Maternal oral probiotic use is associated with decreased breastmilk inflammatory markers, infant fecal microbiome variation, and altered recognition memory responses in infants-a pilot observational study.

Introduction: Early life gut microbiomes are important for brain and immune system development in animal models. Probiotic use has been proposed as a strategy to promote health via modulation of microbiomes. In this observational study, we explore if early life exposure to probiotics via the mother during pregnancy and lactation, is associated with decreased inflammation in breastmilk, maternal and infant microbiome variation, and altered infant neurodevelopmental features. Methods: Exclusively breastfeeding mother-infant dyads were recruited as part of the "Mothers and Infants Linked for Healthy Growth (MILk) Study." Probiotic comparison groups were defined by exposure to maternal probiotics (NO/YES) and by timing of probiotic exposure (prenatal, postnatal, total). C-reactive protein (CRP) and IL-6 levels were determined in breastmilk by immunoassays, and microbiomes were characterized from 1-month milk and from 1- and 6-month infant feces by 16S rDNA sequencing. Infant brain function was profiled via electroencephalogram (EEG); we assessed recognition memory using event-related potential (ERP) responses to familiar and novel auditory (1 month) and visual (6 months) stimuli. Statistical comparisons of study outcomes between probiotic groups were performed using permutational analysis of variance (PERMANOVA) (microbiome) and linear models (all other study outcomes), including relevant covariables as indicated. Results: We observed associations between probiotic exposure and lower breastmilk CRP and IL-6 levels, and infant gut microbiome variation at 1- and 6-months of age (including higher abundances of Bifidobacteria and Lactobacillus). In addition, maternal probiotic exposure was associated with differences in infant ERP features at 6-months of age. Specifically, infants who were exposed to postnatal maternal probiotics (between the 1- and 6-month study visits) via breastfeeding/breastmilk, had larger differential responses between familiar and novel visual stimuli with respect to the late slow wave component of the EEG, which may indicate greater memory updating potential. The milk of mothers of this subgroup of infants had lower IL-6 levels and infants had different 6-month fecal microbiomes as compared to those in the "NO" maternal probiotics group. Discussion: These results support continued research into "Microbiota-Gut-Brain" connections during early life and the role of pre- and postnatal probiotics in mothers to promote healthy microbiome-associated outcomes in infants.

Phlebotomy-induced anemia reduces oxygen-induced retinopathy severity and dampens retinal developmental transcriptomic pathways in rats.

BACKGROUND: Phlebotomy-induced-anemia (PIA), which induces tissue hypoxia and angiogenesis, occurs universally among infants at risk for severe retinopathy of prematurity (ROP). We hypothesized that PIA exacerbates pathologic retinal neovascularization in ROP. METHODS: We induced PIA to a hematocrit of 18% among rats undergoing the established 50/10 oxygen-induced retinopathy (OIR) model. Rats were euthanized at P15 and P20, during the avascular and neovascular phases of OIR, respectively. Retinal vascular morphometry, cytokine/chemokine concentrations, transcriptomes, and mRNA expression of angiogenic and iron-deficiency markers were compared to non-PIA controls. RESULTS: In OIR, PIA decreased percent avascular area at P15 by 35%, percent neovascular area at P20 by 42%, and select pro-inflammatory cytokine/chemokine concentrations at both time points. At P20, PIA increased mRNA expression of angiopoietin 2/ vascular endothelial growth factor-A 2-fold and transferrin and transferrin receptor 5-fold. RNA sequencing showed dampened pathways of angiogenesis, inflammation, and neural development in anemic OIR females. CONCLUSION: Contrary to our hypothesis, PIA decreased OIR severity and retinal cytokine and chemokine levels and dampened transcriptomic pathways central to retinal vascular and neural development in neonatal rats. These data suggest PIA provides a protective effect from OIR. Further investigation into the functional effect of these molecular changes is warranted. IMPACT: This is the first preclinical study to investigate the impact of neonatal anemia on oxygen-induced retinopathy (OIR) outcomes. This study adds to the literature that anemia decreases neovascularization, decreases cytokine and chemokine levels, and dampens angiogenic and neural transcriptomic pathways in the rat 50/10 OIR model. The study identifies a sex-specific transcriptomic response to anemia in the 50/10 OIR model, with females primarily impacted.

Exploring the association between haematological markers of iron and general movements in 4-month-old infants exposed to anaemia in-utero.

