Clinical trial NCT03254329: Establishing Global Reference Values for Human Milk
Official title: Establishing Global Reference Values for Human Milk Composition in a Multi-Center Collaborative Study
Study type: OBSERVATIONAL
Overall status: UNKNOWN
Conditions: Human Milk Nutrient Reference Values
Keywords: human milk
micronutrients
lactation
milk volume
milk composition
reference values
Lead sponsor: USDA, Western Human Nutrition Research Center
Sponsor class: FED
Collaborators: name: University of Copenhagen
name: Medical Research Council Unit, The Gambia
name: International Centre for Diarrhoeal Disease Research, Bangladesh
name: Universidade Federal do Rio de Janeiro
name: Bill and Melinda Gates Foundation
First posted: 2017-08-18
Enrollment: 2000
Sex: FEMALE
Minimum age (years): 18
Maximum age (years): 40
Standard ages: ADULT
Accepts healthy volunteers: yes
Observational model: FAMILY_BASED
Time perspective: PROSPECTIVE
Sampling method: NON_PROBABILITY_SAMPLE
Study population: Well-nourished pregnant women in the third trimester of pregnancy at recruitment, then lactating with their infant from 1-8.5 months postpartum.

Brief summary: The primary purpose of the proposed research is to establish Reference Values (RVs) for micronutrients (MN) and macronutrients in human milk. The investigators' research to date has revealed highly variable milk MN concentrations among populations, and very low values in some low income countries (LICs) likely caused by poor maternal status and/or diet but RVs are needed to interpret these values. During the past year the investigators conducted a review of human milk nutrient composition, and formed a Technical Advisory Group (TAG) which developed the current proposal. Here, the investigators are conducting a longitudinal project on well-nourished women and infants. The Mothers, Infants and Lactation Quality (MILQ) study is a multi-center cohort project, investigating breast milk nutrient composition in well-nourished women across the first 8.5 months of lactation in four different populations. The countries involved are Denmark, Brazil, Bangladesh and The Gambia. Exclusive breastfeeding is an eligibility criterion up until the second post-partum study visit (between 1 - 3.4 months postpartum), with the exception of the first week after delivery. Other data collected on mothers and infants, including maternal and infant nutrient intake and status, morbidity, milk volume, and infant development, will inform interpretation and support application of the results. While the priority is to develop RVs for MN, other analyses will include human milk oligosaccharides (HMOs) and proteins, and free amino acids (FAA) in infant plasma. Thus, with the samples obtained the investigators will perform (a) laboratory analyses of milk, plasma and urine nutrients to construct RVs for global application, (b) analyses of HMOs and proteins in milk, and (c) metabolomic analysis of FAA and other metabolites in infant plasma. By request of the Bill and Melinda Gates Foundation (BMGF), colostrum and fecal microbiota samples will also be collected and stored for later analyses.

Detailed description: The investigators will collect breast milk samples at four visits, at 2-3 days after birth (colostrum) and at 1-3.4, 3.5-5.9, and 6-8.5 months of lactation from well-nourished mothers age ≥18 to ≤40 years, in a systematic, identical way in four countries. The four sites are: Copenhagen, Denmark; Banjul, The Gambia; Rio de Janeiro, Brazil; and Mirpur, Dhaka, Bangladesh. The most important site selection criteria were that multiple micronutrient supplements are not consumed after week 28 of pregnancy; maternal diet is adequate but consumption of highly fortified foods is limited; and exclusive breastfeeding is 60% at 4 months.

In order to have 250 women and infants per site complete the study at 8.5 months, approximately 500 women will be recruited during the third trimester of pregnancy. This should allow for mothers and infants not meeting study eligibility criteria in pregnancy or early lactation, and drop-outs from the study. Recruitment during pregnancy will increase the opportunity to locate and recruit women since recruitment in the immediate post-delivery period will be difficult. It will also enable the women to be counselled on the importance of exclusive breastfeeding (EBF). Colostrum will be collected at 2-3 days postpartum for future analysis. The first collection of mature milk will occur between 1 and 3.4 months postpartum, when maternal and infant blood samples will also be collected on all participants. At the final two visits, in addition to milk collection, blood will be taken from all mothers but only half of the infants each time, primarily for the assessment of micronutrient status.

