Sampling Procedures for Estimating the Infant Intake of Human Milk Leptin, Adiponectin, Insulin, Glucose, and Total Lipid.

Limited attention is given to the efficacy of protocols for the estimation of infant intake of milk components when investigating their impact on infant outcomes. We compared the actual measured intake of human milk components with estimations derived from 15 protocols to determine the most reliable approach for estimating intake of HM leptin, adiponectin, insulin, glucose, and total lipid. Twenty mothers who were 3-5 months postpartum completed a 24 h milk profile study with pre-/post-feed milk samples collection. The true infant intake (control group) based on 24 h milk intake (MI) was compared to estimated infant intakes using concentrations from five sampling protocols that were multiplied by one of true infant MI, considered mean MI (800 mL), or global mean MI (766 mL). The mean measured concentrations of six samples (three sets of pre- and post-feed samples, from morning (06:00-09:00), afternoon (13:00-16:00), and evening (19:00-22:00)) multiplied by the true infant MI, mean considered MI, and global mean MI produced the most accurate estimates of infant intake of these components. Therefore, in the absence of 24 h measurements and sampling, a sampling protocol comprising three sets of pre-/post-feed samples provides the most reliable infant intake estimates of HM leptin, adiponectin, insulin, glucose, and total lipid.

Milk microbiome transplantation: recolonizing donor milk with mother's own milk microbiota.

Donor human milk (DHM) provides myriad nutritional and immunological benefits for preterm and low birthweight infants. However, pasteurization leaves DHM devoid of potentially beneficial milk microbiota. In the present study, we performed milk microbiome transplantation from freshly collected mother's own milk (MOM) into pasteurized DHM. Small volumes of MOM (5%, 10%, or 30% v/v) were inoculated into pasteurized DHM and incubated at 37 °C for up to 8 h. Further, we compared microbiome recolonization in UV-C-treated and Holder-pasteurized DHM, as UV-C treatment has been shown to conserve important biochemical components of DHM that are lost during Holder pasteurization. Bacterial culture and viability-coupled metataxonomic sequencing were employed to assess the effectiveness of milk microbiome transplantation. Growth of transplanted MOM bacteria occurred rapidly in recolonized DHM samples; however, a greater level of growth was observed in Holder-pasteurized DHM compared to UV-C-treated DHM, potentially due to the conserved antimicrobial properties in UV-C-treated DHM. Viability-coupled metataxonomic analysis demonstrated similarity between recolonized DHM samples and fresh MOM samples, suggesting that the milk microbiome can be successfully transplanted into pasteurized DHM. These results highlight the potential of MOM microbiota transplantation to restore the microbial composition of UV-C-treated and Holder-pasteurized DHM and enhance the nutritional and immunological benefits of DHM for preterm and vulnerable infants. KEY POINTS: • Mother's own milk microbiome can be successfully transplanted into donor human milk. • Recolonization is equally successful in UV-C-treated and Holder-pasteurized milk. • Recolonization time should be restricted due to rapid bacterial growth.

Effects of Different Thawing and Warming Processes on Human Milk Composition.

The composition of human milk is influenced by storage and processing practices. The effects of thawing and warming practices on human milk composition remain poorly studied despite their prevalence in home, research, and donor milk bank settings. This review comprehensively examines the impact of different thawing and warming methods on nutritional and bioactive human milk components. While some components such as carbohydrates and minerals remain stable under most typical thawing and warming conditions, others, such as fat, immune proteins, bacterial and human cells, and peptide amine hormones, are sensitive to warming. This review has identified that the data on the effects of milk thawing and warming is limited and often contradictory. Given that numerous important components of milk are diminished during cold storage, it is important that thawing and warming practices do not lead to further loss of or alterations to beneficial milk components. Further work in this field will facilitate greater standardization of thawing methods among researchers and underpin recommendations for thawing and warming of expressed milk for parents.

Causes of Low Milk Supply: The Roles of Estrogens, Progesterone, and Related External Factors.

