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Measurement of Tissue Non-Heme Iron Content using a Bathophenanthroline-Based Colorimetric Assay
Published on: January 31, 2022
Belgrade rats display liver iron loading
Khristy Thompson1, Ramon M Molina, Joseph D Brain
1Department of Genetics and Complex Diseases, Harvard School of Public Health, Boston, MA, 02115, USA.
Abstract:
Patients with mutations in divalent metal transporter-1 (DMT1), an intestinal nonheme iron transporter, suffer from microcytic anemia and hepatic iron loading. DMT1 is also mutated in Belgrade rats, an animal model with a thalassemic-like disorder of microcytic anemia with hyperferrinemia. However, aspects of hepatic iron loading in this genetic model are not well characterized. To more fully define the Belgrade rat's iron status, we compared the characteristics of homozygous (b/b) and heterozygous (b/+) rats fed an iron-supplemented diet for 3 wk postweaning. Dietary supplementation with ferrous iron improved the anemia of b/b rats insofar as hematocrits increased from 0.13 (21-d-old) to 0.31 (42-d-old). However, hematocrits remained significantly lower than those of age-matched b/+ rats (0.36 and 0.41 in 21- and 42-d-old heterozygotes, respectively, P < 0.05). Wright's staining of b/b red cells confirmed the hypochromic microcytic nature of Belgrade rats' anemia. The liver iron concentration of 42-d-old b/b rats was greater than in age-matched b/+ rats (5.97 vs. 2.24 mumol/g, P < 0.05). Whereas Perls' Prussian blue iron staining was evident in both periportal and centrilobular regions in 42-d-old b/b liver sections, no staining was observed in age-matched b/+ tissue sections. Quantitative real-time PCR analysis showed that expression of liver hepcidin mRNA in 42-d-old b/b rats was 3-fold greater than age-matched b/+ rats. These results indicate that, similar to human patients with DMT1 mutations, Belgrade rats also display hepatic iron loading. Our data suggest this condition arises from ineffective erythropoiesis.
Insights
Belgrade rats with divalent metal transporter-1 (DMT1) mutations exhibit microcytic anemia and hepatic iron loading, similar to human patients. These findings suggest ineffective erythropoiesis contributes to iron overload in this genetic model.
Area of Science:
- Biochemistry
- Genetics
- Hematology
Background:
- Mutations in divalent metal transporter-1 (DMT1) cause microcytic anemia and hepatic iron loading in humans.
- Belgrade rats, with a DMT1 mutation, serve as a genetic model for thalassemic-like disorder, exhibiting anemia and hyperferrinemia.
- Hepatic iron loading aspects in Belgrade rats are not fully characterized.
Purpose of the Study:
- To comprehensively define the iron status of Belgrade rats, particularly focusing on hepatic iron loading.
- To compare iron metabolism characteristics between homozygous (b/b) and heterozygous (b/+) Belgrade rats.
- To investigate the link between DMT1 mutations, anemia, and iron accumulation.
Main Methods:
- Comparative analysis of homozygous (b/b) and heterozygous (b/+) Belgrade rats fed an iron-supplemented diet.
- Hematological analysis, including hematocrit measurements and red blood cell morphology (Wright's staining).
- Quantification of liver iron concentration and iron distribution (Perls' Prussian blue staining).
- Gene expression analysis of liver hepcidin mRNA using quantitative real-time PCR.
Main Results:
- Dietary iron improved anemia in b/b rats, but hematocrits remained lower than b/+ rats.
- b/b rats showed significantly higher liver iron concentration compared to b/+ rats.
- Hepatic iron deposition was observed in b/b rats, absent in b/+ rats.
- Liver hepcidin mRNA expression was 3-fold higher in b/b rats than in b/+ rats.
Conclusions:
- Belgrade rats with DMT1 mutations exhibit hepatic iron loading, mirroring human patient conditions.
- The study confirms hepatic iron loading in this animal model.
- Ineffective erythropoiesis is suggested as the underlying cause of hepatic iron loading in Belgrade rats.
