Development of novel bone-like nanocomposite coating of hydroxyapatite/collagen on titanium by modified electrophoretic deposition.

Electrophoretic deposition (EPD) is a simple, rapid, and inexpensive technique to accomplish uniform coatings with controlled thicknesses. The EPD using binders that do not require a thermal degreasing process, which also eliminates the polymer components of the composite, are required for coating polymer-ceramic composites. This study demonstrated the application of a modified EPD technique utilizing Mg2+ ions to coat a bone-like hydroxyapatite/collagen nanocomposite (HAp/Col) on a titanium (Ti) substrate. The coating thickness was successfully controlled by varying the applied voltage and/or the treatment time. The adhesive strength of the modified EPD coating, evaluated by the tape test, showed class 0 (coating was not peeled off) and drastically increased in comparison to that of the non-Mg2+ EPD coating, class 5 (coating was completely peeled off). The MG63 cells on the HAp/Col-coated Ti demonstrated similar proliferation to and superior alkaline phosphatase activity to that on the bare Ti. Thus, the HAp/Col-coated Ti is expected to facilitate the surrounding bone formation than the bare-Ti. The results of the study indicated the HAp/Col-coated Ti prepared by the modified EPD is effective for applications in novel instruments, such as, subperiosteal temporary anchorage devices, which strongly requires rapid osseointegration at the bone-implant surface.

Fabrication of Gentamicin-Loaded Hydroxyapatite/Collagen Bone-Like Nanocomposite for Anti-Infection Bone Void Fillers.

A gentamicin-loaded hydroxyapatite/collagen bone-like nanocomposite (GNT-HAp/Col) was fabricated and evaluated for its absorption-desorption properties, antibacterial efficacy, and cytotoxicity. The hydroxyapatite/collagen bone-like nanocomposite (HAp/Col) powder was mixed with gentamicin sulfate (GNT) in phosphate-buffered saline (PBS) at room temperature. After 6 h mixing, the GNT adsorption in all conditions reached plateau by Langmuir's isotherm, and maximum GNT adsorption amount was 34 ± 7 µg in 250 µg/mL GNT solution. Saturated GNT-loaded HAp/Col powder of 100 mg was soaked in 10 mL of PBS at 37 °C and released all GNT in 3 days. A shaking culture method for a GNT extraction from the GNT-HAp/Col and an inhibition zone assay for the GNT-HAp/Col compact showed antibacterial efficacy to Escherichia coli (E. coli) at least for 2 days. From the release profile of the GNT from the GNT-HAp/Col powder, antibacterial efficacy would affect E. coli at least for 3 days. Further, no cytotoxicities were observed on MG-63 cells. Thus, the GNT-HAp/Col is a good candidate of bioresorbable anti-infection bone void fillers by prevention initial infections, which is the primary cause of implant-associated infection even for rapid bioresorbable materials.

Enhanced bone formation onto the bone surface using a hydroxyapatite/collagen bone-like nanocomposite.

The process of bone formation onto the bone surface using a hydroxyapatite/collagen bone-like nanocomposite (HAp/Col) was investigated. Immersion tests were performed to evaluate the impact of pH on the degradation of the specimens in an aqueous environment. The specimens were soaked in aqueous solutions of pH 4.0, 5.0, and 7.0. Using standardized images, the top-view areas of the specimens were measured. Animal experiments were performed to investigate the bone formation process onto the bone surface. The specimens were placed under the rat calvarial periosteum, and µCT image analysis and histological observation were performed on samples harvested on postoperative Days 3, 5, and 7. In all experiments, ß-tricalciumphosphate (ß-TCP) was adopted as the control. HAp/Col turned to gel in acidic environments below pH 5.0. In contrast to the ß-TCP, the HAp/Col specimens placed under the periosteum expanded and attained a hollow structure with a gel-filled center, accompanied by larger volume of new bone and appearance of TRAP-positive multinucleated cells on postoperative Day 5. Therefore, HAp/Col can enhance bone formation onto the bone surface via induction of TRAP-positive multinucleated cells, and may have clinical applications.

