Lactation and neonatal nutrition: defining and refining the critical questions.

This paper resulted from a conference entitled "Lactation and Milk: Defining and refining the critical questions" held at the University of Colorado School of Medicine from January 18-20, 2012. The mission of the conference was to identify unresolved questions and set future goals for research into human milk composition, mammary development and lactation. We first outline the unanswered questions regarding the composition of human milk (Section I) and the mechanisms by which milk components affect neonatal development, growth and health and recommend models for future research. Emerging questions about how milk components affect cognitive development and behavioral phenotype of the offspring are presented in Section II. In Section III we outline the important unanswered questions about regulation of mammary gland development, the heritability of defects, the effects of maternal nutrition, disease, metabolic status, and therapeutic drugs upon the subsequent lactation. Questions surrounding breastfeeding practice are also highlighted. In Section IV we describe the specific nutritional challenges faced by three different populations, namely preterm infants, infants born to obese mothers who may or may not have gestational diabetes, and infants born to undernourished mothers. The recognition that multidisciplinary training is critical to advancing the field led us to formulate specific training recommendations in Section V. Our recommendations for research emphasis are summarized in Section VI. In sum, we present a roadmap for multidisciplinary research into all aspects of human lactation, milk and its role in infant nutrition for the next decade and beyond

Breastfeeding Concerns at 3 and 7 Days Postpartum and Feeding Status at 2 Months

ObjectiveWe characterized breastfeeding concerns from open-text maternal responses and determined their association with stopping breastfeeding by 60 days (stopping breastfeeding) and feeding any formula between 30 and 60 days (formula use).MethodsWe assessed breastfeeding support, intentions, and concerns in 532 expectant primiparas and conducted follow-up interviews at 0, 3, 7, 14, 30, and 60 days postpartum. We calculated adjusted relative risk (ARR) and adjusted population attributable risk (PAR) for feeding outcomes by concern category and day, adjusted for feeding intentions and education.ResultsIn 2946 interviews, 4179 breastfeeding concerns were reported, comprising 49 subcategories and 9 main categories. Ninety-two percent of participants reported ≥ 1 concern at day 3, with the most predominant being difficulty with infant feeding at breast (52%), breastfeeding pain (44%), and milk quantity (40%). Concerns at any postpartum interview were significantly associated with increased risk of stopping breastfeeding and formula use, with peak ARR at day 3 (eg, stopping breastfeeding ARR [95% confidence interval] = 9.2 [3.0-infinity]). The concerns yielding the largest adjusted PAR for stopping breastfeeding were day 7 "infant feeding difficulty" (adjusted PAR = 32%) and day 14 "milk quantity" (adjusted PAR = 23%).ConclusionsBreastfeeding concerns are highly prevalent and associated with stopping breastfeeding. Priority should be given to developing strategies for lowering the overall occurrence of breastfeeding concerns and resolving, in particular, infant feeding and milk quantity concerns occurring within the first 14 days postpartum

In-Hospital Formula Use Increases Early Breastfeeding Cessation Among First-Time Mothers Intending to Exclusively Breastfeed

ObjectiveTo evaluate in-hospital formula supplementation among first-time mothers who intended to exclusively breastfeed and determined if in-hospital formula supplementation shortens breastfeeding duration after adjusting for breastfeeding intention.Study designWe assessed strength of breastfeeding intentions prenatally in a diverse cohort of expectant primiparae and followed infant feeding practices through day 60. Among mothers planning to exclusively breastfeed their healthy term infants for ≥1 week, we determined predictors, reasons, and characteristics of in-hospital formula supplementation, and calculated the intention-adjusted relative risk (ARR) of not fully breastfeeding days 30-60 and breastfeeding cessation by day 60 with in-hospital formula supplementation (n = 393).ResultsTwo hundred ten (53%) infants were exclusively breastfed during the maternity stay and 183 (47%) received in-hospital formula supplementation. The most prevalent reasons mothers cited for in-hospital formula supplementation were: perceived insufficient milk supply (18%), signs of inadequate intake (16%), and poor latch or breastfeeding (14%). Prevalence of not fully breastfeeding days 30-60 was 67.8% vs. 36.7%, ARR 1.8 (95% CI, 1.4-2.3), in-hospital formula supplementation vs exclusively breastfed groups, respectively, and breastfeeding cessation by day 60 was 32.8% vs. 10.5%, ARR 2.7 (95% CI, 1.7-4.5). Odds of both adverse outcomes increased with more in-hospital formula supplementation feeds (not fully breastfeeding days 30-60, P = .003 and breastfeeding cessation, P = .011).ConclusionsAmong women intending to exclusively breastfeed, in-hospital formula supplementation was associated with a nearly 2-fold greater risk of not fully breastfeeding days 30-60 and a nearly 3-fold risk of breastfeeding cessation by day 60, even after adjusting for strength of breastfeeding intentions. Strategies should be sought to avoid unnecessary in-hospital formula supplementation and to support breastfeeding when in-hospital formula supplementation is unavoidable

RNA sequencing of the human milk fat layer transcriptome reveals distinct gene expression profiles at three stages of lactation.

