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Figure 7 | BMC Developmental Biology

Figure 7

From: β-catenin/cyclin D1 mediated development of suture mesenchyme in calvarial morphogenesis

Figure 7

Expansion of skeletogenic precursors is impaired by the cyclin D1 mutation. (A) Cyclin D1 mutation reduces skeletal precursor proliferation in vitro. Primary calvarial cells, isolated from the CycD1+/+ and CycD1-/- newborns, are analyzed by BrdU incorporation and Ki67 staining assays. The immunostained images are taken randomly to determine the percentage of positive cells (n = 3). (B-G) Double labeling reveals the proliferating skeletogenic precursor populations that are Runx2 negative and positive in the metopic suture. Co-immunostaining of Ki67 (B) and Runx 2 (C), and counterstaining with DAPI (D), show cells undergoing mitotic divisions with (arrow) or without (arrowhead) the expression of Runx2 (E) at E17.5. Enlargements of the insets (E) are shown in F and G. (H-O) Impaired skeletogenic activity at the osteogenic fronts of CycD1-/- during embryogenesis is restored at birth. Immunostaining of the E17.5 (H, I, L, M), or newborn (J, K, N, O), CycD1+/+ (H-K) and CycD1-/- (L-O) metopic sutures with Ki67 (H, J, L, N) and Runx2 (I, K, M, O) identify cells undergoing mitotic divisions and expressing Runx2, respectively. At E17.5, the deletion of cyclin D1 affects the presence of Ki67 (H, L) and Runx2 (I, M) positive cells at both suture mesenchyme (Runx2-, arrowhead) and osteogenic fronts (Runx2+, arrows). At newborn, the expression of Ki67 (J, N) remains affected at the midline suture (arrowhead), but becomes unaffected at the osteogenic fronts (arrows) of CycD1-/- which also contain Runx2-expressing cells (K, O). The suture areas and osteogenic fronts are highlighted by broken lines. Scale bars, 200 μm (B-E); 100 μm (H, L); 50 μm (F, G, I-K, M-O).

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