Low milk supply (LMS) poses a significant challenge to exclusive and continued breastfeeding, affecting ∼10% to 15% of mothers. Milk production is intricately regulated by both endocrine and autocrine control mechanisms, with estrogens and progesterone playing pivotal roles in this process. In addition to endogenously produced hormones, external substances capable of interfering with normal hormonal actions, including phytoestrogens, mycoestrogens, synthetic estrogens, and hormonal contraceptives, can influence milk production. The effects of these extrinsic hormones on milk production may vary based on maternal body mass index. This comprehensive review examines the multifaceted causes of LMS, focusing on the involvement of estrogens, progesterone, and related external factors in milk production. Furthermore, it investigates the interplay between hormonal factors and obesity, aiming to elucidate the endocrine mechanisms underlying obesity-associated LMS. Insights from this review provide valuable perspectives for developing interventions to improve milk production and address the challenges associated with LMS.

Circadian Variation in Human Milk Hormones and Macronutrients.

There is an inadequate understanding of the daily variations in hormones and macronutrients in human milk (HM), and sample collection protocols vary considerably from study to study. To investigate changes in these milk components across 24 h, 22 lactating women collected small milk samples before and after each breastfeed or expression from each breast. Test weighing was used to determine the volume of HM consumed in each feed. The concentrations of leptin, adiponectin, insulin, fat, and glucose were measured, and the intakes were calculated. A linear mixed model was fitted to assess within-feed and circadian variation in HM feed volume and concentration, and intakes of several components. The average infant intake of HM was 879 g/24 h. Significantly higher pre-feed concentrations were found for adiponectin and glucose and lower post-feed concentrations were found for insulin and fat. Significant circadian rhythms were displayed for leptin, adiponectin, insulin, glucose (both concentration and intake), fat concentration, and milk volume. These findings demonstrate the necessity for setting up standardised and rigorous sampling procedures that consider both within-feed and circadian variations in HM components to gain a more precise understanding of the impacts of these components on infant health, growth and development.

Relationships between the Intakes of Human Milk Components and Body Composition of Breastfed Infants: A Systematic Review.

Human milk provides all of the elements necessary for infant growth and development. Previous studies have reported associations between breastfeeding and a reduced risk of developing obesity and late-onset metabolic disorders; however, the underlying mechanisms are poorly understood. Recently, intakes of human milk components have been associated with infant body composition, which is likely partially implicated in the reduced risk of developing childhood obesity among breastfed infants. In this systematic review, we searched electronic bibliographic databases for studies that explored relationships between the 24 h intakes of human milk macronutrients and bioactive components and infant body composition and/or growth parameters. Of 13 eligible studies, 10 assessed relationships of infant body composition and growth outcomes with human milk macronutrients, while 8 studies assessed relationships with human milk bioactive components. Significant time-dependent relationships with infant anthropometrics and body composition were found for intakes and no relationships for concentrations of several human milk components, such as lactose, total protein, and human milk oligosaccharides, suggesting that measuring concentrations of human milk components without quantifying the intake by the infant may provide a limited understanding. Future studies investigating the effect of human milk components on infant growth and body composition outcomes should consider measuring the actual intake of components and employ standardised methods for measuring milk intake.

Oestrogens and progesterone in human milk and their effects on infant health outcomes: A narrative review.

Human milk (HM) is a complex biological system that contains a wide range of bioactive components including oestrogens and progesterone. Whilst maternal oestrogens and progesterone concentrations drop rapidly after birth, they remain detectable in HM across lactation. Phytoestrogens and mycoestrogens, which are produced by plants and fungi, are also present in HM and can interact with oestrogen receptors to interfere with normal hormone functions. Despite the potential impact of HM oestrogens and progesterone on the infant, limited research has addressed their impact on the growth and health of breastfed infants. Furthermore, it is important to comprehensively understand the factors that contribute to these hormone levels in HM, in order to establish effective intervention strategies. In this review, we have summarized the concentrations of naturally occurring oestrogens and progesterone in HM from both endogenous and exogenous sources and discussed both maternal factors impacting HM levels and relationships with infant growth.

Associations of Secretory Activation Breast Milk Biomarkers with Breastfeeding Outcome Measures.

