The Bioavailability and Distribution of Bovine Milk Exosomes is Distinct from that of their Cargos in Mice

Abstract

BackgroundAmong extracellular vesicles, exosomes are of particular interest because of their roles in cell‐to‐cell communication, which is facilitated by the delivery of cargos such as RNAs, to recipient cells. Dietary exosomes in bovine milk are transported by intestinal and vascular endothelial cells in human and rodent cell cultures, and delivered to peripheral tissues in mice. Little is known about the fate of the cargos in milk exosomes following absorption.HypothesisRNAs in bovine milk exosomes are delivered to distinct peripheral tissues in mice.MethodsExosomes were isolated from bovine milk using ultracentrifugation. Exosome membranes were labeled using the fluorophore, DiR, whereas RNA cargos were labeled using EXO‐Glow Red in separate experiments. Identity, integrity, fine dispersion, and count of exosomes were assessed using transmission electron microscopy, western blots, and nanoparticle tracker (not shown). Exosomes were administered orally by gavage (1×1012 exosomes/g) in BALB/c mice. Absorption and distribution of exosomes and their cargos was monitored at timed intervals for up to 24 h using an iBox® Small Animal Imaging system and LiCor Odyssey CLx. At timed intervals, mice were euthanized and tissues were collected for densitometry analysis using VisionWorks®LS and Image Studio software.ResultsAt t = 12 h after administration of DiR‐labeled exosomes by gavage, only a faint DiR signal was detected in peripheral tissues (Fig. 1); labeled RNA was detected in all peripheral tissues with a preference for kidneys, brain, lungs and liver. Figure 2 depicts the densitometry analysis of labeled RNA, corrected by the signal produced by unconjugated Exo‐glow. At t = 24 h, the majority of absorbed exosomes accumulated in liver (Fig. 3), whereas the RNA cargo accumulated preferably in brain and kidneys (Figs. 4,5).ConclusionsBovine milk exosomes and their RNA cargos are bioavailable and accumulate in distinct tissues in mice. Our data suggest that bovine milk exosomes are disassembled in the intestinal mucosa, and RNA cargos are repackaged in endogenous exosomes for transfer to tissues, preferably brain and kidneys.Future studiesWe will conduct a comprehensive analysis of milk whole exosomes and RNA cargo kinetics in mice to obtain an accurate assessment of bioavailability, distribution, and elimination phenomena in health and disease.Support or Funding InformationSupport: NIFA 2015‐67017‐23181, NIFA 2016‐67001‐25301/NIH DK107264, NIH 1P20GM104320, the Gerber Foundation, and USDA Hatch Act and W3002.