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Distraction osteogenesis (DO) is one of the most promising reconstructive methods for repairing large craniofacial defects or growth deficiencies through bone regeneration, but it is also a challenge because of an undesirably long process and its complications, which limit its application in clinical practice. The transplantation of mesenchymal stem cells (MSCs) is regarded as an innovative approach to accelerate bone regeneration. Dental pulp stem cells (DPSCs) have shown some advantages over other human adult MSCs, and DPSCs have been regarded as one of the most promising cell sources used in the endogenous tissue engineering. Furthermore, using stem cells modified by gene engineering in DO has been reported in previous studies. It has been shown that Sirtuin-1 (SIRT1) can directly regulate the differentiation of MSCs into osteoblasts. In this study, DPSCs expressing SIRT1 were prepared and their effects on the new bone formation were further investigated in rabbits with tibia. Rabbits were injected with the adenovirus (Adv)-SIRT1-green fluorescent protein (GFP)-transfected DPSCs (overexpression group, Group OE), Adv-GFP transfected DPSCs (negative control group, Group NC) or physiologic saline (control group, Groups CON) into the distraction gap. The new bone tissues in the distraction gap were harvested 8 weeks later, and subjected to by radiographic examination, micro-CT evaluation, and histological and mechanical testing. The better bone formation, the highest bone mineral density (BMD) and the highest bone mineral content (BMC) were observed in the OE group. In conclusion, SIRT1-modified DPSCs in DO was more effective to promote new bone formation during DO, which provides evidence for further investigation about the role of of SIRT1 in the DO.