Iron is an essential nutrient for humans required for energy production, oxygen transport, cell proliferation, and pathogen destruction. Human infants undergo a critical period of neurodevelopment in the first year and iron availability during this time is crucial, as the imbalance of iron may result in disruptions to neurotransmitter synthesis, oligodendrocyte maturation and myelin production. Disruptions to neurodevelopment due to iron deficiency (ID) in the neonatal period can have lasting adverse effects on cognitive function even after iron repletion later in childhood.
The risk of ID in normal term infants is greatest around 4-6 months, the age when iron stores usually deplete. Unfortunately, human milk provides only 0.2 to 0.4 mg/L iron to sustain exclusively breastfed healthy term infants for the first 6 months of age. To combat the low iron concentration of breast milk and potential consequences from ID and iron deficiency anemia (IDA), infants at 4 to 6 months of age are often given formulas fortified with iron, which in the United States can contain up to 30 times more iron than in breast milk. Because the recommended intake of iron to maintain and meet growing demands is just 1 mg/kg/day of iron for healthy infants in the first 4 months, there is concern that infants are also at risk of developing iron overload from these formulas.
While the consequences of ID have been explored extensively, the exact effects of iron overload on the developing brain remain largely unknown. ID in animal models affects many neurodevelopmental processes, resulting in increased blood-brain-barrier permeability, altered energy metabolism in the brain, reduced of dendritic complexity, synapse dysfunction, impaired myelination, and deficits in spatial learning and memory. Conversely, accumulation of iron in the brain is observed in many neurodegenerative disorders such as Huntington’s Disease and Parkinson’s Disease, though the exact mechanisms of how iron overload contributes to these abnormalities are still unclear.
The current study sought to induce iron-deficiency, iron-repletion, and iron-excess in a pig model to identify effects of developmental ID and iron overload on neuronal morphology in the brain. This study focused on dendritic architecture of hippocampal pyramidal neurons because changes in the dendritic morphology of this neuronal cell types are linked to changes in learning and memory cognitive deficiencies. Pigs were chosen as the animal model because of their brain development, intestinal absorption, and iron storage are similar to those of humans. Thirty newborn piglets (15 male and 15 female) from five different sows were stratified by birth weight and litter and randomized to one of three treatment groups (n=10/treatment) on postnatal day (PD) 1. Piglets assigned to NONE (N), LOW (L) and HIGH (H) treatments received different doses of iron supplement during the pre-weaning (PD1 – 21) and post-weaning (PD22 – 35) periods. Piglets in N, L, and H group received oral iron supplementation at 0, 1 and 30 mg/(kg BW·day) once daily during the pre-weaning period and a diet containing < 30, 125, and 1000 mg iron/kg, respectively, during the post-weaning period. The pigs were weaned on PD22 and euthanized on PD35. Mucosal scrapings from the small intestine, as well as tissue samples from the small intestine, hippocampus, and liver were collected immediately after euthanasia. Blood samples were collected from the jugular vein weekly starting at PD1 until the endpoint. Overall, we successfully induced iron deficiency, repletion, and overload in young piglets by manipulating dietary iron intake. Both hemoglobin (Hb) and hematocrit (Hct) were increased by dietary iron in a dose-dependent manner (P < 0.05). Pigs in the N group were iron-deficient at PD14 and displayed growth retardation in the last week with significantly lower weight than pigs in the H group (P < 0.05), indicative of clinical signs of ID. Sholl analysis of basal dendrites of hippocampal neurons uncovered a significant treatment effect on the basilar dendritic arborization of CA1/3 pyramidal neurons (P ≤ 0.04). Pigs in the L group had fewer branching nodes and dendrites than H pigs (P < 0.05), and the difference in dendritic arborization was more prominent in distal higher order branches (> 3). In the piglet model, the developing hippocampus is susceptible to perturbations in dietary iron status, with iron deficiency and iron overload differentially affecting dendritic arborization. Interestingly, hippocampal iron overload in young pigs was associated with the greatest complexity of dendritic arborization in CA1/3 neurons among the 3 groups. We speculate that the unexpected finding of dendritic overgrowth induced by hippocampal iron overload is unique to the stage of early neurodevelopment. In conclusion, the current study has shown that early-life iron overload and ID have different effects on basal dendrites of CA1/3 pyramidal neurons in a piglet model
KEYWORDS: Iron overload, iron deficiency, hippocampal iron, dendritic arborization, piglet model