Exosome Derived From Gastric Cancer Cells Promotes Vascular Endothelial Cells Activition

Background: Exosomes are a class of extracellular vesicles(EVs)that have received much attention over the past decade.They range in size from 40 to 100 nm in diameter and transport from the inside of cells to the extracellular environment.They play an important role in intercellular communication and mediate diverse biological functions,including angiogenesis,cell proliferation,tumour cell invasion and metastasis,immune response,and antigen presentation,by transferring of proteins,mRNAs and non-coding RNAs to neighbouring or distant cells.Methods: Exosomes from cell culture-conditioned medium were isolated and purified using ultracentrifugation or commercial reagents.The character of exosomes was identified by transmission electron microscopy(TEM)and Western blot.The profile of small RNA from exosomes was characterized by next-generation RNA sequencing.Then the RNA sequencing data were analysed by bioinformatics.In addition,HUVEC were stimulated by cancer exosomes.The cell proliferation assay,cell wound healing assay,transwell migration assay,Matrigel cell tubule formation assay,tumor vasculogenic mimicry formation were carried respectively.HUVECs and cancer cells were cocultured in the Transwell system,to observe the migratory activity of HUVECs after cancer cells secreting exosomes.Results were expressed as mean ± standard deviation(mean ± SD).Results: The exosomes had a characteristic saucer-like shape limited by a lipid bilayer,and their diameters ranged from 40 to 100 nm under TEM.Two of the exosomal membrane proteins,CD63 and CD9,were confirmed by Western blot analysis.Small RNA of exosomes was confirmed using deep sequencing technology.After the bioinformatics analysis,we got the characteristics of small RNA in exosomes.According to the miRNA profiles of exosomes,hsa-miR-21-5p was most abundant.And there is a significant expressional difference between the exosomes from gastric cancer cells and the exosomes from normal cells,as well as the exosomes from cancer-associated fibroblast and the exosomes from normal fibroblast.We also found the exosomes from gastric cancer cells can promote the biology function of HUVEs,such as proliferation,migration,tube formation.In addition,they also promote vasculogenic mimicry formation.When exosomes were inhibited,the functional activity of HUVEC was suppressed.While,when exosomes were added again,the functional activity of HUVEC was got recovery.Conclusion: Our present study provides a profile of small non-coding RNA in exosomes,including miRNA distribution analysis and differential expression.The characteristics of exosomal RNA composition will help us better understanding of exosome-mediated molecular mechanisms and will contribute to the discovery of potential gastric cancer biomarkers.What’s more,the exosomes from gastric cancer cells can promote functional activity of vascular endothelial cells,which is closely related to tumor angiogenesis.