Abstract Text: Breastfeeding provides important benefits to the neonate, including protection against infections and support in immune development. Breastmilk contains a range of immunoactive components, but how and where these contribute to immunity remains poorly understood. Here, we characterized breastmilk T cells using single cell RNA sequencing and flow cytometry. Breastmilk CD4+ and CD8+ T cells exhibited an effector/memory profile, with immune response signaling, proliferation and a Th1/cytotoxic profile with high cytokine production capacities. Activation and regulation appeared well balanced in breastmilk, with a prominent Treg population and co-expression of proliferation and immune regulatory markers in conventional memory T cells. Tissue-related gene expression and surface expression of tissue-resident memory T cell (T-RM) markers indicated that breastmilk T cells represent tissue-adapted rather than circulatory T cells. We showed that an activated effector profile was enriched in the breastmilk CD8+ T-RM population. We hypothesize that these cells played a role in local breast tissue defense against damage and infections introduced by the suckling infant. On the other hand, high expression of a diverse range of homing receptors leaves room for a role of breastmilk T cells in neonatal immune defense, as is supported by previous animal research. We observed a broad homing profile, with (co-)expression of skin-, gut- and inflammatory homing receptors and an enrichment of proliferation, activation markers and cytokine production in these homing cells. Together, our data suggest that breastmilk contains an adapted T cell population with an activated effector profile and potential functionality in maternal and/or neonatal tissues.
