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Details	EV-TRACK ID	Experiment nr.	Species	Sample type	Separation protocol	First author	Year	EV-METRIC
	EV170045	1/1	Homo sapiens	Primary glioblastoma cells	(d)(U)C Filtration	Treps L	2017	55%
Study summary Full title Glioblastoma stem-like cells secrete the pro-angiogenic VEGF-A factor in extracellular vesicles. All authors Treps L, Perret R, Edmond S, Ricard D, Gavard J Journal J Extracell Vesicles Abstract Glioblastoma multiforme (GBM) are mortifying brain tumours that contain a subpopulation of tumour ce (show more...)Glioblastoma multiforme (GBM) are mortifying brain tumours that contain a subpopulation of tumour cells with stem-like properties, termed glioblastoma stem-like cells (GSCs). GSCs largely contribute to tumour initiation, propagation and resistance to current anti-cancer therapies. GSCs are situated in perivascular niches, closely associated with brain microvascular endothelial cells, thereby involved in bidirectional molecular and cellular interactions. Moreover, extracellular vesicles are suspected to carry essential information that can adapt the microenvironment to the tumour's needs, including tumour-induced angiogenesis. In GBM, extracellular vesicles produced by differentiated tumour cells and GSCs were demonstrated to disseminate locally and at distance. Here, we report that the pro-angiogenic pro-permeability factor VEGF-A is carried in extracellular vesicles secreted from ex vivo cultured patient-derived GSCs. Of note, extracellular vesicle-derived VEGF-A contributes to the in vitro elevation of permeability and angiogenic potential in human brain endothelial cells. Indeed, VEGF-A silencing in GSCs compromised in vitro extracellular vesicle-mediated increase in permeability and angiogenesis. From a clinical standpoint, extracellular vesicles isolated from circulating blood of GBM patients present higher levels of VEGF-A, as compared to healthy donors. Overall, our results suggest that extracellular vesicle-harboured VEGF-A targets brain endothelial cells and might impact their ability to form new vessels. Thus, tumour-released EV cargo might emerge as an instrumental part of the tumour-induced angiogenesis and vascular permeability modus operandi in GBM. (hide) EV-METRIC 55% (88th percentile of all experiments on the same sample type) Reported Not reported Not applicable EV-enriched proteins Protein analysis: analysis of three or more EV-enriched proteins non EV-enriched protein Protein analysis: assessment of a non-EV-enriched protein qualitative and quantitative analysis Particle analysis: implementation of both qualitative and quantitative methods. For the quantitative method, the reporting of measured EV concentration is expected. electron microscopy images Particle analysis: inclusion of a widefield and close-up electron microscopy image density gradient Separation method: density gradient, at least as validation of results attributed to EVs EV density Separation method: reporting of obtained EV density ultracentrifugation specifics Separation method: reporting of g-forces, duration and rotor type of ultracentrifugation steps antibody specifics Protein analysis: antibody clone/reference number and dilution lysate preparation Protein analysis: lysis buffer composition Study data Sample type Cell culture supernatant Sample origin Brain cancer Focus vesicles extracellular vesicle Separation protocol Separation protocol Gives a short, non-chronological overview of the different steps of the separation protocol. dUC = (Differential) (ultra)centrifugation DG = density gradient UF = ultrafiltration SEC = size-exclusion chromatography IAF = immuno-affinity capture (d)(U)C Filtration Adj. k-factor 138.6 (pelleting) / 138.6 (washing) Protein markers EV: CD63/ TIMP2/ TIMP1/ VEGF-A/ ANXA5/ MMP1 non-EV: None Proteomics no Show all info Study aim Function Sample Species Homo sapiens Sample Type Cell culture supernatant EV-producing cells Primary glioblastoma cells EV-harvesting Medium Serum free medium Cell viability (%) NA Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Below or equal to 800 g Between 800 g and 10,000 g Between 10,000 g and 50,000 g Between 100,000 g and 150,000 g Pelleting performed Yes Pelleting: time(min) 240 Pelleting: rotor type SW 55 Ti Pelleting: speed (g) 100000 Pelleting: adjusted k-factor 138.6 Wash: time (min) 120 Wash: Rotor Type SW 55 Ti Wash: speed (g) 100000 Wash: adjusted k-factor 138.6 Filtration steps 0.22µm or 0.2µm Characterization: Protein analysis Protein Concentration Method microBCA ELISA Flow cytometry Type of Flow cytometry MACSQuant Analyzer Other 1 Protein array Characterization: Lipid analysis No Characterization: Particle analysis TRPS Report type Mean Reported size (nm) 109 EV concentration Yes EM EM-type Transmission-EM/ Immune-EM EM protein CD63 Image type Close-up, Wide-field Extra information Wash volume not in original publication.

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EV-TRACK ID	EV170045
species	Homo sapiens
sample type	Cell culture
cell type	Primary glioblastoma cells
condition	Brain cancer
separation protocol	(d)(U)C Filtration
Exp. nr.	1
EV-METRIC %	55