Further Analysis of the Crouzon Mouse: Effects of the FGFR2C342Y Mutation Are Cranial Bone–Dependent

Abstract

Crouzon syndrome is a debilitating congenital disorder involving abnormal craniofacial skeletal development caused by mutations in Fibroblast Growth Factor Receptor-2 (FGFR2). Phenotypic expression in humans exhibits an autosomal dominant pattern that commonly involves premature fusion of the coronal suture (craniosynostosis) and severe midface hypoplasia. To further investigate biologic mechanisms by which the Crouzon syndrome associated FGFR2C342Y mutation leads to abnormal craniofacial skeletal development we created congenic BALB/c FGFR2C342Y/+ mice. Here we show that BALB/c FGFR2C342Y/+ mice have a consistent craniofacial phenotype including partial fusion of the coronal and lambdoid sutures, intersphenoidal synchondrosis and multiple facial bones, with minimal fusion of other craniofacial sutures. This phenotype is similar to the classic and less severe form of Crouzon syndrome that involves significant midface hypoplasia with limited craniosynostosis. Linear and morphometric analyses demonstrate that FGFR2C342Y/+ mice on the BALB/c genetic background differ significantly in form and shape from their wild type littermates, and that in this genetic background the FGFR2C342Y mutation preferentially effects some craniofacial bones and sutures over others. Analysis of cranial bone cells indicates that the FGFR2C342Y mutation promotes aberrant osteoblast differentiation and increased apoptosis that is more severe in frontal than parietal bone cells. Additionally, FGFR2C342Y/+ frontal but not parietal bones exhibit significantly diminished bone volume and density compared to wild type mice. These results confirm that FGFR2-associated craniosynostosis occurs in association with diminished cranial bone tissue and may provide a potential biologic explanation for the clinical finding of phenotype consistency that exists between many Crouzon syndrome patients.

Enzyme replacement for craniofacial skeletal defects and craniosynostosis in murine hypophosphatasia

“…Whole skulls were scanned at an 18 µm isotropic voxel resolution using the eXplore Locus SP micro-computed tomography imaging system (GE Healthcare Pre-Clinical Imaging, London, ON, Canada), as previously described [25,38]. Density, volume and mineral content of cranial bones from P15 mice were measured using previously established methods and regions of interest using Microview version 2.2 software (GE Healthcare Pre-Clinical Imaging, London, ON) and established algorithms [38–40].…”

“…Density, volume and mineral content of cranial bones from P15 mice were measured using previously established methods and regions of interest using Microview version 2.2 software (GE Healthcare Pre-Clinical Imaging, London, ON) and established algorithms [38–40]. …”

“…Cranial sutures were viewed using the two-dimensional micro-CT slices in an orthogonal view across the entire length of the coronal suture, as previously described [25,37,38]. Bone fusions identified on micro CT images were verified by visualization of specimens under a dissecting microscope (Leica M60 TL5000; Leica Microsystems Inc., Buffalo Groves, IL).…”

“…P15 (approximately two week-old) vehicle treated Alpl −/− , 8.2 mg/kg/day treated Alpl −/− and control Alpl +/+ mouse skulls were carefully dissected and fixed. Linear measurements were calculated using previously reported craniofacial skeletal landmarks [25,36–38], including five standard measurements currently in use by the Craniofacial Mutant Mouse Resource of Jackson Laboratory (Bar Harbor, ME). Linear measurements were normalized to total skull length (measured from nasale to opisthion) to account for potential size differences between Alpl −/− and Alpl +/+ mice.…”

“…3) [36,38]. Landmarks were placed and landmark coordinate data was generated using Dolphin Imaging 11.0 software (Dolphin Imaging and Management Solutions, Chatsworth, CA), as previously reported [38].…”