Preparation of flexible bone tissue scaffold utilizing sea urchin test and collagen.

Gonads of sea urchin are consumed in Japan and some countries as food and most parts including its tests are discarded as marine wastes. Therefore, utilization of them as functional materials would reduce the waste as well as encourage Japanese fishery. In this study, magnesium containing calcite granules collected from sea urchin tests were hydrothermally phosphatized and the obtained granules were identified as approximately 82% in mass of magnesium containing ß-tricalcium phosphate and 18% in mass of nonstoichiometric hydroxyapatite, i.e., a biphasic calcium phosphate, maintaining the original porous network. Shape-controlled scaffolds were fabricated with the obtained biphasic calcium phosphate granules and collagen. The scaffolds showed good open porosity (83.84%) and adequate mechanical properties for handling during cell culture and subsequent operations. The MG-63 cells showed higher proliferation and osteogenic differentiation in comparison to a control material, the collagen sponge with the same size. Furthermore, cell viability assay proved that the scaffolds were not cytotoxic. These results suggest that scaffold prepared using sea urchin test derived calcium phosphate and collagen could be a potential candidate of bone void fillers for non-load bearing defects in bone reconstruction as well as scaffolds for bone tissue engineering.

Preparation of hydroxyapatite/collagen injectable bone paste with an anti-washout property utilizing sodium alginate. Part 1: influences of excess supplementation of calcium compounds.

The anti-washout property, viscosity, and cytocompatibility to an osteoblastic cell line, MG-63, of anti-washout pastes were investigated. Mixing a hydroxyapatite/collagen bone-like nanocomposite (HAp/Col), an aqueous solution of sodium alginate (Na-Alg), which is a paste hardening and lubricant agent, and supplementation of calcium carbonate or calcium citrate (Ca-Cit) as a calcium resource for the hardening reaction realized an injectable bone paste. Adding Ca-Cit at a concentration greater than eight times the Ca2+ ion concentration to Na-Alg improved the anti-washout property. Although the viscosity test indicated a gradual increase in the paste viscosity as the calcium compounds increased, pastes with excess supplementation of calcium compounds exhibited injectability through a syringe with a 1.8 mm inner diameter, realizing an injectable bone filler. Furthermore, the anti-washout pastes with Ca-Cit had almost the same cell proliferation rate as that of the HAp/Col dense body. Therefore, HAp/Col injectable anti-washout pastes composed of the HAp/Col, Na-Alg, and Ca-Cit are potential candidates for bioresorbable bone filler pastes.

Shape Optimization of Bone-Bonding Subperiosteal Devices with Finite Element Analysis.

Subperiosteal bone-bonding devices have been proposed for less invasive treatments in orthodontics. The device is osseointegrated onto a bone surface without fixation screws and is expected to rapidly attain a bone-bonding strength that successfully meets clinical performance. Hence, the device's optimum shape for rapid and strong bone bonding was examined in this study by finite element analyses. First, a stress analysis was performed for a circular rod device with an orthodontic force parallel to the bone surface, and the estimate of the bone-bonding strength based on the bone fracture criterion was verified with the results of an animal experiment. In total, four cross-sectional rod geometries were investigated: circular (Cr), elliptical (El), semicircular (Sc), and rectangular (Rc). By changing the height of the newly formed bone to mimic the progression of new bone formation, the estimation of the bone-bonding strength was repeated for each geometry. The rod with the Rc cross section exhibited the best performance, followed by those with the Sc, El, and Cr cross sections, from the aspects of the rapid acquisition of strength and the strength itself. Thus, the rectangular cross section is the best for rod-like subperiosteal devices for rapid bone bonding.

Interlaboratory studies on in vitro test methods for estimating in vivo resorption of calcium phosphate ceramics.

