Identification of distinct nanoparticles and subsets of extracellular vesicles by asymmetric flow field-flow fractionation

Haiying Zhang, Daniela Freitas, Han Sang Kim, Kristina Fabijanic, Zhong Li, Haiyan Chen, Milica Tesic Mark, Henrik Molina, Alberto Benito Martin, Linda Bojmar, Justin Fang, Sham Rampersaud, Ayuko Hoshino, Irina Matei, Candia M. Kenific, Miho Nakajima, Anders Peter Mutvei, Pasquale Sansone, Weston Buehring, Huajuan WangJuan Pablo Jimenez, Leona Cohen-Gould, Navid Paknejad, Matthew Brendel, Katia Manova-Todorova, Ana Magalhães, José Alexandre Ferreira, Hugo Osório, André M. Silva, Ashish Massey, Juan R. Cubillos-Ruiz, Giuseppe Galletti, Paraskevi Giannakakou, Ana Maria Cuervo, John Blenis, Robert Schwartz, Mary Sue Brady, Héctor Peinado, Jacqueline Bromberg, Hiroshi Matsui, Celso A. Reis, David Lyden

Research output: Contribution to journalArticle

150 Citations (Scopus)

Abstract

The heterogeneity of exosomal populations has hindered our understanding of their biogenesis, molecular composition, biodistribution and functions. By employing asymmetric flow field-flow fractionation (AF4), we identified two exosome subpopulations (large exosome vesicles, Exo-L, 90-120 nm; small exosome vesicles, Exo-S, 60-80 nm) and discovered an abundant population of non-membranous nanoparticles termed 'exomeres' (~35 nm). Exomere proteomic profiling revealed an enrichment in metabolic enzymes and hypoxia, microtubule and coagulation proteins as well as specific pathways, such as glycolysis and mTOR signalling. Exo-S and Exo-L contained proteins involved in endosomal function and secretion pathways, and mitotic spindle and IL-2/STAT5 signalling pathways, respectively. Exo-S, Exo-L and exomeres each had unique N-glycosylation, protein, lipid, DNA and RNA profiles and biophysical properties. These three nanoparticle subsets demonstrated diverse organ biodistribution patterns, suggesting distinct biological functions. This study demonstrates that AF4 can serve as an improved analytical tool for isolating extracellular vesicles and addressing the complexities of heterogeneous nanoparticle subpopulations.

Original languageEnglish (US)
Pages (from-to)332-343
Number of pages12
JournalNature Cell Biology
Volume20
Issue number3
DOIs
StatePublished - Mar 1 2018

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Field Flow Fractionation
Exosomes
Nanoparticles
Microtubule Proteins
Spindle Apparatus
Secretory Pathway
Glycolysis
Population Characteristics
Glycosylation
Proteomics
Interleukin-2
RNA
Lipids
DNA
Enzymes
Population
Extracellular Vesicles
Proteins

ASJC Scopus subject areas

  • Cell Biology

Cite this

Identification of distinct nanoparticles and subsets of extracellular vesicles by asymmetric flow field-flow fractionation. / Zhang, Haiying; Freitas, Daniela; Kim, Han Sang; Fabijanic, Kristina; Li, Zhong; Chen, Haiyan; Mark, Milica Tesic; Molina, Henrik; Martin, Alberto Benito; Bojmar, Linda; Fang, Justin; Rampersaud, Sham; Hoshino, Ayuko; Matei, Irina; Kenific, Candia M.; Nakajima, Miho; Mutvei, Anders Peter; Sansone, Pasquale; Buehring, Weston; Wang, Huajuan; Jimenez, Juan Pablo; Cohen-Gould, Leona; Paknejad, Navid; Brendel, Matthew; Manova-Todorova, Katia; Magalhães, Ana; Ferreira, José Alexandre; Osório, Hugo; Silva, André M.; Massey, Ashish; Cubillos-Ruiz, Juan R.; Galletti, Giuseppe; Giannakakou, Paraskevi; Cuervo, Ana Maria; Blenis, John; Schwartz, Robert; Brady, Mary Sue; Peinado, Héctor; Bromberg, Jacqueline; Matsui, Hiroshi; Reis, Celso A.; Lyden, David.

In: Nature Cell Biology, Vol. 20, No. 3, 01.03.2018, p. 332-343.

