Enhancing Calvarial Regeneration Through Inhibition Of TGF-Beta 1 Signaling

Background:  Given the significant biomedical burden posed by calvarial defects¹ and the limitations of contemporary craniofacial skeletal reconstructive approaches, a need exists to better understand the biology of calvarial osteoblasts and how to direct them towards endogenous bone for regeneration. The mammalian skull vault is the result of a tightly regulated evolutionary process in which components of disparate embryonic origin are integrated². Studies with transgenic mice with a Wnt1-Cre construct which marks for neural crest cells and a conditional reporter gene R26R have firmly established that the frontal bones arise from neural crest cells whereas the parietal bones are derived from the paraxial mesoderm³. We have previously shown that frontal neural crest-derived osteoblasts possess greater osteogenic potential relative to parietal bone derived cells⁴.  This is, in part, due to increased TGF-β1 signaling in parietal cells which is known to promote apoptosis in a number of different cell types⁵.  We therefore investigated the potential for modulation of TGF-β1 signaling to enhance calvarial regeneration.

 Methods:  Non-critical calvarial defects (2mm) were created in both the frontal and parietal bones of wild-type CD-1 mice.  A collagen sponge was then used to deliver either human recombinant TGF-β1 (400ng) or the TGF-β signaling pathway inhibitor SB431542 (26 mM) into the defects.  Micro-Computed Tomography was employed weekly to evaluate bone regeneration up to a final time point of eight weeks and percent osseous healing was calculated using the GE Microview program for each defect location and treatment group.

 Results:  For parietal bone defects, delivery of TGF-β1 significantly reduced the percentage healing relative to non-treated defects.  In contrast, inhibition of TGF-β1 signaling using SB431542, a small molecule which specifically inhibits TGF-β1 mediated signaling, significantly enhanced bone regeneration in parietal bone defects (*p<0.05) and engendered a more ‘frontal like’ healing capacity at early time-points.

 Conclusions:  We demonstrate that increased TGF-β1 signaling impairs calvarial healing whereas inhibition of this signaling pathway promotes bone regeneration, potentially through inhibition of apoptotic activity.  This study provides an insight into the potential use of specific small molecule inhibitors of TGF-β signaling as a novel therapeutic approach that could be used in conjunction with currently available treatments. This may allow for future treatment of larger, more complex craniofacial skeletal defects.