A requirement is that infants must be EBF in the 1-3.4 month period, and breastfed (BF) in the second and third period. Longitudinal measurements on the same mother after the BF period are not necessary for statistical reasons, as the investigators are not creating RVs for change in milk composition. The investigators expect that due to attrition and cessation of BF, the sample size will need to be augmented in later months; the alternative would have been to recruit and measure many more women in earlier stages in order to have an adequate sample size in the 7-8.5 month interval, which would be a much less efficient approach. Therefore after 6 months, if women are not breastfeeding, additional lactating women will be recruited from the group that was not EBF in the 3.4 month period, or from the local health center or community.

At all three points of mature milk collection the investigators will measure breast milk volume; diet, anthropometry and morbidity of the mother and infant; and infant development at 3.5-5.9 and 6 to 8.5 months; and will collect infant fecal samples for future analyses of the microbiome. Milk volume will be measured at three sites using the International Atomic Energy Agency's protocol that requires dosing the mother with deuterated water and collecting maternal and infant saliva urine samples at 0, 1, 2, 3, 4, 13 and 14 days after the dose. In Denmark breast milk volume will be measured by 24 hour infant weighing.

Most of the laboratory analyses will be conducted at the United States Department of Agriculture, Agricultural Research Service, Western Human Nutrition Research Center (WHNRC) in Davis, California. Other analyses will be performed in the Department of Chemistry at University of California, Davis (HMOs and other bioactive compounds in milk), and the Swiss Federal Institute of Technology in Zurich (iodine status). RVs will be constructed following the methods and principles developed for the World Health Organization (WHO) Child Growth Standards and the Intergrowth-21st Project.

An add-on study "Evaluation of maternal insulin resistance, metabolic and inflammatory biomarkers for prediction of successful initiation and duration of breastfeeding" is being conducted at the Danish site and was approved by The Regional Committee on Health Research Ethics with (H-17015174).

The aims of the add-on study are to evaluate if maternal metabolic and inflammatory biomarkers can predict successful initiation and duration of breastfeeding in a population of healthy non-obese Danish women, to evaluate the impact of maternal inflammatory biomarkers during pregnancy on the offspring's growth and risk factors for later disease, and to relate the impact of maternal metabolic and inflammatory biomarkers during pregnancy on breastfeeding behavior and on the offspring's growth and risk factors for later disease in this population to corresponding outcomes in an established cohort of infants born of obese mothers.

Eligibility criteria: Inclusion Criteria for Mother

* No vitamin-mineral supplements during third trimester of pregnancy or during lactation except for iron + folic acid, also vitamin D and Ca in Denmark.
* Low habitual intake of highly-fortified foods (iodized salt excepted).
* No relevant past or current medical problems including gestational diabetes or pre-eclampsia.
* Singleton delivery, not preterm.
* BMI ≥18.5 to \<30.0 kg/m2, height ≥150 cm.
* Mid-upper arm circumference (MUAC) ≥23 and ≤33 cm in pregnancy
* Consuming a nutritionally adequate diet i.e. not vegan or macrobiotic, ≥5 food groups/d each ≥15g. At screening this information will be collected with a locally-appropriate and validated food frequency questionnaire.
* Non-anemic in pregnancy (Hb \>100 g/L)
* Alcohol intake ≤5 units (50 mL pure alcohol)/week.
* Non-smoker.

Inclusion Criteria for Infant

* Birth weight 2500-4200 g, 37-42 weeks of gestation.
* No congenital malformations that interfere with feeding or growth and development.

Exclusion Criteria at 1 to 3.4 Months Postpartum

* Cessation of or non-exclusive breastfeeding.
* Serious maternal illness.
* Infant length-for-age, weight-for-age or weight-for-length \< -2 Z.

Exclusion criteria 3.5 to 8.5 months postpartum

* Cessation of breastfeeding.
* Serious maternal illness.
* Infant length-for-age, weight-for-age or weight-for-length \< -2 Z.

