Differential response of non-transferrin bound iron uptake in rat liver cells on long-term and short-term treatment with iron

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Abstract

Background: Uptake of non-transferrin-bound iron by the liver is important as a clearance mechanism in iron overload. In contrast to physiological uptake via receptor-mediated endocytosis of transferrin, no regulatory mechanisms for this process are known. This study compares the influence of long-term and shortterm depletion and loading of hepatocytes with iron on the uptake of non-transferrin bound iron, its affinity, specificity and the interaction with the transferrinmediated pathways.

Methods: Rats were fed iron-deficient, normal and 3,5,5-trimethylhexanoyl-ferrocene-containing diets to obtain livers with the corresponding desired status and the hepatocytes from these livers were used for transport studies. Hepatocytes from normal rats were depleted or loaded with iron by short-term treatment with desferrioxamine or ferric ammonium citrate, respectively. Uptake of non-transferrin bound iron was assayed from ferric citrate and from ferric diethylene triammine pentaacetate.

Results: Uptake of non-transferrin-bound iron in hepatocytes could be seen as consisting of a high-affinity (Km=600 nM) and a low-affinity component. Whereas in normal and in iron-starved rats the high-affinity component was more prominent, it disappeared altogether in hepatocytes from rats with iron overload resulting from prolonged feeding with TMH-ferrocene-enriched diet. Overloading also led to loss of inhibition by diferric transferrin, which occured in starved as well as normal cells. In contrast, shortterm iron-depletion of isolated hepatocytes with desferrioxamine had only a weak stimulatory effect, whereas treatment with ferric ammonium citrate strongly increased the uptake rates. However, the inhibition by diferric transferrin also disappeared. In both cases, uptake of non-transferrin bound iron was inhibited by apotransferrin.

Conclusions: Non-transferrin bound iron uptake in liver cells is apparently regulated by the iron status of the liver. The mode of response to iron loading depends on the method of loading in terms of time course and the form of iron used. It cannot be explained by the behavior of the iron regulatory protein, and it is complex, seeming to involve more than one transport system.

Section snippets

Animals and feeding protocol

Female Wistar rats (120–140 g) were obtained from Wiga (Hannover, Germany). One group of rats was fed with Altromin C1000 standard rat diet with an iron content of 160 μg/g of food (normal rats). Four experimental groups of animals receiving different diets on the basis of iron-deficient Altromin C1038 (Altromin, Lage, Germany) were established. One group received this iron-poor diet (6 μg Fe/g diet) for 6 weeks (iron-deficient rats). Two groups of rats were fed a diet of C1038 enriched with

Results

The iron content of hepatocytes from rats fed a normal diet, from iron-depleted rats and from animals overloaded with iron by feeding with TMH-ferrocene or carbonyl-iron was determined and was correlated with properties of NTBI uptake into the hepatocytes isolated from these livers with respect to absolute rates, affinity, specificity and inhibition by apo- and holotransferrin at different concentrations.

Discussion

Iron metabolism in the mammalian organism is tightly regulated under normal conditions. The blood plasma contains an excess of the binding protein transferrin to avoid the occurrence of free, non-protein bound iron with high redox potential, which is highly toxic. Cells acquire iron by receptor-mediated endocytosis of transferrin, which is limited by the concentration of receptors at the cell surface. This, in turn, is regulated at the post-transcriptional level by one or two iron regulatory

Acknowledgements

This work was supported by the Austrian Research Fund, Proj.Nr, P-11594-Med.

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