OBJECTIVE: The objective of this study was to explore associations between day 10 postpartum (D10) secretory activation biomarkers and the breastfeeding outcome measures. STUDY DESIGN: This prospective longitudinal descriptive study collected antepartum, D10, and day 60 postpartum (D60) questionnaire data and D10 milk samples. Protein, lactose, and citrate were analyzed with enzymatic spectrophotometric assays. Sodium and potassium were analyzed with inductively coupled plasma optical emission spectrophotometry. Group comparison data were analyzed using χ2, Fisher exact, and independent sample t tests, as appropriate, using SPSS for Mac (version 28). RESULTS: Participants (n = 92) provided a D10 breastmilk sample and completed D10 questionnaires, and 83 completed D60 questionnaires. Participants with D10 impaired secretory activation sodium (>23.0 mM) were more likely to report D10 perceived insufficient milk supply, χ2 = 7.002, P < .05; and less D10 feeding/pumping frequency a day, P < .05; and partial breastfeeding at D60, P < .05. Additionally, participants with D10 impaired secretory activation sodium-to-potassium ratio (sodium: potassium) > 0.8 were more likely to partially breastfeed at D60, P < .05. CONCLUSION: Elevated milk sodium and sodium: potassium are biomarkers related to variables indicative of low milk supply. Therefore, immediate milk testing can be useful in identifying lactation compromise and improving breastfeeding duration. Because breastfeeding affords maternal and infant health benefits, clinicians should identify ways to measure lactation compromise in conjunction with an examination and clinical history to provide early interventions to increase breastfeeding duration and exclusivity.

Human Milk Microbiome and Microbiome-Related Products: Potential Modulators of Infant Growth.

Infant growth trajectory may influence later-life obesity. Human milk provides a wide range of nutritional and bioactive components that are vital for infant growth. Compared to formula-fed infants, breastfed infants are less likely to develop later-onset obesity, highlighting the potential role of bioactive components present in human milk. Components of particular interest are the human milk microbiota, human milk oligosaccharides (HMOs), short-chain fatty acids (SCFAs), and antimicrobial proteins, each of which influence the infant gut microbiome, which in turn has been associated with infant body composition. SCFAs and antimicrobial proteins from human milk may also systemically influence infant metabolism. Although inconsistent, multiple studies have reported associations between HMOs and infant growth, while studies on other bioactive components in relation to infant growth are sparse. Moreover, these microbiome-related components may interact with each other within the mammary gland. Here, we review the evidence around the impact of human milk microbes, HMOs, SCFAs, and antimicrobial proteins on infant growth. Breastfeeding is a unique window of opportunity to promote optimal infant growth, with aberrant growth trajectories potentially creating short- and long-term public health burdens. Therefore, it is important to understand how bioactive components of human milk influence infant growth.

Human Milk Sodium and Potassium as Markers of Mastitis in Mothers of Preterm Infants.

Introduction: This prospective longitudinal study examined changes in milk sodium concentration (Na) and sodium:potassium ratio (Na:K), microbiological culture, milk production, and breast health in relation to mastitis after preterm birth. Methods: We studied women who gave birth at 29-34 weeks of gestation in a tertiary obstetric hospital in Perth, Western Australia. Milk samples, 24-hour milk production, and breast health data were collected every second day to day 10 postpartum, then every third day until infant discharge from the neonatal unit. Milk Na and K were measured at point of care (POC) using handheld ion selective meters, and Na:K calculated. Cultures were performed on postnatal days 8, 13, and every 6 days thereafter. For episodes of mastitis, milk was cultured at onset, and Na and Na:K measured daily until resolution. Women were followed up at 4 and 8 weeks postpartum. Results: In a sample of 44 women, 4 mastitis cases were detected in 3 women during their infants' neonatal stay; all had elevated milk Na and Na:K that resolved within 48 hours; 2/4 experienced reduced milk production and 1/4 had heavy growth of Staphylococcus epidermidis. A further 2 mastitis cases were reported in 39 women followed up to 8 weeks postpartum. Four women had elevated milk Na and Na:K without clinical signs of mastitis; three also had reduced milk production. Conclusions: POC testing of milk Na and/or Na:K may offer a useful indicator of breast health. Mastitis may cause an acute reduction in milk production regardless of the presence of culture-positive infection.

The Impact of Gestational Diabetes Mellitus on Human Milk Metabolic Hormones: A Systematic Review.

