Characterization and proteomic profiling of pancreatic cancer‐derived serum exosomes

Abstract

Pancreatic cancer (PC) is one of the most lethal cancers known worldwide, and its prognosis is poor in most patients. Exosomes are nanosized extracellular vesicles, which are released from various cell types. They are involved in cellular communication. The diagnosis and treatment of PC were improved substantially with exosomes. In this study, we isolated PC-derived exosomes and investigated their proteomic profile. Then, we conducted bioinformatic analysis on proteomic data. Differential ultracentrifugation was performed to isolate exosomes from human serum samples and four PC cell lines. Transmission electron microscopy and Western blot analysis were used to characterize the isolated exosomes. Liquid chromatography coupled with tandem mass spectrometry was conducted to identify the proteome of serum exosomes. Proteomic analysis demonstrated that all the serum exosomes were derived from three cohorts of human subjects; these serum exosomes contained a total of 655 proteins, out of which 315 proteins overlapped with ExoCarta database. Gene oncology and kyoto encyclopedia of genes and genomes analyses provided the functional annotation of the proteome. Interestingly, 18 or 14 proteins were upregulated and 11 or 14 proteins were downregulated in serum exosomes derived from patients with PC as compared with in serum exosomes derived from healthy volunteers or from pancreatitis patients respectively. Annexin A11, a calcium-dependent phospholipid-binding protein, was expressed in a PC cell line (CFPAC-1)-derived exosomes and in tumor tissues of patients with PC, respectively. Our data provided a basic foundation for further studies on the protein composition of PC-derived exosomes and its involvement in PC biology.

Plasma-Derived Extracellular Vesicles Convey Protein Signatures That Reflect Pathophysiology in Lung and Pancreatic Adenocarcinomas

“…Notably, serum-derived EV-associated LBP (Lipopolysaccharide-binding protein) has previously been shown to be elevated in PDAC cases as compared to healthy controls. Consistent with this study, we also observed that LBP is elevated in EVs of PDAC cases as compared to controls, thus providing independent validation [29].…”

“…We note that a novel component of this study in relation to other studies, which have similarly evaluated utility of EV-associated protein cargo for early detection or prognostication [29,30], is the direct comparison protein features in plasma-derived EVs versus unfractionated plasma from the same individual for utility in distinguishing case from control. This approach enables us to address a fundamental question and conclude that there are indeed EV-associated features that exhibited improved classifier performance specifically in the plasma sEV compartment.…”

Investigating the Immune Function and Proteomic Profiles of Plasmal Exosomes in Lactobacillus Plantarum-treated Immunosuppressive Broilers

“…We observed that the ultrastructure and size of plasma exosomes complied with the typical morphology of exosomes [31,32]. The surface markers of exosomes mainly included CD9, CD63, CD81, CD82, HSP27, HSP90, TSG101 and ALIX [33]. Here, the presence of exosomes was con rmed with the detection of CD63 and TSG101, and the purity of exosomes was con rmed by the absence of Calnexin.…”

Proteomics of Extracellular Vesicles: Update on Their Composition, Biological Roles and Potential Use as Diagnostic Tools in Atherosclerotic Cardiovascular Diseases

“…At this point, a supernatant filtration is sometimes carried out to remove particles larger than 200 nm. Finally, the recovery of exosomes is performed by ultracentrifugation at 100,000–200,000× g for hours [ 80 , 81 , 82 ] and the pellet is washed by resuspension with PBS and centrifuged again to remove contaminants and improve the purity.…”