Apert syndrome is caused by mutations in
fibroblast growth factor receptor 2 (Fgfr2) and is characterized by
craniosynostosis and other skeletal abnormalities. The
Apert syndrome Fgfr2+/S252W mouse model exhibits perinatal lethality. A 3D
hydrogel culture model, derived from tissue engineering strategies, was used to extend the study of the effect of the Fgfr2+/S252W mutation in differentiating osteoblasts postnatally. We isolated cells from the long bones of Apert Fgfr2+/S252W mice (n=6) and cells from the wild-type sibling mice (n=6) to be used as controls. During monolayer expansion, Fgfr2+/S252W cells demonstrated increased proliferation and ALP activity, as well as altered responses of these cellular functions in the presence of FGF
ligands with different binding specificity (
FGF2 or FGF10). To better mimic the in vivo disease development scenario, cells were also encapsulated in 3D
hydrogels and their phenotype in 3D in vitro culture was compared to that of in vivo tissue specimens. After 4 weeks in 3D culture in osteogenic medium, Fgfr2+/S252W cells expressed 2.8-fold more
collagen type I and 3.3-fold more
osteocalcin than did wild-type controls (p<0.01). Meanwhile, Fgfr2+/S252W cells showed decreased bone matrix remodeling and expressed 87% less Metalloprotease-13 and 71% less Noggin (p<0.01). The S252W mutation also led to significantly higher production of
collagen type I and II in 3D as shown by immunofluorescence staining. In situ hybridization and
alizarin red S staining of postnatal day 0 (P0) mouse limb sections demonstrated significantly higher levels of
osteopontin expression and mineralization in Fgfr2+/S252W mice. Complementary to in vivo findings, this 3D
hydrogel culture system provides an effective in vitro venue to study the pathogenesis of
Apert syndrome caused by the analogous mutation in humans.