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Enrichment, Characterization, and Proteomic Profiling of Small Extracellular Vesicles Derived from Human Limbal Mesenchymal Stromal Cells and Melanocytes

Authors :
Sebastian Kistenmacher
Melanie Schwämmle
Gottfried Martin
Eva Ulrich
Stefan Tholen
Oliver Schilling
Andreas Gießl
Ursula Schlötzer-Schrehardt
Felicitas Bucher
Günther Schlunck
Irina Nazarenko
Thomas Reinhard
Naresh Polisetti
Source :
Cells, Vol 13, Iss 7, p 623 (2024)
Publication Year :
2024
Publisher :
MDPI AG, 2024.

Abstract

Limbal epithelial progenitor cells (LEPC) rely on their niche environment for proper functionality and self-renewal. While extracellular vesicles (EV), specifically small EVs (sEV), have been proposed to support LEPC homeostasis, data on sEV derived from limbal niche cells like limbal mesenchymal stromal cells (LMSC) remain limited, and there are no studies on sEVs from limbal melanocytes (LM). In this study, we isolated sEV from conditioned media of LMSC and LM using a combination of tangential flow filtration and size exclusion chromatography and characterized them by nanoparticle tracking analysis, transmission electron microscopy, Western blot, multiplex bead arrays, and quantitative mass spectrometry. The internalization of sEV by LEPC was studied using flow cytometry and confocal microscopy. The isolated sEVs exhibited typical EV characteristics, including cell-specific markers such as CD90 for LMSC-sEV and Melan-A for LM-sEV. Bioinformatics analysis of the proteomic data suggested a significant role of sEVs in extracellular matrix deposition, with LMSC-derived sEV containing proteins involved in collagen remodeling and cell matrix adhesion, whereas LM-sEV proteins were implicated in other cellular bioprocesses such as cellular pigmentation and development. Moreover, fluorescently labeled LMSC-sEV and LM-sEV were taken up by LEPC and localized to their perinuclear compartment. These findings provide valuable insights into the complex role of sEV from niche cells in regulating the human limbal stem cell niche.

Details

Language :
English
ISSN :
20734409
Volume :
13
Issue :
7
Database :
Directory of Open Access Journals
Journal :
Cells
Publication Type :
Academic Journal
Accession number :
edsdoj.3bcdc523f747d3bd412ebb516efc7d
Document Type :
article
Full Text :
https://doi.org/10.3390/cells13070623