Ultrastructural cytochemistry and radioautography of hemoglobin—iron absorption

The subcellular route of hemoglobin-iron absorption by canine intestinal epithelial cells was investigated with cytochemical and radioautographic methods and compared to that observed for inorganic iron absorption. A solution of rabbit [⁵⁹Fe]hemoglobin or an inorganic-iron solution was injected into closed duodenal loops of beagle dogs and mucosal biopsies were obtained 15, 60, and 120 min thereafter. Mucosal retention of radioactivity in dogs given hemoglobin iron was 6.9–10.2% after 120 min, and portal venous blood radioactivity was confined to the inorganic iron fraction. In the same animals diaminobenzidine (DAB)-reactive heme was visualized in microendocytic caveolae at the bases of microvilli and in membrane-bound tubulovesicular structures and granules of the apical cytoplasm. DAB-reactive heme was not identified in the lateral intercellular space or the basal extracellular space. In the same specimens, acid ferrocyanide stained ferric iron in a few apical cytoplasmic vesicles as well as along the outer lateral and basal plasmalemma but failed to identify inorganic iron in microvilli. Both the microvilli and the lateral plasmalemma appeared stained in specimens from animals given inorganic iron. In radioautographic specimens of animals given [⁵⁹Fe]hemoglobin, silver grains were most frequently associated with the apical cytoplasm and the lateral plasmalemma. We conclude that at least one mechanism of hemoglobin-iron absorption involves endocytosis and degradation of hemoglobin in membrane-bound organelles with subsequent conversion of its iron to an inorganic form, which is then transported to the lateral intercellular space where subsequent processing is similar to that observed for absorbed inorganic iron.