CHARACTERIZATION OF HUMAN MILK PROTEOME AND DETERMINATION OF FACTORS UNDERLYING GROWTH AND NUTRITIONAL STATUS AMONG INFANTS HOSPITALIZED WITH ACUTE ILLNESS

Abstract

There have been significant efforts to reduce childhood deaths such as increased access to health care, childhood vaccination, better hygiene and sanitation, and improved nutrition. Despite these efforts infants and young children with poor nutritional status from low- and middle-income countries are susceptible to infectious diseases and are at increased risk of mortality. Breastfeeding reduces the risk of infectious disease-related mortality and promotes infant growth and WHO recommends exclusive breastfeeding for the first six months of life. However, the composition of breast milk may have considerable individual variability and changes during lactation. It is not known whether the composition of breast milk varies among mothers with children of different nutritional status, which may impact recovery from acute illness and growth. This study aims to determine the association between breast milk proteome composition and infant nutritional status and growth following an acute illness. This study was nested within the Breast Milk Composition cohort study that aimed to determine the role of breast milk composition in recovery from infant illness and malnutrition. The study utilized breast milk samples (N=250) from Kenya and Pakistan collected from breastfeeding mothers with infants admitted to hospitals with acute illness and non-hospitalized infants from the same communities. Defatted and casein-depleted breast milk samples were analyzed using liquid chromatography-tandem mass spectrometry and protein identification and quantification were done using the MaxQuant software. Data was preprocessed through filtering, imputing missing values, normalization and correction of batch effects. Elastic net and Random Forest models were used to perform dimension reduction and feature selection. Biological pathway enrichment analysis of the differentially expressed proteins was conducted using ClusterProfiler and DAVID software. Crude linear regression and adjusted mixed-effect linear regression models were used to determine the association between identified proteins and infant’s nutritional status and growth. Correction for multiple testing was carried out using the Benjamin Hochberg’s false discovery rate method. Hospitalized undernourished infants were younger than non-hospitalized infants (P<0.001). Differential protein expression analysis showed that mothers of hospitalized and non-hospitalized infants have different milk proteomic profiles. Mothers of hospitalized infants had upregulation of immune related biological processes and downregulation of body fluids regulation and lactation processes when compared with mothers of non-hospitalized infants. Beta-casein was positively while S1008A, lactadherin and transthyretin were negatively associated with nutritional status including MUAC, weight-for-age, and length-for-age Z scores among infants at hospital admission. Further, selenium-binding-protein1, chordin-like-protein2 and tenascin-c-hexabrachion were among the proteins positively associated with growth as depicted by change in infant’s MUAC from admission to day 45 after hospital discharge. This study has demonstrated that breast milk proteome composition differs between mothers with hospitalized and non-hospitalized infants. Further, the study has shown that breast milk proteins are associated with infants’ nutritional status and growth. While this study requires validation in larger cohorts, the study findings provide mechanistic understanding that can inform interventions to enhance convalescence and improve nutritional status and growth among acutely ill infants. Further studies should be undertaken to validate the findings from the current study.