A potential standard method for measuring the relative dissolution rate to estimate the resorbability of calcium-phosphate-based ceramics is proposed. Tricalcium phosphate (TCP), magnesium-substituted TCP (MgTCP) and zinc-substituted TCP (ZnTCP) were dissolved in a buffer solution free of calcium and phosphate ions at pH 4.0, 5.5 or 7.3 at nine research centers. Relative values of the initial dissolution rate (relative dissolution rates) were in good agreement among the centers. The relative dissolution rate coincided with the relative volume of resorption pits of ZnTCP in vitro. The relative dissolution rate coincided with the relative resorbed volume in vivo in the case of comparison between microporous MgTCPs with different Mg contents and similar porosity. However, the relative dissolution rate was in poor agreement with the relative resorbed volume in vivo in the case of comparison between microporous TCP and MgTCP due to the superimposition of the Mg-mediated decrease in TCP solubility on the Mg-mediated increase in the amount of resorption. An unambiguous conclusion could not be made as to whether the relative dissolution rate is predictive of the relative resorbed volume in vivo in the case of comparison between TCPs with different porosity. The relative dissolution rate may be useful for predicting the relative amount of resorption for calcium-phosphate-based ceramics having different solubility under the condition that the differences in the materials compared have little impact on the resorption process such as the number and activity of resorbing cells. STATEMENT OF SIGNIFICANCE: The evaluation and subsequent optimization of the resorbability of calcium phosphate are crucial in the use of resorbable calcium phosphates. Although the resorbability of calcium phosphates has usually been evaluated in vivo, establishment of a standard in vitro method that can predict in vivo resorption is beneficial for accelerating development and commercialization of new resorbable calcium phosphate materials as well as reducing use of animals. However, there are only a few studies to propose such an in vitro method within which direct comparison was carried out between in vitro and in vivo resorption. We propose here an in vitro method based on measuring dissolution rate. The efficacy and limitations of the method were evaluated by international round-robin tests as well as comparison with in vivo resorption studies for future standardization. This study was carried out as one of Versailles Projects on Advanced Materials and Standards (VAMAS).

Hydroxyapatite/collagen bone-like nanocomposite.

Our group has succeeded to synthesize material with bone-like nanostructure and bone-like inorganic and organic composition via self-organization mechanism between them using simultaneous titration method under controlled pH and temperature. The hydroxyapatite/collagen (HAp/Col) bone-like nanocomposite completely incorporated into bone remodeling process to be substituted by new bone. Cells cultured on the HAp/Col revealed very interesting reactions. Osteoblast-like MG63 cells showed upregulation of alkaline phosphatase >3 times greater than MG63 cells cultured on tissue culture polystyrene (TCPS). MG63 cells 3-dimensionally cultured in a "HAp/Col sponge," a porous HAp/Col having sponge-like viscoelasticity, accumulated calcium phosphate nodules on extracellular matrices they secreted. Bone marrow cells co-cultured with osteoblasts on HAp/Col differentiated to osteoclasts without differentiation supplements. This phenomenon is not found in cells cultured on hydroxyapatite ceramics and TCPS, and rarely in cells cultured on dentin. These results suggest that HAp/Col is a good candidate for tissue engineering of bone as well as bone filler. In a clinical test as a bone filler, the HAp/Col sponge was significantly better than porous ß-tricalcium phosphate. The HAp/Col sponge has been approved by the Japanese government and will be used as greatly needed bone filler in patients. In addition to the above, HAp/Col coating on titanium revealed higher osteo-conductivity than HAp-coated titanium and bare titanium and improved direct bonding between titanium and newly formed bone. The HAp/Col coating may be used for metal devices requiring osseointegration.

Distribution of antimicrobial-resistant lactic acid bacteria in natural cheese in Japan.

