Local application of alendronate controls bone formation and ß-tricalcium phosphate resorption induced by recombinant human bone morphogenetic protein-2.

This study examined the ability of local alendronate (ALN) administration to control ß-tricalcium phosphate (ß-TCP) resorption as well as the induction of bone formation by recombinant human bone morphogenetic protein-2 (rhBMP-2). A 15-mm critical-sized bone defect was created in the diaphysis of rabbit ulnae. Nine female rabbits (4 to 5 months-old) were divided into 3 groups. Group 1 (n = 6 ulnae) animals received implants consisting of ß-TCP granules and 25 µg of rhBMP-2 in 6.5% collagen gel. Group 2 (6 ulnae) and Group 3 (6 ulnae) animals received the same implants, but with 10-6 M and 10-3 M ALN-treated TCP granules, respectively. Two weeks postsurgery, tartrate-resistant acid phosphatase-positive cell counts, new bone formation, and residual ß-TCP were evaluated. This study showed that a high dose of ALN strongly reduced osteoclastic resorption of ß-TCP induced by rhBMP-2, resulting in decreased bone formation. In contrast, a low dose of ALN slightly reduced the bone resorptive effect but increased bone formation. These results suggest that osteoclast-mediated resorption plays an important role in bone formation and a coupling-like phenomenon could occur in the ß-TCP-implanted area, and that administration of a low dose of ALN may solve clinical bone resorptive problems induced by rhBMP-2.

Effects of micro-porosity and local BMP-2 administration on bioresorption of ß-TCP and new bone formation.

BACKGROUND: It has been reported that the microporous structure of calcium phosphate (CaP) ceramics is important to osteoconduction. Bone morphogenetic protein-2 (BMP-2) has been shown to be a promising alternative to bone grafting and a therapeutic agent promoting bone regeneration when delivered locally. The aim of this study was to evaluate the effects of micro-porosity within beta-tricalcium phosphate (ß-TCP) cylinders and local BMP-2 administration on ß-TCP resorption and new bone formation. METHODS: Bilateral cylindrical bone defects were created in rabbit distal femora, and the defects were filled with ß-TCP. Rabbits were divided into 3 groups; defects were filled with a ß-TCP cylinder with a total of approximately 60% porosity (Group A: 13.4% micro- and 46.9% macropore, Group B: 38.5% micro- and 20.3% macropore, Group C: the same micro- and macro-porosity as in group B supplemented with BMP-2). Rabbits were sacrificed 4, 8, 12, and 24 weeks postoperatively. RESULTS: The number of TRAP-positive cells and new bone formation in group B were significantly greater than those in group A at every period. The amount of residual ß-TCP in group C was less than that in group B at all time periods, resulting in significantly more new bone formation in group C at 8 and 12 weeks. The number of TRAP-positive cells in group C was maximum at 4 weeks. CONCLUSIONS: These results suggest that the amount of submicron microporous structure and local BMP-2 administration accelerated both osteoclastic resorption of ß-TCP and new bone formation, probably through a coupling-like phenomenon between resorption and new bone formation.

3D electrochemical and ion current imaging using scanning electrochemical-scanning ion conductance microscopy.

Local cell-membrane permeability and ionic strength are important factors for maintaining the functions of cells. Here, we measured the spatial electrochemical and ion concentration profile near the sample surface with nanoscale resolution using scanning electrochemical microscopy (SECM) combined with scanning ion-conductance microscopy (SICM). The ion current feedback system is an effective way to control probe-sample distance without contact and monitor the kinetic effect of mediator regeneration and the chemical concentration profile. For demonstrating 3D electrochemical and ion concentration mapping, we evaluated the reaction rate of electrochemical mediator regeneration on an unbiased conductor and visualized inhomogeneous permeability and the ion concentration 3D profile on a single fixed adipocyte cell surface.

Densified electrochemical sensors based on local redox cycling between vertically separated electrodes in substrate generation/chip collection and extended feedback modes.

A new local redox cycling-based electrochemical (LRC-EC) device integrated with many electrochemical sensors has been developed into a small chip device. The LRC-EC chip device was successfully applied for detection of alkaline phosphatase and horseradish peroxidase activity in substrate generation/chip collection (SG/CC) and extended feedback modes, respectively. The new imaging approach with extended feedback mode was particularly effective for sharpening of the image, because this mode uses feedback signals and minimizes the undesired influence of diffusion. The LRC-EC chip device is considered to be a useful tool for bioanalysis.

Bone formation and resorption in patients after implantation of beta-tricalcium phosphate blocks with 60% and 75% porosity in opening-wedge high tibial osteotomy.

