Distinct RNA profiles in subpopulations of extracellular vesicles: apoptotic bodies, microvesicles and exosomes.

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Crescitelli, Rossella,1985Gothenburg University,Göteborgs universitet,Krefting Research Centre (författare)

Distinct RNA profiles in subpopulations of extracellular vesicles: apoptotic bodies, microvesicles and exosomes.

Artikel/kapitelEngelska2013

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2013-09-12
Wiley,2013

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LIBRIS-ID:oai:gup.ub.gu.se/186785
https://gup.ub.gu.se/publication/186785URI
https://doi.org/10.3402/jev.v2i0.20677DOI

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Språk:engelska

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INTRODUCTION: In recent years, there has been an exponential increase in the number of studies aiming to understand the biology of exosomes, as well as other extracellular vesicles. However, classification of membrane vesicles and the appropriate protocols for their isolation are still under intense discussion and investigation. When isolating vesicles, it is crucial to use systems that are able to separate them, to avoid cross-contamination. METHOD: EVS RELEASED FROM THREE DIFFERENT KINDS OF CELL LINES: HMC-1, TF-1 and BV-2 were isolated using two centrifugation-based protocols. In protocol 1, apoptotic bodies were collected at 2,000×g, followed by filtering the supernatant through 0.8 µm pores and pelleting of microvesicles at 12,200×g. In protocol 2, apoptotic bodies and microvesicles were collected together at 16,500×g, followed by filtering of the supernatant through 0.2 µm pores and pelleting of exosomes at 120,000×g. Extracellular vesicles were analyzed by transmission electron microscopy, flow cytometry and the RNA profiles were investigated using a Bioanalyzer(®). RESULTS: RNA profiles showed that ribosomal RNA was primary detectable in apoptotic bodies and smaller RNAs without prominent ribosomal RNA peaks in exosomes. In contrast, microvesicles contained little or no RNA except for microvesicles collected from TF-1 cell cultures. The different vesicle pellets showed highly different distribution of size, shape and electron density with typical apoptotic body, microvesicle and exosome characteristics when analyzed by transmission electron microscopy. Flow cytometry revealed the presence of CD63 and CD81 in all vesicles investigated, as well as CD9 except in the TF-1-derived vesicles, as these cells do not express CD9. CONCLUSIONS: Our results demonstrate that centrifugation-based protocols are simple and fast systems to distinguish subpopulations of extracellular vesicles. Different vesicles show different RNA profiles and morphological characteristics, but they are indistinguishable using CD63-coated beads for flow cytometry analysis.

Ämnesord och genrebeteckningar

NATURVETENSKAP Biologi Cellbiologi hsv//swe
NATURAL SCIENCES Biological Sciences Cell Biology hsv//eng
NATURVETENSKAP Biologi Biokemi och molekylärbiologi hsv//swe
NATURAL SCIENCES Biological Sciences Biochemistry and Molecular Biology hsv//eng
apoptotic bodies; microvesicles; exosomes; extracellular vesicles; ultracentrifugation; characterization; RNA; electron microscopy

Biuppslag (personer, institutioner, konferenser, titlar ...)

Lässer, Cecilia,1981Gothenburg University,Göteborgs universitet,Krefting Research Centre(Swepub:gu)xlasce (författare)
Szabó, Tamas G (författare)
Kittel, Agnes (författare)
Eldh, Maria,1980Gothenburg University,Göteborgs universitet,Krefting Research Centre(Swepub:gu)xeldhm (författare)
Dianzani, Irma (författare)
Buzás, Edit I (författare)
Lötvall, Jan,1956Gothenburg University,Göteborgs universitet,Krefting Research Centre(Swepub:gu)xlotja (författare)
Göteborgs universitetKrefting Research Centre (creator_code:org_t)

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Ingår i:Journal of extracellular vesicles: Wiley22001-3078

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Av lärosätet: Göteborgs universitet

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