To determine and compare the extent of contamination caused by antimicrobial-resistant lactic acid bacteria (LAB) in imported and domestic natural cheeses on the Japanese market, LAB were isolated using deMan, Rogosa and Sharpe (MRS) agar and MRS agar supplemented with six antimicrobials. From 38 imported and 24 Japanese cheeses, 409 LAB isolates were obtained and their antimicrobial resistance was tested. The percentage of LAB resistant to dihydrostreptomycin, erythromycin, and/or oxytetracycline isolated from imported cheeses (42.1%) was significantly higher than that of LAB resistant to dihydrostreptomycin or oxytetracycline from cheeses produced in Japan (16.7%; P=0.04). Antimicrobial resistance genes were detected in Enterococcus faecalis (tetL, tetM, and ermB; tetL and ermB; tetM) E. faecium (tetM), Lactococcus lactis (tetS), Lactobacillus (Lb.), casei/paracasei (tetM or tetW), and Lb. rhamnosus (ermB) isolated from seven imported cheeses. Moreover, these E. faecalis isolates were able to transfer antimicrobial resistance gene(s). Although antimicrobial resistance genes were not detected in any LAB isolates from Japanese cheeses, Lb. casei/paracasei and Lb. coryniformis isolates from a Japanese farm-made cheese were resistant to oxytetracycline (minimal inhibitory concentration [MIC], 32 µg/mL). Leuconostoc isolates from three Japanese farm-made cheeses were also resistant to dihydrostreptomycin (MIC, 32 to >512 µg/mL). In conclusion, the present study demonstrated contamination with antimicrobial-resistant LAB in imported and Japanese farm-made cheeses on the Japanese market, but not in Japanese commercial cheeses.

Hydroxyapatite/collagen nanocomposite-coated titanium rod for achieving rapid osseointegration onto bone surface.

This article proposes less-invasive subperiosteal bone-bonding devices capable of realizing rapid osseointegration and the acquisition of fundamental knowledge required for their development. Three candidates were prepared: titanium rod specimens with a machined surface (Bare), hydroxyapatite (HAp) coating, and hydroxyapatite/collagen (HAp/Col) nanocomposite coating. To investigate bone formation around these rods, each specimen was placed under the periosteum of a male Sprague-Dawley rat calvarium. Four weeks after surgery, the samples were evaluated via histomorphometrical analyses and bonding strength tests. All the Bare specimens and more than half of the HAp specimens were encapsulated with fibrous tissue, whereas all the HAp/Col specimens were almost completely surrounded by new bone tissue without encapsulation. Histomorphometrical analyses showed that the HAp/Col group had the greatest bone contact ratio among all candidates (p < 0.05). Further, a bonding strength test indicated that the HAp/Col group exhibited the greatest bonding strength to bone (p < 0.05). Thus, HAp/Col-coated rods are considered as the best candidate materials for achieving rapid osseointegration onto a bone surface.

Comparing the osteogenic potential of canine mesenchymal stem cells derived from adipose tissues, bone marrow, umbilical cord blood, and Wharton's jelly for treating bone defects.

Alternative sources of mesenchymal stem cells (MSCs) for replacing bone marrow (BM) have been extensively investigated in the field of bone tissue engineering. The purpose of this study was to compare the osteogenic potential of canine MSCs derived from adipose tissue (AT), BM, umbilical cord blood (UCB), and Wharton's jelly (WJ) using in vitro culture techniques and in vivo orthotopic implantation assays. After canine MSCs were isolated from various tissues, the proliferation and osteogenic potential along with vascular endothelial growth factor (VEGF) production were measured and compared in vitro. For the in vivo assay, MSCs derived from each type of tissue were mixed with ß-tricalcium phosphate and implanted into segmental bone defects in dogs. Among the different types of MSCs, AT-MSCs had a higher proliferation potential and BM-MSCs produced the most VEGF. AT-MSCs and UCB-MSCs showed greater in vitro osteogenic potential compared to the other cells. Radiographic and histological analyses showed that all tested MSCs had similar osteogenic capacities, and the level of new bone formation was much higher with implants containing MSCs than cell-free implants. These results indicate that AT-MSCs, UCB-MSCs, and WJ-MSCs can potentially be used in place of BM-MSCs for clinical bone engineering procedures.

