Analysis of Craniomaxillofacial Malformations in Mice Using Three-dimensional Microcomputed Tomography

Summary

Here, we describe in depth a method, which is based on microcomputed tomography, to segment and measure 3D models of craniomaxillofacial bones in mice, for better assessment of craniomaxillofacial bone development in mice than what is possible with current methods.

Abstract

To model craniofacial malformations caused by vitamin A deficiency (VAD), we expressed a dominant-negative retinoid receptor mutation in osteoblasts to specifically inhibit RAR transcriptional activity in mice. This approach allowed us to investigate the effects of VAD on cranial hypomineralization, mandibular deformity, and clavicular hypoplasia in clinical cases. In this study, microcomputed tomography (microCT) scanning of the craniomaxillofacial region of mice represented a valuable tool for studying the growth and development of this animal model. The manual estimation of images is both time-consuming and inaccurate. Hence, here, we present a straightforward, efficient, and accurate approach for segmenting and quantifying the microCT images of each craniomaxillofacial bone. MicroCT software was used to slice the mandible, frontal bone, parietal bone, nasal bone, premaxilla, maxilla, interparietal bone, and occipital bone of mice and measure their corresponding lengths and widths. This segmentation method can be applied to study growth and development in developmental biology, biomedicine, and other related sciences and allows researchers to analyze the effects of genetic mutations on individual craniofacial bones.

Introduction

The intricate development of the human skull and face encompasses a sophisticated 3D morphogenetic process, intricately orchestrated by numerous genes. These genes play a pivotal role in regulating the intricate patterns, proliferation, and differentiation of tissues derived from diverse embryonic sources. This highly coordinated process underscores the complexity of human craniofacial growth and development. Craniofacial malformations (including cleft lip and palate, cranial suture closure, and facial hypoplasia) occurring as a result of developmental abnormalities account for more than one-third of all congenital birth defects. As a commonly used model animal in bio....

Protocol

We have complied with all relevant ethical regulations for animal testing and research. All experimental animal procedures were approved by the Institutional Animal Care and Research Advisory Committee of the Shanghai Ninth People's Hospital, School of Medicine, Shanghai Jiaotong University.

Both the Rosa26-loxp-stop-loxp-dnRARα403 strain (R26^dn/dn) and the Osterix-Cre (Osx^Cre) (No.006361) strain of mice used here were maintained on the C57BL/6 .......

Discussion

MicroCT is a powerful tool for obtaining realistic and isotropic 3D information from dense and opaque biological samples with micrometer resolution. The data obtained from microCT are calibrated for geometry and intensity, making it especially useful for quantitative studies^{13,14,15,16,17}. It is used to study bone and dental microstructure^18<.......

Materials

Name	Company	Catalog Number	Comments
GraphPad Prism 6.01 Software	GraphPad Software Inc., La Jolla, CA, USA	/
Micro-CT	Quantum GX micro CT, PerkinElmer, Waltham, MA, USA	/
Mimics Medical 19.0	Materialise, Leuven, Belgium	/
Osterix-Cre (OsxCre)	/	/	from the Jackson Laboratory
Rosa26-loxp-stop-loxp-dnRARα403 strain	/	/	from the Columbia University, USA