Pathways for the regulation of body iron homeostasis in response to experimental iron overload
Introduction
Secondary iron overload is a frequent clinical condition which can arise from inborn errors of haemoglobin synthesis, myelodysplastic syndromes or chemotherapy induced anaemia, which all require multiple blood transfusions [1], [2]. In addition, an inherited form of secondary iron overload is found in Southern Africa which has been linked to a defect in duodenal iron transport together with the consumption of iron rich traditional beer [3].
Secondary iron overload leads to progressive iron accumulation in the reticulo-endothelial system [1] with subsequent negative effects on cell mediated immune function, since iron inhibits interferon-γ inducible pathways in macrophages [4]. Accordingly, subjects with secondary iron overload are at a higher risk for tuberculosis and cancer [3]. In being a catalyst for the formation of highly toxic radicals, iron overload leads to tissue damage and organ failure [5], which is well known in primary iron overload, hereditary hemochromatosis, although with this condition iron is primarily deposited within parenchymal cells [6], [7], [8]. However, information of adaptive changes of body iron homeostasis upon secondary or experimental iron overload is rare.
Maintenance of body iron homeostasis is mainly regulated by duodenal iron absorption. Thereby, ferric iron is reduced at the luminal site by duodenal cytochrome b (Dcytb) [9], and ferrous iron is then transferred into the enterocyte by means of the transmembrane protein divalent metal transporter-1 (DMT-1) [10], [11]. At the basolateral site ferrous iron is exported from the enterocyte to the circulation via ferroportin [12], [13], [14], and after being oxidised by the membrane bound ferroxidase hephaestin [15] iron is incorporated into transferrin. In addition, up to 50% of absorbed iron may be in the heme form, which is taken up by a yet not characterised duodenal heme receptor [16], [17].
The expression of these iron transport genes is strongly regulated by body iron homeostasis, since the duodenal expression of DMT-1, Dcytb, ferroportin and to a lesser extent of hephaestin are increased with iron deficiency anaemia [9], [11], [15], [18], [19], [20], [21], [22]. The presence of iron responsive elements (IREs) within the untranslated regions of DMT-1 and ferroportin mRNA suggested that the expression of these proteins may be susceptible to posttranscriptional regulation by iron regulatory proteins (IRP) [23]. However, transcriptional regulation of DMT-1 and ferroportin expression by iron has also been anticipated [24], [25], [26].
In terms of regulation of duodenal iron absorption [27] transferrin receptor (TfR1) mediated uptake of circulating iron from the basolateral site of enterocytes and the subsequent modulation of IRP activity was believed to be the pivotal mechanism for sensing the body's needs for iron to the enterocyte [16], [17]. The membrane bound protein HFE which is mutated in approximately 80% of subjects with hereditary hemochromatosis [28], modulates the affinity of transferrin for TfR1 but its role in iron absorption remains elusive [29]. Importantly, the recent identification of the liver cationic peptide hepcidin has suggested that this molecule may be the principal regulator of iron absorption. [30], [31], [32]. The expression of hepcidin is increased when body iron stores are high [32], [33], [34]. Hepcidin over-expression resulted in reduced duodenal iron uptake and the development of anaemia [33], [35] while reduced hepcidin expression causes iron overload [36], [37], [38], [39]. This, may be traced back to a direct interaction of hepcidin with ferroportin expression, thus affecting iron transfer from the duodenal enterocyte to the circulation [40], [41]. The liver derived protein hemojuvelin may have additive effects to those of hepcidin [42].
In the study presented here, we used a mouse model of experimental iron overload to study adaptive changes of iron transport molecules in duodenum, liver and spleen in order to get new insights into regulation of iron homeostasis upon secondary iron overload.
Section snippets
Animal care
Male C57BL/6 mice were kept according to institutional and governmental guidelines on a standard rodent diet in the animal quarter at the University of Innsbruck.
Animals used for experiments were all between 12 and 16 weeks of age, with an average weight of 25 g. For induction of iron overload animals were daily intraperitoneally injected with 1 mg of iron dextran (venofer®) for up to 5 days.
Mice were anaesthetized, blood was collected by orbital puncture, and the animals were then sacrificed.
Results
When investigating tissue iron content following iron loading over time we found progressive iron accumulation in liver (Table 1). In the spleen, iron content first increased significantly over 3 days before it descended in spite of continuing iron loading (Table 1).
Discussion
We herein investigated the dynamic changes in the expression patterns of regulatory iron genes in different organs following experimental iron overload.
As duodenal DMT-1 mRNA and protein levels are surprisingly up regulated with prolonged experimental iron overload, DMT-1 appeared unlikely to be the gate keeper for the control of duodenal iron absorption. Accordingly, an in vivo radioactive iron uptake assay demonstrated that iron uptake into enterocytes is even higher in iron overloaded than
Acknowledgements
This study was supported by grants from the Austrian Research Fund, P-14215 and P-15943.
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