Joji Fukunaga

Osteogenic potential of primed periosteum graft in the rat calvarial model.

Repair of bone defects remains a major concern in plastic and maxillofacial surgery. Based on modern concepts of tissue engineering, periosteum has gained attention as a suitable osteogenic material. We tested the hypothesis that surgically released and immediately repositioned periosteum would exhibit high osteogenic capacity upon grafting in a rat calvarial defect. Seven days after periosteum was released from the tibia and immediately repositioned, the “primed periosteum graft” (PPG; n = 15) was placed into a critical-sized defect of rat calvaria and the process of bone formation was evaluated histologically, immunohistologically, and radiographically at 7, 14, and 21 days after grafting. Findings were compared with a nonprimed periosteal graft (NPG; n = 15). Endochondral ossification was observed in both the PPG and NPG. The PPG showed higher expression of proliferative cell nuclear antigen, bone morphogenetic protein, and vascular endothelial growth factor than the NPG. Three-dimensional radiographic examination revealed significantly increased bone formation in the PPG than in the NPG (P < 0.01). These findings suggested that surgical stimulation of the periosteum enhanced the osteogenic potential of periosteal cells. This method may be suitable for the clinical repair of bone defects.

Immobilized recombinant human bone morphogenetic protein-2 enhances the phosphorylation of receptor-activated Smads.

Bone morphogenetic protein (BMP)-2 plays an important role in bone growth and regeneration; however, BMP-2 is easily lost by diffusion through body fluid and has some inhibitory pathways. To address this problem, we previously immobilized recombinant human BMP-2 (rhBMP-2) on succinylated type I atelocollagen. Here, we examined the effect of immobilized rhBMP-2 in vitro and vivo. In ST2, MC3T3-E1, and C2C12 cells, alkaline phosphatase activity, which is a marker of osteoblast differentiation, was enhanced more by immobilized than nonimmobilized rhBMP-2. In addition, the phosphorylation of receptor-activated Smads, part of the signaling pathway activated by BMP-2, was prolonged by immobilized rhBMP-2 in these cells. Furthermore, implantation of immobilized rhBMP-2 into the backs of rats promoted the formation of mature bone-like structure. These results demonstrate that immobilized rhBMP-2 has higher bioactivity than nonimmobilized rhBMP-2, and, therefore, immobilization of rhBMP-2 can prolong BMP signaling.

Evaluation of osteogenic/chondrogenic cellular proliferation and differentiation in the xenogeneic periosteal graft.

To determine whether grafted young periosteum can induce new bone formation in elderly patients, this preliminary study evaluated cell proliferation and differentiation in xenogeneic periosteal grafts in old rats radiographically, histologically, and immunohistochemically. Periosteum harvested from the tibia of young Japanese white rabbits were grafted into old Sprague–Dawley rats with or without administration of 1.0 mg per kilogram per day immunosuppressant FK506. Autogenous old periosteal tissue grafts were also evaluated as a control. Grafted tissue was extirpated after 7, 14, 21, and 45 days. In the xenogeneic group, proliferative cell nuclear antigen-positive cells were observed 7 days after surgery, which differentiated into chondroblasts with bone morphogenetic protein-2 expression and finally formed cartilage by 14 days. Endochondral ossification was observed at 21 days, and bone replacement was completed by 45 days. No osteogenic cell activity was observed in the two other groups. Xenogeneic young periosteum thus maintained its osteogenic/chondrogenic potentiality in older rats.

Immunolocalization of vascular endothelial growth factor during heterotopic bone formation induced from grafted periosteum

Vessel invasion is an important step in cartilage replacement that leads to bone formation, and vascular endothelial growth factor (VEGF) has been implicated as a key player in this process. Although grafted periosteum undergoes endochondral ossification, little is known about the role of VEGF in this process. In the current study the authors investigated by immunohistochemical, histochemical, and ultrastructural techniques the localization of VEGF during bone formation in periosteal grafts. At day 14 after grafting the tibias of Japanese white rabbits, periosteal cells in the grafted tissue had differentiated into chondrocytes to form cartilage. Some chondrocytes were immunopositive for VEGF expression, and subsequent vessel invasion occurred predominantly in these VEGF-positive areas. At day 45, the cartilage invaded by blood vessels had been replaced by newly formed bone. These findings suggest that VEGF is associated with the process of blood vessel invasion into cartilage before bone replacement in endochondral ossification from grafted periosteum.

