Fig. 2. The generation of ROS by mitochondria. Electrons released from mitochondria reduce oxygen molecules, thereby producing such ROS as superoxides (O2-). Superoxide dismutase (SOD) catalyzes H2O2 formation from superoxides. H2O2 might be deactivated by catalase (CAT). However, when H2O2 reacts with iron or copper ions, hydroxyl radicals (OH•), the most reactive form of ROS, are produced. Excessive antioxidants (AO) can inhibit production of O2- and other ROS.
Fig. 3. (a) ROS generated by microsomal monooxygenases, which have cytochrome P450 as a central link. Oxidation is the way to transform hydrophobic toxic substances, drugs, steroids etc., and thereby remove them. Excessive antioxidants can inhibit this protective function.
(b) ROS generated by phagocytes kill infectious microorganisms and cancer cells. Excessive antioxidants can inhibit this protective mechanism.
Fig. 5. Schematic representation of apoptosis. ROS generated by mitochondria are essential mediators of apoptosis. Together with cytochrome C, Apaf-1, and ATP, released from mitochondria, ROS activate proteolytic enzymes, termed caspases, which promote deoxyribonuclease, and thereby destroy targeted "bad" cells.
Fig. 6. The anticancer drug, cisplatin, kills breast cancer cells by the induction of apoptosis. The antioxidant vitamin E inhibits the cisplatin-induced apoptotic death of cancer cells by scavenging ROS that are essential for carrying out apoptosis (see Fig. 7). MCF-7 breast cancer cells were grown in Eagle’s MEM in 6-well plates (4 x 104 cells per well) and incubated for 24 hours with 15 µM cisplatin. Vitamin E (15 µM) was added to the medium simultaneously with cisplatin or separately. Apoptosis was determined by TUNEL assay and morphological cell patterns [31].
Fig. 7. The antioxidant vitamin E inhibits cisplatin-induced ROS generation in cancer cells. The conditions of the experiment were as in Fig. 6. ROS generation was determined using the avidin-FITC which reacts specifically in apoptotic cells with 8-oxo-deoxyguanine, the biomarker of ROS generation, as described in reference [23].
Fig. 8. Increased oxidative stress results in enhanced apoptosis in the brain tumors of mice fed an antioxidant-devoid diet. The distribution of TUNEL-positive (black label, arrow) apoptotic cells in brain tumor for control (A) and in mice on the antioxidant-devoid diet (B) is shown. Oxidized guanine residues (8-oxo-Gua), biomarkers of ROS generation, were detected in brain tumors, using specific monoclonal antibodies (C and D) or an avidin-FITC conjugate (E and F). By both methods, cells in tumors of antioxidant-depleted mice (D and F) exhibit higher levels of 8-oxo-Gua residues than do cells in the tumors of control brains (C and E).
Fig. 9. The tumors of mice fed an antioxidant-depleted diet are reduced in size. Compared with tumors in mice fed a standard diet (A and B), the tumors in mice fed an antioxidant-depleted diet (C) were significantly smaller. Magnification: A= 60x; B and C= 120x.
