Effect of circadian rhythm on NAD and other metabolites in human brain.

Nicotinamide Adenine Dinucleotide (NAD) plays a central role in the master circadian clock of the brain (the suprachiasmatic nuclei, SCN) as demonstrated in many model organisms. NAD acts as an enzyme co-factor and substrate and its modulation was found to be tightly regulated to the periodicity of the cycles. However, in human brain, the effect of the circadian rhythm (CR) on the metabolism of the SCN and other brain regions is poorly understood. We conducted a magnetic resonance spectroscopy (MRS) study at a high magnetic field, measuring the occipital brain NAD levels and other metabolites in two different morning and afternoon diurnal states in 25 healthy participants. Salivary cortisol levels were determined to confirm that the experiment was done in two chronologically different physiological conditions, and a behavioral test of risk-taking propensity was administered. Overall, we found that the CR did not significantly affect NAD levels in the occipital brain region. The other brain metabolites measured, including lactate, were not significantly affected by the CR either, except for taurine. The CR did impact risk-taking behavior and salivary cortisol level, confirming that the participants were in two circadian different behavioral and physiological states in the morning and in the afternoon. Measurement of the CR effect on NAD and taurine levels in other brain regions might provide stronger effects.

Impact of a Nutrient Formulation on Longitudinal Myelination, Cognition, and Behavior from Birth to 2 Years: A Randomized Clinical Trial.

Observation studies suggest differences in myelination in relation to differences in early life nutrition. This two-center randomized controlled trial investigates the effect of a 12-month nutritional intervention on longitudinal changes in myelination, cognition, and behavior. Eighty-one full-term, neurotypical infants were randomized into an investigational (N = 42) or a control group (N = 39), receiving higher versus lower levels of a blend of nutrients. Non-randomized breastfed infants (N = 108) served as a reference group. Main outcomes were myelination (MRI), neurodevelopment (Bayley-III), social-emotional development (ASQ:SE-2), infant and toddler behavior (IBQ-R and TBAQ), and infant sleep (BISQ) during the first 2 years of life. The full analysis set comprised N = 67 infants from the randomized groups, with 81 myelin-sensitive MRI sequences. Significantly higher myelination was observed in the investigational compared to the control group at 6, 12, 18, and 24 months of life, as well as significantly higher gray matter volume at 24 months, a reduced number of night awakenings at 6 months, increased day sleep at 12 months, and reduced social fearfulness at 24 months. The results suggest that brain development may be modifiable with brain- and age-relevant nutritional approaches in healthy infants and young children, which may be foundational for later learning outcomes.

Temporal evolution of fatty acid content in human milk of lactating mothers from the Philippines.

Fatty acids (FA) play a key role in infant growth and development. The aim of this study was to study the temporal evolution of FA from 3 or 4 weeks to 4 months postpartum in human milk (HM) from Filipino mothers. Mid-morning HM samples (n = 41) were collected after full expression from a single breast and FA were assessed using gas-liquid chromatography coupled to flame ionization detector. The total FA content remained relatively constant over the study period. The most abundant FA in HM were oleic acid (OA), palmitic acid (PA) and linoleic acid (LA), a trend similarly reported in HM from European and Chinese mothers. The former two were unchanged over the course of lactation while there was a slight increase in LA content over time. Similarly, the saturated fatty acid (SFA) and monounsaturated FA (MUFA) contents did not vary over the first four months of lactation. The SFA content was much higher than that reported in HM from Europe and China, mainly driven by PA, lauric and myristic acids. The MUFA content on the other hand, while comparable to that reported in HM from Chinese populations was lower than that reported in Europe. There was a small increase in the polyunsaturated FA (PUFA) content over the study duration. The levels of essential FA, linoleic acid (LA) and α-linolenic acid (ALA) were found to be much lower than that reported in other populations. The concentrations of arachidonic acid (AA), docosahexaenoic acid (DHA) and eicosapentaenoic acid (EPA) remained stable over the study duration. AA and DHA in HM from Filipino mothers were comparable to global averages, however in case of the latter the concentration was found to be lower than in previous reports. DHA is of great clinical significance as it plays a key role in infant growth and development. In our study, we observed a wide inter- and intra-individual variability in the levels of DHA in HM, presumably reflecting diverse intakes of DHA rich foods and bioconversion in vivo. Personalized recommendations may help achieve recommended levels of DHA amongst population with levels below global averages. This may help achieve HM sufficiency and therefore be linked to clinical benefits for the mother and the baby. SUMMARY: This study details the temporal evolution of human milk (HM) fatty acids (FA) in Filipino mothers up to four months postpartum. The total FA content remained relatively constant over the study period. The most abundant FA were oleic, palmitic and linoleic acids. HM from Filipino mothers had relatively higher saturated FA content driven by palmitic, lauric and myristic acids, while the levels of essential FA, linoleic and α-linoleic acids were lower compared to other populations. Similarly, the concentration of monounsaturated FA were also lower than that reported in HM from European mothers. Arachidonic acid and docosahexaenoic acid (DHA) concentrations were comparable to global averages however the HM DHA levels were seen to have decreased when compared to previous reports from the Philippines. Additionally, a wide variability was seen in HM DHA levels suggesting a need for strategies such as personalized recommendations in order to ensure HM DHA sufficiency.

