Commercial Cow Milk Contains Physically Stable Extracellular Vesicles Expressing Immunoregulatory TGF-β

Abstract

ScopeExtracellular vesicles, including exosomes, have been identified in all biological fluids and rediscovered as an important part of the intercellular communication. Breast milk also contains extracellular vesicles and the proposed biological function is to enhance the antimicrobial defense in newborns. It is, however, unknown whether extracellular vesicles are still present in commercial milk and, more importantly, whether they retained their bioactivity. Here, we characterize the extracellular vesicles present in semi-skimmed cow milk available for consumers and study their effect on T cells.Methods and ResultsExtracellular vesicles from commercial milk were isolated and characterized. Milk-derived extracellular vesicles contained several immunomodulating miRNAs and membrane protein CD63, characteristics of exosomes. In contrast to RAW 267.4 derived extracellular vesicles the milk-derived extracellular vesicles were extremely stable under degrading conditions, including low pH, boiling and freezing. Milk-derived extracellular vesicles were easily taken up by murine macrophages in vitro. Furthermore, we found that they can facilitate T cell differentiation towards the pathogenic Th17 lineage. Using a (CAGA)12-luc reporter assay we showed that these extracellular vesicles carried bioactive TGF-β, and that anti-TGF-β antibodies blocked Th17 differentiation.ConclusionOur findings show that commercial milk contains stable extracellular vesicles, including exosomes, and carry immunoregulatory cargo. These data suggest that the extracellular vesicles present in commercial cow milk remains intact in the gastrointestinal tract and exert an immunoregulatory effect.

Characterization of Membrane Integrity and Morphological Stability of Human Salivary Exosomes

“…More recently, Pieters et al reported that bovine milk-derived extracellular vesicles/exosomes were more stable after freeze-thaw cycles than those of cellular origin, from murine RAW 264.7 macrophages. 19) It is possible that exosomes derived from different sources vary in their stabilities to freezing and thawing. Further work to compare the stabilities of exosomes derived from various sources is needed.…”

“…Several studies described the stability of exosomes derived from urine, 16) plasma 17,18) and bovine milk. 19) These studies demonstrated high stability of proteins or microRNA in exosomes under a variety of storage conditions. However, the characteristic features of salivary exosomes are still not known.…”

Mesenchymal stem cells-derived exosomal microRNAs contribute to wound inflammation

“…Pieters et al demonstrated that extracellular vesicles, including exosomes, which were isolated from commercial bovine milk, could induce Th17 differentiation in murine splenocytes, and miR-21 was present in the immunoregulatory cargo carried by these extracellular vesicles (Pieters et al, 2015). In addition, data analyses of the human MSC secretome confirmed that miR-21 protects MSCs under serum-deprived conditions and can trigger key biological functions in recipient cells (Nie et al, 2011).…”

Functions of Exosomes and Microbial Extracellular Vesicles in Allergy and Contact and Delayed-Type Hypersensitivity

“…In addition, incubation of the milk vesicle preparations with peripheral blood mononuclear cells (PBMC) increased the number of suppressive Foxp3 pos Treg cells. In studies extending these findings, breast milk exosomes were shown to contain functional inhibitory TGF-beta [169], along with miRNA that can promote thymic Treg cell maturation [170], and that their immunosuppressive action likely inhibits innate immune functions of macrophages that may be acting as APC [171]. Further, mother’s milk exosomes contain miRNA-17, part of the miRNA-17–19 cluster, which is important for the activity of myeloid suppressor cells, as well as miRNA-181a, a modulator of TCR sensitivity to Ag.…”