Pathology of the temporomandibular joint of patients with rheumatoid arthritis–case reports of secondary amyloidosis and macrophage populations

INTRODUCTION The pathogenetic features of rheumatoid arthritis of the temporomandibular joint (TMJ) are not well defined. In this paper the histological features of TMJs affected by rheumatoid arthritis, and the detection of secondary amyloidosis and macrophage populations in the TMJs of two patients with progressive rheumatoid arthritis are described. METHODS In two patients (64-year-old man and 61-year-old woman) with rheumatoid arthritis total TMJ replacement were performed. The surgical specimens were studied histologically. RESULTS It was found that the articular cartilage had been completely replaced by proliferating fibrous tissue. Congo red staining and polarizing microscopy revealed amyloid deposition in the connective tissue of the joint space. Immunohistochemical staining showed CD 68 positive macrophages around the amyloid deposition in the proliferating soft tissue. CONCLUSION TMJ involvement in rheumatoid arthritis followed the same destructive pathway as in other joints. Amyloid deposition and macrophage populations were detected in two TMJs affected by rheumatoid arthritis.

Microcystic adnexal carcinoma in the mental region

Abstract Microcystic adnexal carcinoma is a rare tumour, first described in 1982. Prior to the recognition of microcystic adnexal carcinoma, instances of unusual basal cell carcinoma, sweat gland carcinoma, and trichofolliculoma had been reported. Microcystic adnexal carcinomas are slow growing and demonstrate aggressive local behaviour and a high recurrence rate. The mainstay of treatment is surgical excision using frozen sections to check the peripheral and deep margins. This report is of a microcystic adnexal carcinoma in the mental region of a 69-year-old woman.

Presence of Human β-Defensin-2 in Oral Lichen Planus and its Histamine Releasing Effect

Abstract Objective: To determine by immunohistochemical means the localization of human β-defensin-2, a peptide with antimicrobial activity, in oral lichen planus. The release of histamine from mast cells elicited by human β-defensin-2 was also investigated. Materials and Methods: Biopsy specimens of oral lichen planus, synthetic human β-defensin-2, human α-defensin-1, and mast cells isolated from Sprague Dawley rats were used. Tissue sections were embedded in paraffin and immunostained by the streptavidin-biotin-coupled peroxidase method. Isolated rat mast cells were injected with synthetic human β-defensin-2 and human α-defensin-1. Evans blue dye was injected into the tail vein of Sprague Dawley rats, followed by various doses of histamine, human β-defensin-2, human α- defensin-1, and saline. Results: Epithelial cells in lichen planus from the corneal layer to the spinous layer, were positively stained with anti-human β-defensin-2 antibody. Mast cells in subepithelial areas were also stained by anti-human β-defensin-2 antibody. Human β-defensin-2-induced histamine release from the isolated rat mast cells occurs in a dose-dependent manner. When human β-defensin-2 was injected into rat skin intradermally, the vascular permeability increased. These responses were completely abolished upon injection of the antihistamine drug diphenhydramine hydrochloride. Conclusion: Human β-defensin-2 and histamine may play an important role in the formation of oral lichen planus.

Histamine Release from Rat Mast Cells Induced by Human α-Defensin-1 Present in Jaw Cyst Fluid

Abstract Objective: To investigate the release of histamine from rat mast cells elicited by human α-defensin-1 contained in jaw cyst fluid. Patients and Methods: Human α-defensin-1 was purified by reversed-phase high-performance liquid chromatography from human jaw cyst fluid, and the mast cells were collected from the peritoneal cavities of rats. Results: Histamine release was induced in isolated rat mast cells by both extracted and synthetic human α-defensin-1. Changes in vascular permeability induced by human α-defensin-1 were also estimated by the passive skin test in rats. Extracted and synthetic human α-defensin-1 induced histamine release from isolated rat mast cells in a dose-dependent manner over a concentration range of 1 to 15 μg/mL; the histamine release induced by the extracted and synthetic human α-defensin-1 (15 μg/mL) was 39.4 ± 2.3 and 37.6 ± 1.8%, respectively. When human α-defensin-1 was injected into rat skin intradermally, the vascular permeability increased. This response occurred with as little as 0.1 μg of human α-defensin-1, and a strong response was observed with 0.5 and 1.0 μg. These responses were completely abolished upon injection with 1 μg of the antihistamine drug diphenhydramine hydrochloride. Conclusion: Human α-defensin-1 in human jaw cyst fluid can act as an inducer of histamine release from mast cells both in vivo and in vitro.