Low cardiorespiratory fitness is associated with higher extracellular vesicle counts in obese adults

Figures

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  Figure 1. EV Phenotyping with Imaging Flow Cytometry. Gating strategy based on low scatter (A) and Annexin V intensity positivity (B) on intensity histogram according to our previous published methods, (C) and (D) Controls: Buffer with only reagents, no EVs (C) unlabeled EVs without reagents (D). Example of dot plots of EV labeling: (E): CD31/Annexin V density plot and (F) CD105/Annexin V density plot.

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  Figure 2. Characterization of EV size, concentration, and morphology. Cryo‐electron microscopy images of EVs of different sizes (<100 nm to 1000 nm) (A); Tunable resistive pulse sensing (TRPS, qNano® by Izon, using 200 nm and 400 nm with 2 pressures). Concentration of EV particles in this example using a 200 nm pore size for qNANO with two pressures at 4A and 8A is 5.7e9 Particles/ml. Mean size is 161 nm and mode size is 116 nm taking average of both pressure measurement. Concentration of EV particles in the same example using a 400 nm pore size for qNANO with 2 pressures at 5A and 8A is 4.5e9 Particles/mL. Mean size is 203 nm and mode size is 170 nm taking average of both pressure measurements (B). Particle tracking of EVs with Nanosight tracking analysis (Zetaview® by Particle Metrix). Concentration of EV particles in this example is 5.1e9 particles/mL. Mean size is 142 nm and mode size is 130 nm (C); Western blotting of vesicle proteins. A (protein pattern) and B (Western blotting) show vesicle protein TSG101 in reducing condition and C (protein pattern) and D (Western blotting) show vesicle protein CD9 in nonreducing condition at expected band length (D).

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  Figure 3. Correlation between CD31+/CD41− endothelial EVs and pulse pressure (A) and CD105 + endothelial EVs with 2‐h glucose (B). EV data were log‐transformed.

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  Figure 4. Comparison of Annexin V+ (A) and Annexin V− EV subtypes (B) in obese individuals with very poor fitness (VPF) and poor fitness (PF). EV data were log‐transformed. Subtypes: CD41 (platelets), CD105 (S‐endoglin, endothelial), CD31 (PECAM, platelet endothelial cell adhesion molecule). Age and body fat were included as covariates.