Maternal stress is associated with higher protein-bound amino acid concentrations in human milk.

Background: Maternal stress in the postpartum period affects not only the mother but also her newborn child, who is at increased risk of developing metabolic and mental disorders later in life. The mechanisms by which stress is transmitted to the infant are not yet fully understood. Human milk (HM) is a potential candidate as maternal stress affects various components of HM, e.g., fat and immunoglobulin concentrations. To date, it is unknown whether maternal stress also affects the amino acids (AAs) in HM, even though this nutrient is of extreme importance to child health and development. This study aimed to investigate whether and how maternal stress is associated with the AA composition of HM. Methods: In this observational cohort study (Amsterdam, The Netherlands), lactating women were recruited in two study groups: a high-stress (HS) group; women whose child was hospitalized (n = 24), and a control (CTL) group; women who gave birth to a healthy child (n = 73). HM was collected three times a day, on postpartum days 10, 17, and 24. Perceived psychological stress was measured using validated questionnaires, while biological stress measures were based on hair, saliva, and HM cortisol concentrations. HM protein-bound and free AAs were analyzed by liquid chromatography and compared between groups. Results: Maternal perceived stress scores were higher in the HS group (p < 0.01). The concentrations of protein-bound AAs in HM were higher in the HS group compared to the CTL group (p = 0.028) and were positively associated with HM cortisol concentrations (p = 0.024). The concentrations of free AAs did not differ between study groups and were unrelated to cortisol concentrations. Conclusion: Findings from this prospective cohort study suggest that maternal stress in the postpartum period is associated with an altered human milk amino acid composition, which could play a role in the transmission of maternal stress effects to her child. The physiological implications of these stress-induced changes for infant development await future research.

Sex-dependence and comorbidities of the early-life adversity induced mental and metabolic disease risks: Where are we at?

Early-life adversity (ELA) is a major risk factor for developing later-life mental and metabolic disorders. However, if and to what extent ELA contributes to the comorbidity and sex-dependent prevalence/presentation of these disorders remains unclear. We here comprehensively review and integrate human and rodent ELA (pre- and postnatal) studies examining mental or metabolic health in both sexes and discuss the role of the placenta and maternal milk, key in transferring maternal effects to the offspring. We conclude that ELA impacts mental and metabolic health with sex-specific presentations that depend on timing of exposure, and that human and rodent studies largely converge in their findings. ELA is more often reported to impact cognitive and externalizing domains in males, internalizing behaviors in both sexes and concerning the metabolic dimension, adiposity in females and insulin sensitivity in males. Thus, ELA seems to be involved in the origin of the comorbidity and sex-specific prevalence/presentation of some of the most common disorders in our society. Therefore, ELA-induced disease states deserve specific preventive and intervention strategies.

Rapid quantification of insulin in human milk by immunoassay.

Human milk (HM) contains numerous non-nutritive bioactive factors, amongst which the peptide hormone insulin. HM insulin has been suggested to accelerate intestinal maturation, thereby promoting feeding tolerance. Therefore, recombinant human insulin for enteral administration has been developed which might serve as supplement to HM or formula for preterm infants. However, the natural course of the HM insulin concentration directly following delivery is unknown, which hampers the development of dosage schedules in clinical trials. The aim of this study was to validate a method for insulin determination in small volumes of HM, and to assess the stability of HM insulin. The results showed that the HM insulin concentration can be measured rapidly and reliably by using an automated immunoassay. In addition, HM insulin is stable at 4 °C for at least 72 h, at room temperature for a maximum of 12 h, at -20 °C for at least 2.5 years, and during at least five freeze-thaw cycles.

Enteral Bioactive Factor Supplementation in Preterm Infants: A Systematic Review.

Feeding preterm infants with mother's own milk is associated with a reduction in postnatal complications and an improved neurocognitive outcome. Therefore, the bioactive factor composition of human milk has been used as a tool for the development of nutritional supplements with a potential prophylactic or therapeutic effect. The aim of this systematic review was to provide an overview on bioactive factors which have been studied as supplement to enteral nutrition in randomized controlled trials, and to provide an overview of ongoing trials. MEDLINE, EMBASE, CENTRAL, and clinical trial registers were searched. Studies on the antimicrobial protein lactoferrin were excluded as these were summarized very recently in three separate systematic reviews. Studies on vitamins D, K and iron were also excluded as they are already incorporated in most international guidelines. We identified 17 different bioactive factors, which were investigated in 26 studies. Despite the encouraging potential effects of several bioactive factors, more high-quality studies with a sufficient number of preterm infants are required before a certain factor may be implemented into clinical practice. Three large trials (n > 500) that investigate the effects of either enteral insulin or vitamin A are currently ongoing and could provide more definite answers on these specific supplements.

