Effects of flutamide therapy on craniofacial growth and development in a model of craniosynostosis.

Research has implicated the faulty regulation of transforming growth factor beta signaling as one mechanism for premature calvaria suture fusion. Androgens have been shown to increase the expression and activity of the transforming growth factor beta, resulting in increased osteoblast proliferation and differentiation and possibly premature suture fusion. The present study was designed to test the hypothesis that flutamide, an androgen receptor-blocking agent, would "rescue" a coronal suture destined to fuse and improve craniofacial growth in a familial rabbit model of craniosynostosis. Thirty rabbits with delayed-onset, coronal suture synostosis were examined via longitudinal cephalometry. The rabbits were divided into 4 groups: (1) sham surgical controls (n = 10), (2) bovine serum albumin (500 ng) protein controls (n = 6), (3) flutamide diluent controls (n = 6), and (4) flutamide (15 mg dissolved in ethanol) experimental group (n = 8). At 10 days of age, radiopaque amalgam markers were implanted in all rabbits on either side of the coronal suture to monitor sutural growth. At 25 days of age, the bovine serum albumin, ethanol, and flutamide were combined with a slow-resorbing collagen vehicle and injected subperiosteally above the coronal suture into the respective groups. Although results revealed a slight but significant increase in coronal suture marker separation in flutamide-treated rabbits compared with controls at 42 days of age, few significant differences were noted for craniofacial growth and intracranial volume among groups. Results suggest that androgen receptor-blocking using flutamide may only provide a transient rescue to suture fusion in this model. Further research is needed to investigate the effects of hormones on suture development and maintenance.

BMP-4 response in wild-type and craniosynostotic rabbit bone cells.

BACKGROUND: Craniosynostosis results from improper regulation of bone formation. Investigations of cells derived from patients with craniosynostosis suggest that craniosynostotic bone-derived cells have increased osteogenic or proliferative capacities compared with other cells. Research into the pathogenesis of craniosynostosis using cells derived from children has been hindered by small sample sizes and inappropriate control cell populations. The authors hypothesized that cells derived from suture-associated regions of bone from craniosynostotic rabbits were more osteogenic and proliferative than bone cells derived from wild-type rabbits. METHODS: This study used cells derived from a colony of rabbits with congenital, nonsyndromic craniosynostosis (n = 20) or from age-matched wild-type rabbits (n = 20). Bone cells derived from either suture-associated or non-suture-associated bone were challenged with osteogenic stimuli and assessed for osteogenic differentiation. RESULTS: The results suggest a high level of variability among cells derived from different individual rabbits. Also, craniosynostotic bone cells have a larger response to recombinant human bone morphogenetic protein 4 stimulation relative to baseline expression of alkaline phosphatase, although overall alkaline phosphatase expression was higher in wild-type bone cells. Cell proliferation showed some differences at 3 days in culture, but no differences were found at 7 days in culture. CONCLUSIONS: This study suggests that bone cells in this rabbit model of craniosynostosis are generally similar to wild-type cells. Also, because of variability, it is necessary to have larger sample sizes than are normally available in human studies. Therefore, cells from the rabbit model may be a powerful in vitro model for further craniosynostosis research.

Inkjet-based biopatterning of bone morphogenetic protein-2 to spatially control calvarial bone formation.

The purpose of this study was to demonstrate spatial control of osteoblast differentiation in vitro and bone formation in vivo using inkjet bioprinting technology and to create three-dimensional persistent bio-ink patterns of bone morphogenetic protein-2 (BMP-2) and its modifiers immobilized within microporous scaffolds. Semicircular patterns of BMP-2 were printed within circular DermaMatrix human allograft scaffold constructs. The contralateral halves of the constructs were unprinted or printed with BMP-2 modifiers, including the BMP-2 inhibitor, noggin. Printed bio-ink pattern retention was validated using fluorescent or (125)I-labeled bio-inks. Mouse C2C12 progenitor cells cultured on patterned constructs differentiated in a dose-dependent fashion toward an osteoblastic fate in register to BMP-2 patterns. The fidelity of spatial restriction of osteoblastic differentiation at the boundary between neighboring BMP-2 and noggin patterns improved in comparison with patterns without noggin. Acellular DermaMatrix constructs similarly patterned with BMP-2 and noggin were then implanted into a mouse calvarial defect model. Patterns of bone formation in vivo were comparable with patterned responses of osteoblastic differentiation in vitro. These results demonstrate that three-dimensional biopatterning of a growth factor and growth factor modifier within a construct can direct cell differentiation in vitro and tissue formation in vivo in register to printed patterns.