Improved Neurodevelopmental Outcomes at 5.5 Years of Age in Children Who Received Bovine Milk Fat Globule Membrane and Lactoferrin in Infant Formula Through 12 Months: A Randomized Controlled Trial.

OBJECTIVE: To evaluate the neurodevelopmental outcomes at 5.5 years of age in children who were previously randomized to cow milk-based infant formula (control) or similar formula (milk fat globule membrane + lactoferrin) with added sources of bovine milk fat globule membrane and bovine lactoferrin through 12 months of age. DESIGN: Children who completed study feeding were invited to participate in follow-up assessments: cognitive development across multiple domains (primary outcome; Wechsler Preschool & Primary Scale of Intelligence, 4th Edition), inhibitory control/rule learning (Stroop Task), flexibility/rule learning (Dimensional Change Card Sort), and behavior/emotion (Child Behavior Checklist). RESULTS: Of 292 eligible participants (control: 148, milk fat globule membrane + lactoferrin: 144), 116 enrolled and completed assessments (control: 59, milk fat globule membrane + LF: 57). There were no group demographic differences except family income (milk fat globule membrane + lactoferrin significantly higher). Wechsler Preschool & Primary Scale of Intelligence, 4th Edition composite scores (mean ± standard error) for Visual Spatial (100.6 ± 1.7 vs 95.3 ± 1.7; P = .027), Processing Speed (107.1 ± 1.4 vs 100.0 ± 1.4; P < .001), and Full-Scale IQ (98.7 ± 1.4 vs 93.5 ± 1.5; P = .012) were significantly higher for milk fat globule membrane + lactoferrin versus control, even after controlling for demographic/socioeconomic factors. Stroop Task scores were significantly higher in milk fat globule membrane + lactoferrin versus control (P < .001). Higher Dimensional Change Card Sort scores (P = .013) in the border phase (most complex/challenging) were detected, and more children passed the border phase (32% vs 12%; P = .039) for milk fat globule membrane versus control. No group differences in Child Behavior Checklist score were detected. CONCLUSIONS: Children who received infant formula to 12 months of age with added bovine milk fat globule membrane and bovine lactoferrin versus standard formula demonstrated improved cognitive outcomes in multiple domains at 5.5 years of age, including measures of intelligence and executive function. TRIAL REGISTRATION: Clinicaltrials.gov: https://clinicaltrials.gov/ct2/show/NCT04442477.

DHA Supplementation During Pregnancy Enhances Maternal Vagally Mediated Cardiac Autonomic Control in Humans.

BACKGROUND: DHA is an essential omega-3 (ω-3; n-3) fatty acid that has well-established benefits for the fetus. DHA also has the potential to influence the health of the mother, but this area is understudied. OBJECTIVES: The objective of this secondary analysis was to determine if DHA was related to maternal heart rate (HR) and heart rate variability (HRV) metrics in a large cohort of pregnant women. METHODS: In the parent trial (1R01HD086001) eligible participants (≥18 y old, English speaking, carrying a singleton pregnancy, 12-20 wk of gestation) were randomly assigned to consume 200 mg/d or 800 mg/d DHA for the duration of their pregnancy (n = 300). Weight, blood pressure, and magnetocardiograms (MCGs) were collected at 32 wk and 36 wk of gestation (n = 221). Measures of HR and HRV in time-, frequency-, and nonlinear-domains were determined from the isolated maternal MCG. Treatment group and timepoint were examined as predictors in association with HR and HRV metrics using random-intercept mixed-effects ANOVA unadjusted and adjusted models accounting for weight and dietary DHA intake. RESULTS: Women receiving the higher dose of DHA (800 mg/d) during pregnancy had lower HR, lower sympathetic index, higher vagally mediated HRV indices, and greater HRV complexity when compared with the women who received the lower dose (200 mg/d; all P < 0.05). All the dose relations remained significant even after controlling for the effect of time, maternal weight, and dietary DHA intake. CONCLUSIONS: DHA supplementation increases vagal tone in pregnant women. Longitudinal studies examining the potential link between DHA, enhanced vagal tone, and reported reduction in early preterm birth are warranted.