BACKGROUND: Iron is a vital micronutrient for brain development, influencing myelination, neurotransmitter balance, and the maturation of specific brain cells. Hence iron insufficiency in the foetal, neonatal and infancy period has the potential to influence the neuromotor development. AIMS: We aimed to describe haematological markers of iron at 4 months of age in infants exposed to prenatal anaemia and explore the association with their quality of general movements. STUDY DESIGN: Cross sectional study nested within the RAPIDIRON-KIDS trial. SUBJECTS: All infants whose mothers were part of RAPIDIRON-KIDS trial, were eligible to participate in this study when the infants were 4 months old. Children suffering from fever or acute illness on the day of assessment, or with a history of either surgery, or admission to hospital in the first month were excluded. OUTCOME MEASURES: Haematological markers of iron (Haemoglobin and Ferritin level) and quality of general movements in infants at 4 months of age. RESULTS: 120 infants were assessed with mean birth weight of 2685.5 g (±384.5) and median gestational age of 39 weeks [Q1, Q3:38,40]. There was no significant association between haemoglobin or ferritin levels with fidgety movements (p = 0.18 and p = 0.27, respectively). The combined effect of haemoglobin and ferritin estimates also did not show any significant association with the study groups (p = 0.21). CONCLUSION: A majority of infants still had low iron indices at 4 months of age and this was not associated with the quality of general movements. A prospective longitudinal study needs to be considered in infants exposed to prenatal anaemia rather than assessing the outcomes at a single time point.

Sex differences in the association of pretransfusion haemoglobin and cognition in preterm infants.

OBJECTIVES: To assess sex-specific differences in the association between pre-transfusion haemoglobin values and early neurodevelopmental function. DESIGN: Observational follow-up of infants with birth weights <1000 g and gestational ages 22-28 weeks who were enrolled in the NICHD Neonatal Research Network Transfusion of Prematures (TOP) Trial at 19 U.S. sites, 2012-2017. MAIN OUTCOME MEASURES: Pretransfusion haemoglobin values were obtained longitudinally through 36 weeks' postmenstrual age. The infant's mean pretransfusion haemoglobin was used as a marker of degree of anaemia (n=1655 measures). Measures of brain function were obtained at 22-26 months' corrected age using the Bayley Scales of Infant & Toddler Development, third edition (BSID-III) (n=1290 BSID-III scores). Sex-specific estimates for the linear relation between pretransfusion haemoglobin and BSID-III scores were obtained from repeated-measures regression analysis, adjusted for gestational age, birth weight, study site, clinical characteristics, and demographic covariates. RESULTS: The relation of pretransfusion haemoglobin with 24-month BSID-III scores showed significant, independent interactions with both (1) sex (p=0.046) and (2) retinopathy of prematurity (ROP; p=0.004). In 614 males, BSID-III scores were higher by 1.07 points per g/dL (95% CI 1.58 to 4.33; p=0.008), not differing significantly among the three subscales (cognitive, language and motor; p=0.94). In 247 infants with ROP, BSID-III scores were higher by 2.95 points per g/dL (95% CI 0.28 to 1.87; p<0.0001), uniformly across subscales (p=0.73). These associations were non-significant in 676 females (p=0.96) and 1043 infants without ROP (p=0.81). CONCLUSIONS: This study demonstrates sex-specific associations between mean pretransfusion haemoglobin (a marker of the severity of anaemia throughout the neonatal intensive care unit [NICU] hospitalisation) and early neurodevelopmental function at 22-26 months' corrected age.

Chromatin accessibility and H3K9me3 landscapes reveal long-term epigenetic effects of fetal-neonatal iron deficiency in rat hippocampus.

BACKGROUND: Iron deficiency (ID) during the fetal-neonatal period results in long-term neurodevelopmental impairments associated with pervasive hippocampal gene dysregulation. Prenatal choline supplementation partially normalizes these effects, suggesting an interaction between iron and choline in hippocampal transcriptome regulation. To understand the regulatory mechanisms, we investigated epigenetic marks of genes with altered chromatin accessibility (ATAC-seq) or poised to be repressed (H3K9me3 ChIP-seq) in iron-repleted adult rats having experienced fetal-neonatal ID exposure with or without prenatal choline supplementation. RESULTS: Fetal-neonatal ID was induced by limiting maternal iron intake from gestational day (G) 2 through postnatal day (P) 7. Half of the pregnant dams were given supplemental choline (5.0 g/kg) from G11-18. This resulted in 4 groups at P65 (Iron-sufficient [IS], Formerly Iron-deficient [FID], IS with choline [ISch], and FID with choline [FIDch]). Hippocampi were collected from P65 iron-repleted male offspring and analyzed for chromatin accessibility and H3K9me3 enrichment. 22% and 24% of differentially transcribed genes in FID- and FIDch-groups, respectively, exhibited significant differences in chromatin accessibility, whereas 1.7% and 13% exhibited significant differences in H3K9me3 enrichment. These changes mapped onto gene networks regulating synaptic plasticity, neuroinflammation, and reward circuits. Motif analysis of differentially modified genomic sites revealed significantly stronger choline effects than early-life ID and identified multiple epigenetically modified transcription factor binding sites. CONCLUSIONS: This study reveals genome-wide, stable epigenetic changes and epigenetically modifiable gene networks associated with specific chromatin marks in the hippocampus, and lays a foundation to further elucidate iron-dependent epigenetic mechanisms that underlie the long-term effects of fetal-neonatal ID, choline, and their interactions.