Circadian Variation in Human Milk Composition, a Systematic Review.

BACKGROUND: Breastfeeding is considered the most optimal mode of feeding for neonates and mothers. Human milk changes over the course of lactation in order to perfectly suit the infant's nutritional and immunological needs. Its composition also varies throughout the day. Circadian fluctuations in some bioactive components are suggested to transfer chronobiological information from mother to child to assist the development of the biological clock. This review aims to give a complete overview of studies examining human milk components found to exhibit circadian variation in their concentration. METHODS: We included studies assessing the concentration of a specific human milk component more than once in 24 h. Study characteristics, including gestational age, lactational stage, sampling strategy, analytical method, and outcome were extracted. Methodological quality was graded using a modified Newcastle-Ottawa Scale (NOS). RESULTS: A total of 83 reports assessing the circadian variation in the concentration of 71 human milk components were included. Heterogeneity among studies was high. The methodological quality varied widely. Significant circadian variation is found in tryptophan, fats, triacylglycerol, cholesterol, iron, melatonin, cortisol, and cortisone. This may play a role in the child's growth and development in terms of the biological clock.

The Effects of Early Life Stress, Postnatal Diet Modulation, and Long-Term Western-Style Diet on Later-Life Metabolic and Cognitive Outcomes.

Early life stress (ES) increases the risk to develop metabolic and brain disorders in adulthood. Breastfeeding (exclusivity and duration) is associated with improved metabolic and neurocognitive health outcomes, and the physical properties of the dietary lipids may contribute to this. Here, we tested whether early life exposure to dietary lipids mimicking some physical characteristics of breastmilk (i.e., large, phospholipid-coated lipid droplets; Concept Nuturis® infant milk formula (N-IMF)), could protect against ES-induced metabolic and brain abnormalities under standard circumstances, and in response to prolonged Western-style diet (WSD) in adulthood. ES was induced by exposing mice to limited nesting material from postnatal day (P) 2 to P9. From P16 to P42, male offspring were fed a standard IMF (S-IMF) or N-IMF, followed by either standard rodent diet (SD) or WSD until P230. We then assessed body composition development, fat mass, metabolic hormones, hippocampus-dependent cognitive function, and neurogenesis (proliferation and survival). Prolonged WSD resulted in an obesogenic phenotype at P230, which was not modulated by previous ES or N-IMF exposure. Nevertheless, ES and N-IMF modulated the effect of WSD on neurogenesis at P230, without affecting cognitive function, highlighting programming effects of the early life environment on the hippocampal response to later life challenges at a structural level.

The Importance of Maternal Folate Status for Brain Development and Function of Offspring.

The importance of an adequate periconceptional maternal folate status to prevent fetal neural tube defects has been well demonstrated and resulted in the recommendation for women to use folic acid supplements during the periconception period. The importance of maternal folate status for offspring neurodevelopment and brain health is less well described. We reviewed the current evidence linking maternal folate status before conception and during pregnancy with neurodevelopment and cognition of the offspring. We discuss both animal and human studies. Preclinical research revealed the importance of maternal folate status for several key processes required for normal neurodevelopment and brain functioning in the offspring, including DNA synthesis, regulation of gene expression, synthesis of phospholipids and neurotransmitters, and maintenance of healthy plasma homocysteine concentrations. Human observational studies are inconclusive; about half have shown a positive association between maternal folate status and cognitive performance of offspring. Whereas some studies suggest a positive association between maternal folate intake and cognition of offspring during childhood, data from interventional studies are too limited to conclude that there is a direct effect. Future preclinical studies are needed to help us characterize the behavioral effects, understand the underlying mechanisms, and to establish an optimal dosage and time window of folate supplementation. Moreover, more conclusive data from well-designed human observational studies and randomized controlled trials are needed to determine whether current recommendations for folic acid supplementation during pregnancy cover the needs for normal cognitive development in the offspring.

Increasing availability of ω-3 fatty acid in the early-life diet prevents the early-life stress-induced cognitive impairments without affecting metabolic alterations.

Exposure to early-life stress (ES) is associated with cognitive and metabolic deficits in adulthood. The role of early nutrition in programming these long-term effects is largely unknown. We focused on essential ω-3 and ω-6 long-chain polyunsaturated fatty acids (LCPUFA) and investigated whether ES affects central and peripheral FA profiles, as well as if and how an early diet with increased availability of ω-3 LCPUFA ( via lowering ω-6/ω-3 ratio) protects against ES-induced impairments. ES exposure [limited nesting and bedding paradigm from postnatal day (P)2 to P9] altered central and peripheral FA profiles in mice. An early diet with low ω-6/ω-3 ratio from P2 to P42 notably prevented the ES-induced cognitive impairments, and the alterations in hippocampal newborn cell survival and in CD68+ microglia, without affecting the ES-induced metabolic alterations. Other markers for hippocampal plasticity, apoptosis, and maternal care were unaffected by ES or diet. Our findings highlight the importance of early dietary lipid quality for later cognition in ES-exposed populations.-Yam, K.-Y., Schipper, L., Reemst, K., Ruigrok, S. R., Abbink, M. R., Hoeijmakers, L., Naninck, E. F. G., Zarekiani, P., Oosting, A., Van der Beek, E. M., Lucassen, P. J., Korosi, A. Increasing availability of ω-3 fatty acid in the early-life diet prevents the early-life stress-induced cognitive impairments without affecting metabolic alterations.

