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Details	EV-TRACK ID	Experiment nr.	Species	Sample type	Separation protocol	First author	Year	EV-METRIC
	EV180013	1/1	Homo sapiens	Blood plasma	(d)(U)C	Malin, Steven	2018	55%
Study summary Full title Low cardiorespiratory fitness is associated with higher extracellular vesicle counts in obese adults. All authors Eichner NZM, Gilbertson NM, Gaitan JM, Heiston EM, Musante L, LaSalvia S, Weltman A, Erdbrügger U, Malin SK Journal Physiol Rep Abstract Low cardiorespiratory fitness (CRF) is associated with cardiovascular disease (CVD) independent of o (show more...)Low cardiorespiratory fitness (CRF) is associated with cardiovascular disease (CVD) independent of obesity. Extracellular vesicles (EVs) are a novel target of CVD, however, it remains unknown if obese individuals with very poor fitness (VPF) have elevated EVs versus people with poor fitness (PF). Thus, we tested whether VPF was associated with greater EV subtypes in obese adults. Subjects with VPF (n = 13, VO2 peak: 15.4 ± 0.6 mL/kg/min, BMI: 34.1 ± 1.7 kg/m2 ) and PF (n = 13, VO2 peak: 25.9 ± 3.0 mL/kg/min, BMI: 32.1 ± 1.2 kg/m2 ) were compared in this cross-sectional study. After an overnight fast, AnnexinV (AV) +/- platelet (CD31+ /CD41+ ), leukocyte (CD45+ /CD41- ), and endothelial EVs (CD105+ , CD31+ /CD41- ) were analyzed from fresh platelet poor plasma via imaging flow cytometry. Body fat, blood pressure (BP), and glucose tolerance (OGTT) were also tested. Body weight, BP, and circulating glucose were similar between groups, although VPF subjects were older than PF (64.0 ± 2.1 vs. 49.8 ± 4.2 year; P < 0.05). People with VPF, compared with PF, had higher total AV- EVs (P = 0.04), AV- platelet EVs (CD31+ /CD41+ ; P = 0.006), and AV- endothelial EVs (CD31+ /CD41- ; P = 0.005) independent of age and body fat. Higher AV- platelet and endothelial EVs were associated with lower VO2 peak (r = -0.56, P = 0.006 and r = -0.55, P = 0.005, respectively). Endothelial-derived AV- /CD31+ /CD41- EVs were also related to pulse pressure (r = 0.45, P = 0.03), whereas AV- /CD105 was linked to postprandial glucose (r = 0.41, P = 0.04). VPF is associated with higher AnnexinV- total, endothelial, and platelet EVs in obese adults, suggesting that subtle differences in fitness may reduce type 2 diabetes and CVD risk through an EV-related mechanism. (hide) EV-METRIC 55% (87th 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 Blood plasma Sample origin Prediabetes 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 Adj. k-factor 113.7 (pelleting) / 379.2 (washing) Protein markers EV: TSG101/ CD105/ CD45/ CD31/ CD41/ CD9 non-EV: None Proteomics no Show all info Study aim Biomarker Sample Species Homo sapiens Sample Type Blood plasma Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Between 10,000 g and 50,000 g Pelleting performed Yes Pelleting: time(min) 15 Pelleting: rotor type FA-45-24-11 Pelleting: speed (g) 50000 Pelleting: adjusted k-factor 113.7 Wash: time (min) 10 Wash: Rotor Type FA-45-24-11 Wash: speed (g) 15000 Wash: adjusted k-factor 379.2 Characterization: Protein analysis PMID previous EV protein analysis Western blot Protein Concentration Method Not determined Western Blot Antibody details provided? Yes Lysis buffer provided? Yes Detected EV-associated proteins CD9, TSG101 Flow cytometry Type of Flow cytometry ImageStream MKII Imaging Flow Cytometer Hardware adaptation to ~100nm EV's Hardware adaptation is not needed for ImageStream. Controls included single stained EV samples, buffer only controls (collected for 2 min after filtering with a 0.1 µm filter), buffer plus antibodies Calibration bead size 0.22 Antibody details provided? No Characterization: Lipid analysis No Characterization: Particle analysis NTA Report type Mean Reported size (nm) 168 TRPS Report type Mean Reported size (nm) 182 EM EM-type Cryo-EM Image type Wide-field

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EV-TRACK ID	EV180013
species	Homo sapiens
sample type	Blood plasma
condition	Prediabetes
separation protocol	(d)(U)C
Exp. nr.	1
EV-METRIC %	55