Fig. 2

Dual Epithelial/Mesenchymal Nature of Osteoblasts during Fin Regeneration

(A–H) Whole-mount imaging of a mosaic bone ray before amputation (A) and at 72 hpa (B) showing osteoblasts permanently labeled by Cre-mediated expression of mCherry (magenta, indicated by magenta arrows). Sections of the same ray at 72 hpa immunostained for Runx2 (C and F) and mCherry (D and G). Overlays are shown in (E) and (H). Yellow arrows show distal mCherry⁺/Runx2⁺ cells, and the green arrow points to a mCherry/Runx2⁺ distal osteoblast (C–H). Amputation sites are indicated with a dashed line. (F)–(H) are high-magnification images of another section through the same ray in (C)–(E).

(I–L) Antibody-stained transverse sections of nonregenerating fins showing α-catenin (green) and zns-5 (red). Yellow arrows indicate epithelial α-catenin⁺/zns-5⁺ osteoblasts.

(M–R) A section from a 96 hpa Tg(sp7:EGFP) fin showing Runx2⁺ (white), sp7:EGFP (green), and α-catenin (red) expression. Regions bounded by dashed white boxes in (P) are shown at higher magnification in (Q) and (R). White arrows point to Runx2⁺ preosteoblasts and green arrows show sp7:EGFP⁺ osteoblasts with membrane-localized α-catenin.

(S and T) Expression of Runx2 (red) and α-catenin (green) on 24 hpa sections. Red arrows indicate Runx2⁺/α-catenin mesenchymal cells.

(U and V) Whole-mount RNA in situ hybridizations of runx2a (U) and twist2 (V) at 24 hpa. Red arrows denote the specific expression of runx2a and twist2 in regenerating tissue.

(W–Y) Runx2 immunostaining (W, red) and twist2 in situ hybridization (X, green) is shown on 72 hpa frozen sections and overlaid (Y). Yellow arrows point to twist2⁺/Runx2⁺ preosteoblasts. Hoechst-labeled nuclei are in blue.

Scale bars represent 50 µm except in H, which represents 5 μm.