Transport of MicroRNA-Containing, Milk-Borne Extracellular Vesicles by Human Colon Carcinoma Caco-2 Cells

We have reported that microRNAs (miRNAs, miRs) in bovine milk regulate human genes. In milk, miRNAs are encapsulated in extracellular vesicles (EVs), thereby conferring protection against degradation and a pathway for intestinal uptake of miRNAs. We hypothesized that the uptake of bovine EVs in human intestinal colon carcinoma Caco-2 cells is mediated by endocytosis. Transport studies were carried out using fluorophore-labeled EVs purified from bovine milk. The transport of bovine EVs exhibited saturation kinetics at 37°C (Km= 55.5±48.6 µg/200 µL, Vmax= 0.083±0.057 ng exosomal protein ⋅ 81,750 cells−1 ⋅ hr−1) and decreased the intestinal transport of miR-29b by 80% if transport was measured at 4°C, consistent with carrier-mediated transport. If EVs or cells were treated with proteinase K, the intestinal transport of miR-29b decreased by 80%, suggesting that proteins on membrane surfaces are essential for EV uptake. The ratio of miR-200c to miR-29b decreased 20-fold during cellular transit, which suggests ...