Identification of Genes Responding to Iron or Choline Treatment for Early-Life Iron Deficiency in the Male Rat Hippocampal Transcriptomes.

BACKGROUND: Developmental iron deficiency (ID) is associated with long-term cognitive and affective behavioral impairments in humans. Preclinical studies have shown that developmental ID has short- and long-term effects on gene regulation. Prenatal choline supplementation partially rescues early-life ID-induced cognitive deficits in adult male rats. OBJECTIVES: To identify acute and long-term changes in biological processes regulated by developmental ID and modifiable by choline. METHODS: This study compares the hippocampal transcriptomes of postnatal day (P) 15 iron-deficient (acute) and P65 formerly ID (persistent) rats with or without prenatal choline treatment. Pregnant rats were fed an ID (4 mg/kg Fe) or iron-sufficient (IS) (200 mg/kg Fe) diet from gestational day (G) 2 to P7 with or without choline supplementation (5 g/kg choline) from G11 to G18. Hippocampi were collected from P15 or P65 offspring and analyzed for gene expression by RNA sequencing. RESULTS: Developmental ID-induced changes suggested modified activity of oxidative phosphorylation and fatty acid metabolism. Prenatal choline supplementation induced robust changes in gene expression, particularly in iron-deficient animals, where it partially mitigated the early-life ID-dysregulated genes. Choline supplementation also altered the hippocampal transcriptome in the IS rats, with indications for both beneficial and adverse effects. CONCLUSIONS: This study provided global assessments of gene expression regulated by iron and choline. Our new findings highlight genes responding to iron or choline treatments, including a potentially novel choline-regulated transporter (IPO7), with shared effects on neuroinflammation in the male rat hippocampus.

Prognostic Performance of Hematological and Serum Iron and Metabolite Indices for Detection of Early Iron Deficiency Induced Metabolic Brain Dysfunction in Infant Rhesus Monkeys.

BACKGROUND: The current pediatric practice of monitoring for infantile iron deficiency (ID) via hemoglobin (Hgb) screening at one y of age does not identify preanemic ID nor protect against later neurocognitive deficits. OBJECTIVES: To identify biomarkers of iron-related metabolic alterations in the serum and brain and determine the sensitivity of conventional iron and heme indices for predicting risk of brain metabolic dysfunction using a nonhuman primate model of infantile ID. METHODS: Simultaneous serum iron and RBC indices, and serum and cerebrospinal fluid (CSF) metabolomic profiles were determined in 20 rhesus infants, comparing iron sufficient (IS; N = 10) and ID (N = 10) infants at 2 and 4 mo of age. RESULTS: Reticulocyte hemoglobin (RET-He) was lower at 2 wk in the ID group. Significant IS compared with ID differences in serum iron indices were present at 2 mo, but Hgb and RBC indices differed only at 4 mo (P < 0.05). Serum and CSF metabolomic profiles of the ID and IS groups differed at 2 and 4 mo (P < 0.05). Key metabolites, including homostachydrine and stachydrine (4-5-fold lower at 4 mo in ID group, P < 0.05), were altered in both serum and CSF. Iron indices and RET-He at 2 mo, but not Hgb or other RBC indices, were correlated with altered CSF metabolic profile at 4 mo and had comparable predictive accuracy (area under the receiver operating characteristic curve scores, 0.75-0.80). CONCLUSIONS: Preanemic ID at 2 mo was associated with metabolic alterations in serum and CSF in infant monkeys. Among the RBC indices, only RET-He predicted the future risk of abnormal CSF metabolic profile with a predictive accuracy comparable to serum iron indices. The concordance of homostachydrine and stachydrine changes in serum and CSF indicates their potential use as early biomarkers of brain metabolic dysfunction in infantile ID.

Cellular Iron Deficiency Disrupts Thyroid Hormone Regulated Gene Expression in Developing Hippocampal Neurons.