Gestational diabetes mellitus (GDM) is a common pregnancy complication with short- and long-term health consequences for the infant and mother. Breastfeeding is the recommended mode of feeding as it offers an opportunity to reduce the risk of GDM consequences, likely partially mediated through changes in human milk (HM) composition. This review systematically reviewed 12 identified studies that investigated the impact of GDM on concentrations of HM metabolic hormones. Meta-analysis was not possible due to significant heterogeneity in study designs and hormone measurement techniques. The risk of bias was assessed using the National Institute for Clinical Excellence (NICE) tool. The methodological qualities were medium in half of the studies, while 25% (3/12) of studies carried a high risk of bias. Significant relationships were reported between GDM and concentrations of HM ghrelin (3/3 studies), insulin (2/4), and adiponectin (2/6), which may play an integral role in infant growth and development. In conclusion, preliminary evidence suggests that GDM may alter HM metabolic hormone concentrations; however, these relationships may be limited to the early lactation stage.

Delayed secretory activation and low milk production in women with gestational diabetes: a case series.

BACKGROUND: Gestational diabetes mellitus (GDM) is major pregnancy complication that is associated with short- and long-term consequences for both mother and infant, including increased risk of diabetes later in life. A longer breastfeeding duration has been associated with a reduced risk of diabetes, however, women with GDM are less likely to exclusively breastfeed and have shorter breastfeeding duration. While the timing of breastfeeding initiation and milk removal frequency affects subsequent breastfeeding outcomes, little is known about early infant feeding practices and milk production in women with GDM. This case series offers detailed prospective breastfeeding initiation data, as well as the first report of objective measures of milk production in women with GDM. CASE PRESENTATION: In this case series, we present the early infant feeding practices of eight women with GDM that gave birth at term gestation. Women recorded the timing of initiation of breastfeeding and secretory activation, as well as their breastfeeding, expression and formula feeding frequencies on postpartum days 1, 7 and 21. Measurement of 24 h milk production volume was performed at 3 weeks postpartum using the test weight method. We observed a delayed first breastfeed (> 1 h) in 6 (75%) cases, formula use in hospital in 5 (63%) cases and delayed secretory activation in 3 (38%) cases. At 3 weeks postpartum, 2 cases had measured milk productions that were insufficient to sustain adequate infant weight gain. CONCLUSIONS: Our data suggest that despite early and frequent milk removal, women with GDM are at greater risk of delayed secretory activation and low milk supply. Cohort studies that consider co-morbidities such as obesity are needed to determine the lactation outcomes of women with GDM.

Human Milk Macronutrients and Bioactive Molecules and Development of Regional Fat Depots in Western Australian Infants during the First 12 Months of Lactation.

We investigated associations between intakes of human milk (HM) components (macronutrients and biologically active molecules) and regional fat depots development in healthy term infants (n = 20) across the first year of lactation. Infant limb (mid-arm and mid-thigh) lean and fat areas were assessed by ultrasound imaging at 2, 5, 9 and 12 months of age. Concentrations of HM total protein, whey protein, casein, adiponectin, leptin, lysozyme, lactoferrin, secretory IGA, total carbohydrates, lactose, HM oligosaccharides (total HMO, calculated) and infant 24-h milk intake were measured, and infant calculated daily intakes (CDI) of HM components were determined. This pilot study shows higher 24-h milk intake was associated with a larger mid-arm fat area (p = 0.024), higher breastfeeding frequency was associated with larger mid-arm (p = 0.008) and mid-thigh (p < 0.001) fat areas. Lysozyme (p = 0.001) and HMO CDI (p = 0.004) were time-dependently associated with the mid-arm fat area. Intakes of HM components and breastfeeding parameters may modulate infant limb fat depots development during the first year of age and potentially promote favorable developmental programming of infant body composition; however, further studies are needed to confirm these findings.

Exclusively Breastfed Infant Microbiota Develops over Time and Is Associated with Human Milk Oligosaccharide Intakes.