Effect of an enteral amino acid blend on muscle and gut functionality in critically ill patients: a proof-of-concept randomized controlled trial.

BACKGROUND: A defining feature of prolonged critical illness is muscle wasting, leading to impaired recovery. Supplementation with a tailored blend of amino acids may bolster the innate gut defence, promote intestinal mucosa repair and limit muscle loss. METHODS: This was a monocentric, randomized, double-blind, placebo-controlled study that included patients with sepsis or acute respiratory distress syndrome. Patients received a specific combination of five amino acids or placebo mixed with enteral feeding for 21 days. Markers of renal function, gut barrier structure and functionality were collected at baseline and 1, 2, 3 and 8 weeks after randomization. Muscle structure and function were assessed through MRI measurements of the anterior quadriceps volume and by twitch airway pressure. Data were compared between groups relative to the baseline. RESULTS: Thirty-five critically ill patients were randomized. The amino acid blend did not impair urine output, blood creatinine levels or creatinine clearance. Plasma citrulline levels increased significantly along the treatment period in the amino acid group (difference in means [95% CI] 5.86 [1.72; 10.00] nmol/mL P = 0.007). Alanine aminotransferase and alkaline phosphatase concentrations were lower in the amino acid group than in the placebo group at one week (ratio of means 0.5 [0.29; 0.86] (P = 0.015) and 0.73 [0.57; 0.94] (P = 0.015), respectively). Twitch airway pressure and volume of the anterior quadriceps were greater in the amino acid group than in the placebo group 3 weeks after randomization (difference in means 10.6 [0.99; 20.20] cmH20 (P = 0.035) and 3.12 [0.5; 5.73] cm3/kg (P = 0.022), respectively). CONCLUSIONS: Amino acid supplementation increased plasma citrulline levels, reduced alanine aminotransferase and alkaline phosphatase levels, and improved twitch airway pressure and anterior quadriceps volume. Trial registration ClinicalTrials.gov, NCT02968836. Registered November 21, 2016.

Dynamics of human milk oligosaccharides in early lactation and relation with growth and appetitive traits of Filipino breastfed infants.

Human milk oligosaccharides play a key role in the maturation of the infant gut microbiome and immune system and are hypothesized to affect growth. This study examined the temporal changes of 24 HMOs and their associations to infant growth and appetitive traits in an exploratory, prospective, observational, study of 41 Filipino mother-infant dyads. Exclusively breastfed, healthy, term infants were enrolled at 21-26 days of age (≈ 0.75 mo) and followed for 6 months. Infant growth measures and appetitive traits were collected at visit 1 (V1) (≈ 0.75 mo), V2 (≈ 1.5 mo), V3 (2.5 mo), V4 (2.75 mo), V5 (4 mo), and V6 (6 mo), while HMOs were measured at V1, V2, V3 and V5. Overall exposure to each HMO was summarized as area under the curve from baseline to 4 months of age and examined in association with each measure of growth at 6 months using linear regression adjusted for maternal age at birth, infant sex, birth weight, and mode of delivery. We saw modest associations between several HMOs and infant growth parameters. Our results suggest that specific HMOs, partly as proxy for milk groups (defined by Secretor and Lewis status), may be associated with head circumference and length, increasing their relevance especially in populations at the lower end of the WHO growth curve. We did not identify the same HMOs associated with infant appetitive traits, indicating that at least in our cohort, changes in appetite were not driving the observed associations between HMOs and growth.Clinical trial registration: NCT03387124.

