Cranial neural crest-derived cells participate in craniofacial skeletal repair

Abstract Introduction: Similarities between skeletal formation and repair have led to the theory that adult fracture repair may parallel development. We hypothesize that since the mandible is derived from cranial neural crest (CNC) and forms via intramembranous ossification, non-stabilized mandibular fractures will heal in a similar fashion. Methods: Non-stabilized osteotomies were created in the right hemi-mandible of adult male mice. Mandibles were harvested at 7, 14, 21, and 28 days and processed for histology. Ossification was assessed and the presence of cartilage determined using histology and in situ hybridization (ISH). To determine the embryologic origins of the regenerate, transgenic mice were generated where CNC cells express GFP constitutively under the Wnt-1 promoter. Fractures were created as described above and processed for frozen section. CNC cells were localized using immunohistochemistry (IHC) for GFP. Closed, non-stabilized tibial fractures were also performed to determine the presence, if any, of CNC-derived cells in appendicular skeletal fractures. Results: In contrast to non-stabilized fractures of the appendicular skeleton, mandibular fractures healed predominantly via intramembranous ossification, with minimal and delayed cartilage production seen via histology and collagen2 ISH. IHC for GFP revealed that the mandibular fracture callous and regenerate was comprised primarily of GFP+, CNC-derived cells. In contrast, the tibial fracture callous was devoid of any CNC-derived cells. Conclusions: These data suggest that mandibular fractures heal by recapitulating their developmental program. These findings have important implications for designing appropriate cell-based approaches to craniofacial regeneration.