Efficient enzyme-free isolation of brain-derived extracellular vesicles

Extracellular vesicles (EVs) have gained significant attention as pathology mediators and potential diagnostic tools for neurodegenerative diseases. However, isolation of brain-derived EVs (BDEVs) from tissue remains challenging, often involving enzymatic digestion steps that may compromise the inte...

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Autores: Matamoros-Anglès, Andreu, Karadjuzovic, Emina, Mohammadi, Behnam, Song, Feizhi, Brenna, Santra, Meister, Susanne Caroline, Siebels, Bente, Voß, Hannah, Seuring, Carolin, Ferrer, Isidro (Ferrer Abizanda), Schlüter, Hartmut, Kneussel, Matthias, Altmeppen, Hermann Clemens, Schweizer, Michaela, Puig, Berta, Shafiq, Mohsin, Glatzel, Markus
Tipo de recurso: artículo
Estado:Versión publicada
Fecha de publicación:2024
País:España
Institución:Universidad de Barcelona
Repositorio:Dipòsit Digital de la UB
OAI Identifier:oai:diposit.ub.edu:2445/220022
Acceso en línea:https://hdl.handle.net/2445/220022
Access Level:acceso abierto
Palabra clave:Marcadors bioquímics
Metal·loproteïnases
Cervell
Biochemical markers
Metalloproteinases
Brain
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spelling Efficient enzyme-free isolation of brain-derived extracellular vesiclesMatamoros-Anglès, AndreuKaradjuzovic, EminaMohammadi, BehnamSong, FeizhiBrenna, SantraMeister, Susanne CarolineSiebels, BenteVoß, HannahSeuring, CarolinFerrer, Isidro (Ferrer Abizanda)Schlüter, HartmutKneussel, MatthiasAltmeppen, Hermann ClemensSchweizer, MichaelaPuig, BertaShafiq, MohsinGlatzel, MarkusMarcadors bioquímicsMetal·loproteïnasesCervellBiochemical markersMetalloproteinasesBrainExtracellular vesicles (EVs) have gained significant attention as pathology mediators and potential diagnostic tools for neurodegenerative diseases. However, isolation of brain-derived EVs (BDEVs) from tissue remains challenging, often involving enzymatic digestion steps that may compromise the integrity of EV proteins and overall functionality. Here, we describe that collagenase digestion, commonly used for BDEV isolation, produces undesired protein cleavage of EV-associated proteins in brain tissue homogenates and cell-derived EVs. In order to avoid this effect, we studied the possibility of isolating BDEVs with a reduced amount of collagenase or without any protease. Characterization of the isolated BDEVs from mouse and human samples (both female and male) revealed their characteristic morphology and size distribution with both approaches. However, we show that even minor enzymatic digestion induces 'artificial' proteolytic processing in key BDEV markers, such as Flotillin-1, CD81, and the cellular prion protein (PrPC), whereas avoiding enzymatic treatment completely preserves their integrity. We found no major differences in mRNA and protein content between non-enzymatically and enzymatically isolated BDEVs, suggesting that the same BDEV populations are purified with both approaches. Intriguingly, the lack of Golgi marker GM130 signal, often referred to as contamination indicator (or negative marker) in EV preparations, seems to result from enzymatic digestion rather than from its actual absence in BDEV samples. Overall, we show that non-enzymatic isolation of EVs from brain tissue is possible and avoids artificial pruning of proteins while achieving an overall high BDEV yield and purity. This protocol will help to understand the functions of BDEV and their associated proteins in a near-physiological setting, thus opening new research approaches.Taylor & Francis2024info:eu-repo/semantics/articleinfo:eu-repo/semantics/publishedVersionapplication/pdfhttps://hdl.handle.net/2445/220022Articles publicats en revistes (Patologia i Terapèutica Experimental)reponame:Dipòsit Digital de la UBinstname:Universidad de BarcelonaInglésReproducció del document publicat a: https://doi.org/10.1002/jev2.70011Journal of Extracellular Vesicles, 2024, vol. 13, num.11https://doi.org/10.1002/jev2.70011cc-by (c) Matamoros-Angles, A. et al., 2024http://creativecommons.org/licenses/by/4.0/info:eu-repo/semantics/openAccessoai:diposit.ub.edu:2445/2200222026-05-27T06:46:51Z
dc.title.none.fl_str_mv Efficient enzyme-free isolation of brain-derived extracellular vesicles
title Efficient enzyme-free isolation of brain-derived extracellular vesicles
spellingShingle Efficient enzyme-free isolation of brain-derived extracellular vesicles
Matamoros-Anglès, Andreu
Marcadors bioquímics
Metal·loproteïnases
Cervell
Biochemical markers
Metalloproteinases
Brain
title_short Efficient enzyme-free isolation of brain-derived extracellular vesicles
title_full Efficient enzyme-free isolation of brain-derived extracellular vesicles
title_fullStr Efficient enzyme-free isolation of brain-derived extracellular vesicles