Aware of the important benefits of human milk, most U.S. women initiate breastfeeding but difficulties with milk supply lead some to quit earlier than intended. Yet, the contribution of maternal physiology to lactation difficulties remains poorly understood. Human milk fat globules, by enveloping cell contents during their secretion into milk, are a rich source of mammary cell RNA. Here, we pair this non-invasive mRNA source with RNA-sequencing to probe the milk fat layer transcriptome during three stages of lactation: colostral, transitional, and mature milk production. The resulting transcriptomes paint an exquisite portrait of human lactation. The resulting transcriptional profiles cluster not by postpartum day, but by milk Na:K ratio, indicating that women sampled during similar postpartum time frames could be at markedly different stages of gene expression. Each stage of lactation is characterized by a dynamic range (10(5)-fold) in transcript abundances not previously observed with microarray technology. We discovered that transcripts for isoferritins and cathepsins are strikingly abundant during colostrum production, highlighting the potential importance of these proteins for neonatal health. Two transcripts, encoding β-casein (CSN2) and α-lactalbumin (LALBA), make up 45% of the total pool of mRNA in mature lactation. Genes significantly expressed across all stages of lactation are associated with making, modifying, transporting, and packaging milk proteins. Stage-specific transcripts are associated with immune defense during the colostral stage, up-regulation of the machinery needed for milk protein synthesis during the transitional stage, and the production of lipids during mature lactation. We observed strong modulation of key genes involved in lactose synthesis and insulin signaling. In particular, protein tyrosine phosphatase, receptor type, F (PTPRF) may serve as a biomarker linking insulin resistance with insufficient milk supply. This study provides the methodology and reference data set to enable future targeted research on the physiological contributors of sub-optimal lactation in humans

Cluster trajectories and functional enrichment across stages of lactation.

<p><b>Panels A-H</b>: Each X-axis indicates the lactation state (colostrum, transitional, mature); Y-axis shows expression changes. Trajectory clustering analyses (see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0067531#s3" target="_blank"><b>Methods</b></a>) show that gene expression does not steadily increase or decrease in the progression from colostral to mature lactation. Eight clusters (<b>Panels </b><b>A-H</b>) represent the major transcriptional trajectories. To the right of each cluster are shown the statistically enriched KEGG pathways and the top 10 significantly enriched biologic process GO terms (n = total number of significantly enriched biologic processes).</p

Relative mRNA abundances in the milk fat layer during transitional lactation.

<p>Each pie slice represents the proportion of the total mRNA pool attributed to expression of the labeled gene during transitional lactation. For direct comparison to <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0067531#pone-0067531-g001" target="_blank">Figure 1</a>, the pie slice representing the most abundant transcript in the <b>Mature</b> transcriptome (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0067531#pone-0067531-g001" target="_blank">Figure 1</a>) is placed at the 3′o-clock position, with subsequent transcripts presented counterclockwise according to decreasing order in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0067531#pone-0067531-g001" target="_blank">Figure 1</a>. The color of the pie slice for any individual transcript is also consistent with <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0067531#pone-0067531-g001" target="_blank">Figure 1</a>.</p

Top 20 Expressed Genes in the Milk Fat Layer of Mature Human Milk^*.

*<p>Source: <a href="http://www.ncbi.nlm.nih.gov/gene/" target="_blank">http://www.ncbi.nlm.nih.gov/gene/</a>.</p

Heat map of differentially expressed genes between all stages of lactation.

<p>Columns are clustered by sample and rows are clustered by gene. Dendogram height indicates distances between clusters in gene expression profiles. The heat map illustrates lower (white/yellow) to higher (orange/red) gene expression levels with distinct transcriptional profiles across lactation stages. Blue bars at the top of each column indicate lactation stage: dark blue = colostral; blue = transitional; and light blue = mature. From left to right, starting with ID 187, postpartum timing of sample collections are 41, 52, 52, 39, 56, and 49 hours; and 130, 33, 35, 40, 24, and 45 days. Similarly, starting with ID 187, Na:K ratios are 9.6, 5.5, 0.71, 0.70, 0.98, 1.15, 0.19, 0.30, 0.57, 0.41, 0.33, and 0.45.</p