Periodontal healing with a preameloblast-conditioned medium in dogs.

Abstract

The predictability of conventional periodontal treatments for damaged periodontal tissue is limited, particularly on the regeneration of new cementum. As signaling molecules, a range of growth factors has been used to promote periodontal regeneration on periodontal ligament (PDL) and cementum defects. A preameloblast-conditioned medium (PA-CM) was prepared from cultured murine apical bud cells, which can differentiate into ameloblasts. We examined the effect of PA-CM on PDL cells and cementoblasts invitro and evaluated histologically the effects of PA-CM on the regeneration of experimentally induced periodontal defects invivo. In vitro, the effects of PA-CM on the migration of human PDL cells were examined using a scratch wound healing assay and a transwell assay. The differentiation and mineralization potential of PA-CM-treated human PDL cells and murine cementoblastic OCCM-30 cells was examined by real-time polymerase chain reaction and Alizarin red-S staining. In vivo, six mongrel dogs (12-16kg; 6-8mo old) were used. Twenty-four roots were replanted with either, (i) only periodontal defects (n=12; control group), or (ii) periodontal defects and PA-CM treatment (n=12; experimental group). In the experimental group, the PDL and cementum between notches was removed using a Gracey curette and soaked in 0.08mL water containing 80μg of a PA-CM for 2min. The dogs were killed at 4 and 8wk post-surgery. The invitro results showed that PA-CM stimulated the migration of PDL cells and promoted the differentiation and mineralization of PDL cells and cementoblasts. Real-time polymerase chain reaction analysis revealed stronger expression of Runx2, Osx, OC, Bsp and Cap mRNAs in the PA-CM-treated PDL cells and cementoblasts than those in the control cells. In vivo, newly formed PDL-like tissue and cementum-like tissue were observed partially between the root surfaces and newly formed bone in the experimental group. The regenerated PDL-like tissue in the experimental group was significantly higher than that in the control group at 8wk (p<0.05). The replacement resorption on the experimental group was significantly lower than that in the control group at 8wk (p<0.05). In addition, the amount of newly formed cementum-like tissue in the experimental group was significantly higher than that in the control group at 4 and 8wk (p<0.05). These results suggest that PA-CM has the potential to regenerate periodontal tissues in PDL and cementum defects.