Defining a critical period in calvarial development for Hedgehog pathway antagonist-induced frontal bone dysplasia in mice

Yuanjing Jiang,Di Wu,Chuanqing Mao,Hu Zhao,Shixian Zhang,Yongzhen Lai,Caiyu Liao,Weihui Chen

doi:10.1038/s41368-018-0040-z

Yuanjing Jiang, Di Wu + Show 6 more

Open Access

https://doi.org/10.1038/s41368-018-0040-z

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Abstract

The Hedgehog (Hh) signalling pathway is essential for cellular proliferation and differentiation during embryonic development. Gain and loss of function of Hh signalling are known to result in an array of craniofacial malformations. To determine the critical period for Hh pathway antagonist-induced frontal bone hypoplasia, we examined patterns of dysmorphology caused by Hh signalling inhibition. Pregnant mice received a single oral administration of Hh signalling inhibitor GDC-0449 at 100 mg•kg−1 or 150 mg•kg−1 body weight at preselected time points between embryonic days (E)8.5 and 12.5. The optimal teratogenic concentration of GDC-0449 was determined to be 150 mg•kg−1. Exposure between E9.5 and E10.5 induced frontal bone dysplasia, micrognathia and limb defects, with administration at E10.5 producing the most pronounced effects. This model showed decreased ossification of the frontal bone with downregulation of Hh signalling. The osteoid thickness of the frontal bone was significantly reduced. The amount of neural crest-derived frontal bone primordium was reduced after GDC-0449 exposure owing to a decreased rate of cell proliferation and increased cell death.

Highlights

Craniofacial malformations are common congenital defects, accounting for more than three-quarters of all congenital malformations, and they require extensive medical intervention.They can be caused by a variety of stimulating factors during the period of embryonic development.[1]The Hedgehog (Hh) signalling pathway plays a vital role in calvarial growth and patterning.[2,3] In mammals, patterning and morphogenesis of the frontal bone, a major craniofacial structure, begins with the migration of frontal bone precursors derived from cranial neural crest cells (CNCCs) into the frontal bone primordium; in the mouse, this migration proceeds in a caudal-to-rostral direction beginning at E11.5, followed by apical migration at E13.5.4 The mesenchymal cells in the initial condensation differentiate into osteoblasts
The results showed a slight reduction of Patched[1] (Ptch1) and Gli[1] expression levels in GD10.5 embryos compared with vehicle-exposed embryos (Fig. 5a, b, e, f), and Gli[3] levels were further reduced in the frontal bone primordium (Fig. 5i, j)
We suggest that the compromised condensation of the CNC-derived mesenchyme in the frontal bone primordium due to disruption of the Hh pathway at E10.5 is a trigger for the defect observed in early frontal bone development

Summary

INTRODUCTION

Craniofacial malformations are common congenital defects, accounting for more than three-quarters of all congenital malformations, and they require extensive medical intervention They can be caused by a variety of stimulating factors during the period of embryonic development.[1]. Preosteoblasts continue to proliferate to form the bone anlage These osteoblasts within the primordium subsequently synthesize bone matrix through intramembranous ossification.[5,6] Sonic Hedgehog (Shh) has been shown to be necessary in craniofacial and limb development. Loss of Ihh decreases the proliferation of preosteoblasts, leading to a reduction of cranial bone size and widened cranial sutures. Both chemical and genetic disruptions of the Hh pathway have been proposed to result in frontal bone dysplasia. A very large gap was detected between the frontal bones and parietal bones, especially in the GD9.5 and GD10.5

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MATERIALS AND METHODS