivatives of molecular oxygen, e.g. O2•–, HO• and hydrogen peroxide. ROS are unstable and react rapidly with other free radicals and macromolecules in chain reactions to generate increasingly harmful oxidants.1 Acute T. cruzi infection induces H 2 O 2 release by peripheral blood monocytes and spleen macrophages in rats infected with the Y strain.2 During the course of development of Chagas’ disease, high levels of ROS can be produced as a consequence of tissue destruction caused by toxic secretions from the parasite, immune-mediated cytotoxic reactions and secondary damage to mitochondria.1 All aerobic organisms are continuously subjected to ROS generated by oxidative metabolism, the detoxification of xenobiotics or the action of ultraviolet radiation and several other mechanisms, because anaerobic organisms do not need such antioxidant adaptations, and there are many other mechanisms that generate ROS in animals, fungi, algae, protists and plants. In response, organisms have developed a variety of antioxidant defense systems to cope with ROS, although it is important to note that the defense strategies of parasitic protozoa exhibit major differences one from another and also in comparison with their mammalian hosts. Many antioxidant defense strategies depend on the action of antioxidant enzymes, including superoxide dismutase (SOD), glutathione peroxidase (GPx) and catalase, or low molecular weight antioxidants due to their importance in ROS (hydrogen peroxide) detoxification3. SOD catalyses the conversion of the superoxide anion into hydrogen peroxide and oxygen, and exists in a number of different isoforms. Unlike other eukaryotic parasites, the trypanosomatids (Leishmania spp. and Trypanosoma spp.) together with Plasmodium spp. have an iron-containing SOD isoform that is normally found only in bacteria.4,5 GPx exists in five isoforms and catalyses reduction by glutathione of hydrogen peroxide or alkyl peroxide to water or alcohol, respectively.1 A number of assays are available that enable assessment of the level of oxidative stress induced by 186 Redox Report 2010 Vol 15 No 4 different biological agents within an organism. The determination of lipid peroxidation in terms of thiobarbituric acid reactive substances (TBARS) is considered to be a very useful, cheap and easy assay for evaluating oxidative stress. However, this assay, despite common use in evaluating oxidative stress, has some limitations, especially when measured by spectrophotometry and not by fluorescence or HPLC. Additionally, the direct attack of protein by ROS leads to the formation of carbonyl groups, such that the estimation of protein carbonyl (PC) provides an additional (or complementary) measure of the effect of oxidative stress. Desferrioxamine (DFO; Desferal®, Novartis, Basel, Switzerland) is a hexadentate iron chelator that complexes with iron in a 1:1 molar ratio to yield the stable complex ferrioxamine (stability constant 1031).6 DFO is believed to access the pools of iron in the liver and in the reticulo-endothelial system that are together responsible for the major proportion of the body iron burden. However, since DFO permeates into cells relatively slowly, because of its large molecular size and hydrophilic nature, its efficiency as an intracellular chelator is limited.5,7 Various studies2,8–11 have established that administration of DFO in T. cruzi infection modifies the course of Chagas’ disease, reducing parasitemia and mortality. Results obtained using a murine model have shown, however, that whilst there were reductions in the number of circulating parasites and in mortality, the animals did not suffer from anemia.8,9 It was concluded, therefore, that DFO had a direct effect on the parasite and not on the reduction of iron supply in the host. The aim of the present study was to evaluate the relationship between oxidative stress (assessed by TBARS, PC and nitrate/nitrite assays) and antioxidant defenses (evaluated in terms of SOD and GPx levels) in T. cruzi-infected mice that had been treated with DFO in order to clarify the mode of action of the drug. Materials and methods Details of the project were submitted to, and approved by, the Ethical Committee on Animal Research of the Universidade Federal de Ouro Preto and by the institutional Animal Care and Use Committee (CETEA 153/07) of the Universidade Federal de Minas Gerais. All procedures were carried out in compliance with current Brazilian regulations relating to Experimental Biology and Medicine as described in the guidelines issued by the Colégio Brasileiro de Experimentaçao Animal (COBEA) in 2006.