Fig. 6. Forced expression of Sox9 influences the fate of neural crest cells in the periphery. (A-F) Transverse sections of the neural tube electroporated with GFP alone (A-C) and Sox9 IRES GFP (D-F) analysed 48 hpt. (A,B) GFP⁺ neural crest cells migrate into core and peripheral regions of dorsal root ganglion (DRG) and sympathetic ganglion (SG), and some cells express the neuronal marker Islet1/2 (blue arrow). (C) GFP⁺ cells are also observed along the peripheral nerve (pn) identified by ß-tubulin (TuJ1) expression. (D,E) In a Sox9-electroporated embryo, GFP⁺ cells are not observed coexpressing Isl1/2 in either the DRG (D) or SG (E) and cells are predominantly located in the periphery of the ganglia (D,E). (F) Sox9-transfected cells are also observed along the TuJ1⁺ peripheral nerve. (G,H) Embryos electroporated with GFP alone (G) or Sox9 (H) analysed 24 hpt. Cells transfected with only GFP migrate exclusively along the medial-lateral migration route leading to the DRG (G). By contrast, cells expressing Sox9 migrate along both medial-lateral and dorsal-lateral migration routes (H) (white arrow). (I) Co-expression of Sox9 and the gene encoding the Schwann cell marker protein zero (P0) is observed in neural crest cells (yellow arrow, inset) after 72 hpt. (J-L) Quantitative analysis of the effects of Sox9 expression on neural crest cells fate (n=9, ±s.e.m.). Following electroporation of Sox9-IRES-GFP, the number of cells in dorsal-lateral migratory route increased compared with the GFP control (J), whereas the number of cells co-producing Islet1/2 was reduced (K), Moreover a small increase in the number of cells co-producing P0 was evident in Sox9-IRES-GFP-transfected embryos (L). (M) The experimental approach. Following electroporation of GFP alone or Sox9-IRES-GFP, neural plate explants [i] are dissected and transplanted into the region between the neural tube and somite at stage 12-14 of the chick embryos at forelimb level and incubated for 72 hours before processing. (N-Q) Cells from [i] regions transfected with GFP alone do not migrate away from the position of transplantation. Many of these cells express Pax7 (N) and Islet1/2 (Q) but few if any GFP⁺ cells were observed expressing HNK-1 (O) or P0 (P). Blue arrows in (N-Q) indicate the high magnification of GFP⁺ cells shown in the insets. (R-U) By contrast, Sox9-expressing cells show extensive migration from sites of transplantation. These cells do not express Pax7 (R) or Islet1/2 (U) but many express HNK-1 (S) and P0 (T). White arrows in (R-U) indicate the high magnification of the Sox9⁺ cells shown in the insets. nc, notochord; da, dorsal aorta.

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