Human milk oligosaccharides in breast milk and 2-year outcome in preterm infants: An exploratory analysis.

BACKGROUND & AIMS: The health benefit of human milk (HM) for preterm infant development is known but the role of human milk oligosaccharides (HMOs) contained in HM remains underexplored. We explored the relationship between exposure to HMOs contained in mother's milk and growth and neurodevelopment at 2-years corrected age in preterm infants. METHODS: Exclusively breastfed preterm infants born between 27 and 34 weeks of gestation were enrolled in a monocentric prospective observational study, LACTACOL. Samples of breast milk were collected once a week for 7 weeks after birth. HMOs and sialic acid were measured by liquid chromatography. Age and Stages questionnaire (ASQ) version 2 was used to assess 2-year neurodevelopmental outcome. We analyzed the relationship between HMO content and (i) infant neurodevelopment at 2-years, and (ii) growth outcome at discharge and at 2 years. A secondary analysis was performed among Secretor(+) Lewis(+) mothers. Only associations with a false discovery rate of 10% or less according to the Benjamini-Hochberg procedure were considered significant. RESULTS: 137 preterm infants (mean gestational age of 31.3 ± 1.7 weeks, mean birth weight of 1494 g ± 336 g) born to 117 mothers (mean age of 30.8 ± 5.0 years) were enrolled. Total HMOs and most individual HMOs and sialic acid concentrations decreased with advancing postnatal age, except for lacto-N-fucopentaose-III and 3-fucosyllactose, which increased. Total HMOs were positively correlated with neonatal length growth (adjusted p = 0.012). Neither total HMOs nor any individual HMO correlated with ASQ score in the overall cohort. However, lacto-N-fucopentaose-III (LNFP-III) was significantly associated with total ASQ score (adjusted p ≤ 0.015) among the 104 infants born to Secretor(+) Lewis(+) mothers. CONCLUSIONS: In this exploratory study in very preterm infants, total HMOs and most individual HMOs, except LNFP-III, decreased with advancing postnatal age. Neither the concentration of total HMOs nor that of any individual HMO were associated with ASQ score at 2 years, except for LNFP-III in Secretor(+) Lewis(+) mothers.

Use of a Liquid Supplement Containing 2 Human Milk Oligosaccharides: The First Double-Blind, Randomized, Controlled Trial in Pre-term Infants.

There is growing evidence supporting the benefit of human milk oligosaccharides (HMOs) on reducing risk of illnesses and improving immune function in newborn infants, but evidence in pre-term infants is lacking. This randomized, double-blind, placebo-controlled trial (NCT03607942) of pre-term infants evaluated the effects of HMO supplementation on feeding tolerance, growth, and safety in 7 neonatal units in France. Pre-term infants (27-33 weeks' gestation, birth weight <1,700 g) were randomized early after birth to receive HMO supplement (n = 43) [2'-fucosyllactose (2'FL) and lacto-N-neotetraose (LNnT) in a 10:1 ratio (0.374 g/kg body weight/day)] or an isocaloric placebo (n = 43) consisting of only glucose (0.140 g/kg/day) until discharge from the neonatal unit. Anthropometric z-scores were calculated using Fenton growth standards. Primary outcome was feeding tolerance, measured by non-inferiority (NI) in days to reach full enteral feeding (FEF) from birth in HMO vs. placebo group (NI margin = 4+ days). Mean number of days on intervention prior to FEF was 8.9 and 10.3 days in HMO and placebo, respectively. Non-inferiority in time to reach FEF in HMO (vs. placebo) was achieved [LS mean difference (95% CI) = -2.16 (-5.33, 1.00); upper bound of 95% CI < NI margin] in full analysis set and similar for per protocol. Adjusted mean time to reach FEF from birth was 2 days shorter in HMO (12.2) vs. placebo (14.3), although not statistically significant (p = 0.177). There was no difference in weight-for-age z-scores between groups throughout the FEF period until discharge. Length-for-age z-scores were higher in HMO at FEF day 14 [0.29 (0.02, 0.56), p = 0.037] and 21 [0.31 (0.02, 0.61), p = 0.037]. Head circumference-for-age z-score was higher in HMO vs. placebo at discharge [0.42 (0.12, 0.71), p = 0.007]. Occurrence of adverse events (AEs) was similar in both groups and relatively common in this population, whereas 2.3 and 14.3%, respectively, experienced investigator-confirmed, related AEs. HMO supplementation is safe and well-tolerated in pre-term infants. After 9 days of supplementation, the HMO group reached FEF 2 days earlier vs. placebo, although the difference was not statistically significant. In addition, HMO supplementation supports early postnatal growth, which may have a positive impact on long-term growth and developmental outcomes.

