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Details	EV-TRACK ID	Experiment nr.	Species	Sample type	Separation protocol	First author	Year	EV-METRIC
	EV190058	1/1	Homo sapiens	primary spindle-shaped amniotic fluid stem cells	(d)(U)C qEV UF	Takov K	2019	67%
Study summary Full title Small extracellular vesicles secreted from human amniotic fluid mesenchymal stromal cells possess cardioprotective and promigratory potential. All authors Takov K, He Z, Johnston HE, Timms JF, Guillot PV, Yellon DM, Davidson SM Journal Basic Res Cardiol Abstract Mesenchymal stromal cells (MSCs) exhibit antiapoptotic and proangiogenic functions in models of myoc (show more...)Mesenchymal stromal cells (MSCs) exhibit antiapoptotic and proangiogenic functions in models of myocardial infarction which may be mediated by secreted small extracellular vesicles (sEVs). However, MSCs have frequently been harvested from aged or diseased patients, while the isolated sEVs often contain high levels of impurities. Here, we studied the cardioprotective and proangiogenic activities of size-exclusion chromatography-purified sEVs secreted from human foetal amniotic fluid stem cells (SS-hAFSCs), possessing superior functional potential to that of adult MSCs. We demonstrated for the first time that highly pure (up to 1.7 × 1010 particles/µg protein) and thoroughly characterised SS-hAFSC sEVs protect rat hearts from ischaemia-reperfusion injury in vivo when administered intravenously prior to reperfusion (38 ± 9% infarct size reduction, p < 0.05). SS-hAFSC sEVs did not protect isolated primary cardiomyocytes in models of simulated ischaemia-reperfusion injury in vitro, indicative of indirect cardioprotective effects. SS-hAFSC sEVs were not proangiogenic in vitro, although they markedly stimulated endothelial cell migration. Additionally, sEVs were entirely responsible for the promigratory effects of the medium conditioned by SS-hAFSC. Mechanistically, sEV-induced chemotaxis involved phosphatidylinositol 3-kinase (PI3K) signalling, as its pharmacological inhibition in treated endothelial cells reduced migration by 54 ± 7% (p < 0.001). Together, these data indicate that SS-hAFSC sEVs have multifactorial beneficial effects in a myocardial infarction setting. (hide) EV-METRIC 67% (94th 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 Control condition Focus vesicles small extracellular vesicles (sEVs) 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 qEV UF Protein markers EV: CD81/ CD63/ CD9 non-EV: ACTN4 Proteomics yes Show all info Study aim Function/Identification of content (omics approaches)/Technical analysis comparing/optimizing EV-related methods Sample Species Homo sapiens Sample Type Cell culture supernatant EV-producing cells primary spindle-shaped amniotic fluid stem cells EV-harvesting Medium Serum free medium Cell viability (%) 86 Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Below or equal to 800 g Between 10,000 g and 50,000 g Pelleting performed No Ultra filtration Cut-off size (kDa) 30 Membrane type Hydrosart (stabilised cellulose);Other Commercial kit qEV Other Name other separation method qEV EV-subtype Distinction between multiple subtypes Used subtypes Characterization: Protein analysis Protein Concentration Method microBCA;Bradford Proteomics database No Other 1 Immunoassay (DELFIA) Detected EV-associated proteins CD9/ CD63/ CD81 Other 2 Dot blot Detected EV-associated proteins CD81/ CD63 Not detected EV-associated proteins CD9 Not detected contaminants ACTN4 Characterization: Lipid analysis No Characterization: Particle analysis NTA Report type Mode Reported size (nm) 100.5 EV concentration Yes EM EM-type Transmission-EM Image type Close-up, Wide-field

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EV-TRACK ID	EV190058
species	Homo sapiens
sample type	Cell culture
cell type	primary spindle-shaped amniotic fluid stem cells
condition	Control condition
separation protocol	(d)(U)C qEV UF
Exp. nr.	1
EV-METRIC %	67