Human vascular endothelial cells transport foreign exosomes from cow's milk by endocytosis

Abstract

Encapsulation of microRNAs in exosomes confers protection against degradation and a vehicle for shuttling of microRNAs between cells and tissues, and cellular uptake by endocytosis. Exosomes can be found in foods including milk. Humans absorb cow's milk exosomes and deliver the microRNA cargo to peripheral tissues, consistent with gene regulation by dietary nucleic acids across species boundaries. Here, we tested the hypothesis that human vascular endothelial cells transport milk exosomes by endocytosis, constituting a step crucial for the delivery of dietary exosomes and their cargo to peripheral tissues. We tested this hypothesis by using human umbilical vein endothelial cells and fluorophore-labeled exosomes isolated from cow's milk. Exosome uptake followed Michaelis-Menten kinetics (Vmax = 0.057 ± 0.004 ng exosome protein × 40,000 cells/h; Km = 17.97 ± 3.84 μg exosomal protein/200 μl media) and decreased by 80% when the incubation temperature was lowered from 37°C to 4°C. When exosome surface proteins were removed by treatment with proteinase K, or transport was measured in the presence of the carbohydrate competitor d-galactose or measured in the presence of excess unlabeled exosomes, transport rates decreased by 45% to 80% compared with controls. Treatment with an inhibitor of endocytosis, cytochalasin D, caused a 50% decrease in transport. When fluorophore-labeled exosomes were administered retro-orbitally, exosomes accumulated in liver, spleen, and lungs in mice. We conclude that human vascular endothelial cells transport bovine exosomes by endocytosis and propose that this is an important step in the delivery of dietary exosomes and their cargo to peripheral tissues.

Extracellular Vesicles from a Helminth Parasite Suppress Macrophage Activation and Constitute an Effective Vaccine for Protective Immunity

“…H. polygyrus EV uptake over time was also verified in BMDMs by confocal microscopy (Figure 1B; Figure S1C). To test whether uptake was occurring by an active process, we co-treated BMDMs with EVs and cytochalasin D, a potent inhibitor of actin polymerization that blocks endocytosis and phagocytosis and prevented EV uptake in other studies (Escrevente et al., 2011, Kusuma et al., 2016). Cytochalasin D treatment blocked EV uptake at 1 and 24 hr post-incubation, with 1% and 8% of cells being PKH67+, respectively (Figures 1C and 1D).

Figure 1 H. polygyrus EVs Are Efficiently Taken up by Bone Marrow-Derived Macrophages in a Cytochalasin D-Sensitive Manner(A) Percentage uptake of 2.5 μg PKH67-labeled EVs incubated with 2.5 × 10 5 BMDMs, RAW264.7, or MODE-K cells from 1 to 24 hr, as assessed by flow cytometry using the gating strategy in Figure S1A.

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Exosomes derived from platelet-rich plasma promote the re-epithelization of chronic cutaneous wounds via activation of YAP in a diabetic rat model

“…Extracellular vesicles derived from LX2, a human liver stellate cell line, have been used in a rat model 56, while therapeutic effects of human MSC-derived extracellular vesicles have been verified using a mouse 57 and a rat model 58. Humans can absorb cow's milk exosomes and transport them to peripheral tissues across species boundaries 59. The immune evasion mechanisms of exosomes remain elusive and will require further study.…”

Roles of MicroRNA across Prenatal and Postnatal Periods

“…Exogenous food-derived miRNAs appear to be stable in the oral cavity and in the GI tract and are transferred to the blood of adults, thought to influence gene expression in different tissues [81,82,83]. Recent evidence noted that bovine milk exosomes are taken up by human intestinal cells and vascular endothelial cells via endocytosis [84]. However, more studies are required to demonstrate that milk miRNAs, once ingested, can cross the GI tract of the infant [83,85].…”