Prenatal docosahexaenoic acid effect on maternal-infant DHA-equilibrium and fetal neurodevelopment: a randomized clinical trial.

INTRODUCTION: Maternal-infant equilibrium occurs when cord blood docosahexaenoic acid (DHA) is less than or equal to maternal DHA at delivery. Equilibrium may be an indicator of sufficient DHA for optimal fetal and infant neurodevelopment. The purpose of this study was to test the effect of maternal DHA supplementation on equilibrium status and fetal neurodevelopment. METHODS: Women enrolled between 12 and 20 weeks gestation and were randomized to 200 or 800 mg DHA/day until delivery. Maternal red blood cell (RBC) phospholipids were measured at enrollment, 32 weeks, delivery, and in cord blood at delivery. Fetal neurodevelopment was measured at 32 and 36 weeks gestation. Intent-to-treat analyses were conducted to test differences in equilibrium status by group. Fetal outcomes were assessed by equilibrium status and group. RESULTS: Three hundred women enrolled and 262 maternal-infant dyads provided blood samples at delivery. No maternal-infant dyads with maternal RBC-DHA ≤ 6.96% at delivery achieved equilibrium. The incidence of equilibrium was significantly higher in the 800 mg group. There was no effect of maternal group or equilibrium status on fetal neurodevelopment. CONCLUSION: The significance of maternal-infant DHA equilibrium remains unknown. Ongoing research will test the effect of treatment group, equilibrium, and nutrient status on infant behavior and brain function. IMPACT: Pregnant women who received a higher dose of docosahexaenoic acid (DHA) were more likely to achieve maternal-infant DHA equilibrium at delivery. Equilibrium status had no effect on fetal neurodevelopment in this sample. While DHA is crucial for early life neurodevelopment, the significance of achieving maternal-infant equilibrium above the lower threshold is uncertain. There is a lower threshold of maternal DHA status where maternal-infant DHA equilibrium never occurs. The lack of equilibrium associated with low maternal DHA status may indicate insufficient maternal status for optimal placental transfer.

Prenatal docosahexaenoic acid supplementation has long-term effects on childhood behavioral and brain responses during performance on an inhibitory task.

Introduction: Offsprings from a prenatal docosahexaenoic acid (DHA) supplementation trial, in which pregnant women were assigned to placebo or 600mg DHA/day, were followed to determine the effect of prenatal DHA supplementation on the behavior and brain function at 5.5 years (n=81 placebo, n=86 supplemented).Methods: Event-related potentials (ERP) were recorded during a visual task requiring a button press (Go) to frequent target stimuli and response inhibition to the rare stimuli (No-Go). Univariate ANOVAs were used to test differences between group and sex for behavioral measures. ERP differences were tested using a three-way mixed-design multivariate analysis of variance (MANOVA).Results: There was a significant sex × group interaction for hit rate and errors of omission; there was no difference between males and females in the placebo group, but DHA males outperformed DHA females. Males overall and the placebo group made more errors requiring response inhibition; DHA females were significantly better than placebo females and DHA males. ERP P2 amplitude was larger in the DHA group. A significant N2 amplitude condition effect was observed in females and DHA group males, but not in placebo group males.Discussion: Prenatal DHA supplementation improved inhibitory performance overall, especially for females in the DHA group, possibly accounting for their conservative behavior during Go trials. Development of brain regions responsible for visual processing may be sensitive to maternal DHA status, evidenced by greater P2 amplitude. Males may benefit more from maternal DHA supplementation, indicated by the N2 condition effect seen only in males in the DHA group.

Developmental effects on sleep-wake patterns in infants receiving a cow's milk-based infant formula with an added prebiotic blend: a Randomized Controlled Trial.

