Single‐extracellular vesicle (EV) analyses validate the use of L1 Cell Adhesion Molecule (L1CAM) as a reliable biomarker of neuron‐derived EVs

Isolation of neuron‐derived extracellular vesicles (NDEVs) with L1 Cell Adhesion Molecule (L1CAM)‐specific antibodies has been widely used to identify blood biomarkers of CNS disorders. However, full methodological validation requires demonstration of L1CAM in individual NDEVs and lower levels or ab...

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Veröffentlicht in:Journal of extracellular vesicles Jg. 13; H. 6; S. e12459 - n/a
Hauptverfasser: Nogueras‐Ortiz, Carlos J, Eren, Erden, Yao, Pamela, Calzada, Elizabeth, Dunn, Christopher, Volpert, Olga, Delgado‐Peraza, Francheska, Mustapic, Maja, Lyashkov, Alexey, Rubio, F Javier, Vreones, Michael, Cheng, Lesley, You, Yang, Hill, Andrew F, Ikezu, Tsuneya, Eitan, Erez, Goetzl, Edward J, Kapogiannis, Dimitrios
Format: Journal Article
Sprache:Englisch
Veröffentlicht: United States John Wiley & Sons, Inc 01.06.2024
John Wiley and Sons Inc
Wiley
Schlagworte:
Alzheimer's disease
Biomarkers
Biomarkers - blood
Biomarkers - metabolism
blood biomarkers
Brain diseases
Cell adhesion & migration
Cell adhesion molecules
Cerebrospinal fluid
Epitopes
Extracellular vesicles
Extracellular Vesicles - metabolism
Humans
L1CAM
Ligands
Microscopy
Molecules
Neural Cell Adhesion Molecule L1 - metabolism
Neurons
Neurons - metabolism
neuron‐derived extracellular vesicles
Pathology
Plasma
Proteins
Tubulin
Tubulin - metabolism
Vesicle-Associated Membrane Protein 2 - metabolism
blood biomarkers
extracellular vesicles
neuron‐derived extracellular vesicles
L1CAM
Alzheimer's disease
ISSN:2001-3078, 2001-3078
Online-Zugang:Volltext
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Zusammenfassung:Isolation of neuron‐derived extracellular vesicles (NDEVs) with L1 Cell Adhesion Molecule (L1CAM)‐specific antibodies has been widely used to identify blood biomarkers of CNS disorders. However, full methodological validation requires demonstration of L1CAM in individual NDEVs and lower levels or absence of L1CAM in individual EVs from other cells. Here, we used multiple single‐EV techniques to establish the neuronal origin and determine the abundance of L1CAM‐positive EVs in human blood. L1CAM epitopes of the ectodomain are shown to be co‐expressed on single‐EVs with the neuronal proteins β‐III‐tubulin, GAP43, and VAMP2, the levels of which increase in parallel with the enrichment of L1CAM‐positive EVs. Levels of L1CAM‐positive EVs carrying the neuronal proteins VAMP2 and β‐III‐tubulin range from 30% to 63%, in contrast to 0.8%–3.9% of L1CAM‐negative EVs. Plasma fluid‐phase L1CAM does not bind to single‐EVs. Our findings support the use of L1CAM as a target for isolating plasma NDEVs and leveraging their cargo to identify biomarkers reflecting neuronal function. Previous research suggest that extracellular vesicles (EVs) secreted by brain neurons carry L1CAM and circulate in peripheral blood, coexisting with L1CAM‐positive EVs from peripheral cellular sources and free L1CAM peptides (top left). These other versions of L1CAM in blood challenge the approach of targeting L1CAM for the immunoaffinity isolation of neuronal EVs (top right), a methodology widely adopted to derive blood biomarkers for brain disorders. To further understand L1CAM‐positive EVs, we sought to establish their neuronal origin and determine the abundance of L1CAM‐positive neuronal EVs in human blood using single‐, intact‐EV techniques including flow cytometry, confocal microscopy and a novel Simoa® assay specific for L1CAM in EVs carrying tetraspanins CD9, CD63 and CD81 (bottom left). Results demonstrate that L1CAM ectodomain epitopes are co‐expressed on blood‐derived EVs with the neuronal proteins β‐III‐tubulin, GAP43, and VAMP2, the levels of which increase in parallel with the enrichment of L1CAM‐positive EVs via L1CAM immunocapture (bottom right). Results further validate the use of L1CAM as a target for the immunoaffinity isolation of neuron‐derived EVs from blood.
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ISSN:2001-3078
2001-3078
DOI:10.1002/jev2.12459