A Nutrient Formulation Affects Developmental Myelination in Term Infants: A Randomized Clinical Trial.

BACKGROUND AND OBJECTIVES: Observational studies suggest differences between breast-fed and formula-fed infants in developmental myelination, a key brain process for learning. The study aims to investigate the efficacy of a blend of docosahexaenoic acid (DHA), arachidonic acid (ARA), iron, vitamin B12, folic acid, and sphingomyelin (SM) from a uniquely processed whey protein concentrate enriched in alpha-lactalbumin and phospholipids compared with a control formulation on myelination, cognitive, and behavioral development in the first 6 months of life. METHODS: These are 6-month results from an ongoing two-center, randomized controlled trial with a 12-month intervention period (completed for all participants). In this study, full term, neurotypical infants of both sexes (N = 81) were randomized into investigational (N = 42) or control groups (N = 39). In addition, non-randomized breast-fed children (N = 108) serve as a natural reference group. Main outcomes are myelination (MRI), cognitive (Bayley Scales of Infant and Toddler Development, 3rd edition [Bayley-III]), social-emotional development (Ages and Stages Questionnaires: Social-Emotional, 2nd edition [ASQ:SE-2]), sleep (Brief Infant Sleep Questionnaire [BISQ]), and safety (growth and adverse events [AEs]). RESULTS: The full analyses set comprises N = 66 infants. Significant differences in myelin structure, volume, and rate of myelination were observed in favor of the investigational myelin blend at 3 and 6 months of life. Effects were demonstrated for whole brain myelin and for cerebellar, parietal, occipital, and temporal regions, known to be functionally involved in sensory, motor, and language skills. No statistically significant differences were found for early behavior and cognition scores. CONCLUSIONS: This is the first study demonstrating the efficacy of a myelin nutrient blend in well-nourished, term infants on developmental myelination, which may be foundational for later cognitive and learning outcomes. CLINICAL TRIAL REGISTRATION: ClinicalTrials.gov, identifier: NCT03111927.

Effects of higher protein formula with improved fat blend on growth, feeding tolerance and nutritional biomarkers in preterm infants: A double-blind, randomized, controlled clinical trial.

BACKGROUND: Preterm formulas containing greater protein:energy ratio are beneficial for non-breastfed infants, since protein is critical for promoting catch-up growth and synthesis of lean body mass. Additionally, formulas containing enriched sn-2 palmitate (sn-2) and reduced medium chain triglycerides (MCTs) may support better feeding tolerance and nutrient utilization. METHODS: The objective of this randomized, controlled, double-blinded clinical trial is to evaluate growth, feeding tolerance and nutritional biomarkers of preterm infants with birth weight ≤2000g and gestational age ≤33wks from one neonatal unit in Vietnam receiving experimental formula (EF, n = 80) containing higher protein level of 3.4 g/100 kcal and improved fat blend with enriched sn-2 and modified level of MCTs or isocaloric control formula (CF, n = 80) containing protein level of 2.9 g/100 kcal and standard fat blend. The differences in weight gain (g/d; primary endpoint) from day 1 (D1) of full enteral feeding (FEF) until D21 between groups was evaluated for non-inferiority (margin = -2.5  g/d) and superiority (margin = 0  g/d). RESULTS: Mean weight gain was 3.09  g/d greater in EF than CF; the lower limit of 95% CI (0.31  g/d) exceeded both non-inferiority and superiority margins. There was no significant difference in length-for-age and head circumference-for-age z-score. By D79, the mean change in weight-for-age z-scores from D1 in EF group (+0.76 SDs) surpassed the criteria for catch-up growth (+0.67 SDs). Infants in the EF group (vs. CF) tended to have softer stools (EF = 3.2 ± 0.59 vs. CF = 3.4 ± 0.58; P = 0.07) based on 5-point scale (1 = watery, 5 = hard). Difference in blood urea nitrogen and biomarkers for bone mineral status (i.e., plasma phosphorus, alkaline phosphatase and urinary calcium/phosphorus ratio) between EF and CF on FEF Day 21 reached statistical significance (P < 0.05) but all mean values stayed within normal clinical ranges for both groups. CONCLUSION: Preterm formula with greater protein:energy ratio and new fat blend is safe, nutritionally suitable, well-tolerated, and improves catch-up weight gain of preterm infants. Clinical trial registry identifier is NCT03055052 (ClinicalTrials.gov).

