Delta 12-PGJ 2 stimulates bone formation by PDGE-mediated mechanism

Abstract

The aims of the present study were to establish a new model for the study of biological events during bone regeneration using the principle of guided tissue regeneration and to study the effect of delta12-Prostaglandin J2 (delta12-PGJ2) on bone formation. Half-cylindrical shaped titanium tubes (8x1.5x2 mm) were placed and secured over transcortical defects (5x1.5 mm) at femoral bone in male Wistar rats. Healing periods of 3, 7, 21, 28 days were studied. Animals were injected with 5-Bromo-2'deoxy-uridine (BrdU) one hour before sacrifice in order to characterize cellular mitotic activity. Samples were processed for routine paraffin embedding, then stained with either H&E or immunostained for COX-2, PDGF-B, IGF and BMP-6 expression. Bone regeneration occurred in both cortical defect area and supracortical area at day 7. The amount of the area of new bone was increased during healing period and reached about 56-60% of the defect area by day 28. Mitotic activity was found dominantly in each defect area before a large amount of bone would take placed. COX-2, PDGE-B and BMP-6, highly expressed early at day 3, were found to be important in the early stage of bone regeneration. COX-2 inhibitor (NS-398) decreased the area of new bone by 25% (p<0.05) and both COX-2 and BMP-6 expression by 50-60% (p<0.05) in this model. This study shows the early role of prostaglandins and some key growth factors in bone regeneration. The effect of a single application of delta12-PGJ2 on bone formation was studied in this model of bone regeneration. Four groups of the animals received a different concentration of delta12-PGJ2 (10 -9, 10 -7, 10 -5 and 10 -3 M). Ten mul of delta12-PGJ2 were loaded into a collagen sponge and then was placed inside the titanium chamber before placing on top of the defect. The contralateral defect served as a control without delta12-PGJ2. The animals were sacrificed 10 days after surgery. The amount of new boneformed and the proliferation rate in the cortical defects were determined histomorphometrically. The expression of PDGF, IGF, BMP-2 and BMP-6 was examined immunohistochemically in new bone regeneration. Bone formation in the defect area was significantly promoted by delta12-PGJ2 in a dosedependent manner. It was increased by 35% when the 10 -5 M concentration (54% of the defect area) or the 10 -3 M concentration (53%) were used, as compared with the bone formation observed with the control (40%). Statistically significant increases (2-3 folds) inthe area of PDGF-A and -B expression were seen at high doses of delta12-PGJ2. A significant increase in BMP-2 and BMP-6 expression (2-3 folds) was seen in the group receiving 10 -3 M of delta12-PGJ2. No significant difference was found in IGF expression and mitotic activity between the treatment and control groups. Single dose of delta12-PGJ2 (1x10 -5 and 1x10 -3 M) was also investigated for potential use in bone regeneration around implant. Screw-shaped titanium implants (1.35 mm in diameter and 1.5 mm in length) were inserted to femoral bones (2 implants for each femur). Five mul of carboxymethylcellulose, as carrier, alone (for control) or with delta12-PGJ2 were injected into surgically prepared holes before the insertion of implants. Rats were sacrificed at 3 and 8 weeks after surgery. The femoral bones with implants were embedded in methyl methacrylate resin and were prepared to make undecalcified sections. delta12-PGJ2 increased about 20% more new bone area in the space between thread at 8 week after surgery. Both vertical distance and the thickness of extended new bone regeneration surrounding the implant from endosteal surface were significantly increase (34% and 59% respectively) with delta12-PGJ2 at 8 week after surgery. The present findings indicate that a single application of delta12-PGJ2 promotes new bone formation in this model system. Moreover, PDGF-A and-B may be involved in the mechanism of delta12-PGJ2-induced bone formation as well as BMP-2 and -6