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2022, Vol. 26 ›› Issue (19): 3036-3042Extraction, identification and proteomic analysis of exosomes derived from human umbilical cord mesenchymal stem cellsShan Zhengming, Tao Shuchun, Hu Chunmei, Zhang Zhiyuan, Ding Yinan, He Mengcheng, Tang Qiusha
Abstract: BACKGROUND: Current proteomics studies of exosomes only have proteomic comparisons between exosomes from different sources or exosomes secreted under different conditions. There is no comparative analysis between exosomes and their parent cells. OBJECTIVE: To extract, identify and analyze the exosomes of human umbilical cord mesenchymal stem cells. METHODS: Human umbilical cord mesenchymal stem cells were cultured and exosomes were extracted using sequential ultrafiltration and identified using transmission electron microscope, nanoparticle tracking analysis, bicinchoninic acid, western blot assay, and proteomics analysis. RESULTS AND CONCLUSION: (1) The extracted exosomes had good dispersion and uniformity. Most of them were cup-shaped or round-like membranous vesicles. The double membranous structure of the vesicles could be seen, with low electron density components in the center, which were more concentrated and distributed. The boundary was clear. (2) The peak size distribution of exosomes was (129.5±8.7) nm. The membrane surface was negatively charged; the concentration was 8.375×1010 particles/mL; and the average zeta potential was (-28.1±3.6) mV. (3) Exosomes showed the expression of characteristic membrane proteins CD9 and CD63. (4) Proteomics analysis showed that most of the highly expressed proteins in exosomes were involved in biological processes, such as RNA splicing, mRNA processing, and protein folding, involved in the synthesis, processing, and degradation of RNA/DNA and other geneticmaterial and proteins, and participated in the composition of organelles and subcellular membrane structures, involved in multiple signal pathways, cell adhesion, extracellular matrix receptor interactions, and were also closely related to a variety of diseases and viral carcinogenesis. (5) It is concluded that after proteomics analysis, it is believed that the exosomes have certain homogeneity and differences compared with their parent cells. Differential protein function has nothing to do with immunity. Therefore, the low immunogenicity and safety of exosomes derived from human umbilical cord mesenchymal stem cells are verified. Key words:stem cells, human umbilical cord mesenchymal stem cells, exosomes, proteomics, immunogenicity, safety |