RNA-containing Exosomes in Human Nasal Secretions

Abstract

Background Exosomes are nanovesicles of endocytic origin released by cells and present in human body fluids such as plasma, breast milk, and bronchoalveolar lavage fluid. These vesicles take part in communication between cells. Recently, it was shown that exosomes contain both mRNA and microRNA. This RNA can be shuttled between cells (exosomal shuttle RNA), which is a new route of communication between cells. The aim of this study was to determine whether nasal secretions harbor exosomes and furthermore, whether these exosomes contain RNA. Methods Human nasal lavage fluid (NLF) underwent centrifugation and filtration to discard cells and debris, followed by a final ultracentrifugation at 120,000 × g to pellet the exosomes. Exosomes were detected using electron microscopy (EM), flow cytometry, and Western blot. RNA was extracted and analyzed using a Bioanalyzer. Results Exosomes were visualized as 40–80 nm, CD63+ vesicles using EM. Flow cytometry of exosomes using anti–major histocompatibility complex class II beads revealed exosomes positive for the tetraspanins CD9, CD63, and CD81. Western blot confirmed the presence of exosomal protein and absence of proteins from the endoplasmic reticulum (ER), because the exosomes were positive for Tsg101, but negative for the ER marker, calnexin. Bioanalyzer analysis revealed that, these exosomes contain RNA. Conclusion This study shows for the first time that NLF contains exosomes and that these exosomes contain RNA. Further characterization of the exosomal RNA and proteins may provide important information about communication in the nose and potentially provide a source of biomarkers for upper airway diseases.