BACKGROUND: Developing neurons have high thyroid hormone and iron requirements to support their metabolically demanding growth. Early-life iron and thyroid-hormone deficiencies are prevalent and often coexist, and each independently increases risk of permanently impaired neurobehavioral function in children. Early-life dietary iron deficiency reduces thyroid-hormone concentrations and impairs thyroid hormone-responsive gene expression in the neonatal rat brain, but it is unclear whether the effect is cell-intrinsic. OBJECTIVES: This study determined whether neuronal-specific iron deficiency alters thyroid hormone-regulated gene expression in developing neurons. METHODS: Iron deficiency was induced in primary mouse embryonic hippocampal neuron cultures with the iron chelator deferoxamine (DFO) beginning at 3 d in vitro (DIV). At 11DIV and 18DIV, thyroid hormone-regulated gene messenger ribonucleic acid (mRNA)concentrations indexing thyroid hormone homeostasis (Hairless, mu-crystallin, Type II deiodinase, solute carrier family member 1c1, and solute carrier family member 16a2) and neurodevelopment (neurogranin, Parvalbumin, and Krüppel-like factor 9) were quantified. To assess the effect of iron repletion, DFO was removed at 14DIV from a subset of DFO-treated cultures, and gene expression and adenosine 5'-triphosphate (ATP) concentrations were quantified at 21DIV. RESULTS: At 11DIV and 18DIV, neuronal iron deficiency decreased neurogranin, Parvalbumin, and mu-crystallin, and by 18DIV, solute carrier family member 16a2, solute carrier family member 1c1, Type II deiodinase, and Hairless were increased, suggesting cellular sensing of a functionally abnormal thyroid hormone state. Dimensionality reduction with Principal component analysis reveals that thyroid hormone homeostatic genes strongly correlate with and predict iron status. Iron repletion from 14-21DIV did not restore ATP concentration, and Principal component analysis suggests that, after iron repletion, cultures maintain a gene expression signature indicative of previous iron deficiency. CONCLUSIONS: These novel findings suggest there is an intracellular mechanism coordinating cellular iron/thyroid hormone activities. We speculate this is a part of the homeostatic response to acutely match neuronal energy production and growth signaling. However, the adaptation to iron deficiency may cause permanent deficits in thyroid hormone-dependent neurodevelopmental processes even after recovery from iron deficiency.

Differences in Oxygen-Induced Retinopathy Susceptibility Between Two Sprague Dawley Rat Vendors: A Comparison of Retinal Transcriptomes.

PURPOSE: To determine the retinal transcriptomic differences underlying the oxygen-induced retinopathy phenotypes between Sprague Dawley rat pups from two commonly used commercial vendors. This will allow us to discover genes and pathways that may be related to differences in disease severity in similarly aged premature babies and suggest possible new treatment approaches. METHODS: We analyzed retinal vascular morphometry and transcriptomes from Sprague Dawley rat pups from Charles River Laboratories and Envigo (previously Harlan). Room air control and oxygen-induced retinopathy groups were compared. Oxygen-induced retinopathy was induced with the rat 50/10 model. RESULTS: Pups from Charles River Laboratories developed a more severe oxygen-induced retinopathy phenotype, with 3.6-fold larger percent avascular area at P15 and twofold larger % neovascular area at P20 than pups from Envigo. Changes in retinal transcriptomes of rat pups from both vendors were substantial at baseline and in response to oxygen-induced retinopathy. Baseline differences centered on activated pathways of neuronal development in Charles River Laboratories pups. In response to oxygen-induced retinopathy, during the neovascular phase, retinas from Charles River Laboratories pups exhibited activation of pathways regulating necrosis, neuroinflammation, and interferon signaling, supporting the observed increase of neovascularization. Conversely, retinas from Envigo pups showed decreased necrosis and increased focal adhesion kinase signaling, supporting more normal vascular development. Comparing oxygen-induced retinopathy transcriptomes at P15 to those at P20, canonical pathways such as phosphate and tensin homolog, interferon, and coordinated lysosomal expression and regulation element signaling were identified, highlighting potential novel mechanistic targets for future research. CONCLUSION: Transcriptomic profiles differ substantially between rat pup retinas from Charles River Laboratories and Envigo at baseline and in response to oxygen-induced retinopathy, providing insight into vascular morphologic differences. Comparing transcriptomes identified new pathways for further research in oxygen-induced retinopathy pathogenesis and increased scientific rigor of this model.

RAPIDIRON Trial follow-up study - the RAPIDIRON-KIDS Study: protocol of a prospective observational follow-up study.

