Past, Present and Future of Human Milk Research

Introduction

Human milk is the best source of infant nutrition, and it is recognized as a biological fluid vital for optimal growth and development of the infant ^[1]. One of the most powerful practices for promoting child survival and well-being is through exclusive breastfeeding during the first 6 months of infant life ^[2]. 

Providing optimal nutrition in the first 6 months of infant life is critical as the consequences of inadequate nutrition can lead to health concerns both in short and long term ^[1]. The World Health Organization (WHO) and United Nations Children’s Fund (UNICEF) recommend initiation of breastfeeding within an hour after birth and exclusively for first 6 months of life, without any additional food, not even water: breastfeed on demand and continued until 2 years or as long as mutually agreeable by mother–infant dyad with provision of safe and nutritious complementary feeding around 6 months of life ^{[2, 3]}. 

Benefits of Breastfeeding for the Mother–Infant Dyad

Aside from bonding experience between the mother and infant, breastfeeding has established short- and long-term benefits ^[1]. One of the most important health benefits of breastfeeding is the preventive effect on infections in the babies ^[4]. Human milk feeding decreases the incidence of a wide range of infectious diseases. Infants who were breastfed exclusively for more than 4 months had 72% reduced risk of hospitalization for lower respiratory tract infections and 74% reduced duration of hospitalization and oxygen requirements of respiratory syncytial virus bronchiolitis. Breastfeeding is also associated with reduced incidence of otitis media and nonspecific gastrointestinal tract infections with a protective role against necrotizing enterocolitis ^[4]. Breastfeeding could prevent 72% of hospital admissions due to diarrhea ^[5]. 

Moreover, exclusive breastfeeding for 3–4 months was found to offer a protective effect in lowering incidence of asthma, atopic dermatitis, and eczema ^[4]. Lower infectious morbidity and mortality and lesser dental malocclusions were evident among those who are breastfed for longer periods ^[5]. Breastfeeding also supports healthy brain development and is associated with higher performance in intelligence among children and adolescents ^[2]. On the other hand, long-term positive effects of breastfeeding have been linked to reducing risk of obesity and type 2 diabetes mellitus later in life ^{[4, 5]}. Breastfeeding for longer periods was associated with 26% reduction of overweight and obesity. With regard to type 2 diabetes, studies indicate 35% reduction, supporting the protective effect of breastfeeding ^[5]. Breastfeeding also provides important health benefits to mothers that include decreased postpartum bleeding and menstrual blood loss and lactational amenorrhea and may contribute to risk reduction for breast cancer and ovarian cancers, type 2 diabetes, and osteoporosis ^{[4, 5]}. 

Evolution of Human Milk: Millions of Years in the Making

As early as the Pennsylvanian period, approximately 310 million years ago, milk originated as a glandular skin secretion in synapsids. The early synapsids laid their eggs with parchment-like shells that are intolerant to desiccation and seemingly dependent on glandular skin secretions for moisture. The glands incorporated elements of the innate immune system in providing protection to the skin and eggs. The secretory fluid and the producing glands became more complex, volumes became greater, and the extent of dependence by the young increased. The skin secretions intended for the moisture and as antimicrobial agents at that time have evolved to this remarkable secretory product as the foundation of nutrient-rich milk long before mammals evolved ^[6]. 

Human Milk Composition

Human milk is described as a complex fluid that is rich in nutrients and non- nutritional bioactive components ^[7]. The nutritional composition and nonnutritive bioactive factors in human milk are uniquely appropriate for the infant, which promote survival and healthy development ^[8]. Human milk also comprises many hundreds to thousands of distinctive bioactive molecules that protect against infection and inflammation and contribute to immune maturation, organ development, and healthy microbial colonization ^[8]. 

