STUDIES IN IRON METABOLISM

Abstract

An enzyme system incorporating inorganic iron into protoporphyrin has been demonstrated in human and rat liver, kidney, spleen and in human bone marrow and measured, using radioiron as a tracer. Studies on the enzyme content of the intracellular fractions of human and rat liver showed that while there was some activity in all fractions, the mitochondrial fraction was the most active. Dialysis of tissue homogenates led to a loss of enzymic activity which could be restored by the addition of ascorbic acid, GSH, cysteine or DPNH. The effect of these substances was not entirely due to an effect of reduction of the ferric iron since potentiation of activity resulted when these substances were added to a system containing iron in the ferrous form. Observations have also been made on the in vitro biosynthesis of haem from protoporphyrin and iron bound to siderophilin or to rat liver homogenate. Rat liver homogenate or a preparation from it of mitochondria served as the source of iron-incorporating enzyme. The reducing substances namely ascorbic acid, GSH, cysteine and DPNH, which were effective in increasing the incorporation into protoporphyrin of inorganic ferric iron also substantially increased the transfer of protein bound iron from haem biosynthesis. ATP enhanced the transfer of iron bound to siderophilin for incorporation into protophyrin but had no effect on the transfer of iron bound to rat liver protein. From this it is clear that physiological reducing substances play an important role in the transfer of iron from its protein-bound form for incorporation into protoporphyrin in the process of haem biosynthesis. A technique has been established for the measurement of iron incorporating enzyme activity in haemolysates of rabbit bone marrow containing a known number of red cell precursors. The method has been applied to marrows of normal and bled rabbits. In the bled animal, the enzymic activity of a unit number of cells was greatly decreased as compared to normal but was increased by the addition of reducing substances. This would indicate that the marrow responds to blood loss by an increase in the production of cells rather than an increased capacity of each cell to form haemoglobin. The enzymic activity of liver homogenates was also measured in normal and bled rabbits. In this tissue there was no difference in the enzymic activity per gram of liver between the normal and the bled animals. There was, however, a marked decrease in iron incorporation by liver cells from animals which had been injected with 300 mgs. Imferon. In two rabbits made iron deficient and anaemic there was an increase in iron incorporation as compared to normal. From this it is clear that gross changes in the size of the iron pool will vitiate the correct interpretation of results on the measurement of haem synthesis using radio-iron as a tracer. Nakao et al (1960) have suggested that plasma samples from a variety of haematological disorders possess a factor (or factors) stimulating haem synthesis as compared to normal plasma. The 'haem-stimulating activity' was measured by an in vitro technique in which radio-iron incorporation was used as a measure of haem synthesis. This work has been repeated using the same technique and results show that any marked increase in haem formation, measured in this way, could be explained by the diminution of the residual iron-binding capacity of the plasma sample. It is concluded that no definite evidence has yet been obtained for the presence of a specific haem-stimulating factor in human plasma. A statistically significant difference has been shown between the absorption of radio-iron given along with a standard meal, between patients with acid present in their gastric juice and patients with a histamine-fast achlorhydria. Both of these groups of patients were anaemic and there was an absence of stainable iron in the bone marrow of both groups. The mean absorption from the meal of the patients with acid in their gastric juice was 57.5 per cent while the mean absorption of those patients with histamine-fast achlorhyrdria was 18.5 per cent.