BACKGROUND: Anemia is a worldwide problem with iron deficiency being the most common cause. When anemia occurs in pregnancy, it increases the risk of adverse maternal, fetal, and postnatal outcomes. It induces preterm births and low birth weight (LBW) deliveries, long-term neurodevelopmental sequelae, and an increased risk of earlier onset of postnatal iron deficiency. Anemia rates are among the highest in South Asia, and India's National Family Health Survey (NFHS-5) for 2019-2021 indicated that over half of pregnant women, and more than 65% of children, in the country are classified as anemic (Sciences IIfP, National Family Health Survey-5, 2019-21, India Fact Sheet). In 2021, the parent RAPIDIRON Trial (Derman et al., Trials 22:649, 2021) was initiated in two states in India, with the goal of assessing whether a dose of intravenous (IV) iron given to anemic women during early pregnancy results in a greater proportion of participants with normal hemoglobin concentrations in the third trimester and a lower proportion of participants with LBW deliveries compared to oral iron. As a follow-up to the RAPIDIRON Trial, the RAPIDIRON-KIDS Study will follow the offspring of previously randomized mothers to assess, neurobehavioral, hematological, and health outcomes. METHODS: This prospective observational cohort study will follow a subset of participants previously randomized as part of the RAPIDIRON Trial and their newborns. Study visits occur at birth, 6 weeks, 4 months, 12 months, 24 months, and 36 months and include blood sample collection with both maternal and infant participants and specific neurobehavioral assessments conducted with the infants (depending on the study visit). The primary outcomes of interest are (1) infant iron status as indicated by both hemoglobin and ferritin (a) at birth and (b) at 4 months of age and (2) the developmental quotient (DQ) for the cognitive domain of the Bayley Scales of Infant Development Version IV (BSID-IV) at 24 months of age. DISCUSSION: This RAPIDIRON-KIDS Study builds upon its parent RAPIDIRON Trial by following a subset of the previously randomized participants and their offspring through the first 3 years of life to assess neurodevelopmental and neurobehavioral (infants, children), hematological, and health outcomes. TRIAL REGISTRATION: ClinicalTrials.gov NCT05504863 , Registered on 17 August 2022. Clinical Trials Registry - India CTRI/2022/05/042933 . Registered on 31 May 2022.

Anemia, Iron Supplementation, and the Brain.

The developing brain is particularly vulnerable to extrinsic environmental events such as anemia and iron deficiency during periods of rapid development. Studies of infants with postnatal iron deficiency and iron deficiency anemia clearly demonstrated negative effects on short-term and long-term brain development and function. Randomized interventional trials studied erythropoiesis-stimulating agents and hemoglobin-based red blood cell transfusion thresholds to determine how they affect preterm infant neurodevelopment. Studies of red blood cell transfusion components are limited in preterm neonates. A biomarker strategy measuring brain iron status and health in the preanemic period is desirable to evaluate treatment options and brain response.

The Interaction between Psychological Stress and Iron Status on Early-Life Neurodevelopmental Outcomes.

This review presents evidence from animal and human studies demonstrating the possible connection and significant impact of poor iron status and psychological distress on neurocognitive development during pregnancy and the neonatal period, with implications for long-term cognition. Stress and iron deficiency are independently prevalent and thus are frequently comorbid. While iron deficiency and early-life stress independently contribute to long-term neurodevelopmental alterations, their combined effects remain underexplored. Psychological stress responses may engage similar pathways as infectious stress, which alters fundamental iron metabolism processes and cause functional tissue-level iron deficiency. Psychological stress, analogous to but to a lesser degree than infectious stress, activates the hypothalamic-pituitary-adrenocortical (HPA) axis and increases proinflammatory cytokines. Chronic or severe stress is associated with dysregulated HPA axis functioning and a proinflammatory state. This dysregulation may disrupt iron absorption and utilization, likely mediated by the IL-6 activation of hepcidin, a molecule that impedes iron absorption and redistributes total body iron. This narrative review highlights suggestive studies investigating the relationship between psychological stress and iron status and outlines hypothesized mechanistic pathways connecting psychological stress exposure and iron metabolism. We examine findings regarding the overlapping impacts of early stress exposure to iron deficiency and children's neurocognitive development. We propose that studying the influence of psychological stress on iron metabolism is crucial for comprehending neurocognitive development in children exposed to prenatal and early postnatal stressors and for children at risk of early iron insufficiency. We recommend future directions for dual-exposure studies exploring iron as a potential mediating pathway between early stress and offspring neurodevelopment, offering opportunities for targeted interventions.

Establishing Normative Values for Healthy Term Infant Feeding Performance: Neonatal Eating Assessment Tool-Mixed, Oral Feeding Scale, and Early Feeding Skills Assessment.

