Iron-fortified formulas and iron drops (both usually ferrous sulfate, FS) prevent early life iron deficiency, but may delay growth and adversely affect neurodevelopment by providing excess iron. We ...used a rat pup model to investigate iron status, growth, and development outcomes following daily iron supplementation (10 mg iron/kg body weight, representative of iron-fortified formula levels) with FS or an alternative, bioavailable form of iron, ferrous bis-glycinate chelate (FC). On postnatal day (PD) 2, sex-matched rat litters (
= 3 litters, 10 pups each) were randomly assigned to receive FS, FC, or vehicle control until PD 14. On PD 15, we evaluated systemic iron regulation and CNS mineral interactions and we interrogated iron loading outcomes in the hippocampus, in search of mechanisms by which iron may influence neurodevelopment. Body iron stores were elevated substantially in iron-supplemented pups. All pups gained weight normally, but brain size on PD 15 was dependent on iron source. This may have been associated with reduced hippocampal oxidative stress but was not associated with CNS mineral interactions, iron regulation, or myelination, as these were unchanged with iron supplementation. Additional studies are warranted to investigate iron form effects on neurodevelopment so that iron recommendations can be optimized for all infants.
Infants are frequently supplemented with iron to prevent iron deficiency, but iron supplements may have adverse effects on infant health. Although iron supplements can be highly effective at ...improving iron status and preventing iron deficiency anemia, iron may adversely affect growth and development, and may increase risk for certain infections. Several reviews exist in this area; however, none has fully summarized all reported outcomes of iron supplementation during infancy. In this review, we summarize the risks and benefits of iron supplementation as they have been reported in controlled studies and in relevant animal models. Additionally, we discuss the mechanisms that may underly beneficial and adverse effects.
Iron supplements are frequently provided to infants in high-income countries despite low incidence of iron deficiency. There is growing concern regarding adverse health and development outcomes of ...excess iron provision in early life. Excess iron may directly damage developing organs through the formation of reactive oxygen species, alter systemic inflammatory signaling, and/or dysregulate trace mineral metabolism. To better characterize the in vivo effects of excess iron on development, we utilized a pre-weanling rat pup model. Lewis rat litters were culled to eight pups (four males and four females) and randomly assigned to daily supplementation groups receiving either vehicle control (CON; 10% w/v sucrose solution) or ferrous sulfate (FS) iron at one of the following doses: 10, 30, or 90 mg iron/kg body weight—FS-10, FS-30, and FS-90, respectively—from postnatal day (PD) 2 through 9. FS-90 litters, but not FS-30 or FS-10, failed to thrive compared to CON litters and had smaller brains on PD 10. Among the groups, FS-90 liver iron levels were highest, as were white blood cell counts. Compared to CON, circulating MCP-1 and liver zinc were increased in FS-90 pups, whereas liver copper was decreased. Growth defects due to excess FS provision in pre-weanling rats may be related to liver injury, inflammation, and altered trace mineral metabolism.
The gut microbiota is implicated in the adverse developmental outcomes of postnatal iron supplementation. To generate hypotheses on how changes to the gut microbiota by iron adversely affect ...development, and to determine whether the form of iron influences microbiota outcomes, we characterized gut microbiome and metabolome changes in Sprague-Dawley rat pups given oral supplements of ferrous sulfate (FS), ferrous bis-glycinate chelate (FC), or vehicle control (CON) on postnatal day (PD) 2−14. Iron supplementation reduced microbiome alpha-diversity (p < 0.0001) and altered short-chain fatty acids (SCFAs) and trimethylamine (TMA) in a form-dependent manner. To investigate the long-term effects of iron provision in early life, an additional cohort was supplemented with FS, FC, or CON until PD 21 and then weaned onto standard chow. At ~8 weeks of age, young adult (YA) rats that received FS exhibited more diverse microbiomes compared to CON (p < 0.05), whereas FC microbiomes were less diverse (p < 0.05). Iron provision resulted in 10,000-fold reduced abundance of Lactobacilli in pre-weanling and YA animals provided iron in early life (p < 0.0001). Our results suggest that in pre-weanling rats, supplemental iron form can generate differential effects on the gut microbiota and microbial metabolism that persist into adulthood.
Milk fat globule membrane (MFGM) is a glycosylated, protein-embedded, phospholipid fraction that surrounds triglycerides in milk. Commercial bovine sources have recently come to the market as a novel ...food ingredient and have been added to various products, including infant formula. Considering that MFGM is a heterogeneous mixture of fat, protein, and carbohydrate, it can be expected that variations among MFGM products exist. For this reason, our aim was to characterize the composition of commercial MFGM samples through a combination of proteomic and lipidomic analyses. Six bovine milk fractions, represented as MFGM fractions or phospholipid fractions, were obtained from various commercial sources. Additionally, the MFGM samples were compared with 2 infant formulas, a standard formula as well as a premium formula containing MFGM. For proteomic analysis, bottom-up data-dependent liquid chromatography–tandem mass spectrometry (LC-MS/MS) was performed on each MFGM fraction, and nearly a thousand proteins were identified across all samples, with 364 of them having different abundance across the samples tested. One hundred twelve proteins differed by a fold-change of 10 or greater, 14 by a fold-change of 50, and 2 by a fold-change of 100 in at least 1 pair, suggesting large differences in the proteins present in these fractions. Even though the classical MFGM proteins were enriched in the MFGM fractions, the relative protein composition varied considerably, and all contain an abundance of milk (casein and whey) proteins. Lipidomic analysis identified a total of 393 lipid species across both positive and negative ionization modes, with the major classes detected being triglycerides, sphingomyelins, and several phospholipids. Across all samples, triglycerides comprised at least 50% of total lipids, with phosphatidylcholine and sphingomyelin being the second and third most abundant lipid classes, respectively. These findings demonstrate the heterogeneous nature of various bovine commercial MFGM fractions. This variation must be considered when evaluating and describing potential functional benefits of these products shown in clinical trials.