Understanding the Past of Human Milk Research

Research over the past 2–3 decades focused on increasing the understanding of the composition of human milk and different factors that influence the composition ^[7]. Human milk is composed of the essential nutrients required by the newborn ^[7]. Nutrients such as proteins, lipids, carbohydrates, minerals, vitamins, and trace elements are of great importance to fulfill the nutritional needs and ensure normal growth and development of the infant ^[4]. Human milk derives its nutritional components from three sources: some of the nutrients of milk originate by synthesis in the lactocyte, some are dietary in origin, and some originate from maternal stores ^[8]. After delivery, the first fluid produced by the mother is colostrum, a fluid rich in immunologic components such as secretory IgA, lactoferrin, leukocytes, and epidermal growth factor. Five days to 2 weeks postpartum, transitional milk is produced to support the nutritional and developmental needs of the rapidly growing infant. Mature milk is produced about 2–4 weeks postpartum ^[8]. Human milk macronutrient composition varies within mothers and across lactation ^[8]. Human milk composition is dynamic and changes over time, adapting to the changing needs of the growing infant ^[1]. The mean macronutrient composition of mature term milk is estimated to be approximately 0.9–1.2 g/dL for protein, 3.2–3.6 g/dL for lipid, and 6.7–7.8 g/dL for lactose ^[8]. 

Human Milk Nutrients

Carbohydrates

Disaccharide lactose is the principal carbohydrate of human milk. Lactose is found in higher concentrations in the milk of mothers producing high quantities of milk ^[8]. The average content of lactose increases slightly from colostrum to transitional milk and mature milk ^[9]. A stable lactose concentration is important in maintaining a constant osmotic pressure in human milk. Lactose also facilitates the absorption of minerals including calcium ^[1]. Other significant carbohydrates found in human milk are the oligosaccharides. These are nonnutritive bioactive factors that are approximately 1 g/dL in human milk and depend on the stage of lactation and maternal genetic factors ^[8]. Human milk oligosaccharides have anti-infective properties against pathogens in the gastro-intestinal tract by acting as soluble decoy binding sites for pathogenic microbes. It also plays a significant role in the development of a diverse and balanced microbiota that are essential for appropriate innate and adaptive immune responses ^[1].

Proteins 

The third most abundant solid in human milk are proteins. They provide not only the indispensable amino acids for growth but also bioactive proteins and peptides that are highly essential for many functions. The concentration of protein in human milk is highest during the first weeks of lactation and decreases throughout the first year (fig. 1) ^[10]. During the first 2 weeks of lactation, concentration of whey proteins is very high, while that of caseins is low, resulting in a whey:casein ratio as high as 80:20 ^[10]. The quality and quantity of protein in human milk are crucial for healthy growth and long-term development. Human milk contains a multitude of bioactive proteins that are highly concentrated in early lactation and play immunomodulatory and antimicrobial effects that provide insights on why breastfed infants have lower morbidity and shorter periods of infection ^[10].

Lipids 

The most highly variable macronutrient in human milk is the lipid content ^[8]. Human milk contains over 200 fatty acids, and long-chain polyunsaturated fatty acids (PUFAs) constitute around 2% of total fatty acids in human milk ^[11]. Colostrum has the lower percentage of lipid relative to transitional milk; however, mature milk has, on average, 3.6% lipid, which may vary depending on time of the day, diet, duration of lactation, and even the length of time elapsed between feeds ^[11]. The long chain PUFAs of human milk vary in relation to maternal diet ^[8]. The lipid content in human milk is highly influenced by the diet of the mother and is also positively related to weight gain during pregnancy ^[1]. 

Vitamins and Minerals 

Human milk contains adequate amounts of most vitamins and minerals to support the growth of a normal infant, except vitamin D and vitamin K ^[1]. Thus, supplementation with vitamin D, vitamin K, and vitamin B12 early in lactation and with iron after 6 months of age may be needed ^[12]. Minerals in human milk contribute to different physiological functions and as important parts of many enzymes ^[1]. 

