Fig. 7: NRP1 overexpression enhances vascular sprouting in MEECs.
From: NRP1 interacts with endoglin and VEGFR2 to modulate VEGF signaling and endothelial cell sprouting
MEEC+/+ and MEEC-/- cells were transfected with myc-NRP1 or empty vector as in Fig. 5. At 24 h post-transfection, they were lifted in full medium and cultured as hanging drops (2 × 105 cells/drop; 24 h). The spheroids were collected, and allowed to sprout on growth factor-reduced Matrigel with or without VEGF-A (100 ng/ml) for another 24 h (Methods). Phase-contrast images were taken with 10x magnification, and Image Pro Plus was used to outline the sprouts and quantify the area covered by them for each spheroid (Methods). a Typical images of spheroids analysed for sprouts area. Each original phase contrast image (left) is shown alongside the image with red-outlined sprouts. Scale bar, 100 µm. b Quantification of the sprouting experiments. Sprouting was enhanced by overexpression of NRP1 either in the absence (MEEC-/-) or presence (MEEC+/+) of ENG, while VEGF-A-mediated sprouting required ENG. Data are mean ± SEM of n = 37−61 spheroids per condition (the number of spheroids measured are indicated under each bar) from 5 independent experiments. The area of the sprouts in VEGF-A-stimulated MEEC+/+ was normalized to 100%, and the sprouts area under all other conditions was calculated relative to this value. **p < 0.01; ***p < 10-3; ****p < 10-4 (one-way ANOVA and Bonferroni post-hoc test). ns = not significant.
