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Fig. 2. LSox5 misexpression in the cephalic neural tube augments the neural crest population. (A) Dorsal view of an embryo 20 hours after co-electroporation with pCX-LSox5 and PCX-EGFP on the right-hand side of the cephalic neural tube. The EGFP expression is seen on the treated side and in the emerging streams of migratory neural crest cells (white arrowheads). (B,C) Transverse sections of an electroporated embryo showing the coincident expression of EGFP and ectopic LSox5 all along the dorsoventral axis of the right hand side of the neural tube. (D,E) Lateral views of an electroporated embryo immunostained for HNK1 where the increase in HNK1-positive migratory cells in the electroporated (ep) side can be observed, particularly in the circumpharyngeal stream (black arrowhead). (F,G) A transverse section at the level of the caudal hindbrain shows the supernumerary HNK1-positive migratory cells (red) in the electroporated side of the embryo. (H) A magnification of G shows the appearance of ectopic HNK1-positive cells within the treated side of the neural tube (arrowheads). (I,J) At the hindbrain level, in a double immunostained transverse section, the overexpression of Pax7 (red) in cells expressing EGFP (green) can be seen in both neuroepithelial and migratory cells on the LSox5 transfected side. (K,L) An overlay of RhoB hybridisation and double immunostaining for Pax7 (green) and laminin 1 (red) shows that severe damage of the basement membrane coincides with the expanded region of neural crest delamination on the electroporated side of the neural tube. (M,N) Transverse sections of electroporated embryos hybridised for LSox5 and Slug (M), and N-cadherin (N). The territory competent for neural crest generation extends ventrally on the electroporated side, as judged by the expanded expression of Slug (M) and the coincident inhibition of N-cadherin expression (compare the length of the brackets on both sides of the neural tube). (O) At more ventral regions of the neural tube, scattered cells could be detected that ectopically expressed RhoB on the treated side of the neural tube, breaking the basal lamina and invading into the lateral mesenchyme (open arrowhead). ov, otic vesicle.





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