Nrecon v1

μCT images were obtained on all collected skulls with a SkyScan 1174 (Kontich, Belgium) at a 19.17 μm voxel resolution (Table 1). Skulls were stabilized using a custom cradle, and reconstructed with NRecon v1.6.4.8 (BrukermicroCT, Kontich, Belgium) as previously described [38 ]. Threshold settings were set to capture bone volume within the skull. Cranial base length was measured from basion to foramen cecum, anterior and posterior base lengths were determined from spheno-occipital synchondrosis (SOS) to foramen cecum and basion to SOS respectively. Measurements of the lengths, widths, and heights of the cartilaginous regions between ossified centers, SOS and inter-sphenoidal synchondrosis (ISS) at the midline of the cartilaginous segment were recorded. A standardized region of interest that included cartilaginous synchondrosis and abutting bone was used to assess the Bone Volume compared to Tissue volume (BV/TV) (Figure 1A). Measures were compared at each time point by split-plot Kruskal-Wallis by postnatal time-point effects by dose; p≤0.05 was considered significant for post-hoc Bonferonni analyses. All statistical analyses were completed using SPSS 23.0 (IBM, Armonk, NY, USA).

μCT images were obtained on 15, 20, and 25 day post-natal
Twist 1 +/− mouse pup skulls with a SkyScan 1174
(Kontich, Belgium) at a 22.57 μm voxel resolution. Scans were obtained on
142 animals (Male=42.8%; Female=43.3%; Undetermined=13.9%) (Table 1). Mouse skulls were reconstructed
with NRecon v1.6.4.8 (BrukermicroCT, Kontich, Belgium) as previously described
and imported into Amira v5.0 where it was exposed to a Gaussian Smoothing image
filter (r50.3 in X, Y, and Z dimensions; isometric kernel size53) to reduce
extraneous noise in the images (Parsons et al.,
2014). Threshold settings were then set to only visualize bone volume
within the skull. Measurements of the length and width of the cranial vault were
collected by a single experienced rater (TEP) from each reconstructed mouse
skull. Landmarks can be visualized at the following website: http://getahead.psu.edu/viewer.html?id=Adult_Mouse_Skull.
Measures were compared at each time point by split-plot ANOVA or Kruskal-Wallis
(if data was not normally distributed) where appropriate for effects by dose,
p<0.05 was considered significant for post-hoc Bonferonni analyses. Sex
of the pups was recorded for future post-hoc investigation but was not
considered a factor in the current analyses. All statistical analyses were
completed using SPSS 23.0 (IBM, Armonk, NY).

μCT images were obtained on postnatal day 15 mouse skulls with a SkyScan 1174 (Kontich, Belgium) at a 22.57 μm voxel resolution. Scans were obtained on 129 animals. Mouse skulls were reconstructed with NRecon v1.6.4.8 (BrukermicroCT, Kontich, Belgium) as previously described (Parsons et al., 2014 ; Howie et al., 2016 (link)). Threshold settings were then set to only visualize bone volume within the skull. The width of the coronal and posterior interfrontal sutures were measured per published methodology at 25, 50, and 75% of its length as previously described (Howie et al., 2016 (link)). Width was defined as the distance between bony fronts.