Research output: Contribution to journalArticle

Zhang, H, Freitas, D, Kim, HS, Fabijanic, K, Li, Z, Chen, H, Mark, MT, Molina, H, Martin, AB, Bojmar, L, Fang, J, Rampersaud, S, Hoshino, A, Matei, I, Kenific, CM, Nakajima, M, Mutvei, AP, Sansone, P, Buehring, W, Wang, H, Jimenez, JP, Cohen-Gould, L, Paknejad, N, Brendel, M, Manova-Todorova, K, Magalhães, A, Ferreira, JA, Osório, H, Silva, AM, Massey, A, Cubillos-Ruiz, JR, Galletti, G, Giannakakou, P, Cuervo, AM, Blenis, J, Schwartz, R, Brady, MS, Peinado, H, Bromberg, J, Matsui, H, Reis, CA & Lyden, D 2018, 'Identification of distinct nanoparticles and subsets of extracellular vesicles by asymmetric flow field-flow fractionation', Nature Cell Biology, vol. 20, no. 3, pp. 332-343. https://doi.org/10.1038/s41556-018-0040-4
Zhang, Haiying ; Freitas, Daniela ; Kim, Han Sang ; Fabijanic, Kristina ; Li, Zhong ; Chen, Haiyan ; Mark, Milica Tesic ; Molina, Henrik ; Martin, Alberto Benito ; Bojmar, Linda ; Fang, Justin ; Rampersaud, Sham ; Hoshino, Ayuko ; Matei, Irina ; Kenific, Candia M. ; Nakajima, Miho ; Mutvei, Anders Peter ; Sansone, Pasquale ; Buehring, Weston ; Wang, Huajuan ; Jimenez, Juan Pablo ; Cohen-Gould, Leona ; Paknejad, Navid ; Brendel, Matthew ; Manova-Todorova, Katia ; Magalhães, Ana ; Ferreira, José Alexandre ; Osório, Hugo ; Silva, André M. ; Massey, Ashish ; Cubillos-Ruiz, Juan R. ; Galletti, Giuseppe ; Giannakakou, Paraskevi ; Cuervo, Ana Maria ; Blenis, John ; Schwartz, Robert ; Brady, Mary Sue ; Peinado, Héctor ; Bromberg, Jacqueline ; Matsui, Hiroshi ; Reis, Celso A. ; Lyden, David. / Identification of distinct nanoparticles and subsets of extracellular vesicles by asymmetric flow field-flow fractionation. In: Nature Cell Biology. 2018 ; Vol. 20, No. 3. pp. 332-343.
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abstract = "The heterogeneity of exosomal populations has hindered our understanding of their biogenesis, molecular composition, biodistribution and functions. By employing asymmetric flow field-flow fractionation (AF4), we identified two exosome subpopulations (large exosome vesicles, Exo-L, 90-120 nm; small exosome vesicles, Exo-S, 60-80 nm) and discovered an abundant population of non-membranous nanoparticles termed 'exomeres' (~35 nm). Exomere proteomic profiling revealed an enrichment in metabolic enzymes and hypoxia, microtubule and coagulation proteins as well as specific pathways, such as glycolysis and mTOR signalling. Exo-S and Exo-L contained proteins involved in endosomal function and secretion pathways, and mitotic spindle and IL-2/STAT5 signalling pathways, respectively. Exo-S, Exo-L and exomeres each had unique N-glycosylation, protein, lipid, DNA and RNA profiles and biophysical properties. These three nanoparticle subsets demonstrated diverse organ biodistribution patterns, suggesting distinct biological functions. This study demonstrates that AF4 can serve as an improved analytical tool for isolating extracellular vesicles and addressing the complexities of heterogeneous nanoparticle subpopulations.",
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AU - Zhang, Haiying

AU - Freitas, Daniela

AU - Kim, Han Sang

AU - Fabijanic, Kristina

AU - Li, Zhong

AU - Chen, Haiyan

AU - Mark, Milica Tesic

AU - Molina, Henrik

AU - Martin, Alberto Benito

AU - Bojmar, Linda

AU - Fang, Justin

AU - Rampersaud, Sham

AU - Hoshino, Ayuko

AU - Matei, Irina

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AU - Nakajima, Miho

AU - Mutvei, Anders Peter

AU - Sansone, Pasquale

AU - Buehring, Weston

AU - Wang, Huajuan

AU - Jimenez, Juan Pablo

AU - Cohen-Gould, Leona

AU - Paknejad, Navid

AU - Brendel, Matthew

AU - Manova-Todorova, Katia

AU - Magalhães, Ana

AU - Ferreira, José Alexandre

AU - Osório, Hugo

AU - Silva, André M.

AU - Massey, Ashish

AU - Cubillos-Ruiz, Juan R.

AU - Galletti, Giuseppe

AU - Giannakakou, Paraskevi

AU - Cuervo, Ana Maria

AU - Blenis, John

AU - Schwartz, Robert

AU - Brady, Mary Sue

AU - Peinado, Héctor

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N2 - The heterogeneity of exosomal populations has hindered our understanding of their biogenesis, molecular composition, biodistribution and functions. By employing asymmetric flow field-flow fractionation (AF4), we identified two exosome subpopulations (large exosome vesicles, Exo-L, 90-120 nm; small exosome vesicles, Exo-S, 60-80 nm) and discovered an abundant population of non-membranous nanoparticles termed 'exomeres' (~35 nm). Exomere proteomic profiling revealed an enrichment in metabolic enzymes and hypoxia, microtubule and coagulation proteins as well as specific pathways, such as glycolysis and mTOR signalling. Exo-S and Exo-L contained proteins involved in endosomal function and secretion pathways, and mitotic spindle and IL-2/STAT5 signalling pathways, respectively. Exo-S, Exo-L and exomeres each had unique N-glycosylation, protein, lipid, DNA and RNA profiles and biophysical properties. These three nanoparticle subsets demonstrated diverse organ biodistribution patterns, suggesting distinct biological functions. This study demonstrates that AF4 can serve as an improved analytical tool for isolating extracellular vesicles and addressing the complexities of heterogeneous nanoparticle subpopulations.

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