title_full_unstemmed Efficient enzyme-free isolation of brain-derived extracellular vesicles
title_sort Efficient enzyme-free isolation of brain-derived extracellular vesicles
dc.creator.none.fl_str_mv Matamoros-Anglès, Andreu
Karadjuzovic, Emina
Mohammadi, Behnam
Song, Feizhi
Brenna, Santra
Meister, Susanne Caroline
Siebels, Bente
Voß, Hannah
Seuring, Carolin
Ferrer, Isidro (Ferrer Abizanda)
Schlüter, Hartmut
Kneussel, Matthias
Altmeppen, Hermann Clemens
Schweizer, Michaela
Puig, Berta
Shafiq, Mohsin
Glatzel, Markus
author Matamoros-Anglès, Andreu
author_facet Matamoros-Anglès, Andreu
Karadjuzovic, Emina
Mohammadi, Behnam
Song, Feizhi
Brenna, Santra
Meister, Susanne Caroline
Siebels, Bente
Voß, Hannah
Seuring, Carolin
Ferrer, Isidro (Ferrer Abizanda)
Schlüter, Hartmut
Kneussel, Matthias
Altmeppen, Hermann Clemens
Schweizer, Michaela
Puig, Berta
Shafiq, Mohsin
Glatzel, Markus
author_role author
author2 Karadjuzovic, Emina
Mohammadi, Behnam
Song, Feizhi
Brenna, Santra
Meister, Susanne Caroline
Siebels, Bente
Voß, Hannah
Seuring, Carolin
Ferrer, Isidro (Ferrer Abizanda)
Schlüter, Hartmut
Kneussel, Matthias
Altmeppen, Hermann Clemens
Schweizer, Michaela
Puig, Berta
Shafiq, Mohsin
Glatzel, Markus
author2_role author
author
author
author
author
author
author
author
author
author
author
author
author
author
author
author
dc.subject.none.fl_str_mv Marcadors bioquímics
Metal·loproteïnases
Cervell
Biochemical markers
Metalloproteinases
Brain
topic Marcadors bioquímics
Metal·loproteïnases
Cervell
Biochemical markers
Metalloproteinases
Brain
description Extracellular vesicles (EVs) have gained significant attention as pathology mediators and potential diagnostic tools for neurodegenerative diseases. However, isolation of brain-derived EVs (BDEVs) from tissue remains challenging, often involving enzymatic digestion steps that may compromise the integrity of EV proteins and overall functionality. Here, we describe that collagenase digestion, commonly used for BDEV isolation, produces undesired protein cleavage of EV-associated proteins in brain tissue homogenates and cell-derived EVs. In order to avoid this effect, we studied the possibility of isolating BDEVs with a reduced amount of collagenase or without any protease. Characterization of the isolated BDEVs from mouse and human samples (both female and male) revealed their characteristic morphology and size distribution with both approaches. However, we show that even minor enzymatic digestion induces 'artificial' proteolytic processing in key BDEV markers, such as Flotillin-1, CD81, and the cellular prion protein (PrPC), whereas avoiding enzymatic treatment completely preserves their integrity. We found no major differences in mRNA and protein content between non-enzymatically and enzymatically isolated BDEVs, suggesting that the same BDEV populations are purified with both approaches. Intriguingly, the lack of Golgi marker GM130 signal, often referred to as contamination indicator (or negative marker) in EV preparations, seems to result from enzymatic digestion rather than from its actual absence in BDEV samples. Overall, we show that non-enzymatic isolation of EVs from brain tissue is possible and avoids artificial pruning of proteins while achieving an overall high BDEV yield and purity. This protocol will help to understand the functions of BDEV and their associated proteins in a near-physiological setting, thus opening new research approaches.
publishDate 2024
dc.date.none.fl_str_mv 2024
dc.type.none.fl_str_mv info:eu-repo/semantics/article
info:eu-repo/semantics/publishedVersion
format article
status_str publishedVersion
dc.identifier.none.fl_str_mv https://hdl.handle.net/2445/220022
url https://hdl.handle.net/2445/220022
dc.language.none.fl_str_mv Inglés
language_invalid_str_mv Inglés
dc.relation.none.fl_str_mv Reproducció del document publicat a: https://doi.org/10.1002/jev2.70011
Journal of Extracellular Vesicles, 2024, vol. 13, num.11
https://doi.org/10.1002/jev2.70011
dc.rights.none.fl_str_mv cc-by (c) Matamoros-Angles, A. et al., 2024
http://creativecommons.org/licenses/by/4.0/
info:eu-repo/semantics/openAccess
rights_invalid_str_mv cc-by (c) Matamoros-Angles, A. et al., 2024
http://creativecommons.org/licenses/by/4.0/
eu_rights_str_mv openAccess
dc.format.none.fl_str_mv application/pdf
dc.publisher.none.fl_str_mv Taylor & Francis
publisher.none.fl_str_mv Taylor & Francis
dc.source.none.fl_str_mv Articles publicats en revistes (Patologia i Terapèutica Experimental)
reponame:Dipòsit Digital de la UB
instname:Universidad de Barcelona
instname_str Universidad de Barcelona
reponame_str Dipòsit Digital de la UB
collection Dipòsit Digital de la UB
repository.name.fl_str_mv
repository.mail.fl_str_mv
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score 15,812429

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