BACKGROUND: Few studies have evaluated nutritive effects of prebiotics on infant behavior state, physiology, or metabolic status. METHODS: In this double-blind randomized study, infants (n = 161) received cow's milk-based infant formula (Control) or similar formula with an added prebiotic blend (polydextrose and galactooligosaccharides [PDX/GOS]) from 14-35 to 112 days of age. Infant wake behavior (crying/fussing, awake/content) and 24-h sleep-wake actograms were analyzed (Baseline, Days 70 and 112). Salivary cortisol was immunoassayed (Days 70 and 112). In a subset, exploratory stool 16S ribosomal RNA-sequencing was analyzed (Baseline, Day 112). RESULTS: One hundred and thirty-one infants completed the study. Average duration of crying/fussing episodes was similar at Baseline, significantly shorter for PDX/GOS vs. Control at Day 70, and the trajectory continued at Day 112. Latency to first and second nap was significantly longer for PDX/GOS vs. Control at Day 112. Cortisol awakening response was demonstrated at Days 70 and 112. Significant stool microbiome beta-diversity and individual taxa abundance differences were observed in the PDX/GOS group. CONCLUSIONS: Results indicate faster consolidation of daytime waking state in infants receiving prebiotics and support home-based actigraphy to assess early sleep-wake patterns. A prebiotic effect on wake organization is consistent with influence on the gut-brain axis and warrants further investigation. IMPACT: Few studies have evaluated nutritive effects of prebiotics on infant behavior state, cortisol awakening response, sleep-wake entrainment, and gut microbiome. Faster consolidation of daytime waking state was demonstrated in infants receiving a prebiotic blend in infant formula through ~4 months of age. Shorter episodes of crying were demonstrated at ~2 months of age (time point corresponding to age/developmental range associated with peak crying) in infants receiving formula with added prebiotics. Results support home-based actigraphy as a suitable method to assess early sleep-wake patterns. Prebiotic effect on wake organization is consistent with influence on the gut-brain axis and warrants further investigation.

DHA supplementation in infants born preterm and the effect on attention at 18 months' corrected age: follow-up of a subset of the N3RO randomised controlled trial.

Infants born preterm miss out on the peak period of in utero DHA accretion to the brain during the last trimester of pregnancy which is hypothesised to contribute to the increased prevalence of neurodevelopmental deficits in this population. This study aimed to determine whether DHA supplementation in infants born preterm improves attention at 18 months' corrected age. This is a follow-up of a subset of infants who participated in the N3RO randomised controlled trial. Infants were randomised to receive an enteral emulsion of high-dose DHA (60 mg/kg per d) or no DHA (soya oil - control) from within the first days of birth until 36 weeks' post-menstrual age. The assessment of attention involved three tasks requiring the child to maintain attention on toy/s in either the presence or absence of competition or a distractor. The primary outcome was the child's latency of distractibility when attention was focused on a toy. The primary outcome was available for seventy-three of the 120 infants that were eligible to participate. There was no evidence of a difference between groups in the latency of distractibility (adjusted mean difference: 0·08 s, 95 % CI -0·81, 0·97; P = 0·86). Enteral DHA supplementation did not result in improved attention in infants born preterm at 18 months' corrected age.

Synthesis and characterization of novel calcium phosphate glass-derived cements for vital pulp therapy.

Evaluation of the physicochemical behavior and setting reactions of a novel inorganic pulp capping cement which makes use of the unique corrosion properties of sodium metasilicate (NaSi) glass. NaSi and calcium phosphate (CaP) glass powders were synthesized through a melt-quench method. Cements were created by mixing various amounts of the glasses with deionized water at a powder-to-liquid ratio of 2.5 g mL-1. Working and setting times were measured using the indentation standard ISO 9917-1. Sealing ability was tested by placing set samples of each composition in methylene blue dye solution for 24 h. Set samples were also submerged in phosphate buffered saline and incubated at 37 °C for one week. X-ray diffraction was used to identify mature crystalline phases after incubation. Infrared spectroscopy and scanning electron microscopy were used to characterize cements before and after setting and after incubation. Working and setting times measured in the ranges of 2-5 and 10-25 min, respectively. Working and setting time generally decrease with increased NaSi concentration. Cements with compositions of 25 and 33 wt% NaSi were found to resist the infiltration of dye and maintain their shape. Compositions outside this range absorbed dye and collapsed. Infrared spectroscopy provided insight into the setting mechanism of these cements. After one week in vitro, cements were found to contain crystalline phases matching chemically stable, bioactive phases. The combination of NaSi and CaP glasses has favorable setting behavior, sealing ability, and mature phases for pulp capping while relying on a relatively simple, inorganic composition.

