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Fig. 3. Distinct effect of exogenous FGF-2 and VEGF-A on vascular development. EBs were cultured in the absence (basal) or presence of 20 ng/ml VEGF-A or FGF-2 from day 0 to day 8 and stained immunohistochemically or by fluorescence using antibodies against CD31 (A) or {alpha}-smooth muscle actin (ASMA) (B) as indicated. Each growth factor produced distinct morphologies of the endothelial and smooth muscle cell pools. The bottom panels in A and B are higher magnification images of part of the boxed regions in the upper panels. Scale bars: A, 300 µm; B, upper panel 200 µm, lower panel 100 µm. (B) Co-localization (arrowheads) of CD31-positive cells (red) with ASMA cells (green) is shown in the far right image. (C) Quantification of CD31 staining as area (i.e. area without holes) in 8-day EBs (mean±s.d., n=6). *P<0.0001 FGF-2 compared to basal and FGF-2 compared to VEGF-A. (D) Quantification of CD31 staining as vessel length in 8-day EBs (mean±s.d., n=6). *P<0.0001 FGF-2 compared to basal, **P=0.0002 VEGF-A compared to basal.





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