PURPOSE: Infants with perceived feeding problems are frequently referred for assessment of their feeding abilities. However, little is known regarding how healthy nondysphagic infants perform on commonly used assessments, making determination of impairment difficult. The aim of this investigation was to elucidate the characteristics of healthy term infant feeding performance using three commonly employed clinical assessments: Neonatal Eating Assessment Tool-Mixed (NeoEat-Mixed), Oral Feeding Scale, and Early Feeding Skills (EFS). METHOD: In this prospective case-control study, we recruited 30 infants without feeding impairments to undergo video-monitored bottle feeds under their normal feeding conditions. Caregiver perception of infant feeding was evaluated using the NeoEat-Mixed. Milk ingestion was monitored real time using the Oral Feeding Scale for rate of milk transfer and modified proficiency as characterized by the total volume consumed out of the total volume the caregiver provided. Videos were analyzed by two speech pathologists using the EFS assessment. Descriptive statistics were used to characterize performance. RESULTS: Participants underwent feeding monitoring at an average chronological age of 4 ± 2 months. Caregivers primarily reported normal, nonconcerning feeding patterns across all of the NeoEAT-Mixed outcomes. Infants consumed milk at an average rate of transfer of 7 ± 3 ml/min, a modified proficiency of 50 ± 21%, and achieved the highest OFS score of 4 (93%, n = 28). The majority of infants scored the best EFS score (mature-3) as it related to the absence of color changes during the feed (97%, n = 29), although commonly scored in the worst EFS score (immature-1) in their presentation of one or more compelling stress cues (63%, n = 19). CONCLUSION: Establishing healthy term infant normative values for commonly used feeding assessments is critical in accurately distinguishing infants with feeding impairments from those with normal developmental variants.

The importance of iron deficiency in pregnancy on fetal, neonatal, and infant neurodevelopmental outcomes.

The role of iron in neurodevelopment has long been recognized, and the adverse effects of early-life iron deficiency on brain development and subsequent function across the lifespan continue to be a subject of research. A greater appreciation of the contribution of maternal preconceptional iron status and fetal iron accretion to offspring, postnatal iron status, and brain health across the lifespan has occurred over the past decade. This paradigm shift in thinking links two previously relatively siloed literatures: neonatal iron deficiency and postnatal iron deficiency. The understanding that iron accretion during the fetal period strongly influences postnatal iron balance has led to an appreciation of the importance and value of ensuring proper fetal iron loading. This article reviews the dynamics of fetal iron metabolism, the role of iron in the developing fetal brain, the short- and long-term neurobehavioral consequences of fetal iron underloading, and the potential mechanisms that account for the long-term effects of fetal/neonatal iron deficiency.

Maternal gestational iron status and infant haematological and neurodevelopmental outcomes.

Prevention of iron deficiency (ID), the most common micronutrient deficiency in infants and children, begins prenatally by ensuring adequate fetal loading. Adequate intrauterine iron status is crucial for normal fetal brain development, postnatal brain performance and prevention of early postnatal iron deficiency, particularly in infants fed exclusively human milk. Adequate fetal loading may be achieved in some cases through adequate maternal iron levels prior to pregnancy and oral iron supplementation during pregnancy. However, because so many women are iron-deficient leading up to pregnancy, coupled with the negative iron balance induced by pregnancy, a large number of women remain iron-deficient during pregnancy. More consistent iron-specific early screening and more effective iron delivery approaches are needed to solve this global problem.

Blood Levels of Environmental Heavy Metals are Associated with Poorer Iron Status in Ugandan Children: A Cross-Sectional Study.

BACKGROUND: Iron deficiency (ID) and environmental exposure to metals frequently co-occur among Ugandan children, but little is known about their associations, although iron and other divalent metals share the same intestinal absorption transporter, divalent metal transporter 1 (DMT1). OBJECTIVES: We examined associations between iron status and blood concentrations of lead, manganese (Mn), cobalt (Co), and cadmium, both singly and as a mixture. METHODS: We used data on sociodemographic status, iron biomarkers, and blood concentrations of heavy metals collected from a cross-sectional survey of 100 children aged 6-59 mo in Kampala, Uganda. We compared blood concentrations of metals in ID with iron-sufficient children. We examined associations between a metal mixture and iron biomarkers using multiple linear regression and weighted quintile sum regression. RESULTS: The median (interquartile range) blood Mn (µg/L) was higher in ID children defined by soluble transferrin receptor (sTfR) and ferritin (ID compared with iron-sufficient children): (sTfR [21.3 {15.1, 28.8}, 11.2 {8.6, 18.5}], ferritin [19.5 {15.0, 27.2}, 11.2 {8.8, 19.6}]; P < 0.001 for both). Similarly, the median (interquartile range) blood Co (µg/L) was higher in ID children by ferritin ([0.5 {0.4, 0.9}, 0.4 {0.3, 0.5}], P = 0.05). Based on the multiple linear regression results, higher blood Co and Mn were associated with poorer iron status (defined by all 4 iron indicators for Co and by sTfR for Mn). The weighted quintile sum regression result showed that higher blood concentrations of a metal mixture were associated with poorer iron status represented by sTfR, ferritin, and hepcidin, mainly driven by Co and Mn. CONCLUSIONS: Our study findings suggest that poorer iron status is associated with overall heavy metal burden, predominantly Co and Mn, among Ugandan children. Further prospective studies should confirm our primary findings and investigate the combined effects of coexposures to neurotoxicants on the neurodevelopment of young children.

