Involvement of the endocannabinoid system in periodontal healing.

Endocannabinoids including anandamide (AEA) and 2-arachidonoylglycerol (2-AG) are important lipid mediators for immunosuppressive effects and for appropriate homeostasis via their G-protein-coupled cannabinoid (CB) receptors in mammalian organs and tissues, and may be involved in wound healing in some organs. The physiological roles of endocannabinoids in periodontal healing remain unknown. We observed upregulation of the expression of CB1/CB2 receptors localized on fibroblasts and macrophage-like cells in granulation tissue during wound healing in a wound-healing model in rats, as well as an increase in AEA levels in gingival crevicular fluid after periodontal surgery in human patients with periodontitis. In-vitro, the proliferation of human gingival fibroblasts (HGFs) by AEA was significantly attenuated by AM251 and AM630, which are selective antagonists of CB1 and CB2, respectively. CP55940 (CB1/CB2 agonist) induced phosphorylation of the extracellular-regulated kinases (ERK) 1/2, p38 mitogen-activated protein kinase (p38MAPK), and Akt in HGFs. Wound closure by CP55940 in an in-vitro scratch assay was significantly suppressed by inhibitors of MAP kinase kinase (MEK), p38MAPK, and phosphoinositol 3-kinase (PI3-K). These findings suggest that endocannabinoid system may have an important role in periodontal healing.

Effectable application of vascular endothelial growth factor to critical sized rat calvaria defects.

OBJECTIVE: An early vascular response for angiogenesis is essential for the normal progression of bone defect healing. Vascular endothelial growth factor (VEGF) is a potent inducer of angiogenesis. The aim of this study was to evaluate the effects of a poly (L,D-lactic-co-glycolic acid) (PLGA) membrane with VEGF encapsulated into PLGA microspheres on bone regeneration at bone defects in rat calvaria. STUDY DESIGN: Microspheres of PLGA incorporating VEGF(165) (VEGF microspheres) were prepared, and critical-size bone defects were created in rat calvaria. The VEGF microspheres, PLGA microspheres, or VEGF microspheres plus PLGA membrane were applied to the defects. Bone regeneration was evaluated using image analysis based on soft radiographic and histologic examination. RESULTS: Mature thick bone regeneration was observed in selected sites at bone defects that had been applied with VEGF microspheres/PLGA membrane compared with those that had been applied with the other treatments. CONCLUSION: A combination of VEGF microspheres and a PLGA membrane effectively enhances bone regeneration.

Favorable periodontal healing of 1-wall infrabony defects after application of calcium phosphate cement wall alone or in combination with enamel matrix derivative: a pilot study with canine mandibles.

BACKGROUND: Although various periodontal regenerative therapies are used, their effects on non-contained infrabony defects are unpredictable. Our previous studies showed that injectable, moldable, fast-setting calcium phosphate cement (CPC) promoted histocompatible periodontal healing in 3-wall intrabony defects. The present study evaluated healing patterns after surgical application of CPC walls with and without an enamel matrix derivative (EMD) in 1-wall infrabony defects in dogs. METHODS: One-wall infrabony defects (5 x 5 x 4 mm) were created surgically on the mesial and distal sides of bilateral mandibular fourth premolars in four beagle dogs. After elevating a full-thickness flap, exposed root surfaces were planed thoroughly. The 16 defects were assigned randomly to one of the following experimental conditions: CPC, CPC+EMD, EMD, and open flap debridement (OFD). Ten weeks post-surgery, the animals were sacrificed, and histologic specimens were prepared for histomorphometric evaluation. RESULTS: Defect sites treated with EMD only exhibited varying degrees of new cementum and new bone formation, whereas the OFD group presented only limited new cementum and bone formation. Defect sites where a CPC wall was implanted (CPC and CPC+EMD groups) revealed significantly greater regeneration of new bone and new cementum than in the EMD and OFD groups. No significant differences were observed between the CPC and CPC+EMD groups. CONCLUSIONS: CPC walls with and without EMD promoted regeneration of alveolar bone and cementum in 1-wall infrabony defects. Space and stable wound healing are believed to be crucial for periodontal regeneration in non-contained infrabony defects.

Recombinant human growth/differentiation factor-5 (rhGDF-5) induced bone formation in murine calvariae.

OBJECTIVES: Growth/differentiation factor-5 (GDF-5), a member of the transforming growth factor-beta superfamily, shows a close structural relationship to bone morphogenetic proteins and plays crucial roles in skeletal morphogenesis. Recombinant human (rh) GDF-5 was reported as a suitable factor for enhancing healing in bone defect and inducing ectopic bone formation. The purpose of the present study was to investigate the mechanism of bone formation induced by rhGDF-5 in murine calvariae by radiological, histological and immunohistochemical methods. Cell proliferation was also examined in vitro. MATERIAL AND METHODS: Cells including primary osteoblasts, periosteum cells and connective tissue fibroblasts were isolated enzymatically from neonatal murine calvariae or head skin. In the presence or absence of rhGDF-5, cell proliferation was estimated by tetrazolium reduction assay. To examine the mechanism of osteoinduction, rhGDF-5/atelocollagen (AC) composite or 0.01 N HCl/AC composite were injected into murine calvariae subcutaneously. Tissue was examined radiologically, histologically and immunohistochemically. RESULTS: In the presence of rhGDF-5, proliferation of primary osteoblasts, periosteum cells, and connective tissue fibroblasts was increased significantly in culture. Immunohistochemical observations showed cells at the site injected with rhGDF-5/AC displayed immunoreactivity for proliferating cell nuclear antigen (PCNA). Newly formed bone- and cartilage-like tissue contained chondrocyte osteocyte and osteoclastic cells, and were immunoreactive for both type I and II collagen. CONCLUSION: Exposure to GDF-5 promotes proliferation and differentiation of calvarial cells, which give rise to ectopic bone formation.

[Clinical study of congenital cholesteatoma of the middle ear].

We studied 35 ears of 34 patients with congenital cholesteatoma who were operated between June 1992 and May 2003, focusing on the localization and progression of congenital cholesteatoma. Patients were 2 to 55 years of age. Congenital cholesteatoma was diagnosed based on two intraoperative findings: 1) no continuity between the tympanic membrane and cholesteatoma, 2) no presence of perforation, retraction, or granulation of the tympanic membrane. All patients had closed-type cholesteatoma, and none formed open-type cholesteatoma, which grows as a flat sheet of epidermis. The primary site of congenital cholesteatoma was classified into 3 types; 1) anterior-superior quadrant, 2) posterior-superior quadrant, and 3) epitympanic, and the origin of these three types of congenital cholesteatoma was difficult to explain by a single theory. We operated on 31 ears by canal wall up tympanoplasty and on 4 ears by canal wall down tympanoplasty. On all but 4 ears, we performed planned-staged operation, including second-look operations, in case of recurrence or residual cholesteatoma. Improvement in hearing after surgery was seen in 22 of the 26 ears treated and followed up for more than 6 months after surgery. By drawing all localization of congenital cholesteatoma in 35 ears, we studied the progression of congenital cholesteatoma and speculated on the original primary site. Congenital cholesteatoma in restricted areas consequently implies good results in hearing after surgery, making earlier diagnosis and treatment desirable.