Preterm infants in the NICU are fed human milk from their own mothers or donors, and exposed to a variety of handling practices, including fresh mother’s milk, frozen and thawed mother’s milk and Holder pasteurized donor milk. Freeze-thaw cycles and pasteurization can alter the structure of milk proteases and proteins, and these alterations can affect the release of milk peptides. Milk peptides present in the infant intestine have an array of activities including antimicrobial, and enterocyte- and macrophage-immunomodulatory. Differences in milk sources, processing, and storage may yield differential release of bioactive peptides, and therefore differential activities in the gastrointestinal tract of infants. There is a critical need to evaluate effects of milk handling practices on release of peptides within the infant and their bioactivities. Our long-term goal is to determine feeding practices for preterm infants that promote optimal ex utero development and growth. The overall objective of this proposal is to identify how common milk handling practices affect the release of gut health-promoting peptides within the preterm infant intestine. Our central hypothesis is that the identity and amounts of milk peptides released within the intestine of preterm infants fed with these milks will be markedly different and therefore will have different profiles of antimicrobial, and enterocyte- and macrophage-immunomodulatory actions. Our specific aims are to determine the 1) antimicrobial and bifidogenic activities, 2) enterocyte-immunomodulatory activity, and 3) macrophage- immunomodulatory activity of peptides in the intestinal contents of preterm infants fed fresh mother’s milk (MM), frozen and thawed mother’s milk (FTMM), Holder-pasteurized mother’s milk (HPMM) and Holder- pasteurized donor milk (HPDM). Our approach will be to collect intestinal samples from preterm neonates within the neonatal intensive care unit (NICU) after feeding milk exposed to different handling practices, extract the peptide component from these samples, test their antibacterial, bifidogenic and enterocyte- and macrophage-immunomodulatory activity in vitro, and identify the peptides via mass spectrometry and database searching techniques. This research is innovative because it identifies the bioactivities of peptides released during in vivo intestinal digestion and examines the effect of human milk handling practices. At the conclusion of this project, we expect to have determined 1) the extent to which milk peptides released in the preterm infant intestine express gut health-related bioactivity; and 2) how different milk handling practices affect peptide release and bioactivity. The positive impact of this research is that it will help determine optimal milk handling practices for preterm infants, and possibly supplementation strategies for milk that will lead to improved bioactivity within the infant gut.