Human Milk Feeding and Direct Breastfeeding Improve Outcomes for Infants With Single Ventricle Congenital Heart Disease: Propensity Score-Matched Analysis of the NPC-QIC Registry.

Background Infants with single ventricle congenital heart disease undergo 3 staged surgeries/interventions, with risk for morbidity and mortality. We estimated the effect of human milk (HM) and direct breastfeeding on outcomes including necrotizing enterocolitis, infection-related complications, length of stay, and mortality. Methods and Results We analyzed the National Pediatric Cardiology Quality Improvement Collaborative (NPC-QIC) registry (2016-2021), examining HM/breastfeeding groups during stage 1 and stage 2 palliations. We calculated propensity scores for feeding exposures, then fitted Poisson and logistic regression models to compare outcomes between propensity-matched cohorts. Participants included 2491 infants (68 sites). Estimates for all outcomes were better in HM/breastfeeding groups. Infants fed exclusive HM before stage 1 palliation (S1P) had lower odds of preoperative necrotizing enterocolitis (odds ratio [OR], 0.37 [95% CI, 0.17-0.84]; P=0.017) and shorter S1P length of stay (rate ratio [RR], 0.87 [95% CI, 0.78-0.98]; P=0.027). During the S1P hospitalization, infants with high HM had lower odds of postoperative necrotizing enterocolitis (OR, 0.28 [95% CI, 0.15-0.50]; P<0.001) and sepsis (OR, 0.29 [95% CI, 0.13-0.65]; P=0.003), and shorter S1P length of stay (RR, 0.75 [95% CI, 0.66-0.86]; P<0.001). At stage 2 palliation, infants with any HM (RR, 0.82 [95% CI, 0.69-0.97]; P=0.018) and any breastfeeding (RR, 0.71 [95% CI, 0.57-0.89]; P=0.003) experienced shorter length of stay. Conclusions Infants with single ventricle congenital heart disease in high-HM and breastfeeding groups experienced multiple significantly better outcomes. Given our findings of improved health, strategies to increase the rates of HM/breastfeeding in these patients should be implemented. Future research should replicate these findings with granular feeding data and in broader congenital heart disease populations, and should examine mechanisms (eg, HM components, microbiome) by which HM/breastfeeding benefits these infants.

Cellular Iron Deficiency Disrupts Thyroid Hormone Regulated Gene Expression in Developing Hippocampal Neurons.

Background: Developing neurons have high thyroid hormone and iron requirements to support their metabolism and growth. Early-life iron and thyroid hormone deficiencies are prevalent, often coexist, and increase the risk of permanently impaired neurobehavioral function in children. Early-life dietary iron deficiency reduces thyroid hormone levels and impairs thyroid hormone-responsive gene expression in the neonatal rat brain. Objective: This study determined whether neuronal-specific iron deficiency alters thyroid hormone-regulated gene expression in developing neurons. Methods: Iron deficiency was induced in primary mouse embryonic hippocampal neuron cultures with the iron chelator deferoxamine (DFO) beginning at 3 days in vitro (DIV). At 11DIV and 18DIV, mRNA levels for thyroid hormone-regulated genes indexing thyroid hormone homeostasis (Hr, Crym, Dio2, Slco1c1, Slc16a2) and neurodevelopment (Nrgn, Pvalb, Klf9) were quantified. To assess the effect of iron repletion, DFO was removed at 14DIV from a subset of DFO-treated cultures and gene expression and ATP levels were quantified at 21DIV. Results: At 11DIV and 18DIV, neuronal iron deficiency decreased Nrgn, Pvalb, and Crym, and by 18DIV, Slc16a2, Slco1c1, Dio2, and Hr were increased; collectively suggesting cellular sensing of a functionally abnormal thyroid hormone state. Dimensionality reduction with Principal Component Analysis (PCA) reveals that thyroid hormone homeostatic genes strongly correlate with and predict iron status (Tfr1 mRNA). Iron repletion from 14-21DIV restored neurodevelopmental genes, but not all thyroid hormone homeostatic genes, and ATP concentrations remained significantly altered. PCA clustering suggests that cultures replete with iron maintain a gene expression signature indicative of previous iron deficiency. Conclusions: These novel findings suggest there is an intracellular mechanism coordinating cellular iron/thyroid hormone activities. We speculate this is a part of homeostatic response to match neuronal energy production and growth signaling for these important metabolic regulators. However, iron deficiency may cause permanent deficits in thyroid hormone-dependent neurodevelopmental processes even after recovery from iron deficiency.

