Surface landmark quantification of embryonic mouse craniofacial morphogenesis

Abstract

BackgroundMorphometric quantification of subtle craniofacial variation in studies of experimentally modified embryonic mice has proved valuable in determining the effects of developmental perturbations on craniofacial morphogenesis. The direct comparison of landmark coordinate data from embryos of many different mouse strains and mouse models can advance our understanding of the bases for craniofacial variation. We propose a standard set of craniofacial surface landmarks, for use with embryonic day (E) 10.5-12.5 mice, to serve as the foundation for this type of data compilation and analysis. We quantify the intra- and inter-observer landmark placement variation associated with each landmark and determine how the results of a simple ontogenetic analysis might be influenced by selection of landmark set.ResultsIntraobserver landmark placement error for experienced landmarkers generally remains below 0.1 mm, with some landmarks exhibiting higher values at E11.5 and E12.5. Interobserver error tends to increase with embryonic age and those landmarks defined on wide inflections of curves or facial processes exhibit the highest error. Landmarks with highest intra- or inter-observer are identified and we determine that their removal from the dataset does not significantly change the vectors of craniofacial shape change associated with an ontogenetic regression.ConclusionsOur quantification of landmark placement error demonstrates that it is preferable for a single observer to identify all landmark coordinates within a single study and that significant training and experience are necessary before a landmarker can produce data for use in larger meta-analyses. However, we are confident that this standard landmark set, once landmarks with higher error are removed, can serve as a foundation for a comparative dataset of facial morphogenesis across various mouse populations to help identify the developmental bases for phenotypic variation in the craniofacial complex.

Morphometrics, 3D Imaging, and Craniofacial Development

“…Leaving aside the debate over homology as representations of discrete, biologically defined, developmental entities versus resemblance caused by continuity of information (Hall 2007; Jamniczky 2008; Roth 1984; Roth 1991; Van Valen 1982; Wagner 2007), homology is a difficult problem for morphometrics (Klingenberg 2008). This is particularly the case for applications of morphometrics to morphogenesis in embryos (Percival et al 2014). Figure 6 shows a standard landmark set that we use for mouse embryonic craniofacial morphology from E10 to E12.5.…”

“…Yet, as morphogenesis proceeds, this initially obvious anatomical feature moves in relation to the position of the eye, while descendants of the cells found at this landmark at E9.5 are probably buried as the prominences fuse and the whisker field develops by E12.5. Since our landmarking scheme is intended to track the growth and movement of the facial prominences, we define this landmark such that it remains at the border between cells derived from the maxillary and lateral nasal prominences rather than maintaining a standard geometric relationship with other cranial features like the eye (Percival et al 2014). In this case we have made a conscious choice to attempt to follow the specific cell population rather a point defined by its geometric relationship with other features.…”

“…Manual landmarks vary among and can drift over time for landmarkers. Landmarking protocols are also exceedingly difficult to standardize across labs (Percival et al 2014). For this reason, development of automated methods that can handle the complexity of real datasets is crucial.…”

“…Computed microtomography (μCT) is the technique that has most commonly been used in the quantitative study of craniofacial morphogenesis (Billington et al 2015; Boughner et al 2008; Chong et al 2011; Green et al 2015; Hu et al 2015b; Parsons et al 2008; Percival et al 2014; Schmidt et al 2010a; Young et al 2010a; Young et al 2014). Although μCT was initially used mostly for visualizing skeletal tissues, it has proven to be an effective modality for 3D volumetric imaging of embryonic soft tissues (Boughner et al 2008; Metscher 2009; Parsons et al 2008; Wong et al 2014).…”

“…As previously defined (Percival et al ., 2014) the homologous location of a landmark between the maxillary and the lateral nasal prominences is shown. We consciously defined it so that it landmark such that it remains at the border between these cell populations rather than maintaining a standard geometric relationship with other cranial features like the eye.…”