Fig. 4. Endoneurial fibroblasts are neural crest derived. Sciatic nerves were dissected from P11 Wnt1-Cre⁺loxpRosa⁺ pups and processed for immunohistochemistry (A,B), or electron microscopy (C-H). Nerves were stained with antibodies against ß-gal (red) to identify neural crest-derived cells, and S100ß (green) to identify Schwann cells. The same field of view is shown in A and B, with DAPI staining (blue) added to panel B. In addition to ß-gal⁺S100ß⁺ glia (green or yellow), we also observed ß-gal⁺S100ß^– endoneurial cells (white arrows) that were neither associated with axons nor blood vessels. By electron microscopy, we consistently observed endoneurial fibroblasts stained with bluo-gal, indicating neural crest-derivation (C-F). (C,D) Bluo-gal-stained endoneurial fibroblasts (f), surrounded by myelinating Schwann cells, most of which are also stained with bluo-gal. Red arrows indicate bluo-gal crystals in the fibroblast; green arrows indicate bluo-gal crystals in Schwann cells. (E) A lower magnification image of a large bluo-gal stained fibroblast, with a very long process, surrounded by Schwann cells. The boxed area is shown at a higher magnification in F. Note the presence of a basal lamina around the Schwann cells (black arrows), but the lack of a basal lamina around the fibroblast process. (G,H) Low and high power images of sections through control littermates that lack any bluo-gal staining in perineurial cells (G), Schwann cells (G,H) or fibroblasts (H).