Paracrine effect of canine allogenic umbilical cord blood-derived mesenchymal stromal cells mixed with beta-tricalcium phosphate on bone regeneration in ectopic implantations.

BACKGROUND AIMS: The aim of this study was to evaluate the paracrine effects of canine umbilical cord blood (cUCB) mesenchymal stromal cells (MSC) mixed with beta-tricalcium phosphate (beta-TCP) on bone regeneration in ectopic implantation. METHODS: beta-TCP mixed with cUCB MSC (UCB-MSC group), cell lysates (cell lysate group) or a control (control group) were respectively implanted in a subcutaneous pouches in the back of beagle dogs . The implants were harvested 1, 4, 7, 14, 28, 56, 84 days after implantation. Histological findings and stain analyzes of tartrate-resistant acid phosphatase (TRACP) and assays of alkaline phosphatase (ALP) and TRACP were evaluated. The mRNA expression levels of interleukin-1 (IL-1), interleukin-6 (IL-6), tumor necrosis factor-alpha (TNF-alpha), cyclooxygenase-2 (COX-2), vascular endothelial growth factor (VEGF) and transforming growth factor-beta (TGF-beta) were analyzed using semi-quantitative reverse transcription - polymerase chain reactions (RT-PCR). An enzyme-linked immunosorbent assay (ELISA) was used to confirm the protein expression levels of IL-6, COX-2, VEGF and TGF-beta. RESULTS: TRACP-positive cells were observed in all groups 7 days after implantation. ALP and TRACP activities in the UCB-MSC group 84 days after implantation were significantly higher than those of the control (P>0.05). Histologic findings after 84 days showed that the osteoid matrix area in the UCB-MSC group was significantly larger than that of the control (P<0.05). The mRNAs levels of IL-1, IL-6 and VEGF in UCB-MSC and cell lysate groups on day 1 were up-regulated compared with the control. The protein levels of IL-6 and VEGF in the UCB-MSC group at day 1 were significantly higher than that of the other groups (P<0.05). CONCLUSIONS: It is suggested that a significant release of cytokines by cUCB MSC, 1 day following implantation, could enhance bone regeneration.

Characterization of recombinant beta-fructofuranosidase from Bifidobacterium adolescentis G1.

BACKGROUND: We have previously reported on purification and characterization of beta-fructofuranosidase (beta-FFase) from Bifidobacterium adolescentis G1. This enzyme showed high activity of hydrolysis on fructo-oligosaccharides with a low degree of polymerization. Recently, genome sequences of B. longum NCC2705 and B. adolescentis ATCC 15703 were determined, and cscA gene in the both genome sequences encoding beta-FFase was predicted. Here, cloning of cscA gene encoding putative beta-FFase from B. adolescentis G1, its expression in E. coli and properties of the recombinant protein are described. RESULTS: Using the information of cscA gene from Bifidobacterium adolescentis ATCC 15703, cscA gene from B. adolescentis G1 was cloned and sequenced. The N-terminal amino acid sequence of purified beta-FFase from B. adolescentis G1 was identical to the deduced amino acid sequences of cscA gene from B. adolescentis G1. To confirm the translated product of the cscA gene, the recombinant protein was expressed in Escherichia coli. Molecular mass of the purified recombinant enzyme was estimated to be about 66,000 by SDS-PAGE and 60,300 by MALDI TOF-MS. The optimum pH of the enzyme was 5.7 and the enzyme was stable at pH 5.0-8.6. The thermostability of the enzyme was up to 50 degrees C. The K(m) (mM), Vmax (micromol/mg of protein/min), k0 (sec(-1)) and k0/K(m)(mM(-1) sec(-1)) for 1-kestose, neokestose, nystose, fructosylnystose, sucrose and inulin were 1.7, 107, 107.5, 63.2, and 1.7, 142, 142.7, 83.9, and 3.9, 152, 152.8, 39.2, and 2.2, 75, 75.4, 34.3, and 38, 79, 79.4, 2.1, and 25.9, 77, 77.4, 3.0, respectively. The hydrolytic activity was strongly inhibited by AgNO3, SDS, and HgCl2. CONCLUSION: The recombinant enzyme had similar specificity to the native enzyme, high affinity for 1-kestose, and low affinity for sucrose and inulin, although properties of the recombinant enzyme showed slight difference from those of the native one previously described.

