Comprehensive analysis and comparison of proteins in salivary exosomes of climacteric and adolescent females.

Currently, it is difficult to extract exosomes with stable physicochemical properties from saliva. Furthermore, due to inadequate availability of basic data, the application of salivary exosomes as a diagnostic material is limited. In this study, we aimed to investigate an easier method for extraction of exosomes from whole saliva and compared proteins in salivary exosomes derived from subjects of two age groups. Salivary exosomes were extracted from nine females (56.7 ± 1.17 years old; climacteric or 19.9 ± 0.20 years old; adolescent) using commercial reagents and kits and detected using western blotting with anti-exosome marker antibodies. Exosome particle size and exosome-containing proteins were identified using NanoSight® and liquid chromatography with tandem mass spectrometry, respectively. In addition, an efficient method of exosome extraction from saliva using a reagent and without the use of an ultracentrifuge was shown. Our results showed a higher total protein content and larger particle size in climacteric exosomes than in adolescent exosomes. However, adolescent exosomes showed a larger variety of proteins (780 proteins) than the climacteric exosomes (573 proteins). Altogether, 893 proteins were identified in the salivary exosomes. Although viral process-, ribosome- and structural molecule-related proteins were higher in the adolescent exosomes, the levels of major salivary proteins such as immunoglobulins and amylase, were higher in the climacteric exosomes than in the adolescent exosomes. The data presented, which show the fundamental protein composition of salivary exosomes and the changes that occur with age, are beneficial in both diagnostic and biotechnological applications.