VEGF-A121 fails to rescue ISV sprouting and caudal plexus remodeling defect in vegfa knockdown zebrafish. (A) Vegfa MO (9 ng) alone alternatively vegfa MO (9 ng) combined with vegfa mRNA encoding VEGF-A165, VEGF-A121 (100 pg each), or VEGF-E-NZ2 (200 pg, which was required to reach full effect), was injected into fli1-GFP transgenic zebrafish. At 2.5 dpf, wild-type (WT) zebrafish embryos showed axial vessels, ISV (yellow arrow), and DLAV (dorsal longitudinal anastomotic vessel). The caudal plexus in the tail appeared as a mesh-like structure (red arrow). Vegfa MO-injected zebrafish showed decreased ISV formation and lack of plexus formation. Coinjection of vegfa MO with vegfa121 mRNA resulted in partial ISV rescue, although vessels appeared thicker and stunted compared with WT. Coinjection of vegfa MO and vegfeNZ2 resulted in normal size ISV and caudal plexus, whereas injection of vegfa165 tended to result in hypervascularization. (B) Quantification of vascular defects in zebrafish lacking VEGF-A expression (vegfa MO), and after rescue by expression of VEGF-A165, VEGF-A121, or VEGF-E-NZ2. (C) Coinjection of vegfa MO and vegfa121 mRNA at 3.5 dpf showed thick, less branched vessels in the SIV (red arrow), compared with rescue by vegfa165 or vegfeNZ2. (D) Quantification of SIV branch points in the different conditions.