STUDIES OF IRON OVERLOAD - RAT-LIVER SIDEROSOME FERRITIN

Abstract

To investigate storage of ferritin and its transition to hemosiderin under conditions of Fe overload, rats were either given multiple injections of iron dextran over 4-5 wk or fed a diet containing 1.3% Fe as ferric ammonium citrate for 60 days. Then, preparations of liver siderosomes (heavily Fe-laden lysosomes) were examined for content of buffer-soluble ferritin and buffer-insoluble, ferritin-related protein, total nonheme Fe and protein, cathepsin D activity and ability to incorporate 14C-leucine into ferritin. Total liver nonheme Fe, ferritin protein and Fe and cathepsin D activity were also determined. Although parenteral Fe loading produced higher total nonheme Fe in livers than dietary loading, the Fe content of ferritin was .apprx. 20% in both groups, reflecting saturation of ferritin with Fe. Siderosome nonheme Fe content was > 40% in relation to protein. The siderosomes contained little buffer-soluble ferritin; on isoelectric focusing this was composed of isoferritins present also in cytosol ferritin. Buffer-insoluble ferritin protein, identified in siderosomes by immunofluorescence, was solubilized and contained immunoreactive material corresponding to L and H subunits of buffer-soluble ferritin. Transmission electron microscopy indicated the presence of relatively large quantities of ferritin in siderosomes, and it is argued that this was mostly buffer insoluble (denatured) or represented ferritin [FeOOH], cores divested of protein shells. Although siderosomes had substantial cathepsin D activity, the known resistance of ferritin to this and other proteases makes it unlikely that proteolysis is an early event in the decomposition of ferritin in siderosomes. Heavily Fe-laden siderosomes did not take up newly labeled ferritin or ferritin protein or 14C-precursor within 24 h of labeling, when 14C-labeled ferritin was abundant in cytosol. A sequence of steps leading from sequestration of buffer-soluble cytosol ferritin to storage of insoluble hemosiderin is proposed.