Improved Neurodevelopmental Outcomes Associated with Bovine Milk Fat Globule Membrane and Lactoferrin in Infant Formula: A Randomized, Controlled Trial.

OBJECTIVE: To evaluate neurodevelopment, growth, and health outcomes in infants receiving bovine milk fat globule membrane (MFGM) and lactoferrin in infant formula. STUDY DESIGN: Healthy term infants were randomized to a cow's milk-based infant formula or MFGM + LF (a similar infant formula, with an added source of bovine milk fat globule membrane [bMFGM; whey protein-lipid concentrate, 5 g/L] and bovine lactoferrin [0.6 g/L]) through 365 days of age. The Bayley Scales of Infant Development, 3rd edition cognitive composite score at day 365 was the primary outcome. Secondary outcomes included tolerance measures through day 365, additional neurodevelopmental and language outcomes, growth, and medically confirmed adverse events through day 545. RESULTS: Of 451 infants enrolled (control, 228; MFGM + LF, 223), 291 completed study feeding and Bayley-III testing at day 365 (control, 148; MFGM + LF, 143). The mean cognitive (+8.7), language (+12.3), and motor (+12.6) scores were higher (P < .001) for the MFGM + LF group; no differences were observed at day 545. Global development scores from day 120 to day 275 and attention at day 365 were significantly improved. Few group differences in day 545 neurodevelopmental outcomes were detected, however scores of some subcategories of the MacArthur-Bates Communicative Development Inventories were higher (P < .05) in the MFGM + LF group. The overall incidence of respiratory-associated adverse events and diarrhea were significantly lower for the MFGM + LF group through day 545. CONCLUSIONS: Infants receiving formula with added bovine MFGM and bovine lactoferrin had an accelerated neurodevelopmental profile at day 365 and improved language subcategories at day 545. Formulas were associated with age-appropriate growth and significantly fewer diarrhea and respiratory-associated adverse events through 545 days of age. TRIAL REGISTRATION CLINICALTRIALS.GOV:: NCT02274883.

Critical and Sensitive Periods in Development and Nutrition.

Critical or sensitive periods in the life of an organism during which certain experiences or conditions may exert disproportionate influence (either for harm or benefit) on long-term developmental outcomes have been the subject of investigation for over a century. This chapter reviews research in the context of the development of social preferences and sensory systems, with a summary of the criteria for defining such a period and the evidence necessary to establish its existence. The notion of nutritional programming, central to the Barker/Developmental Origins hypotheses of health and disease, represents a variant of the critical/sensitive period concept. It is implicit in these hypotheses that the fetal period is a time during which metabolic and physiological systems are malleable and thus susceptible to either insult or enhancement by nutrient intake. Evidence for critical/sensitive periods or nutritional programming requires a systematic manipulation of the age at which nutritional conditions or supplements are implemented. While common in research using animal models, the approach is difficult to establish in epidemiological studies and virtually nonexistent in human clinical trials. Future work seeking to establish definitive evidence for critical/sensitive periods or programming may be advanced by harmonized outcome measures in experimental trials across which the timing, duration, and dose of nutrients is varied.

Effects of multimodal synchrony on infant attention and heart rate during events with social and nonsocial stimuli.

Attention is a state of readiness or alertness, associated with behavioral and psychophysiological responses, that facilitates learning and memory. Multisensory and dynamic events have been shown to elicit more attention and produce greater sustained attention in infants than auditory or visual events alone. Such redundant and often temporally synchronous information guides selectivity and facilitates perception, learning, and memory of properties of events specified by redundancy. In addition, events involving faces or other social stimuli provide an extraordinary amount of redundant information that attracts and sustains attention. In the current study, 4- and 8-month-old infants were shown 2-min multimodal videos featuring social or nonsocial stimuli to determine the relative roles of synchrony and stimulus category in inducing attention. Behavioral measures included average looking time and peak look duration, and convergent measurement of heart rate (HR) allowed for the calculation of HR-defined phases of attention: Orienting (OR), sustained attention (SA), and attention termination (AT). The synchronous condition produced an earlier onset of SA (less time in OR) and a deeper state of SA than the asynchronous condition. Social stimuli attracted and held attention (longer duration of peak looks and lower HR than nonsocial stimuli). Effects of synchrony and the social nature of stimuli were additive, suggesting independence of their influence on attention. These findings are the first to demonstrate different HR-defined phases of attention as a function of intersensory redundancy, suggesting greater salience and deeper processing of naturalistic synchronous audiovisual events compared with asynchronous ones.