Predictors of Human Milk Feeding and Direct Breastfeeding for Infants with Single Ventricle Congenital Heart Disease: Machine Learning Analysis of the National Pediatric Cardiology Quality Improvement Collaborative Registry.

OBJECTIVE: To identify factors that support or limit human milk (HM) feeding and direct breastfeeding (BF) for infants with single ventricle congenital heart disease at neonatal stage 1 palliation (S1P) discharge and at stage 2 palliation (S2P) (∼4-6 months old). STUDY DESIGN: Analysis of the National Pediatric Cardiology Quality Improvement Collaborative (NPC-QIC) registry (2016-2021; 67 sites). Primary outcomes were any HM, exclusive HM, and any direct BF at S1P discharge and at S2P. The main analysis involved multiple phases of elastic net logistic regression on imputed data to identify important predictors. RESULTS: For 1944 infants, the strongest predictor domain areas included preoperative feeding, demographics/social determinants of health, feeding route, clinical course, and site. Significant findings included: preoperative BF was associated with any HM at S1P discharge (OR = 2.02, 95% CI = 1.74-3.44) and any BF at S2P (OR = 2.29, 95% CI = 1.38-3.80); private/self-insurance was associated with any HM at S1P discharge (OR = 1.91, 95% CI = 1.58-2.47); and Black/African-American infants had lower odds of any HM at S1P discharge (OR = 0.54, 95% CI = 0.38-0.65) and at S2P (0.57, 0.30-0.86). Adjusted odds of HM/BF practices varied among NPC-QIC sites. CONCLUSIONS: Preoperative feeding practices predict later HM and BF for infants with single ventricle congenital heart disease; therefore, family-centered interventions focused on HM/BF during the S1P preoperative time are needed. These interventions should include evidence-based strategies to address implicit bias and seek to minimize disparities related to social determinants of health. Future research is needed to identify supportive practices common to high-performing NPC-QIC sites.

Human milk feeding and direct breastfeeding improve outcomes for infants with single ventricle congenital heart disease: Propensity score matched analysis of the NPC-QIC registry.

Background: Infants with single ventricle (SV) congenital heart disease (CHD) undergo three staged surgeries/interventions, with risk for morbidity and mortality. We estimated the effect of human milk (HM) and direct breastfeeding (BF) on outcomes including necrotizing enterocolitis (NEC), infection-related complications, length of stay (LOS), and mortality. Methods: We analyzed the National Pediatric Cardiology Quality Improvement Collaborative registry (2016-2021), examining HM/BF groups during stage 1 (S1P) and stage 2 (S2P) palliations. We calculated propensity scores for feeding exposures, then fitted Poisson and logistic regression models to compare outcomes between propensity-matched cohorts. Results: Participants included 2491 infants (68 sites). Estimates for all outcomes were better in HM/BF groups. Infants fed exclusive HM before S1P had lower odds of preoperative NEC (OR=0.37, 95% CI=0.17-0.84, p=0.017) and shorter S1P LOS (RR=0.87, 0.78-0.98, p=0.027). During the S1P hospitalization, infants with high HM had lower odds of postoperative NEC (OR=0.28, 0.15-0.50, p<0.001) and sepsis (0.29, 0.13-0.65, p=0.003), and shorter S1P LOS (RR=0.75, 0.66-0.86, p<0.001). At S2P, infants with any HM (0.82, 0.69-0.97, p=0.018) and any BF (0.71, 0.57-0.89, p=0.003) experienced shorter LOS. Conclusions: Infants with SV CHD in high HM and BF groups experienced multiple significantly better outcomes. Given our findings of improved health, strategies to increase the rates of HM/BF in these patients should be implemented. Future research should replicate these findings with granular feeding data and in broader CHD populations, and should examine mechanisms (eg, HM components; microbiome) by which HM/BF benefits these infants. Clinical Perspective: What is new?: This is the first large, multisite study examining the impact of human milk and breastfeeding on outcomes for infants with single ventricle congenital heart disease.All outcome estimates were better in high human milk and breastfeeding groups, with significantly lower odds of necrotizing enterocolitis, sepsis, and infection-related complications; and significantly shorter length of stay at both the neonatal stage 1 palliation and the subsequent stage 2 palliation.All estimates of all-cause mortality were substantially lower in human milk and breastfeeding groups, with clinically important estimates of 75%-100% lower odds of mortality in direct breastfeeding groups.What are the clinical implications?: There is a critical need for improved, condition-specific lactation support to address the low prevalence of human milk and breastfeeding for infants with single ventricle congenital heart disease.Increasing the dose and duration of human milk and direct breastfeeding has strong potential to substantially improve the health outcomes of these vulnerable infants.