Early micronutrient supplementation protects against early stress-induced cognitive impairments.

Early-life stress (ES) impairs cognition later in life. Because ES prevention is problematic, intervention is needed, yet the mechanisms that underlie ES remain largely unknown. So far, the role of early nutrition in brain programming has been largely ignored. Here, we demonstrate that essential 1-carbon metabolism-associated micronutrients (1-CMAMs; i.e., methionine and B vitamins) early in life are crucial in programming later cognition by ES. ES was induced in male C57Bl/6 mice from postnatal d (P)2-9. 1-CMAM levels were measured centrally and peripherally by using liquid chromatography-mass spectroscopy. Next, we supplemented the maternal diet with 1-CMAM only during the ES period and studied cognitive, neuroendocrine, neurogenic, transcriptional, and epigenetic changes in adult offspring. We demonstrate that ES specifically reduces methionine in offspring plasma and brain. Of note, dietary 1-CMAM enrichment during P2-9 restored methionine levels and rescued ES-induced adult cognitive impairments. Beneficial effects of this early dietary enrichment were associated with prevention of the ES-induced rise in corticosterone and adrenal gland hypertrophy did not involve changes in maternal care, hippocampal volume, neurogenesis, or global/Nr3c1-specific DNA methylation. In summary, nutrition is important in brain programming by ES. A short, early supplementation with essential micronutrients can already prevent lasting effects of ES. This concept opens new avenues for nutritional intervention.-Naninck, E. F. G., Oosterink, J. E., Yam, K.-Y., de Vries, L. P., Schierbeek, H., van Goudoever, J. B., Verkaik-Schakel, R.-N., Plantinga, J. A., Plosch, T., Lucassen, P. J., Korosi, A. Early micronutrient supplementation protects against early stress-induced cognitive impairments.

Regulation of Adult Neurogenesis and Plasticity by (Early) Stress, Glucocorticoids, and Inflammation.

Exposure to stress is one of the best-known negative regulators of adult neurogenesis (AN). We discuss changes in neurogenesis in relation to exposure to stress, glucocorticoid hormones, and inflammation, with a particular focus on early development and on lasting effects of stress. Although the effects of acute and mild stress on AN are generally brief and can be quickly overcome, chronic exposure or more severe forms of stress can induce longer lasting reductions in neurogenesis that can, however, in part, be overcome by subsequent exposure to exercise, drugs targeting the stress system, and some antidepressants. Exposure to stress, particularly during the sensitive period of early life, may (re)program brain plasticity, in particular, in the hippocampus. This may increase the risk to develop cognitive or anxiety symptoms, common to brain diseases like dementia and depression in which plasticity changes occur, and a normalization of neurogenesis may be required for a successful treatment response and recovery.

Early-life adversity programs emotional functions and the neuroendocrine stress system: the contribution of nutrition, metabolic hormones and epigenetic mechanisms.

Clinical and pre-clinical studies have shown that early-life adversities, such as abuse or neglect, can increase the vulnerability to develop psychopathologies and cognitive decline later in life. Remarkably, the lasting consequences of stress during this sensitive period on the hypothalamic-pituitary-adrenal axis and emotional function closely resemble the long-term effects of early malnutrition and suggest a possible common pathway mediating these effects. During early-life, brain development is affected by both exogenous factors, like nutrition and maternal care as well as by endogenous modulators including stress hormones. These elements, while mostly considered for their independent actions, clearly do not act alone but rather in a synergistic manner. In order to better understand how the programming by early-life stress takes place, it is important to gain further insight into the exact interplay of these key elements, the possible common pathways as well as the underlying molecular mechanisms that mediate their effects. We here review evidence that exposure to both early-life stress and early-life under-/malnutrition similarly lead to life-long alterations on the neuroendocrine stress system and modify emotional functions. We further discuss how the different key elements of the early-life environment interact and affect one another and next suggest a possible role for the early-life adversity induced alterations in metabolic hormones and nutrient availability in shaping later stress responses and emotional function throughout life, possibly via epigenetic mechanisms. Such knowledge will help to develop intervention strategies, which gives the advantage of viewing the synergistic action of a more complete set of changes induced by early-life adversity.