Long-chain polyunsaturated fatty acid supplementation in the first year of life affects brain function, structure, and metabolism at age nine years.

The present study sought to determine whether supplementation of long-chain polyunsaturated fatty acids (LCPUFA) during the first year of life influenced brain function, structure, and metabolism at 9 years of age. Newborns were randomly assigned to consume formula containing either no LCPUFA (control) or formula with 0.64% of total fatty acids as arachidonic acid (ARA; 20:4n6) and variable amounts of docosahexaenoic acid (DHA; 22:6n3) (0.32%, 0.64%, or 0.96% of total fatty acids) from birth to 12 months. At age 9 years (±0.6), 42 children enrolled in a follow-up multimodal magnetic resonance imaging (MRI) study including functional (fMRI, Flanker task), resting state (rsMRI), anatomic, and proton magnetic resonance spectroscopy (1 H MRS). fMRI analysis using the Flanker task found that trials requiring greater inhibition elicited greater brain activation in LCPUFA-supplemented children in anterior cingulate cortex (ACC) and parietal regions. rsMRI analysis showed that children in the 0.64% group exhibited greater connectivity between prefrontal and parietal regions compared to all other groups. In addition, voxel-based analysis (VBM) revealed that the 0.32% and 0.64% groups had greater white matter volume in ACC and parietal regions compared to controls and the 0.96% group. Finally, 1 H MRS data analysis identified that N-acetylaspartate (NAA) and myo-inositol (mI) were higher in LCPUFA groups compared to the control group. LCPUFA supplementation during infancy has lasting effects on brain structure, function, and neurochemical concentrations in regions associated with attention (parietal) and inhibition (ACC), as well as neurochemicals associated with neuronal integrity (NAA) and brain cell signaling (mI).

Event-related potential differences in children supplemented with long-chain polyunsaturated fatty acids during infancy.

Long-chain polyunsaturated fatty acids (LCPUFA) have been shown to be necessary for early retinal and brain development, but long-term cognitive benefits of LCPUFA in infancy have not been definitively established. The present study sought to determine whether LCPUFA supplementation during the first year of life would result in group differences in behavior and event-related potentials (ERPs) while performing a task requiring response inhibition (Go/No-Go) at 5.5 years of age. As newborns, 69 children were randomly assigned to infant formulas containing either no LCPUFA (control) or formula with 0.64% of total fatty acids as arachidonic acid (ARA; 20:4n6) and various concentrations of docosahexaenoic acid (DHA; 22:6n3) (0.32%, 0.64% or 0.96%) for the first 12 months of life. At 5.5 years of age, a task designed to test the ability to inhibit a prepotent response (Go/No-Go) was administered, yielding both event-related potentials (ERPs) and behavioral data. Behavioral measures did not differ between groups, although reaction times of supplemented children were marginally faster. Unsupplemented children had lower P2 amplitude than supplemented children to both Go and No-Go conditions. N2 amplitude was significantly higher on No-Go trials than Go trials, but only for supplemented children, resulting in a significant Group × Condition interaction. Topographical analysis of the ERPs revealed that the LCPUFA-supplemented group developed a novel period of synchronous activation (microstate) involving wider anterior brain activation around 200 ms; this microstate was not present in controls. These findings suggest that LCPUFA supplementation during the first 12 months of life exerts a developmental programming effect that is manifest in brain electrophysiology. A video abstract of this article can be viewed at: https://www.youtube.com/watch?v=oM2leg4sevs.

Prenatal DHA supplementation and infant attention.

BACKGROUND: Results of randomized trials on the effects of prenatal docosahexaenoic acid (DHA) on infant cognition are mixed, but most trials have used global standardized outcomes, which may not be sensitive to effects of DHA on specific cognitive domains. METHODS: Women were randomized to 600 mg/d DHA or a placebo for the last two trimesters of pregnancy. Infants of these mothers were then followed on tests of visual habituation at 4, 6, and 9 mo of age. RESULTS: DHA supplementation did not affect look duration or habituation parameters but infants of supplemented mothers maintained high levels of sustained attention (SA) across the first year; SA declined for the placebo group. The supplemented group also showed significantly reduced attrition on habituation tasks, especially at 6 and 9 mo. CONCLUSION: The findings support with the suggestion that prenatal DHA may positively affect infants' attention and regulation of state.