Metabolism of Exogenous D-Beta-Hydroxybutyrate, an Energy Substrate Avidly Consumed by the Heart and Kidney.

There is growing interest in the metabolism of ketones owing to their reported benefits in neurological and more recently in cardiovascular and renal diseases. As an alternative to a very high fat ketogenic diet, ketones precursors for oral intake are being developed to achieve ketosis without the need for dietary carbohydrate restriction. Here we report that an oral D-beta-hydroxybutyrate (D-BHB) supplement is rapidly absorbed and metabolized in humans and increases blood ketones to millimolar levels. At the same dose, D-BHB is significantly more ketogenic and provides fewer calories than a racemic mixture of BHB or medium chain triglyceride. In a whole body ketone positron emission tomography pilot study, we observed that after D-BHB consumption, the ketone tracer 11C-acetoacetate is rapidly metabolized, mostly by the heart and the kidneys. Beyond brain energy rescue, this opens additional opportunities for therapeutic exploration of D-BHB supplements as a "super fuel" in cardiac and chronic kidney diseases.

Modulation of cerebral ketone metabolism following traumatic brain injury in humans.

Adaptive metabolic response to injury includes the utilization of alternative energy substrates - such as ketone bodies (KB) - to protect the brain against further damage. Here, we examined cerebral ketone metabolism in patients with traumatic brain injury (TBI; n = 34 subjects) monitored with cerebral microdialysis to measure total brain interstitial tissue KB levels (acetoacetate and ß-hydroxybutyrate). Nutrition - from fasting vs. stable nutrition state - was associated with a significant decrease of brain KB (34.7 [10th-90th percentiles 10.7-189] µmol/L vs. 13.1 [6.5-64.3] µmol/L, p < 0.001) and blood KB (668 [168.4-3824.9] vs. 129.4 [82.6-1033.8] µmol/L, p < 0.01). Blood KB correlated with brain KB (Spearman's rho 0.56, p = 0.0013). Continuous feeding with medium-chain triglycerides-enriched enteral nutrition did not increase blood KB, and provided a modest increase in blood and brain free medium chain fatty acids. Higher brain KB at the acute TBI phase correlated with age and brain lactate, pyruvate and glutamate, but not brain glucose. These novel findings suggest that nutritional ketosis was the main determinant of cerebral KB metabolism following TBI. Age and cerebral metabolic distress contributed to brain KB supporting the hypothesis that ketones might act as alternative energy substrates to glucose. Further studies testing KB supplementation after TBI are warranted.

Comparative study of preterm infants fed new and existing human milk fortifiers showed favourable markers of gastrointestinal status.

AIM: This study examined the influence of different human milk fortifiers on biomarkers of gastrointestinal immaturity and inflammation in preterm infants. METHODS: We report secondary outcomes from a controlled, double-blind, randomised, parallel group study conducted from 2011 to 2014 in neonatal intensive care units at 11 metropolitan hospitals in France, Belgium, Germany, Switzerland and Italy. Preterm infants born at up to 32 weeks or weighing up to 1500 g were randomised to a new powdered human milk fortifier (n = 77) or a control fortifier (n = 76) for a minimum of 21 days. We analysed faecal markers of gut inflammation, namely alpha-1 antitrypsin and calprotectin, and maturity, namely elastase-1. RESULTS: Faecal alpha-1 antitrypsin was slightly lower in the new than control fortifier group after 21 days of full enteral feeding, with a geometric mean and standard deviation of 1.52 ± 1.32 vs 1.82 ± 1.44 mg/g stools (P = .01). There was no significant difference in faecal calprotectin (median [Q1-Q3] of 296 [136-565] µg/g stools in both groups combined at study day 21). Faecal elastase-1 was lower in the new fortifier than control fortifier group (202.5 ± 1.6 vs 257.7 ± 1.5 µg/g stools, P = .016). CONCLUSION: Mean values for each parameter were within the ranges in healthy term infants, indicating favourable markers of gastrointestinal status in both groups. In addition, for faecal calprotectin, the relatively high concentration observed in preterm infants fed fortified human milk suggests that the threshold level for detecting necrotising enterocolitis should be revised.