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. 2016 Jun 3;25(14):2997–3010. doi: 10.1093/hmg/ddw153

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© The Author 2016. Published by Oxford University Press.

This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/4.0/), which permits non-commercial re-use, distribution, and reproduction in any medium, provided the original work is properly cited. For commercial re-use, please contact journals.permissions@oup.com

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Figure 4. — NVP-BGJ398 corrects primary and secondary cartilages defects in ex vivo cultures of mandibles from Fgfr3^Y367C/+embryos. (A) Representative macroscopic views of hemi-mandibles from E16.5 WT or Fgfr3^Y367C/+embryos, treated with DMSO or NVP-BGJ398 and cultured for 6 days, following alcian blue and alizarin red staining. Symphyseal (s), condylar (c) and Meckel’s (m) cartilages are indicated. (B) Mandibular body and condylar neck length of cultured hemi-mandibles (n ≥ 5 individuals for each genotype and treatment). Total length of the hemi-mandible was measured as well as the length of the condylar and symphyseal cartilages (identified with the alcian blue staining). The length of the body was calculated as the total length minus the condylar and symphyseal cartilages. (C) Histological staining (Safranin’O) and immunostaining for ColX of cultured hemi-mandibles. An enlargement of the ColX immunostaining highlights the modification in the size of hypertrophic chondrocytes in MC of Fgfr3^Y367C/+embryos and the correction of the defect with NVP-BGJ398 (scale bar = 200 μm). (D) Measurement of the ColX positive zone inside MC of cultured hemi-mandibles (n ≥ 6 individuals for each genotype and treatment). (E) Mean percentage of hypertrophic chondrocytes for different size categories (expressed in μm²) inside MC of cultured hemi-mandibles (n ≥ 50 cells from n ≥ 5 individuals for each genotype and treatment). (F) Mean number of immune positive cells for Fgfr3 or Ki67 inside MC of WT, Fgfr3^Y367C/+DMSO and Fgfr3^Y367C/+NVP-BGJ398 E16.5 embryos (n ≥ 5 individuals for each genotype). (G) Histological staining (Safranin’O) and immunostaining for Collagen X of cultured hemi-mandibles condylar cartilage (scale bar = 100 μm). Meckel’s cartilage (m) is indicated. (H) Mean percentage of hypertrophic chondrocytes for different size categories (expressed in μm²) inside condylar cartilage of cultured hemi-mandibles (n ≥ 50 cells from n ≥ 5 individuals for each genotype and treatment). Data shown as mean with SD; *P < 0.05, **P < 0.01.