Formula with long-chain polyunsaturated fatty acids reduces incidence of allergy in early childhood.

BACKGROUND: Allergy has sharply increased in affluent Western countries in the last 30 years. N-3 long-chain polyunsaturated fatty acids (n-3 LCPUFAs) may protect the immune system against development of allergy. METHODS: We prospectively categorized illnesses by body system in a subset of 91 children from the Kansas City cohort of the DIAMOND (DHA Intake and Measurement of Neural Development) study who had yearly medical records through 4 years of age. As infants, they were fed either a control formula without LCPUFA (n = 19) or one of three formulas with LCPUFA from docosahexaenoic acid (DHA) and arachidonic acid (ARA) (n = 72). RESULTS: Allergic illnesses in the first year were lower in the combined LCPUFA group compared to the control. LCPUFAs significantly delayed time to first allergic illness (p = 0.04) and skin allergic illness (p = 0.03) and resulted in a trend to reduced wheeze/asthma (p = 0.1). If the mother had no allergies, LCPUFAs reduced the risk of any allergic diseases (HR = 0.24, 95% CI = 0.1, 0.56, p = 0.0.001) and skin allergic diseases (HR = 0.35, 95% CI = 0.13, 0.93, p = 0.04). In contrast, if the mother had allergies, LCPUFAs reduced wheezing/asthma (HR = 0.26, 95% CI = 0.07, 0.9, p = 0.02). CONCLUSIONS: LCPUFA supplementation during infancy reduced the risk of skin and respiratory allergic diseases in childhood with effects influenced by maternal allergies.

Dietary patterns of early childhood and maternal socioeconomic status in a unique prospective sample from a randomized controlled trial of Prenatal DHA Supplementation.

BACKGROUND: Dietary habits established in early childhood and maternal socioeconomic status (SES) are important, complex, interrelated factors that influence a child's growth and development. The aim of this study was to define the major dietary patterns in a cohort of young US children, construct a maternal SES index, and evaluate their associations. METHODS: The diets of 190 children from a randomized, controlled trial of prenatal supplementation of docosahexaenoic acid (DHA) were recorded at 6-mo intervals from 2-4.5 years by 24-h dietary recall. Hierarchical cluster analysis of age-adjusted, average daily intake of 24 food and beverage groups was used to categorize diet. Unrotated factor analysis generated an SES score from maternal race, ethnicity, age, education, and neighborhood income. RESULTS: We identified two major dietary patterns: "Prudent" and "Western." The 85 (45%) children with a Prudent diet consumed more whole grains, fruit, yogurt and low-fat milk, green and non-starchy vegetables, and nuts and seeds. Conversely, those with a Western diet had greater intake of red meat, discretionary fat and condiments, sweet beverages, refined grains, French fries and potato chips, eggs, starchy vegetables, processed meats, chicken and seafood, and whole-fat milk. Compared to a Western diet, a Prudent diet was associated with one standard deviation higher maternal SES (95% CI: 0.80 to 1.30). CONCLUSIONS: We found two major dietary patterns of young US children and defined a single, continuous axis of maternal SES that differed strongly between groups. This is an important first step to investigate how child diet, SES, and prenatal DHA supplementation interact to influence health outcomes. TRIAL REGISTRATION: NCT00266825 . Prospectively registered on December 15, 2005.

A 3D ex vivo mandible slice system for longitudinal culturing of transplanted dental pulp progenitor cells.

