Fig. 4

Endothelial cords are required for initial tumor growth in zebrafish.

(A) Simultaneous dual fluorescence imaging showing the xenografted mouse melanoma (red) and endothelial cords (green panel, arrows) in 5d Tg(flk:eGFP) transgenic zebrafish using tumor cells transfected with control siRNA or VEGF siRNA, or using zebrafish recipients treated with SU5416 (2 µM) or control vehicle. (B,C) Quantitative analysis of tumor-induced endothelial sprouts and the tumor volume at the day 5 (n > 50 fish for each group; *p < 0.05, **p < 0.01, error bars show SEM). (D) Slower growth of xenografted microtumor (red) in zebrafish cloche mutant is significantly rescued by exogenous ECs (green) sorted from 24 hpf Tg(flk:eGFP) zebrafish. Dotted lines indicate the location of microtumor. (E) Quantitative analysis of tumor-associated endothelial cells and the xenografted microtumor volume (n > 11 fish for each group, error bars show SEM). (F) Number of proliferating tumor cells (EdU+, green) in microtumor in Tg(flk:mCherry) transgenic zebrafish decreased when solid endothelial cords (red) are blocked by SU5416 (2 µM) treatment. Dotted lines indicate the location of microtumors. (G) Quantitative analysis of tumor-associated endothelial cells and the EdU + proliferating tumor cells density (n > 12 fish for each group, error bars show SEM). (H) Cell proliferation in normal developing organ buds is not affected by SU5416 treatment at the same dosage.