Temporal development of maternal and infant microbiomes during early life impacts short- and long-term infant health. This study aimed to characterize bacterial dynamics within maternal faecal, human milk (HM), infant oral, and infant faecal samples during the exclusive breastfeeding period and to document associations between human milk oligosaccharide (HMO) intakes and infant oral and faecal bacterial profiles. Maternal and infant samples (n = 10) were collected at 2−5, 30, 60, 90 and 120 days postpartum and the full-length 16S ribosomal RNA (rRNA) gene was sequenced. Nineteen HMOs were quantitated using high-performance liquid chromatography. Bacterial profiles were unique to each sample type and changed significantly over time, with a large degree of intra- and inter-individual variation in all sample types. Beta diversity was stable over time within infant faecal, maternal faecal and HM samples, however, the infant oral microbiota at day 2−5 significantly differed from all other time points (all p < 0.02). HMO concentrations and intakes significantly differed over time, and HMO intakes showed differential associations with taxa observed in infant oral and faecal samples. The direct clinical relevance of this, however, is unknown. Regardless, future studies should account for intakes of HMOs when modelling the impact of HM on infant growth, as it may have implications for infant microbiota development.

Breast Milk Production in Women Who Use Nipple Shields for Persistent Nipple Pain.

OBJECTIVE: To examine relationships between nipple pain scores and 24-hour milk production volumes, breastfeeding and pumping frequencies, and breastfeeding duration in women using nipple shields for persistent nipple pain. DESIGN: Secondary outcome analysis of a prospective cohort study. SETTING: Research laboratory and participants' homes. PARTICIPANTS: Twenty-five breastfeeding women (6 ± 4 weeks after birth) who used nipple shields for persistent nipple pain. METHODS: We conducted a randomized trial to investigate the primary outcome of milk transfer with and without nipple shields among participants with and without nipple pain. Here, we report secondary outcomes of associations between 24-hour milk production, breastfeeding and pumping frequencies, breastfeeding durations, and intake in participants using a nipple shield for nipple pain. Participants completed demographic, health and breastfeeding questionnaires and, at two monitored breastfeeding sessions, completed a pain visual analogue scale and Brief Pain Inventory-Short Form (BPI-SF; total and subscale scores for pain interference with General Activity, Mood, Sleep, and Breastfeeding). Milk production (milliliters per 24 hours), feed volumes, and percentage of available milk removed were calculated from data and milk samples obtained by participants over one 24-hour period and at study visits. Participants logged 24-hour data on a customized research website. We used descriptive statistics as well as simple and multiple linear regression for analyses. RESULTS: Milk production and feeding duration were not associated with nipple pain scores (visual analogue scale: p = .80, BPI-SF: p = .44). An increase in BPI-SF Breastfeeding subscale score of 1 unit, indicating pain interference with breastfeeding, was associated with a 0.28 decrease in 24-hour breastfeeding frequency (p = .02) and an 18.8-ml decrease in 24-hour breastfeeding intake (p = .04). CONCLUSION: Persistent nipple pain was associated with reduced breastfeeding frequency; therefore, continuing professional support is required to ensure adequate milk removal and pain management.

Use of a Portable Point-of-Care Instrumentation to Measure Human Milk Sodium and Potassium Concentrations.

Objective: Human milk (HM) sodium (Na) and potassium (K) concentrations, as well as the Na and K ratio (Na:K), are associated with stages of lactation and breast health. Portable point-of-care instruments to measure HM biomarkers related to secretory activation or tight junction disturbances would supply clinicians immediate information for individualized lactation care. This study compared HM concentrations of Na and K and Na:K measured by a biochemist with inductively coupled plasma optical emission spectrometry (ICP-OES) and measured by a clinician with ion-selective electrode probes (ISEPs). Design: HM samples (n = 92) were participant collected and donated on day 10 postpartum through a convenience sample of breastfeeding women in West Michigan. Na and K concentrations were determined using ICP-OES and ISEPs. Bland-Altman plots, paired sample t-test, and logistic regressions were completed using R 3.4.4 and R Studio 1.2.1335 with package Lattice. Results: Na concentrations were not significantly different with ICP-OES (13.0 ± 14.8 mM) and ISEPs (12.6 ± 13.7 mM, p = 0.06), whereas K concentrations (ICP-OES 16.1 ± 1.9 mM, ISEPs 14.4 ± 2.4 mM, p < 0.001) and Na:K (ICP-OES 0.81 ± 0.92, ISEPs 1.01 ± 1.62, p = 0.011) were significantly different. Between both methods, the Na, K, and Na: K linear relationships were statistically significant. Conclusion: With additional longitudinal and clinical usefulness research clinicians could use ISEPs to collect and interpret relevant lactation data to support individualized lactation interventions.