Harnessing mesenchymal stem cells for tissue repair underpins regenerative medicine. However, how the 3D tissue matrix maintains such cells in a quiescent state whilst at the same time primed to respond to tissue damage remains relatively unknown. Developing more physiologically relevant 3D models would allow us to better understand the matrix drivers and influence on cell-lineage differentiation in situ. In this study, we have developed an ex vivo organotypic rat mandible slice model; a technically defined platform for the culture and characterization of dental pulp progenitor cells expressing GFP driven by the ß-actin promoter (cGFP DPPCs). Using confocal microscopy we have characterized how the native environment influences the progenitor cells transplanted into the dental pulp. Injected cGFP-DPPCs were highly viable and furthermore differentially proliferated in unique regions of the mandible slice; in the dentine region, cGFP-DPPCs showed a columnar morphology indicative of expansion and lineage differentiation. Hence, we demonstrated the systematic capacity for establishing a dental pulp cell-micro-community, phenotypically modified in the tooth (the "biology"); and at the same time addressed technical challenges enabling the mandible slice to be accessible on platforms for high-content imaging (the biology in a "multiplex" format).

Zinc supplementation sustained normative neurodevelopment in a randomized, controlled trial of Peruvian infants aged 6-18 months.

A double-blind, randomized clinical trial was conducted to determine the effects of prevention of zinc deficiency on cognitive and sensorimotor development during infancy. At 6 mo of age, infants were randomly assigned to be administered a daily liquid supplement containing 10 mg/d of zinc (zinc sulfate), 10 mg/d of iron (ferrous sulfate), and 0.5 mg/d of copper (copper oxide), or an identical daily liquid supplement containing only 10 mg/d of iron and 0.5 mg/d of copper. Various controls were implemented to ensure adherence to the supplement protocol. A battery of developmental assessments was administered from 6 to 18 mo of age that included a visual habituation/recognition memory task augmented with heart rate at 6, 9, and 12 mo of age; the Bayley Scales of Infant Development, 2nd edition (BSID2) at 6, 12, and 18 mo; the A-not-B error task at 9 and 12 mo; and free-play attention tasks at 12 and 18 mo. Only infants supplemented with zinc had the normative decline in look duration from 6 to 12 mo during habituation and a normative decline in shifting between objects on free-play multiple-object attention tasks from 12 to 18 mo of age. The 2 groups did not differ on any of the psychophysiologic indices, the BSID2, or the A-not-B error task. The findings are consistent with zinc supplementation supporting a profile of normative information processing and active attentional profiles during the first 2 y of life. This trial was registered at clinicaltrials.gov as NCT00589264.

Sequence effects of self-assembling multidomain peptide hydrogels on encapsulated SHED cells.

Here we report three new nanofibrous, self-assembling multidomain peptide (MDP) sequences and examine the effect of sequence on the morphology and expansion of encapsulated Stem cells from Human Exfoliated Deciduous teeth (SHED). We modified our previously reported set of serine-based MDPs, changing the serine residues in the amphiphilic region to threonine. The three new threonine-based sequences self-assemble into antiparallel ß-sheet nanofibers, confirmed by CD and IR. AFM and negative-stained TEM show that the nanofibers formed by the new sequences are more curved than their serine-containing predecessors. Despite this change in nanofiber morphology, SEM illustrates that all three new sequences still form porous hydrogels. K(TL)2SLRG(TL)3KGRGDS, with a designed cleavage site, is able to be degraded by Matrix Metalloprotease 2. We then examine SHED cell response to these new sequences as well as their serine-based predecessors. We observe faster cell attachment and spreading in hydrogels formed by K2(SL)6K2GRGDS and K(SL)3RG(SL)3KGRGDS. By day 3, the SHEDs in all of the serine-based sequences exhibit a fibroblast-like morphology. Additionally, the SHED cells expand more rapidly in the serine-based gels while the cell number remains relatively constant in the threonine-based peptides. In hydrogels formed by K2(TL)6K2GRGDS and K(TL)2SLRG(TL)3KGRGDS, this low expansion rate is accompanied by changes in morphology where SHEDs exhibit a stellate morphology after 3 days in culture; however, by day 7 they appear more fibroblast-shaped. Throughout the duration of the experiment, the SHED cells encapsulated in the K2(TL)6K2 hydrogels remain rounded. These results suggest that the basic MDP structure easily accommodates modifications in sequence and, for SHED cells, the threonine-containing gels require the integrin-binding RGDS sequence for cell attachment to occur, while the serine-based gels are less selective and support an increase in cell number, regardless of the presence or absence of RGDS.