Repair of large osteochondral defects in rabbits using porous hydroxyapatite/collagen (HAp/Col) and fibroblast growth factor-2 (FGF-2).

Articular cartilage has a limited capacity for self-renewal. This article reports the development of a porous hydroxyapatite/collagen (HAp/Col) scaffold as a bone void filler and a vehicle for drug administration. The scaffold consists of HAp nanocrystals and type I atelocollagen. The purpose of this study was to investigate the efficacy of porous HAp/Col impregnated with FGF-2 to repair large osteochondral defects in a rabbit model. Ninety-six cylindrical osteochondral defects 5 mm in diameter and 5 mm in depth were created in the femoral trochlear groove of the right knee. Animals were assigned to one of four treatment groups: porous HAp/Col impregnated with 50 microl of FGF-2 at a concentration of 10 or 100 microg/ml (FGF10 or FGF100 group); porous HAp/Col with 50 microl of PBS (HAp/Col group); and no implantation (defect group). The defect areas were examined grossly and histologically. Subchondral bone regeneration was quantified 3, 6, 12, and 24 weeks after surgery. Abundant bone formation was observed in the HAp/Col implanted groups as compared to the defect group. The FGF10 group displayed not only the most abundant bone regeneration but also the most satisfactory cartilage regeneration, with cartilage presenting a hyaline-like appearance. These findings suggest that porous HAp/Col with FGF-2 augments the cartilage repair process.

Osteogenic activity of MG63 cells on bone-like hydroxyapatite/collagen nanocomposite sponges.

The hydroxyapatite/collagen (HAp/Col) sponge with 95% (v/v) porosity was prepared by freeze-drying of a HAp/Col fiber suspension. MG63 cells were seeded onto the HAp/Col sponge and cultured under a pressure/perfusion condition with osteogenic supplements. A collagen (Col) sponge was used as a control. The cells with sponge were examined by a histology, total DNA content and gene expression. The cells showed good attachment and proliferation everywhere in the HAp/Col sponge, while the cells mainly proliferated at the peripheral part of the Col sponge. Thus, total DNA content in the HAp/Col sponges reached 1.8 times greater than that in the Col sponges at Day 21. Further, the cells and extracellular matrix only in the HAp/Col sponge were calcified, although the cells in both sponge evenly expressed osteogenic gene. These results suggest that the HAp/Col sponge could be useful as a scaffold for bone tissue engineering.

Implantation of canine umbilical cord blood-derived mesenchymal stem cells mixed with beta-tricalcium phosphate enhances osteogenesis in bone defect model dogs.

