Bovine mammary alveolar MAC-T cells afford a tool for studies of bovine milk exosomes in drug delivery

Abstract

Bovine milk exosomes (BMEs) have attracted attention as vehicles for delivering RNA therapeutics. BMEs originate in mammary alveolar cells. Here, we determined whether bovine mammary alveolar MAC-T cells afford a tool to assess RNA delivery by BMEs. MAC-T cells exosomes (MAC-T BMEs) and BMEs were harvested by differential ultracentrifugation. Exosome size, morphology, microRNA content and marker proteins were assessed using nanoparticle tracking analysis, transmission electron microscopy, real-time PCR and immunoblot analysis, respectively. MAC-T cells were genetically engineered to secrete MAC-T BMEs endogenously labeled with a near-infrared fluorescent protein and tissue distribution was compared to fluorophore-labeled BMEs following intravenous injection in C57BL/6 mice. Morphology and size were similar in MAC-T BMEs and BMEs (94 ± 5.8 nm and 101 ± 4.2 nm, p > 0.05). Both preparations expressed miR-320a, miR-200c and let-7a-5p (positive controls) but not miR-1 (negative control). Exosome marker proteins, CD9, CD63, CD81 and Tsg101, were detected in both MAC-T BMEs and BMEs. Distribution in mouse tissues was similar for both preparations, with liver being the primary accumulation site. Collectively, MAC-T BMEs afford a tool for BMEs-based RNA delivery studies.

Keywords: Bovine mammary alveolar MAC-T cells; Bovine milk exosomes; CD81 fusion protein; Drug delivery; RNA; Transgenic.