The Transfer of Iron to Reticulocytes by Synthetic Chelating Agents*

Abstract

Iron, once absorbed from the gastrointestinal tract, is distributed throughout the body by the specific iron-binding protein (IBP, transferrin, siderophilin) (1). Iron binding by IBP is presumed to be of a chelate type in which the hexacoordination capacity of the ferric ion is satisfied by linkage with three phenolic oxygens, one carboxyl group, and a bicarbonate ion on the remaining coordination position (2-4). Since IBP has been shown to have a highly selective ability to deliver iron to the immature red cell (5), we have utilized various synthetic iron chelates in an attempt to delineate the characteristics for iron binding essential to carry out this function of the protein. MethodsPreparation of the reticulocyte-rich blood. Reticulocytosis was produced in adult albino rabbits (3 to 5 kg) by removing approximately 5 to 10%v of the total blood volume (30 to 40 ml) daily for 3 or 4 days by repeated cardiac puncture. Two days thereafter, a final cardiac puncture delivered all available blood, which was immediately placed into iron-free, heparinized glass containers. Reticulocyte count using the new methylene blue stain technique (6) indicated 10 to 25%, immature cells.The red cells were separated from the plasma and leukocytes by centrifugation at 1,000 X g for 15 minutes at 40 C. The harvested cells were washed and centrifuged three times with 5 vol of iron-free, phosphate

Iron-donating properties of transferrin

“…We used this procedure to establish that about 93% of the -Felabeled iron nitrilotriacetate added to preincubated transferrin in dilute solution was actually bound. This is important because iron nitrilotriacetate, itself, is an efficient source of iron for hemoglobin synthesis bv reticulocytes (Princiotto et al, 1964;Morgan, 1971). The demonstration of iron binding in this type of experiment, and especially for in vivo experiments, seems mandatory.…”

Biliary Excretion of Chelated Iron.

“…In hemolytic anemia, for example, the biliary iron in humans increases Sfold (4). We have also shown that uptake of iron by the reticulocyte in an in vitro system can be sharply modified by the nature of the iron chelate (7). We have also shown that uptake of iron by the reticulocyte in an in vitro system can be sharply modified by the nature of the iron chelate (7).…”

“…The significant structural features which brought about this result were the net neutral charge and lipid solubility of the iron chelates (7,16). Support for this view is provided by the fact that introduction of a charged anionic site into the parent molecular structure brings about a complete disappearance of the ability of the compounds to transport iron into the bile.…”

Distribution of Intravenously Administered Ferrioxamine-59 and Ferric N, N'-Ethylenebis (a Imino-2-Hydroxy-5-Chlorophenylacetate) in Rats

“…Ferrioxamine and ferric N,.V' ethylenebis-( a imino-2 -hydroxy-5-chlorophenylacetate) (FeCI-EDDHA) are two iron chelates of differing molecular architecture whose iron binding strength exceeds that of iron transferrin (3,4 ) . The present study suggests that the molecular environment of the metal is a major determinant in its biological fate.…”

“…Information about the composition of meconium in meconium ileus (MI) is accumulating (1)(2)(3)(4)). An early report ( 1) indicated that MI meconium contained similar water, 13 times more trichlmacetic acid (TCA)-insoluble nitrogen, and 4 times less carbohydlrate than is found in no& m m nium.…”