Titania nanotube-based protein delivery system to inhibit cranial bone regeneration in Crouzon model of craniosynostosis

Abstract

Background

Craniosynostosis is a developmental disorder characterized by the premature fusion of skull sutures, necessitating repetitive, high-risk neurosurgical interventions throughout infancy. This study used protein-releasing Titania nanotubular implant (TNT/Ti) loaded with glypican 3 (GPC3) in the cranial critical-sized defects (CSDs) in Crouzon murine model (Fgfr2^c342y/+ knock-in mutation) to address a key challenge of delaying post-operative bone regeneration in craniosynostosis.

Materials and methods

A 3 mm wide circular CSD was created in two murine models of Crouzon syndrome: (i) surgical control (CSDs without TNT/Ti or any protein, n=6) and (ii) experimental groups with TNT/Ti loaded with GPC3, further subdivided into the presence or absence of chitosan coating (on nanotubes) (n=12 in each group). The bone volume percentage in CSDs was assessed 90 days post-implantation using micro-computed tomography (micro-CT) and histological analysis.

Results

Nano-implants retrieved after 90 days post-operatively depicted well-adhered, hexagonally arranged, and densely packed nanotubes with average diameter of 120±10 nm. The nanotubular architecture was generally well-preserved. Compared with the control bone volume percentage data (without GPC3), GPC3-loaded TNT/Ti without chitosan coating displayed a significantly lower volume percent in cranial CSDs (P<0.001). Histological assessment showed relatively less bone regeneration (healing) in GPC3-loaded CSDs than control CSDs.

Conclusion

The finding of inhibition of cranial bone regeneration by GPC3-loaded TNT/Ti in vivo is an important advance in the novel field of minimally-invasive craniosynostosis therapy and holds the prospect of altering the whole paradigm of treatment for affected children. Future animal studies on a larger sample are indicated to refine the dosage and duration of drug delivery across different ages and both sexes with the view to undertake human clinical trials.

Keywords:

• craniosynostosis
• protein delivery
• glypican
• titania nanotube
• murine

Acknowledgments

The authors gratefully acknowledge the financial support provided by the Australian Dental Research Foundation (ADRF 18/2013), the Australian Research Council (FT110100711 and IH150100003), and the Australian Craniomaxillofacial Foundation. Ms Ruth Williams from Adelaide Microscopy provided assistance with micro-CT imaging and histological work. Titania nanotubular implants were fabricated at the School of Chemical Engineering, University of Adelaide and at the OptoFab node of the Australian National Fabrication Facility utilizing Commonwealth and South Australia State Government funding.

Disclosure

The authors declare no conflicts of interest in this work.