Fig. 3

Premature collagen deposition, mineralisation and osteoblast alignment are seen in E18.0 Grhl3 ^−/− embryos (a, b) Verhoeff’s van Gieson’s (collagen) stain showing little evidence of organised collagen deposits at the location of the coronal suture in WT embryos (a), in contrast to Grhl3 ^−/−embryos (arrowheads, b). c, d Masson’s trichrome stain showing diffuse staining in the region of the developing coronal suture in WT embryos (c), compared to overlapping projections of blue staining (indicative of increased mineralisation) in Grhl3 ^−/− (arrowheads, d). e, f Toluidine blue staining showing dark blue nuclei to define a clear row of osteoblasts along the emerging front in Grhl3 ^−/− embryos (red arrows, f); these were not noticeable in WT embryos (e). Inset (boxed) regions in all panels show increased magnifications of the embryonic skull. Note that the dura mater (DM) and underlying cerebrum are in clear contact with the skull in WT (a, c, e) but not Grhl3 ^−/− (b, d, f) embryonic skulls