Serum exosomal hsa_circ_0069313 has a potential to diagnose more aggressive non-small cell lung cancer
Understanding the Role and Clinical Applications of Exosomes in Gynecologic Malignancies: A Review of the Current Literature
Proteomic analysis of serum and serum exosomes, and their application in intrahepatic cholangiocarcinoma
Exosomes are extracellular vesicles with a diameter in the range of 50-200 nm and a double-layer lipid membrane structure that are released by various types of cells under normal or abnormal physiological conditions. At present, according to their extensive biological functions, exosomes have been used in a wide range of research fields and applications, including as potential biomarkers and drug delivery vehicles. Intrahepatic cholangiocarcinoma is a malignant tumor of the biliary epithelium with the characteristics of cholangiocellular differentiation, which accounts for 10%-15% of all types of primary liver cancer. Intrahepatic cholangiocarcinoma has no obvious clinical symptoms in the early stages, which results in a low survival rate. Imaging equipment dependent diagnostic methods and currently commonly used diagnostic markers with low sensitivity/specificity have necessitated the development of new specific markers for intrahepatic cholangiocarcinoma.
In this study, exosomes were isolated from serum using a commercial kit and characterized through nanoparticle tracking analysis, Western blotting analysis, and transmission electron microscopic analysis to prove the successful isolation of exosomes. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) showed that the protein profiles of the serum and serum exosome samples were significantly different. In particular, some high-abundance proteins in the serum samples were significantly reduced or disappeared in the serum exosome sample. Meanwhile, some protein bands (which may belong to exosomes) that did not appear in the serum samples appeared in the serum exosome samples, leading to the conduciveness of subsequent mass spectrometry analysis.
The serum and serum exosome samples of the healthy control and intrahepatic cholangiocarcinoma groups were analyzed by liquid chromatography-mass spectrometry for label-free quantitative proteomics. In total, 547 proteins were identified in the serum exosome samples, of which 341 (more than 60%) could be found in the exosomal protein database. In addition, 271 and 430 credible proteins were screened from the serum and serum exosome samples for multi-dimensional statistical analysis and differential protein discovery. Unsupervised principal component analysis and supervised orthogonal partial least squares discriminant analysis based on the quantitative proteome of the serum and serum exosome samples could distinguish the healthy control and intrahepatic cholangiocarcinoma groups well, which illustrates that the two types of samples both have potential in the diagnosis of intrahepatic cholangiocarcinoma.
There were 15 upregulated and 8 downregulated proteins screened in the intrahepatic cholangiocarcinoma group compared to the healthy control group based on the serum samples, while 33 upregulated and 18 downregulated proteins were screened in the intrahepatic cholangiocarcinoma group compared to the healthy control group based on the serum exosome samples, and only four of the differential proteins screened based on the two types of samples were duplicates. At the same time, 35 of the 51 differentially expressed proteins screened based on serum exosome samples belonged to the exosomal protein database.
Finally, biological information analysis was performed according to these differential proteins. The molecular functions, biological processes, and signal pathways enriched by these differential proteins mainly involved the innate immune responses, inflammatory responses, and blood coagulation. This study provides a reference value for potential biomarker discovery and exploration of the process of occurrence, development, and metastasis of intrahepatic cholangiocarcinoma. Moreover, compared with proteomic analysis based on serum samples, proteomic analysis based on serum exosome samples can be used to identify more differential proteins and biological information, and although these differential proteins and biological information may show big differences, the specificity and sensitivity of exosome-based diagnosis and the superiority of exosomes as samples for proteomic analysis have proven the application value of exosomes.
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