Fig. 2

Fluorescent bead perfusion of engineered microvessels (µVs). a µV+SA construct with perfused anastomotic connections. High magnification views of outlined regions i and ii with corresponding in situ staining for mTm-hESC-ECs (DsRed+, red) and GFP-hESC-ECs (GFP+, green). Scale bars, 100 μm, 40 μm. b High magnification view of fluorescent beads at two time points, t₀ and t₁, 0.27 s apart. Red arrows track the movement of fluorescent beads to calculate velocities. c, d Average bead velocity in the patterned vessel (c) and in sprouts with diameter <50 µm (d) in µV only (blue circles) and µV+SA (green circles) constructs after 4 days and 7 days of culture N = 3, 3, 4, and 3 biological independent samples for D4 µV only, D4 µV+SA, D7 µV only, and D7 µV+SA, respectively. p = 0.004 for D4 µV+SA and D7 µV+SA in sprouts, p = 0.012 for D7 µV and D7 µV+SA in sprouts, p > 0.05 for all others (two-tailed t test). e Scatter plot of bead velocity in relation to sprout diameter N = 15, 20, 37, 53 (number of sprouts analyzed) for D4 µV only, D4 µV+SA, D7 µV only, and D7 µV+SA in 2, 1, 5, and 3 biologically independent samples. Representative images for a, b from three biologically independent samples of D4 µV+SA, with similar results. Hoechst-stained nuclei, blue. Error bars, mean ± SEM. *p < 0.05 determined using two-tailed t test. ns non-significant (p > 0.05). D4 after 4 days of culture, D7 after 7 days of culture