This study was performed to evaluate the osteogenic effect of allogenic canine umbilical cord blood-derived mesenchymal stem cells (UCB-MSCs) mixed with beta-tricalcium phosphate (beta-TCP) in orthotopic implantation. Seven hundred milligrams of beta-TCP mixed with 1 x 10(6) UCB-MSCs diluted with 0.5 ml of saline (group CM) and mixed with the same volume of saline as control (group C) were implanted into a 1.5 cm diaphyseal defect and wrapped with PLGC membrane in the radius of Beagle dogs. Radiographs of the antebrachium were made after surgery. The implants were harvested 12 weeks after implantation and specimens were stained with H&E, toluidine blue and Villanueva-Goldner stains for histological examination and histomorphometric analysis of new bone formation. Additionally, UCB-MSCs were applied to a dog with non-union fracture. Radiographically, continuity between implant and host bone was evident at only one of six interfaces in group C by 12 weeks, but in three of six interfaces in group CM. Radiolucency was found only near the bone end in group C at 12 weeks after implantation, but in the entire graft in group CM. Histologically, bone formation was observed around beta-TCP in longitudinal sections of implant in both groups. Histomorphometric analysis revealed significantly increased new bone formation in group CM at 12 weeks after implantation (p < 0.05). When applied to the non-union fracture, fracture healing was identified by 6 weeks after injection of UCB-MSCs. The present study indicates that a mixture of UCB-MSCs and beta-TCP is a promising osteogenic material for repairing bone defects.

Effects of pore size and implant volume of porous hydroxyapatite/collagen (HAp/Col) on bone formation in a rabbit bone defect model.

A porous hydroxyapatite/collagen composite (HAp/Col) was developed that consists of hydroxyapatite nanocrystals and atelocollagen. In this study, cylindrical (diameter: 5 mm, height: 3 mm) porous HAp/Col implants with different pore sizes (diameter: 160 or 290 microm) were prepared, and the influences of pore size and implanted volume were evaluated using a rabbit bone defect model. In the implant groups, one or three (diameter: 5 mm, total height: 9 mm) implants were transplanted into bone holes created in the anteromedial site of the proximal tibiae, while a group without implantation was used as a control. Histological observation revealed that at two weeks after implantation, bone formation was initiated not only from the periosteum but in regions where the implants bordered on bone marrow. At four weeks, bone formation expanded from the marrow cavity side into the center of the implants, particularly in those implants with large pores. At twelve weeks, four implant groups showed repair of cortical defects and implant absorption, which was thought to be the result of natural bone remodeling mechanisms. The control group showed bone formation developed from the periosteum without bone induction in the marrow cavity, and at four weeks, the bone hole was almost healed. pQCT analysis revealed that the expansion rates of bone tissue were higher in the large-pore implant groups than in the small-pore groups. These data demonstrate the osteoconductivity of porous HAp/Col and the importance of its porous structure.

Material design of bioabsorbable inorganic/organic composites for bone regeneration.

Two new bioabsorbable inorganic/organic composite materials were developed for bone regeneration. One material used was beta-TCP/PLGC in which poly(L-lactide-co-glycolide-co-epsilon-caprolactone) and beta-tricalcium phosphate were used as the matrix and filler, respectively. The other material used was HAp/Col-a soft nanocomposite of hydroxyapatite and type I collagen. Using these composites, two bone implants were designed. The efficacy of these implants was investigated by applying them to the critical-sized bone defects that were created in the canine tibia. Although no tissue engineering techniques such as application of growth factors or stem cells was utilized, successful healing was observed. These results suggested that bone regeneration in the critical-sized defects is possible without the use of growth factors or stem cells if the materials and the bone implants are suitably designed.

Reconstruction of bone fenestration on mandiblar by the guided bone regeneration methods with beta-TCP/PLGC membranes.

Guided Bone Regeneration (GBR) is a method for bone tissue regeneration. In this method, membranes are used to cover bone defects and to block the invasion of the surrounding soft tissues. It would provide sufficient time for the osteogenic cells from bone marrow to proliferate and form new bony tissues. In spite of the potential usefulness of this method, no appropriate materials for the GBR membrane have been developed. Here we design the ideal mechanical properties of the GBR membranes and created novel materials, which is the composite of beta-tricalcium phosphate (beta-TCP) and block copolymer of L-lactide, glycolide, and epsilon-caplolactone (PLGC). In the animal experiments with the use of the GBR membranes for large bone defects, we observed significant enhancement in the bone regeneration after 12 weeks implantation and proved the effectiveness of the materials.