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Metabolic labelling of a subpopulation of small extracellular vesicles using a fluorescent palmitic acid analogue.

Authors :
Barreca V
Boussadia Z
Polignano D
Galli L
Tirelli V
Sanchez M
Falchi M
Bertuccini L
Iosi F
Tatti M
Sargiacomo M
Fiani ML
Source :
Journal of extracellular vesicles [J Extracell Vesicles] 2023 Dec; Vol. 12 (12), pp. e12392.
Publication Year :
2023

Abstract

Exosomes are among the most puzzling vehicles of intercellular communication, but several crucial aspects of their biogenesis remain elusive, primarily due to the difficulty in purifying vesicles with similar sizes and densities. Here we report an effective methodology for labelling small extracellular vesicles (sEV) using Bodipy FL C16, a fluorescent palmitic acid analogue. In this study, we present compelling evidence that the fluorescent sEV population derived from Bodipy C16-labelled cells represents a discrete subpopulation of small exosomes following an intracellular pathway. Rapid cellular uptake and metabolism of Bodipy C16 resulted in the incorporation of fluorescent phospholipids into intracellular organelles specifically excluding the plasma membrane and ultimately becoming part of the exosomal membrane. Importantly, our fluorescence labelling method facilitated accurate quantification and characterization of exosomes, overcoming the limitations of nonspecific dye incorporation into heterogeneous vesicle populations. The characterization of Bodipy-labelled exosomes reveals their enrichment in tetraspanin markers, particularly CD63 and CD81, and in minor proportion CD9. Moreover, we employed nanoFACS sorting and electron microscopy to confirm the exosomal nature of Bodipy-labelled vesicles. This innovative metabolic labelling approach, based on the fate of a fatty acid, offers new avenues for investigating exosome biogenesis and functional properties in various physiological and pathological contexts.<br /> (© 2023 The Authors. Journal of Extracellular Vesicles published by Wiley Periodicals LLC on behalf of International Society for Extracellular Vesicles.)

Details

Language :
English
ISSN :
2001-3078
Volume :
12
Issue :
12
Database :
MEDLINE
Journal :
Journal of extracellular vesicles
Publication Type :
Academic Journal
Accession number :
38072803
Full Text :
https://doi.org/10.1002/jev2.12392