Most of the implanted porous beta-tricalcium phosphate (beta-TCP) can be resorbed. However, beta-TCP block with 75% porosity is inadequate for weight-bearing sites until bone incorporation occurs. Thus, the authors have recently developed beta-TCP block with 60% porosity, which is approximately sevenfold greater in terms of compressive strength than that of beta-TCP with 75% porosity. The authors investigated bone formation and resorption of beta-TCP after implantation in patients of beta-TCP blocks with two different porosities. From May 2003 to November 2004, medial opening high tibial osteotomy was performed in 25 patients with a mean age of 66 years. The opened defect was fixed with a Puddu plate. Then 6-8 cm(3) of beta-TCP block with 75% porosity was used to fill the cancellous bone defect, except on the medial side where 2.83-3.18 cm(3) of wedge-shaped beta-TCP block with 60% porosity was implanted. At least 2 years after surgery, the 25 patients had no correction loss, and bone formation was noted in all cases. Complete or nearly complete resorption of beta-TCP with 60 and 75% porosity was obtained within 3.5 years. Thirteen biopsy samples obtained from the 60% porosity implantation sites showed good lamellar bone formation, and the percentage of beta-TCP remaining relative to the newly formed bone plus beta-TCP ranged from 0.3 to 14.5%, with a mean of 6.7%. The authors suspect that mechanical stress loading to the medial side of the tibia facilitated bone formation and resorption of beta-TCP with 60% porosity.

Repair of segmental bone defects in rabbit tibiae using a complex of beta-tricalcium phosphate, type I collagen, and fibroblast growth factor-2.

The objective of this study was to evaluate the effects of a complex of beta-tricalcium phosphate (beta-TCP) granules, collagen, and fibroblast growth factor-2 (FGF-2) on cortical bone repair in rabbits. Segmental bone defects of 5 mm in length were created in the middle of the tibial shaft. The defect was stabilized with a plate and screws, and was filled with 0.3 ml of a complex of beta-TCP granules and 5% collagen, with or without 200 microg of recombinant human fibroblast growth factor-2 (rhFGF-2). Bone regeneration and beta-TCP resorption were assessed by X-ray and micro-CT scanner. A three-point bending test was also performed. The results showed that the segmental bone defect was not only radiologically, but also mechanically healed with cortical bone 12 weeks after implantation of the complex with rhFGF-2. In contrast, after implantation of the complex without rhFGF-2, most of the defect was filled with beta-TCP and only a small amount of bone formation was found. These results suggest that resorption of beta-TCP is important for bone formation and may be promoted by FGF-2 in the beta-TCP implantation site. In addition, the complex of beta-TCP granules and collagen combined with rhFGF-2 provides a paste-like material that is easy to handle. This material may be of considerable use in the treatment of cortical bone defects.

Use of a biphasic graft constructed with chondrocytes overlying a beta-tricalcium phosphate block in the treatment of rabbit osteochondral defects.

To evaluate the ability of a biphasic construct to repair osteochondral defects in articular cartilage, plugs made of chondrocytes in collagen gel overlying a resorbable porous beta-tricalcium phosphate (TCP) block were implanted into defects in rabbit knees. The repair tissue was evaluated at 8, 12, and 30 weeks. Eight weeks after implantation of the biphasic construct, histologic examination showed hyaline-like cartilage formation that was positive for safranin O and type II collagen. At 12 weeks, most of the beta-TCP was replaced by bone, with a small amount remaining in the underlying cartilage. In the cell-seeded layer, the newly formed middle and deep cartilage adjacent to the subchondral bone stained with safranin O, but no staining was observed in the superficial layer. In addition, cell morphology was distinctly different from the deep levels of the reparative cartilage, with hypertrophic cells at the bottom of the cartilaginous layer. At 30 weeks, beta-TCP had completely resorbed and a tidemark was observed in some areas. In contrast, controls (defects filled with a beta-TCP block alone) showed no cartilage formation but instead had subchondral bone formation. These findings indicate that beta-TCP-supported chondrocytes in collagen gel can partially repair isolated articular cartilage osteochondral defects.

Bone formation and bioresorption after implantation of injectable beta-tricalcium phosphate granules-hyaluronate complex in rabbit bone defects.

The objective of this study was to evaluate the effects of a complex of beta-tricalcium phosphate (beta-TCP) granules and 3.5% hyaluronate (beta-TCP granules-HY complex) compared with a beta-TCP block, in terms of osteoconductivity and biodegradability, to determine whether this complex would be a good candidate for bone void filler. Both materials were implanted into cavities drilled in rabbit femoral condyles. New bone formation and mineral apposition rate were evaluated to analyze osteoconductivity, whereas residual beta-TCP within the defects and tartrate-resistant acid phosphatase (TRAP) cellular activity were studied for beta-TCP resorption. The results show that both the beta-TCP block and the beta-TCP granules-HY complex support bone ingrowth; however, bioresorption was rapid for beta-TCP granules-HY but weak for beta-TCP block. This biodegradation mechanism was considered to be a cell-mediated disintegration by numerous TRAP-positive giant cells. The time lag between the peak value of TRAP-positive giant cell population and that of new bone formation rate suggests that a coupling-like phenomenon could be occurring in the beta-TCP-filled bone defects. In addition, beta-TCP granules-HY complex, which is an injectable, pastelike material, has similar osteoconductive properties to beta-TCP block. Thus, this complex may be useful as a bone filler in clinical application.