Chelation of transferrin iron by desferrioxamine in K562 cells. The partition of iron between ferrioxamine and ferritin

In this study we have determined whether desferrioxamine can chelate iron delivered to human leukaemic cells by the transferrin endocytic cycle. The cellular uptake of desferrioxamine was investigated by an indirect method in which the conversion of repeated pulses of [59Fe]transferrin to [59Fe]ferrioxamine was determined at two concentrations of the drug. Maximum generation of [59Fe]ferrioxamine occurred in cells exposed to either 100 microM- or 500 microM-desferrioxamine after 40-60 min. Thereafter (up to 180 min) [59Fe]ferrioxamine levels remained steady with 20% of a 59Fe pulse partitioning to chelator at 100 microM and 50% at 500 microM. Of the cellular [59Fe]ferrioxamine loss 50% occurred within 90-120 min. In cells preloaded with desferrioxamine for 1 or 4 h the partitioning of iron during a 3 h incubation with [59Fe]transferrin was dependent upon the extracellular concentration of the chelator. Above 1 mM more than 80% of entering iron was converted to ferrioxamine and less than 5% partitioned to ferritin. Below this concentration (50-500 microM) a proportion of the iron became ferritin associated (7-41%). There was a linear increase in the total amount of intracellular [59Fe]ferrioxamine in accordance with cellular iron uptake showing that transferrin continued to cycle in the presence of high concentrations of desferrioxamine. The uptake of iron and generation of ferrioxamine were markedly reduced by 5 mM-methylamine, which prevented endosome acidification and uncoupling of iron from endocytosed transferrin.