Effects of Co-Culture EBV-miR-BART1-3p on Proliferation and Invasion of Gastric Cancer Cells Based on Exosomes
Abstract
:Simple Summary
Abstract
1. Introduction
2. Materials and Methods
2.1. Cell Culture and Reagents
2.2. EBER In Situ Hybridization
2.3. Preparation of Exosome-Free Fetal Bovine Serum
2.4. Isolation of Exosomes
2.5. Exosome Morphology Observed by Transmission Electron Microscopy
2.6. Exosome Size and Concentration by Nanoparticle Tracking Analysis (NTA)
2.7. Extraction of Cell Protein
2.8. Extraction of Exosomal Protein
2.9. Determination of Cellular and Exosomal Protein Concentrations
2.10. Expression Analysis of Exosomal Protein
2.11. Extraction of Ribonucleic Acid
2.12. Library Construction and Small RNA Sequencing
2.13. Expression Profile Analysis of EBV-miRNA
2.14. Construction of SNU-719-Inhibitor Cell Line with Low Expression of EBV-miR-BART1-3p
2.15. Verification of the Expression Level of miR-BART1-3p in Cell Lines and the Expression of Target Genes in AGS Cells after Co-Culture of Exosomes for 24 h
2.16. CCK8 Cell Proliferation Experiment
2.17. Cell Scratch Experiment
2.18. Transwell Migration Assay
2.19. Transwell Invasion
2.20. Absorption of Exosomes by Living Cells Observed by Laser Confocal Microscopy
2.21. Expression of miR-BART1-3p after Co-Culture with GC Cells in Exosomes with High Expression or Silence of miR-BART1-3p and Its Effect on Proliferation and Invasion
2.22. Target Gene Analysis of miR-BART1-3p
2.23. Statistical Analysis
3. Results
3.1. Characterization of miRNA in Exosomes of EBV-Positive GC Cells
3.1.1. Morphology of AGS and SNU-719 GC Cells
3.1.2. Structure of Exosomes Secreted by SNU-719 and AGS Cells
3.1.3. Size of Exosomes Secreted by SNU-719 and AGS Cells
3.1.4. Expression Analysis of Exosomal Proteins Secreted by SNU-719 and AGS Cells
3.1.5. Distribution of Small RNAs in Cells and Exosomes
Annotation of ncRNAs in SNU-719 and AGS Cells and Exosomes
Small RNA Sequence Length Distribution in SNU-719 and AGS Cells and Exosomes
sRNA Analysis of Genomic Repeat Regions in SNU-719 and AGS Cells and Exosomes
Analysis of Functional Elements in SNU-719 and AGS Cells and Exosomes
Common and Unique Sequence Analysis of SNU-719 and AGS Cells and Exosomes
Correlation Analysis between SNU-719 and AGS Cells and Exosomes
Differential Expression of miRNA in SNU-719 and AGS Cells and Exosomes
Expression Analysis of the Top 15 miRNAs in Cells and Exosomes
Differential Expression Analysis of EBV-Related miRNAs in SNU-719 Cells and Their Exosomes
3.2. Effects of High and Low Expression of EBV-miR-BART1-3p on Proliferation and Invasion Ability of GC Cells
3.3. Effects of Co-Culture with Exosomes with High and Low Expression of EBV-miR-BART1-3p on Proliferation and Invasion of GC Cells
3.4. Changes of Target Genes in Gastric Cancer Cells after Co-Culture with High and Low Expression of EBV-miR-BART1-3p in Exosomes
4. Discussion
5. Conclusions
Supplementary Materials
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Conflicts of Interest
References
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Lin, M.; Hu, S.; Zhang, T.; Li, J.; Gao, F.; Zhang, Z.; Zheng, K.; Li, G.; Ren, C.; Chen, X.; et al. Effects of Co-Culture EBV-miR-BART1-3p on Proliferation and Invasion of Gastric Cancer Cells Based on Exosomes. Cancers 2023, 15, 2841. https://doi.org/10.3390/cancers15102841
Lin M, Hu S, Zhang T, Li J, Gao F, Zhang Z, Zheng K, Li G, Ren C, Chen X, et al. Effects of Co-Culture EBV-miR-BART1-3p on Proliferation and Invasion of Gastric Cancer Cells Based on Exosomes. Cancers. 2023; 15(10):2841. https://doi.org/10.3390/cancers15102841
Chicago/Turabian StyleLin, Mengyao, Shun Hu, Tianyi Zhang, Jiezhen Li, Feng Gao, Zhenzhen Zhang, Ke Zheng, Guoping Li, Caihong Ren, Xiangna Chen, and et al. 2023. "Effects of Co-Culture EBV-miR-BART1-3p on Proliferation and Invasion of Gastric Cancer Cells Based on Exosomes" Cancers 15, no. 10: 2841. https://doi.org/10.3390/cancers15102841
APA StyleLin, M., Hu, S., Zhang, T., Li, J., Gao, F., Zhang, Z., Zheng, K., Li, G., Ren, C., Chen, X., Guo, F., & Zhang, S. (2023). Effects of Co-Culture EBV-miR-BART1-3p on Proliferation and Invasion of Gastric Cancer Cells Based on Exosomes. Cancers, 15(10), 2841. https://doi.org/10.3390/cancers15102841