Abstract
Extracellular vesicles (EVs) are membrane surrounded structures released by cells, which have been increasingly recognized as mediators of intercellular communication. Recent reports indicate that EVs participate in important biological processes and could serve as potential source for cancer biomarkers. As an attractive EVs source with merit of non-invasiveness, human saliva is a unique medium for clinical diagnostics. Thus, we proposed a facile approach to prepare salivary extracellular vesicles (SEVs). Affinity chromatography column combined with filter system (ACCF) was developed to efficiently remove the high abundant proteins and viscous interferences of saliva. Protein profiling in the SEVs obtained by this strategy was compared with conventional centrifugation method, which demonstrated that about 70% more SEVs proteins could be revealed. To explore its utility for cancer proteomics, we analyzed the proteome of SEVs in lung cancer patients and normal controls. Shotgun proteomic analysis illustrated that 113 and 95 proteins have been identified in cancer group and control group, respectively. Among those 63 proteins that have been consistently discovered only in cancer group, 12 proteins are lung cancer related. Our results demonstrated that SEVs prepared through the developed strategy are valuable samples for proteomics and could serve as a promising liquid biopsy for cancer.
Highlights
Extracellular vesicles (EVs) are defined as intact, submicron, phospholipid-rich vesicles ranging from 100 nm to 1000 nm in diameters, which shed from the surface of cells[1]
Our human saliva contains high abundant proteins along with many viscous proteins, which interfere in the preparation of salivary extracellular vesicles (SEVs)
To prepare quality SEVs for clinical applications and minimize the interference of salivary proteins, we developed an affinity chromatography coupled with filter system aiming at high quality SEVs separation, which can be further used for cancer proteomics
Summary
Extracellular vesicles (EVs) are defined as intact, submicron, phospholipid-rich vesicles ranging from 100 nm to 1000 nm in diameters, which shed from the surface of cells[1]. Recent research has revealed that these vesicles act as important messengers for intercellular communication[4] They could putatively attach or fuse with the target cell membrane, delivering surface proteins and perhaps cytoplasm to the recipient cell[5,6]. It is of great interest to explore the proteome of EVs that originate from human body fluids, which might carry important biomarkers for the early detection of cancers[11]. Removal of amylase and other viscous proteins from saliva before SEVs’ extraction could benefit downstream proteomic analysis of SEVs and contribute to biomarker discovery for cancer[26]. The developed approach was further applied to harvest SEVs from healthy subjects and lung cancer patients, respectively The proteome of both groups were compared through shotgun proteomics and further used for candidate biomarker discovery for the detection of lung cancer
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