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. 2017 Aug 14;7:8094. doi: 10.1038/s41598-017-06904-7

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© The Author(s) 2017

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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Loss of Ikk2 affects angiogenesis. (A,B) Confocal horizontal section of ventral hindbrain in wild-type and ikk2 ^−/− (zygotic) embryos obtained from heterozygotic parents of Tg(fli:EGFP) background at 48 hpf. CtA (purple arrowhead) stem sprouting from PHBC in each rhombomere (r) was marked accordingly. At this stage, the bilateral CtAs in the r3 formed two connections with lumen to BA (yellow arrow) in wild-type and only one in ikk2 ^−/− embryo. Connections of CtAs in r5 (yellow) and r6 (blue) marked. (C) Quantification of CtA sprouting from PHBC in each rhombomere. P value calculated by paired Student’s t test. (D,E) Confocal projection of CtA angiogenic vessels in Tg(fli:EGFP)/Tg(gata:DsRed) 48 hpf embryos. (J′,K′) Blowup shows the hindbrain of Tg(fli:EGFP)- labelled CtA vessels. Numbers correspond to the rhombomeres containing CtA sprouts. Note that 1*, 2*, 3* marked the CtA vessels that were reduced both in size and connectivity in ikk2 ^−/− embryos. This is the area of hemorrhage (E). (F–H) Quantification of angiogenic index of hindbrain CtAs. P value calculated by Student’s t test. All data are expressed as mean ± SEM; scale bar in all panels: 100 μm. Abbreviations – PHBC: primordial hindbrain channels; BA: basilar artery; CtAs: central arteries (51).