Human Milk Lactose, Insulin, and Glucose Relative to Infant Body Composition during Exclusive Breastfeeding.

Human milk (HM) components may influence infant growth and development. This study aimed to investigate relationships between infant body composition (BC) and HM lactose, insulin, and glucose (concentrations and calculated daily intakes (CDI)) as well as 24-h milk intake and maternal BC at 3 months postpartum. HM samples were collected at 2 months postpartum. Infant and maternal BC was assessed with bioimpedance spectroscopy. Statistical analysis used linear regression accounting for infant birth weight. 24-h milk intake and CDI of lactose were positively associated with infant anthropometry, lean body mass and adiposity. Higher maternal BC measures were associated with lower infant anthropometry, z-scores, lean body mass, and adiposity. Maternal characteristics including BC and age were associated with concentrations and CDI of HM components, and 24-h milk intake. In conclusion, 24-h intake of HM and lactose as well as maternal adiposity are related to development of infant BC.

25 Years of Research in Human Lactation: From Discovery to Translation.

Researchers have recently called for human lactation research to be conceptualized as a biological framework where maternal and infant factors impacting human milk, in terms of composition, volume and energy content are studied along with relationships to infant growth, development and health. This approach allows for the development of evidence-based interventions that are more likely to support breastfeeding and lactation in pursuit of global breastfeeding goals. Here we summarize the seminal findings of our research programme using a biological systems approach traversing breast anatomy, milk secretion, physiology of milk removal with respect to breastfeeding and expression, milk composition and infant intake, and infant gastric emptying, culminating in the exploration of relationships with infant growth, development of body composition, and health. This approach has allowed the translation of the findings with respect to education, and clinical practice. It also sets a foundation for improved study design for future investigations in human lactation.

Development of Visceral and Subcutaneous-Abdominal Adipose Tissue in Breastfed Infants during First Year of Lactation.

This study aimed to investigate relationships between infant abdominal visceral and subcutaneous adiposity and human milk (HM) components and maternal body composition (BC) during first year of lactation. Subcutaneous-abdominal depth (SAD), subcutaneous-abdominal fat area (SFA), visceral depth (VD) and preperitoneal fat area of 20 breastfed infants were assessed at 2, 5, 9 and 12 months using ultrasound. Maternal BC was determined with bioimpedance spectroscopy. HM macronutrients and bioactive components concentrations and infant 24-h milk intake were measured and calculated daily intakes (CDI) determined. Maternal adiposity associated with infant SFA (negatively at 2, 5, 12, positively at 9 months, all overall p < 0.05). 24-h milk intake positively associated with infant SAD (p = 0.007) and VD (p = 0.013). CDI of total protein (p = 0.013), total carbohydrates (p = 0.004) and lactose (p = 0.013) positively associated with SFA. Lactoferrin concentration associated with infant VD (negatively at 2, 12, positively at 5, 9 months, overall p = 0.003). CDI of HM components and maternal adiposity have differential effects on development of infant visceral and subcutaneous abdominal adiposity. Maintaining healthy maternal BC and continuing breastfeeding to 12 months and beyond may facilitate favourable BC development reducing risk of obesity.

Human Milk Metabolic Hormones: Analytical Methods and Current Understanding.

Human milk (HM) contains a wide array of peptide hormones including leptin and adiponectin, which are involved in the regulation of infant growth and development. These essential hormones might play an important role in the regulation of metabolic reprogramming of the new-born infant. However, HM hormone studies are sparse and heterogeneous in regard to the study design, sample collection, preparation and analysis methods. This review discussed the limitations of HM hormone analysis highlighting the gaps in pre-analytical and analytical stages. The methods used to quantify HM metabolic hormones (leptin, adiponectin, ghrelin, insulin, obestatin, resistin and apelin) can be classified as immunoassay, immunosensor and chromatography. Immunoassay methods (ELISA and RIA) have been predominantly used in the measurement of these HM hormones. The relative validity parameters of HM hormones analysis are often overlooked in publications, despite the complexity and differences of HM matrix when compared to that of plasma and urine. Therefore, appropriate reports of validation parameters of methodology and instrumentation are crucial for accurate measurements and therefore better understanding of the HM metabolic hormones and their influences on infant outcomes.