67 Infant Health Status Impacts the Neutrophil Phenotypes and Polyunsaturated Fatty Acids Composition in Human Milk
Authors: Badewy Rana , Glogauer Michael , Azarpazhooh Amir , Tenenbaum Howard , Connor Kristin , Sgro Michael , Bazinet Richard , Fine Noah , Sun Chunxiang , Saha Sourav , Lai Jim 
Published: September 2023

Abstract

Background
The abundance of human milk leukocytes, including neutrophils, changes in response to the health status of mothers and their infants. Elevated leukocyte counts occur during infant infection and return to baseline levels upon recovery. However, it is unclear how infant health status can affect the neutrophil phenotypes and the concentration of fatty acids in their mothers’ human milk.

Objectives
This study investigates the association between infant health status and human milk neutrophil counts and activation state, and fatty acid levels, in human milk.

Design/Methods
This is a prospective cohort study of 50 healthy breastfeeding mothers recruited from St. Michael’s hospital, in Toronto, Ontario, who were followed up from 2-4 weeks until 4 months postpartum. Human milk samples were collected from participants and data regarding infant health status were collected from self-reported questionnaires completed by participants at both timepoints, then imputed based on whether infants had taken antibiotics, had been hospitalized, or had been diagnosed with a medical condition, at least 2 weeks prior to samples collection. Neutrophils were quantified using flow cytometry and labelled with a panel of antibodies to detect specific cluster of differentiation (CD) biomarkers. Fatty acids were identified and quantified using a gas chromatography-flame ionization detector (GC-FID). Linear mixed-effects models were used to evaluate the correlation between infant health status and changes in neutrophil counts, along with their expressed CD markers, and fatty acids composition in human milk during lactation.

Results
The CD markers were classified into 4 categories based on their function: degranulation/activation markers (CD63, CD64, and CD66a), an immunoregulation marker (CD16), adhesion markers (CD11b, CD18), and a lipopolysaccharide receptor (CD14). Human milk from mothers whose infants had a health condition had neutrophils which significantly expressed elevated levels of the CD64 biomarker (β: 85.65, p=0.009 in adjusted models), compared to human milk from mothers of healthy infants. However, there were no significant associations between infant health status, absolute hmPMN counts, andother CD biomarkers (p&gt;0.05). There were also elevated levels of arachidonic acid (β: 0.12, p=0.013) and C22:5n-6 (β: 0.06, p=0.015) fatty acids in the human milk of mothers whose infants had a health condition, compared with human milk from mothers of healthy infants, after adjusting for maternal age, post-pregnancy body mass index, infant sex, and infant feeding pattern.

Conclusion
This study demonstrates that infant health status is associated with changes in human milk immunological components, suggesting an inflammatory protective response mechanism in the mammary glands triggered by infants infection. How these alterations can affect infant health outcomes in the short and longer terms need critical consideration.