Methods: hDPCs were cultured and transfected with adenoviral mediated human Shh gene (AdShh). Overexpression of Shh and cell proliferation was tested by real-time PCR analysis and MTT analysis, respectively, while odontoblastic differentiation was assessed by alkaline phosphatase (ALP) activity and real-time PCR analysis on markers of Patched-1 (Ptc-1), Smoothened (Smo), Gli 1, Gli 2, Gli 3, osteocalcin (OCN), dentin matrix protein-1 (DMP-1), and dentin sialophosphoprotein (DSPP). Finally, AdShh transfected hDPCs were combined with porous CPC and placed subcutaneously in nude mice for 8 and 12 weeks, while AdEGFP transfected and untransfected hDPCs were treated as control groups.
Results: Results indicated that Shh could promote proliferation and odontoblastic differentiation of hDPCs, while Shh/Gli 1 signaling pathway played a key role in this process. Importantly, more mineralized tissue formation was observed in combination with AdShh transfected hDPCs and porous CPC, moreover, the mineralized tissue exhibited dentin-like features such as structures similar to dentin-pulp complex and the postive staining for OCN, DMP-1 and DSPP protein.
Conclusions: These results suggested that Shh could induce odontoblastic differentiation of DPCs, while Shh/Gli 1 signaling pathway might play an important role in this process. More importantly, the constructs with Shh gene modified DPCs and porous CPC may be a better alternative for dental tissue regeneration.