Matrix-Bound Nanovesicles: The Effects of Isolation Method upon Yield, Purity, and Function

Quijano, Lina M.; Naranjo, Juan D.; El-Mossier, Salma O.; Turner, Neill J.; Pineda Molina, Catalina; Bartolacci, Joseph; Zhang, Li; White, Lisa; Li, Hui; Badylak, Stephen F.

Abstract

Identification of matrix-bound nanovesicles (MBV) as ubiquitous components of the extracellular matrix (ECM) raises questions regarding their biologic functions and their potential theranostic application. Unlike liquid-phase extracellular vesicles (e.g., exosomes), MBV are tightly bound to the ECM, which makes their isolation and harvesting more challenging. The indiscriminate use of different methods to harvest MBV can alter or disrupt their structural and/or functional integrity. The objective of the present study was to compare the effect of various MBV harvesting methods upon yield, purity, and biologic activity. Combinations of four methods to solubilize the ECM (collagenase [COL], liberase [LIB], or proteinase K [PK] and nonenzymatic elution with potassium chloride) and four isolation methods (ultracentrifugation, ultrafiltration [UF], density barrier, and size exclusion chromatography [SEC]) were used to isolate MBV from urinary bladder-derived ECM. All combinations of solubilization and isolation methods allowed for the harvesting of MBV, however, distinct differences were noted. The highest yield, purity, cellular uptake, and biologic activity were seen with MBV isolated by a combination of liberase or collagenase followed by SEC. The combination of proteinase K and UF was shown to have detrimental effects on bioactivity. The results show the importance of selecting appropriate MBV harvesting methods for the characterization and evaluation of MBV and for analysis of their potential theranostic application.

Citation

Quijano, L. M., Naranjo, J. D., El-Mossier, S. O., Turner, N. J., Pineda Molina, C., Bartolacci, J., …Badylak, S. F. (2020). Matrix-Bound Nanovesicles: The Effects of Isolation Method upon Yield, Purity, and Function. Tissue Engineering Part C Methods, 26(10), 528-540. https://doi.org/10.1089/ten.tec.2020.0243

• Journal Article Type: Article
• Acceptance Date: Sep 28, 2020
• Online Publication Date: Oct 3, 2020
• Publication Date: Oct 19, 2020
• Deposit Date: Nov 24, 2020
• Publicly Available Date: Oct 4, 2021
• Journal: Tissue Engineering Part C: Methods
• Print ISSN: 1937-3384
• Electronic ISSN: 1937-3392
• Peer Reviewed: Peer Reviewed
• Volume: 26
• Issue: 10
• Pages: 528-540
• DOI: https://doi.org/10.1089/ten.tec.2020.0243
• Keywords: Medicine (miscellaneous); Bioengineering; Biomedical Engineering
• Public URL: https://nottingham-repository.worktribe.com/output/5068856
• Publisher URL: https://www.liebertpub.com/doi/pdf/10.1089/ten.tec.2020.0243
• Additional Information: Final publication is available from Mary Ann Liebert, Inc., publishers http://dx.doi.org/10.1089/ten.tec.2020.0243