Abstract 14050: Characteristics of Extracellular Vesicle miRNAs in the Breast Milk

Abstract

Introduction: An increasing number of obese mothers is a rising issue that may negatively impact the overall benefit of breastfeeding. Extracellular vesicles (EVs) in breast milk are important mediators transferring biological molecules of mothers to children, because EV-miRNAs can survive in the stomach and then might be internalized into gut epithelial cells. In this study, we examined breast milk EV-miRNAs to determine the impact of the mother’s pre-pregnancy BMI on the EV-miRNA profiles. Methods: We recruited healthy nursing mothers who delivered a singleton baby in the past 6 months. Mothers pumped breast milk after 1-hour fasting and froze them. EVs were isolated from breast milk with ultracentrifugation. EV characteristics were examined by flow cytometry using 39 different EV markers. A total of 800 miRNAs were screened using a human miRNA panel (NanoString diagnostics, WA). Differential expression analysis was performed using DESEQ2. Results: We included 27 nursing mothers; the mean age was 31.1±7.6 years old, and 92% were Caucasian. All mothers breastfed their children and 22% of mothers supplemented with formula. Mean pre-pregnancy BMI was 21.6±2.0 kg/m 2 in the lean group (BMI<25kg/m 2 , n=17), and 29.1±4.5 kg/m 2 in the overweight/obese group (BMI≥25kg/m 2 , n=10). Besides BMI, all other demographics were not significantly different between groups. Breast milk EVs expressed not only general EV markers including CD9, CD63, and CD81, but also CD14, CD24, CD133/1, HLA-DRDPDQ and CD326. The most highly expressed marker is CD326 showing more than 2 times higher intensity than the general EV markers. Marker profiles were not different between groups. After normalization with housekeeping genes and spike-in controls, we obtained 10 differentially expressed miRNAs in the overweight/obese group (p<0.05 cut-off). Among those, miR-490-5p, miR-139-3p, and miR-291a-3p were the top 3 deregulated miRNAs downregulated in the overweight/obese group with a fold change >1.5. Conclusion: We found (1) CD326 as a breast milk EV marker, and (2) deregulated miRNAs in the overweight/obese mothers’ breast milk. Our findings will provide the opportunity to improve the quality of breast milk which ultimately promotes the children’s growth and health.