Factors Affecting Human Milk Composition

Stages of Lactation

In early stages of lactation, there is a presence of a relatively high abundance of immune-related factors ^[8]. There is also a higher concentration of total proteins in colostrum and transitional milk when compared with mature milk ^[13]. In contrast to lipids, they are present in higher concentration in mature milk ^[14].

Impact of Maternal Diet

An important factor that influences the volume and composition of human milk is maternal diet. Regardless of the nutritional status of the mother, the macro-nutrient composition of human milk is relatively constant. Evidence showed that the quality of fatty acid intake during pregnancy and lactation was significant. Fish oil supplementation starting at 20 weeks of pregnancy was found to be positively associated with infant docosahexaenoic acid (DHA) status. There are several studies that have highlighted the influence of maternal diet on levels of PUFA, which is a modifiable nutrient ^[15]. However, a good example of a nonmodifiable nutrient is calcium, in which calcium intake of mothers had no significant effect on the calcium concentration in human milk ^[16].

Geographical Location

Geographical location is contributory to the human milk fatty acid composition, and this is frequently attributed to the differences in the diet of the mother. Concentrations of epidermal growth factor in human milk significantly decreased with increase in intake of proteins, total energy, vegetable, fruits, soy products, and dairy foods. On the other hand, transforming growth factor alpha content in human milk significantly increased with increasing intake of carbohydrates and dairy products and decreased with increasing intake of proteins and meat. Human milk oligosaccharides, minerals, specifically phosphorus, and growth factors in human milk also vary depending on the geographical location ^[15]. It is worth noting that geography is often a proxy for underlying habits of mothers, which are specific to each region.

Gestational Age of Infant at Birth

Human milk composition also varies depending on the infant’s gestational age at birth. No differences were observed for lipid and energy content, but lower lactose concentrations were found in preterm compared to term human milk ^[15]. Preterm milk was higher in true protein than term milk with differences up to 35% in colostrum. Compared to mature milk, colostrum has higher protein but lower energy, lipid, and lactose for both preterm and term milk ^[17].

Immunological markers were found to be in higher concentrations in human milk of mothers of preterm babies. Preterm human milk was shown to have a highly variable content of human milk oligosaccharides and was richer with glycosaminoglycans when compared to term human milk ^[15]. In terms of human milk, there are higher copper and zinc concentrations that decline across lactation, while lower calcium concentrations that remain constant over lactation ^[14].

Circadian Rhythm

One of the identified factors that influences human milk composition is circadian rhythm. The circadian variability of human milk macronutrients showed highest content of protein and lipid during the day expressions and the lowest during the night expressions ^[18]. Lipids as well as lipolytic enzymes in human milk show a circadian rhythm and peak at midday ^[15]. On the other hand, there are no significant fluctuations in carbohydrate content in the human milk for 24 h. The fluctuations of the macronutrient content in human milk were more prominent following premature birth ^[18].

Changes within Feed

 The lipid content of foremilk is relatively low and increases with feeding, whereas hindmilk has the higher lipid content ^[19]. When compared to foremilk, hindmilk has two to three times higher lipid content ^[20]. On the other hand, no significant differences are noted between the protein and lactose contents ^[19].

These broad range of factors that include maternal and infant characteristics affect human milk composition and volume across lactation periods. Some of these factors are modifiable and some are nonmodifiable. The infant-related factors known to affect the nutritive and nonnutritive components of human milk are well studied. However, little is known or evidence is limited with regard to different maternal factors ^[15].

For a detailed review of the past knowledge of factors affecting nutritive and nonnutritive components of milk, we encourage readers to refer to one of our earlier publications that summarizes the existing knowledge ^[15].

