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[Pathogenesis and mechanism of iron overload: ferric nitrilotriacetate, hemosiderin, active oxygen, and
Abstract:
Iron overload is found clinically in such conditions as hemochromatosis and sideroblastic anemia, and after long term repeated transfusion in aplastic anemia. An animal model of iron overload was successfully developed in rats and rabbits by repeated intraperitoneal injections of ferric nitrilotriacetate (Fe3+-NTA). This procedure induced a diabetic state with hyperglycemia, ketonemia, glycosuria and ketonuria. Blood venesection on these rats reduced the iron load in the liver and pancreas, and ameliorated the general diabetic symptoms. A single injection of Fe3+-NTA in rats induced a temporary elevation in plasma iron concentration, lipid peroxidation in the perfused liver homogenate expressed by malondialdehyde (MDA) formation, blood GOT, GPT, ALP and gamma-GTP sequentially. Fe3+-NTA uptake in the liver caused membrane lipid peroxidation, and subsequently produced a transit liberation of liver cell enzymes, although the incorporated liver Fe3+-NTA was only 1% of the injected dosage (7.5 mg iron/kg BW) at 3 hr after injection. The direct toxic effect of Fe3+-NTA to living cells was examined using cultured normal rat liver parenchymal cells (RL-34). Marked cytolysis was found in cells exposed to more than 25 micrograms of iron through Fe3+-NTA/ml. At 50 micrograms iron of Fe3+-NTA/ml, most cells were lethally injured and the remaining cells were piled up and aggregated at 15 days. They grew on soft agar culture, and when inoculated subcutaneously to five newly born rats a subcutaneous tumor developed in all animals within three weeks. Lung metastases were found in three of five inoculated rats. A spin trapping technique with electron spin resonance (ESR) on Fe3+-NTA employing 5, 5-dimethyl-l-pyrroline-N-oxide (DMPO) yielded a spin adduct with three doublets (DMPO-Z) which corresponded to singlet oxygen. By ESR in the presence of H2O2, the Fe3+-NTA solution strongly generated hydroxyl radical. The production of active oxygen species by Fe3+-NTA solution may explain the toxicity and carcinogenicity of Fe3+-NTA. The majority of stainable iron in the iron overloaded tissue was hemosiderin (Hs). We tried to purify the Hs from multi-transfused human spleen by the method of Weir et al. The purified Hs did not show a DMPO-OH adducts in the presence of H2O2 and DMPO on ESR measurement. The Hs iron was solubilized with several biological ligands in an acidic state in the presence of a reducing reagent like glutathione. Solubilized Hs iron produced iron chelate complexes which resulted in OH radicals production in the presence of H2O2 in acidic conditions below pH 5.5.(ABSTRACT TRUNCATED AT 400 WORDS)