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. 2015 Oct 8;6:8479. doi: 10.1038/ncomms9479

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(a) Tg(fli1a:EGFP)y1 zebrafish embryos were treated with control (ctr) or nova2 morpholino (MO-nova2) oligos and analysed with anti-Podocalyxin antibody (Podxl2, red) in the ISVs (upper panel). Co-injection of a morpholino-resistant zebrafish nova2 mRNA was able to rescue the apical-membrane staining (arrow) of Podxl2 in nova2 morphant (MO-nova2+nova2; scale bar, 20 μm). Areas (i) are magnified on the right (scale bar, 10 μm). This is also visualized in the optical transverse sections (XZ, scale bar, 5 μm). Asterisk, lumen; arrowheads indicate Podxl2 mislocalization. Lower panel: Podxl2 localization in the dorsal aorta (scale bar, 50 μm). (b) Lateral view of the Tg(fli1a:EGFP)y1 embryo with EGFP under the control of the endothelial-specific promoter fli1a. The same transgenic zebrafish embryos were co-injected at one-cell stage with a vector (pTol2fli1ep:mCherry-nova2) driving the expression of a morpholino-resistant zebrafish nova2 mRNA fused to mCherry fluorescent protein under the fli1a promoter. In this case individual ECs were mosaically labelled with mCherry-nova2 expression (nova2, red) in the nucleus of some cells of the trunk blood vessels (scale bar, 250 μm). (c) Endothelial-autonomous rescue of nova2 morphants. PTol2fli1ep:mCherry-nova2 plasmid was co-injected with the nova2 morpholino oligo (MO-nova2) in one-cell stage Tg(fli1a:EGFP)y1 embryos (bar 40 μm; arrowheads and arrows indicate vessels positive and negative for nova2, respectively). (d) ISV of the trunk expressing nova2 (arrowheads) showed a normal pattern and developed a correct lumen, whereas an adjacent ISV (arrows), negative for nova2 expression, was not formed properly, so that it failed to reach DLAV (scale bar, 30 μm). (e) Lateral view of 28-hpf transgenic embryos injected with the MO-nova2. Embryo showed in the upper row expresses in the vessel endothelium only GFP, whereas the embryo showed in the lower row expresses also a morpholino-resistant nova2 cDNA fused with mCherry. The presence of nova2 in the vessel endothelium is sufficient to preserve vessel morphology of nova2 morphant embryos (scale bar, 50 μm).