The Present of Human Milk Research

Presently, different groups are working on the establishment of reference from mother’s milk in the form of databases that involve cross-continent collaborations using reference and growth standards methodology. A key example is the multicenter collaborative study known as Mothers, Infants, and Lactation Quality (MILQ) Study, in which reference values will be established for nutrients in human milk across the first 8.5 months postpartum. Analyses of milk will include macronutrients, selected vitamins, trace elements and minerals, iodine, metabolomics, amino acids, human milk oligosaccharides, and bioactive peptides. The reference values will be constructed based on methods for the WHO Child Growth Standards and the INTERGROWTH 21st Project ^[21]. Another example is the online database for human milk composition in China. Human milk samples were collected from different provinces in China and the nutrition related composition of human milk across the country was analyzed ^[22]. Collaborative efforts are ongoing in communicating the clinical advantages of human milk composition in relation to public health. Understanding the health benefits of human milk and the long-term impact to infants and mothers will further improve breastfeeding practices. 

Future Directions of Human Milk Research

Toward the future, continuous efforts should be made in exploring the structure–function relationship of human milk, and the focus is directed to preterm infants and small for gestational age infants. With advancements in the analytical methods and technology, the exploration of bioactive factors and nutrients in human milk will continue. It will further recognize temporal modeling as well as interaction networks within human milk. An effective way to understand human milk as a biological system will involve applications of the modern technologies with optimal use of conventional analytical methods in human milk research. It includes chronobiology and systems biology approach ^[23]. Other methods such as computational and modeling methods in understanding milk as a system should be employed. Advanced approaches to uncover human milk as a biological system are highly important, and it is through identification of maternal and infant factors that greatly influence its composition and biology ^[23]. Cellular agriculture is the next exciting field of human milk research. It includes a set of technologies for manufacturing products from livestock farming using culturing techniques ^[24]. This emerging field is taking the challenge to try and replicate growth of breast cells and produced cell-based human milk, since understanding the composition will ultimately affect the infant health outcomes in the near and distant future ^[25].

References

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2. United Nations Children’s Fund (UNICEF). Breastfeeding: A mother’s gift, for every child, 2018. Available from: https://data.unicef.org/ resources/breastfeeding-a-mothers-gift-for-every-child/. 

3. World Health Organization (WHO). Breastfeeding, 2019. Available from: https://www.who.int/ health-topics/breastfeeding#tab=tab_1. 

4. Anatolitou F. Human milk benefits and breastfeeding. J Pediatr Neonat Individual Med. 2012;1:11–8. 

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7. Prentice A. Constituents of human milk. Food Nutr Bull. 1996;17:1–10. 

8. Ballard O, Morrow AL. Human milk composition: nutrients and bioactive factors. Pediatr Clin North Am. 2013;60:49–74. 

9. Yi DY, Kim SY. Human breast milk composition and function in human health: from nutritional components to microbiome and microRNAs. Nutrients. 2021;13:3094. 

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17. Gidrewicz DA, Fenton TR. A systematic review and meta-analysis of the nutrient content of preterm and term breast milk. BMC Pediatr. 2014;14:216. 

18. Paulaviciene IJ, Liubsys A, Molyte A, et al. Circadian changes in the composition of human milk macronutrients depending on pregnancy duration: a cross-sectional study. Int Breastfeed J. 2020;15:49 

19. Kim SY, Yi DY. Components of human breast milk: from macronutrient to microbiome and microRNA. Clin Exp Pediatr. 2020;63:301–9. 

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21. Allen L, Hampel D, Shahab-Ferdows S, et al. The mothers, infants, and lactation quality (MILQ) study: a multi-center collaboration. Curr Dev Nutr. 2021;5:nzab116. 

22. Yin SA, Yang ZY. An on-line database for human milk composition in China. Asia Pac J Clin Nutr. 2016;25:818–25. 

23.  Christian P, Smith ER, Lee SE, et al. The need to study human milk as a biological system. Am J Clin Nutr. 2021;113:1063–72. 

24.  Mendly-Zambo Z, Powell L, Newman L. Dairy 3.0: cellular agriculture and future of milk. Food Cult Soc. 2021;24:675–93. 

25. Beecher C. Human milk the next challenge for cellular ag, 2022. Available from: https://www. foodsafetynews.com/2022/05/human-milk-thenext-challenge-for-cellular-ag/.