Copper supplementation in drinking water reverses dietary iron overload‐induced Cu‐deficiency anemia and cardiac hypertrophy in mice

Our previous studies showed that high‐iron (0.88% carbonyl iron) diet feeding of mice and rats lead to impaired growth, severe anemia, tissue copper depletion and cardiac hypertrophy. 1,2 Interestingly, these physiological perturbations were prevented by adding extra copper to the high Fe diet, proving that they were the consequence of severe copper deficiency. Whether these pathophysiological changes could be reversed by copper treatment, however, is unknown. The current study was thus undertaken to test the hypothesis that Cu supplementation of dietary Fe overload mice could restore systemic copper status, and reverse the anemia and cardiac hypertrophy. The experimental design was to fed weanling, male C57BL/6 mice AIN‐93G‐based diets containing adequate (~8 ppm) Cu in combination with adequate (~80 ppm) or high (~8000 ppm) Fe for 2 weeks. Then, the mice were given 20 mg/L Cu (as Cu sulfate) in drinking water for another 2 weeks. Subsequently, the iron and copper‐related phenotype of the mice was assessed. High‐Fe diet fed mice showed a significant growth impairment, accompanied by severe copper‐deficiency anemia, decreased serum ceruloplasmin (Cp) activity and cardiac hypertrophy, which were consistent with our previous studies. 1,2 Cu replenishment in the drinking water corrected deficits in hemoglobin (Hb) and hematocrit (HCT) levels. Additionally, serum Cp (i.e. amine oxidase) activity, a biomarker of copper status, was also restored after Cu supplementation. Cu treatment also reversed the cardiac hypertrophy in high‐Fe diet fed mice. We thus conclude that Cu supplementation corrects the physiologic perturbations associated with high Fe consumption in mice. These observations have important implications for investigators that utilize the dietary iron overload model in that high‐iron feeding should likely be accompanied by copper supplementation to prevent these noted physiologic perturbations that are not directly related to tissue iron accumulation. Moreover, this study has provided further evidence of physiologically‐relevant iron‐copper interactions in mammals. Support or Funding Information Supported by NIH grant 1R01 DK074867 (to J.F.C.). 1 Body weight, growth rate, and organ weights in C57BL/6 miceMale Weanling mice were fed with diet containing adequate Cu in combination with adequate or high Fe for 2 weeks. Then, the mice were given Cu contained water for 2 more weeks. Body weight (A) and growth rate (B) was measured at sacrifice and during 4 weeks, respectively. The relative heart weight (C), liver weight (D), spleen weight (E) and kidney weight (F) were measured and normalized to body weight.2 Hematological Parameters and Ceruloplasmin Activity in C57BL/6 MiceHemoglobin (A) and hematocrit (B) were determined from whole blood. Ceruloplasmin activity (C) was measure from serum. The trends in data were analyzed using a 2‐factor ANOVA test. Labeled means without a common letter differ (p<0.05, n=6). Values are means ± SDs.