Establishing Normative Embryonic and Early Postnatal Cranial Bone Volume and Density Values for C57BL/6J Laboratory Mice

Abstract

We provide normative data for cranial bone volume and tissue mineral density (TMD) for embryonic and early postnatal C57BL/6J mice that can be used as control data for researchers using experimental mouse models. C57BL/6J mice were selected based on their common usage in experimental research. Our sample includes 3 males and 3 females at four ages: embryonic day (E) 15.5, E17.5, postnatal day (P) 0, and P7. MicroCT images were acquired by the Center for Quantitative Imaging at the Pennsylvania State University using the General Electric v|tom|x L300 nano/microCT system. Prior to data collection, all postcranial bones were electronically removed from the scan using Avizo 9.4, and then background noise was removed from each scan using a 3D median filter. Isosurfaces were reconstructed to represent cranial bone based on a hydroxyapatite phantom that was scanned with each specimen. To maintain consistency between specimens and across age groups, we used 85 mg/cm3 as the minimum threshold to create isosurfaces and segment bone for this study. Twenty cranial bones were individually segmented for each specimen, and volume and TMD were estimated in Avizo. After correcting for multiple testing, no significant differences were found in bone volume or TMD between males and females for any of the examined cranial bones at any age, and data were pooled for analyses. As expected, volume of ossified bone increased with age from E15.5 to P7. The number of ossification centers present in a single individual is highly variable at E15.5 (ranging from 2 – 10), with all bones present by P7. Most of the measured bones consistently increased in TMD with age, with several exceptions. The palatine, vomer, and zygomatic bones each decreased in TMD from P0 to P7. The TMD of the maxilla also decreased slightly from E17.5 to P0, but then increased from P0 to P7. No significant patterns of development related to originating cell population (mesoderm v. neural crest) or mode of ossification (endochondral v. intramembranous) were apparent. These data can serve as a reference standard for future studies that examine craniofacial growth and disease models using mice bred on a C57BL/6J background.Support or Funding InformationNational Science Foundation Grant BCS‐1731909, NIDCR R01DE027677 and R01 DE022988, and the Pennsylvania State University Energy and Environmental Sustainability Laboratories Green Seed Grant