Primary outcomes:
measure: Change in vitamin and mineral concentrations in human breast milk
timeFrame: Breast milk samples collected 1-3.4, 3.5-5.9, and 6-8.5 months postpartum
Secondary outcomes:
measure: Variation in other constituents of human breast milk
timeFrame: Breast milk samples collected at 1-3.4, 3.5-5.9, and 6-8.5 months postpartum
measure: Micronutrient status of mothers and infants
timeFrame: Blood and urine samples surveyed at 1-3.4, 3.5-5.9, and 6-8.5 months postpartum
measure: Milk volume
timeFrame: 1-3.4, 3.5-5.9, and 6-8.5 months postpartum
measure: Maternal and infant iodine status
timeFrame: 1-3.4, 3.5-5.9, and 6-8.5 months postpartum
measure: Milk iodine
timeFrame: 1-3.4, 3.5-5.9, and 6-8.5 months postpartum
measure: Maternal nutrient intake during pregnancy
timeFrame: Measured during gestational week 35-37
measure: Maternal nutrient intake
timeFrame: 1-3.4, 3.5-5.9, and 6-8.5 months postpartum
measure: Infant nutrient intake
timeFrame: 1-3.4, 3.5-5.9, and 6-8.5 months
measure: Infant dietary habits
timeFrame: 1-3.4, 3.5-5.9, and 6-8.5 months
measure: Maternal weight
timeFrame: 1-3.4, 3.5-5.9, and 6-8.5 months postpartum
measure: Infant weight
timeFrame: 1-3.4, 3.5-5.9, and 6-8.5 months
measure: Maternal height
timeFrame: 1-3.4, 3.5-5.9, and 6-8.5 months postpartum
measure: Infant length
timeFrame: 1-3.4, 3.5-5.9, and 6-8.5 months
measure: Maternal body mass index
timeFrame: 1-3.4, 3.5-5.9, and 6-8.5 months postpartum
measure: Infant growth
timeFrame: 1-3.4, 3.5-5.9, and 6-8.5 months postpartum
measure: Maternal body composition
timeFrame: 1-3.4, 3.5-5.9, and 6-8.5 months postpartum
measure: Infant body composition
timeFrame: 1-3.4, 3.5-5.9, and 6-8.5 months
measure: Infant morbidity
timeFrame: 1-3.4, 3.5-5.9, and 6-8.5 months
measure: Infant development
timeFrame: 6-8.5 months
measure: Infant motor development
timeFrame: 3.5-5.9 and 6-8.5 months
measure: Change in infant microbiome
timeFrame: 1-3.4, 3.5-5.9, and 6-8.5 months
measure: Change in maternal microbiome
timeFrame: Collected at gestational week 28-40, and 1-3.4, 3.5-5.9, and 6-8.5 months postpartum
measure: Maternal Glucose Tolerance
timeFrame: Measured once at gestational week 28-30
measure: Maternal Hemoglobin A1c (HbA1c)
timeFrame: Measured once at gestational week 28-30
measure: Maternal markers of insulin sensitivity
timeFrame: Measured once at gestational week 28-30
measure: Maternal lipid panel
timeFrame: Measured once at gestational week 28-30
measure: Maternal inflammatory markers
timeFrame: Measured once at gestational week 28-30
Other outcomes:
Locations:
city: Dhaka
country: Bangladesh
facility: International Centre for Diarrhoeal Disease Research, Bangladesh (icddr,b)
city: Rio de Janeiro
country: Brazil
facility: Federal University of Rio de Janeiro
city: Copenhagen
country: Denmark
facility: University of Copenhagen
state: Frederiksberg
city: Bakau
country: Gambia
facility: Medical Research Council Gambia
state: Kombo Saint Mary's
Study officials:
affiliation: USDA/WHNRC
name: Lindsay H Allen, PhD
role: PRINCIPAL_INVESTIGATOR
References:
citation: Allen LH, Hampel D, Shahab-Ferdows S, Andersson M, Barros E, Doel AM, Eriksen KG, Christensen SH, Islam M, Kac G, Keya FK, Michaelsen KF, de Barros Mucci D, Njie F, Peerson JM, Moore SE. The Mothers, Infants, and Lactation Quality (MILQ) Study: A Multi-Center Collaboration. Curr Dev Nutr. 2021 Sep 20;5(10):nzab116. doi: 10.1093/cdn/nzab116. eCollection 2021 Oct.