Additional file 7: Figure S7..
FoxC2 knock-down induces myocyte differentiation at the expense of vascular fates. (A-B) Dorsal view of whole-mount segments electroporated with control-GFP (A) or with FOXC2-engrailed (En) (B) and incubated for 16 hours. Only a few elongating myocytes are observed in the control (arrows, N = 8), whereas FOXC2-En promoted premature fiber formation (arrows in B, N = 6). (C,D) Transverse sections 40 hours post-electroporation with (C) control-GFP or (D) FOXC2-En. In control embryos cells are apparent in both vascular (as SM cells, see C’ and C”) and myotomal domains (N = 4). (C’,C”) High magnification and channel breakdown of the inset showing the cardinal vein (CV) exhibiting co-localization of a GFP-labeled cell with Des/SMA immunoreactivity (arrow). (D) In contrast, knocking-down FoxC2 promotes myotomal colonization while hindering development of SM cells (N = 5). (E) Quantification (mean ± SEM) of labeled cell distribution. (F,G) Dorsal view of whole-mount segments 16 hours post-electroporation of (F) control dsRNA LacZ or (G) dsRNA FoxC2. In control embryos cells are in the lateral DM (arrows, N = 3) whereas in the presence of dsRNA FoxC2 myotomal cells begun differentiating (arrows in G, N = 3). (H) Transverse section showing that co-electroporation of full-length Foxc2 along with dsRNA FoxC2 rescues the FoxC2 knock-down phenotype by inducing cell migration through the sclerotome and SM fate (arrow, see H’, H”) and lack of myotome colonization (compare to Figures 3A,C and 4C). (H’, H”) High magnification and channel breakdown of the inset in H showing the CV exhibiting co-localization of a labeled cell with Des/SMA + immunoreactivity. Qh1 antibody (blue) labels endothelial cells. (I) Quantification (mean ± SEM) of labeled cell distribution (N = 22 for control dsRNA; N = 16 for Foxc2; N = 16 for dsRNA FoxC2; N = 7 for dsRNA FoxC2 + Foxc2). *P ≤0.05; **P ≤0.01. Bar: (C,D) 50 μm; (H) 35 μm; (C’,C”,H’,H”) 17.5 μm. Des, desmin; dsRNA, double-stranded RNA; M, myotome; SEM, standard error of the mean; SMA, smooth muscle actin.
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Applebaum et al. BMC Biology 2014 12:53 doi:10.1186/s12915-014-0053-9