Accurate measurement of the essential micronutrients methionine, homocysteine, vitamins B6, B12, B9 and their metabolites in plasma, brain and maternal milk of mice using LC/MS ion trap analysis.

Methionine, homocysteine, vitamins B6, B12, B9, and their metabolites are crucial co-factors and substrates for many basic biological pathways including one-carbon metabolism, and they are particularly important for brain function and development and epigenetic mechanisms. These are essential nutrients that cannot be synthesized endogenously and thus need to be taken in via diet. A novel method was developed that enables simultaneous assessment of the exact concentrations of these essential micronutrients in various matrices, including maternal milk, plasma, and brain of neonatal mice. The protocol for analysis of these components in the various matrices consists of a cleanup step (i.e. lipid extraction followed by protein precipitation) combined with a liquid chromatography mass spectrometry (LC/MS) ion trap method with high sensitivity and selectivity (SRM mode). This novel method enables the measurement of these essential nutrients with good recoveries (69-117%), and high intra-day (<10%) and high intra-day precision (defined as <15% for compounds with an isotopologue and <20% for compounds without an isotopologue as internal standard) in plasma, maternal milk, and brain of mice at low and high levels. In addition, lower limits of quantitation (LOQ) were determined for the various matrices in the range for methionine (700-2000nmol/L), homocysteine (280-460-nmol/L), vitamins B6 (5-230nmol/L), B12 (7-11nmol/L), B9 (20-30nmol/L). Degradation of vitamins and oxidation of homocysteine is limited to a minimum, and only small sample volumes (30µL plasma, 20mg brain and maternal milk) are needed for simultaneous measurement. This method can help to understand how these nutrients are transferred from mother to offspring via maternal milk, as well as how these nutrients are absorbed by the offspring and eventually taken up in various tissues amongst the brain in preclinical and clinical research settings. Therefore the method can help to explore critical periods in lactating mothers and developing offspring.

Chronic early life stress alters developmental and adult neurogenesis and impairs cognitive function in mice.

Early life stress (ES) increases vulnerability to psychopathology and impairs cognition in adulthood. These ES-induced deficits are associated with lasting changes in hippocampal plasticity. Detailed information on the neurobiological basis, the onset, and progression of such changes and their sex-specificity is currently lacking but is required to tailor specific intervention strategies. Here, we use a chronic ES mouse model based on limited nesting and bedding material from postnatal day (P) 2-9 to investigate; (1) if ES leads to impairments in hippocampus-dependent cognitive function in adulthood and (2) if these alterations are paralleled by changes in developmental and/or adult hippocampal neurogenesis. ES increased developmental neurogenesis (proliferation and differentiation) in the dentate gyrus (DG) at P9, and the number of immature (NeurD1(+)) cells migrating postnatally from the secondary dentate matrix, indicating prompt changes in DG structure in both sexes. ES lastingly reduced DG volume and the long-term survival of developmentally born neurons in both sexes at P150. In adult male mice only, ES reduced survival of adult-born neurons (BrdU/NeuN(+) cells), while proliferation (Ki67(+)) and differentiation (DCX(+)) were unaffected. These changes correlated with impaired performance in all learning and memory tasks used here. In contrast, in female mice, despite early alterations in developmental neurogenesis, no lasting changes were present in adult neurogenesis after ES and the cognitive impairments were less prominent and only apparent in some cognitive tasks. We further show that, although neurogenesis and cognition correlate positively, only the hippocampus-dependent functions depend on changes in neurogenesis, whereas cognitive functions that are not exclusively hippocampus-dependent do not. This study indicates that chronic ES has lasting consequences on hippocampal structure and function in mice and suggests that male mice are more susceptible to ES than females. Unraveling the mechanisms that underlie the persistent ES-induced effects may have clinical implications for treatments to counteract ES-induced deficits.

Perinatal programming of adult hippocampal structure and function; emerging roles of stress, nutrition and epigenetics.

Early-life stress lastingly affects adult cognition and increases vulnerability to psychopathology, but the underlying mechanisms remain elusive. In this Opinion article, we propose that early nutritional input together with stress hormones and sensory stimuli from the mother during the perinatal period act synergistically to program the adult brain, possibly via epigenetic mechanisms. We hypothesize that stress during gestation or lactation affects the intake of macro- and micronutrients, including dietary methyl donors, and/or impairs the dam's metabolism, thereby altering nutrient composition and intake by the offspring. In turn, this may persistently modulate gene expression via epigenetic programming, thus altering hippocampal structure and cognition. Understanding how the combination of stress, nutrition, and epigenetics shapes the adult brain is essential for effective therapies.