Fig. 6

Ectopic expression of mature VEGFC rescues secondary sprouting in ccbe1 morphants. (A) Confocal projections of Tg(fli1a:EGFP) at 54 hpf. Knock down of ccbe1 or vegfr3 leads to a loss of PLs at the horizontal myoseptum (arrowheads and asterisks). Ectopic expression of the mature form of VEGFC strongly rescues PL formation in ccbe1 morphants but not in vegfr3 morphants. Arrows indicate hyperbranched ISVs. (B) Quantification of PL formation at 54 hpf. In wild type, 98% (n=54/55) of embryos develop PLs, whereas in MO-ccbe1-injected embryos <4% (n=2/52) do. PL development is rescued to 74% (n=29/39) in ccbe1 morphants transiently overexpressing ΔNΔCVEGFC (P<0.0001). This rescue was never observed in vegfr3 morphants with all embryos devoid of PLs (n=23/23). (C) Quantification of ISV hypersprouting at 54 hpf. ISV hypersprouting was observed in wild-type embryos (93%; n=40/43), with mild reductions in ccbe1 morphants (79%; n=31/39) and vegfr3 morphants (65%; n=15/23) after ΔNΔCVEGFC overexpression.