Iron excretion in iron dextran-overloaded mice

Abstract

Background: Iron homeostasis in humans is tightly regulated by mechanisms aimed to conserve iron for reutilisation, with a negligible role played by excretory mechanisms. In a previous study we found that mice have an astonishing ability to tolerate very high doses of parenterally administered iron dextran. Whether this ability is linked to the existence of an excretory pathway remains to be ascertained.

Materials and methods: Iron overload was generated by intraperitoneal injections of iron dextran (1 g/kg) administered once a week for 8 weeks in two different mouse strains (C57bl/6 and B6D2F1). Urinary and faecal iron excretion was assessed by inductively coupling plasma-mass spectrometry, whereas cardiac and liver architecture was evaluated by echocardiography and histological methods. For both strains, 24-hour faeces and urine samples were collected and iron concentration was determined on days 0, 1 and 2 after iron administration.

Results: In iron-overloaded C57bl/6 mice, the faecal iron concentration increased by 218% and 157% on days 1 and 2, respectively (p<0.01). The iron excreted represented a loss of 14% of total iron administered. Similar but smaller changes was also found in B6D2F1 mice. Conversely, we found no significant changes in the concentration of iron in the urine in either of the strains of mice. In both strains, histological examination showed accumulation of iron in the liver and heart which tended to decrease over time.

Conclusions: This study indicates that mice have a mechanism for removal of excess body iron and provides insights into the possible mechanisms of excretion.

Figure 1

Histological effects of iron accumulation on mice liver after iron dextran administration. (a,b) Representative haematoxylin and eosin staining of liver sections. Panel (a) shows the histology of C57bl/6 control mouse liver (×100), with normal lobular architecture and without signs of iron accumulation. Panel (b) shows marked iron deposition in Küppfer cells (arrow) and, to a lesser extent, in hepatocytes of C57bl/6 mice treated with iron dextran (×100). (c, d) Picrosirius red (×100) and Prussian blue (×200) staining of liver sections in C57bl/6 iron-overloaded mice, respectively. Iron accumulation is clearly evident (arrow). However no signs of fibrosis are present. (e, f) Representative haematoxylin and eosin staining (×25) of iron-treated mouse liver at 9 and 13 weeks, respectively (see Material and methods section for more details). The iron stores in the liver appeared reduced in mice evaluated at 13 weeks compared to those at 9 weeks. Vehicle, phosphate buffer saline-treated mice; Iron, iron-treated mice.

Figure 2

Preserved cardiac structure and function in iron dextran-loaded mice after 13 weeks. (a) Representative transthoracic echocardiographic tracings from the vehicle-treated and iron-treated mice; LV, left ventricle. (b) Fractional shortening (FS), (c) left ventricular end-diastolic diameter (LV EDD), and (d) left ventricular posterior wall thickness (LV PWT). Echocardiographic analysis revealed no significant changes between vehicle and iron-treated groups. (e and f) Representative Prussian blue and picrosirius red staining. (e) Prussian blue staining shows visible accumulation of iron both inside and outside of cardiomyocytes in iron-treated mice (bottom panel). (f) Conversely, there is no collagen accumulation in iron-treated mice as evidenced by picrosirius red staining (middle and bottom panels). Vehicle, phosphate buffer saline-treated mice; iron, iron-treated mice.