Iron deficiency (ID) during infancy is harmful to health and development. Iron supplements such as iron drops and fortified infant formula prevent ID effectively but typically provide 10-20x more ...iron than breast milk and may have adverse health and development effects when provided to infants not at high risk for ID. Adverse effects on growth, gut microbiota development, trace mineral status, neurodevelopment, and risk of morbidity have been observed, but these effects have been poorly understood. These effects and the underlying mechanisms were investigated underlying utilizing a pre-weanling rat supplementation model. In the first study, Sprague Dawley rat litters were randomly assigned to receive daily vehicle control (CON) supplementation, or 10 mg iron/kg body weight (BW) (representative of the typical daily iron intake from fortified formula) as either ferrous sulfate (FS) or ferrous bis-glycinate chelate (FC), a novel bioavailable form of iron. FS and FC groups had comparable liver iron, hemoglobin, and hematocit values that were higher than CON (p < 0.0001) at postnatal day (PD) 15 after 2 weeks of supplementation. BW gain was unaffected by group, but FS brains were heavier than FC brains (p < 0.05). In the second study, short- and long-term effects of routine iron levels on gut microbiota development were assessed. Iron supplementation induced over 10,000-fold loss of Lactobacillus commensal bacteria in the gut compared to CON. Gut microbiome composition and diversity depended on iron form: FS and FC gut microbiome communities were distant, and while iron reduced gut microbiota diversity, FC microbiomes were even less diverse than FS as compared to CON. Long-term effects of iron were revealed when an additional cohort of groups were supplemented with FS, FC, or CON up to weaning: adult gut microbiome compositions 6 weeks after weaning depicted a 10,000-fold loss of Lactobacillus if they had received iron prior to weaning, and overall microbiome also which form was provided (FS or FC). The results of the first two studies concluded that iron provision prior to weaning elevates iron status beyond needs and adversely effects long-term microbiome composition. Additionally, it was concluded that microbiome and development effects depend on the form of iron provided. A final FS dose response study sought to identify the role of the excess iron dose in adverse health and development outcomes and found that increasing daily iron supplementation to 90 mg iron/kg BW further elevated liver iron loading, reduced pre-weanling rat weight gain and brain size, elevated inflammation, and altered levels of copper and zinc in the liver. The results of the final experiment suggest that mineral interactions and inflammatory signaling are implicated in adverse growth and development effects of excess iron. In summary, results from all three studies support that excess iron disrupts growth and development, and these effects depend upon iron form and dose. The findings provide novel evidence it is likely that the gut microbiota, inflammatory signaling, and mineral interactions may play important roles in the adverse outcomes of iron provision during infancy.
•Bmp5-mutant mice have low hepcidin in early life and exhibit mild iron loading under low- or high-iron or Bmp6-heterozygous conditions.•Bmp5 mutations exacerbate hemochromatosis and abrogate ...hepcidin responses to iron in Bmp6-knockout mice.
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Hepcidin is the master regulator of systemic iron homeostasis. The bone morphogenetic protein (BMP) signaling pathway is a critical regulator of hepcidin expression in response to iron and erythropoietic drive. Although endothelial-derived BMP6 and BMP2 ligands have key functional roles as endogenous hepcidin regulators, both iron and erythropoietic drives still regulate hepcidin in mice lacking either or both ligands. Here, we used mice with an inactivating Bmp5 mutation (Bmp5se), either alone or together with a global or endothelial Bmp6 knockout, to investigate the functional role of BMP5 in hepcidin and systemic iron homeostasis regulation. We showed that Bmp5se-mutant mice exhibit hepcidin deficiency at age 10 days, blunted hepcidin induction in response to oral iron gavage, and mild liver iron loading when fed on a low- or high-iron diet. Loss of 1 or 2 functional Bmp5 alleles also leads to increased iron loading in Bmp6-heterozygous mice and more profound hemochromatosis in global or endothelial Bmp6-knockout mice. Moreover, double Bmp5- and Bmp6-mutant mice fail to induce hepcidin in response to long-term dietary iron loading. Finally, erythroferrone binds directly to BMP5 and inhibits BMP5 induction of hepcidin in vitro. Although erythropoietin suppresses hepcidin in Bmp5se-mutant mice, it fails to suppress hepcidin in double Bmp5- and Bmp6-mutant males. Together, these data demonstrate that BMP5 plays a functional role in hepcidin and iron homeostasis regulation, particularly under conditions in which BMP6 is limited.
Hepcidin is the master regulator of iron homeostasis and is regulated by bone morphogenetic protein (BMP) signaling in response to iron loading, iron deficiency, and erythropoietic drive. This regulation is largely a reflection of BMP6 and BMP2, but the role of BMP5 has not been defined. Xiao et al probed the contribution of BMP5 signaling to iron homeostasis, demonstrating that it also has a role in hepcidin regulation. BMP6 heterozygous or homozygous-null mice have increased iron loading that is further amplified with concomitant BMP5 knockout. This further elucidates the complex process of hepcidin regulation and iron metabolism.
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