Fig. 3

Adenoviral (ad) vector-mediated expression of vascular endothelial growth factor A (VEGFA) alone and in combination with bone morphogenetic protein 2 (BMP2) was associated with enhanced angiogenesis in scaffold explants. a Representative images at 2 weeks revealed more reddish scaffold explants with multiple capillary/vessel like structures (black arrows) radiating from the periphery in ad-VEGFA and ad-BMP2 + VEGFA groups compared with the other control explants. b–e Frozen sections of scaffold explants at 2 weeks were stained with anti-CD31 targeting mouse CD31 protein and examined for capillary-like structures. Few CD31-positive structures were found in untransduced and ad-GFP scaffold explants (b and c, white arrows). However, in ad-VEGFA and ad-BMP2 + VEGFA explants, many CD31-positive capillary-like structures (white arrows) were seen both at the periphery (not shown in the figure) as well as within the scaffolds (d, e). Red arrowheads indicate remnants of the scaffold material. f Quantification of CD31-positive area in the scaffold explants at 2 weeks demonstrated significantly more CD31-positive structures in ad-VEGFA and ad-BMP2 + VEGFA (ad-B + V) scaffolds than in the untransduced and ad-GFP explants. ANOVA test with Bonferroni post hoc analysis was used for statistical analysis in (e). Error bars represent SEM. ***p < 0.001; **p < 0.01; *P < 0.05. GFP green fluorescent protein, ns not significant