Characterization of Human Breast Milk Leukocytes and Their Potential Role in Cytomegalovirus Transmission to Newborns

Background: Breast milk is the primary source of cytomegalovirus (CMV) transmission to newborns and premature infants. The role of cell-free milk whey in virus transmission is well understood, yet the knowledge about the role of milk cells in this process is scarce. Objective: To preliminarily characterize different breast milk cell types during various stages of lactation to evaluate their potential role in the transmission of CMV. Materials and Methods: Breast milk cells of 18 lactating and 3 CMV-seropositive mothers of preterm infants were isolated and characterized for expression of myeloid markers by flow cytometry. In parallel, cytospin preparations were stained with α-naphthyl acetate esterase to identify milk macrophages and describe the dynamic changes of the macrophage-granulocyte population during lactation. The influence of different time points of lactation was analyzed by FACS analysis of double-stained (CD15/CD66b) milk cells. To characterize CMV target cells in breast milk, we enriched CD14+ cells by MACS (Miltenyi) and monitored cell fractions using CMV IEEx4 nested PCR and pp67 CMV RNA by NASBA. Results: Virolactia, viral DNAlactia, and viral pp67 late mRNA could be detected in breast milk cells only in defined time periods. Granulocytes and macrophages demonstrated an inverse dynamic with neutrophils predominating in the early stages (60 days postpartum) of lactation. Enrichment of CD14-positive cells resulted in viral DNA and pp67 late mRNA detection. Conclusions: Granulocytes and monocytes/macrophages are the predominating cell populations in breast milk with changing frequencies during early lactation. These results demonstrate that CD14-positive breast milk cells seem to be one of the target cells for CMV in breast milk.