3D microenvironment as essential element for osteoinduction by biomaterials

In order to unravel the mechanism of osteoinduction by biomaterials, in this study we investigated the influence of the specific surface area on osteoinductive properties of two types of calcium phosphate ceramics. Different surface areas of the ceramics were obtained by varying their sintering temperatures. Hydroxyapatite (HA) ceramic was sintered at 1150 and 1250 degrees C. Biphasic calcium phosphate (BCP) ceramic, consisting of HA and beta-tricalcium phosphate (beta-TCP), was sintered at 1100, 1150 and 1200 degrees C. Changes in sintering temperature did not influence the chemistry of the ceramics; HA remained pure after sintering at different temperatures and the weight ratio of HA and beta-TCP in the BCP was independent of the temperature as well. Similarly, macroporosity of the ceramics was unaffected by the changes of the sintering temperature. However, microporosity (pore diameter <10 microm) significantly decreased with increasing sintering temperature. In addition to the decrease of the microporosity, the crystal size increased with increasing sintering temperature. These two effects resulted in a significant decrease of the specific surface area of the ceramics with increasing sintering temperatures. Samples of HA1150, HA1250, BCP1100, BCP1150 and BCP1200 were implanted in the back muscles of Dutch milk goats and harvested at 6 and 12 weeks post implantation. After explantation, histomorphometrical analysis was performed on all implants. All implanted materials except HA1250 induced bone. However, large variations in the amounts of induced bone were observed between different materials and between individual animals. Histomorphometrical results showed that the presence of micropores within macropore walls is necessary to make a material osteoinductive. We postulate that introduction of microporosity within macropores, and consequent increase of the specific surface area, affects the interface dynamics of the ceramic in such a way that relevant cells are triggered to differentiate into the osteogenic lineage.     

Transformation of biphasic calcium phosphate ceramics in vivo: ultrastructural and physicochemical characterization

Specially prepared biphasic calcium phosphate (BCP) macroporous ceramics consisting of an intimate association of beta tricalcium phosphate (beta-TCP) and hydroxyapatite (HA) with beta-TCP/HA weight ratios of 15/85, 35/65, and 85/15 were implanted in surgically created periodontal osseous defects in dogs and recovered after 6 months. A decrease in average size of crystals in BCP ceramics and an increase in the size of microporosities in the surface and at the core of the ceramic after implantation were observed, indicating that in vivo dissolution has taken place. The resorbability (reflecting in vivo dissolution) of BCP ceramics depended on their beta-TCP/HA ratios, the higher the ratio, the greater the resorbability. The formation of microcrystals with crystallographic properties and Ca/P ratio similar to those of bone apatite crystals were also observed. The abundance of these crystals were directly related to the beta-TCP/HA ratio of the BCP ceramic before implantation. The formation of the bone apatite-like crystals may be due to the precipitation of calcium and phosphate ions released from the dissolving ceramic crystals (the beta-TCP component dissolving preferentially to the HA component). Results from this study suggested that one of the means of controlling resorbability (in vivo dissolution) of BCP ceramic is by varying its beta-TCP/HA ratio.     

Removal of surface by-products from sintered hydroxyapatite: effect of a chelation treatment on fibronectin adsorption and cell adhesion

It was observed that fibronectin precipitates when deposited on hydroxyapatite (HA) ceramics. Fibronectin's known affinity for calcium and the composition of the ceramic itself suggested that calcium release could be the main cause of this aggregation effect. It was then decided to investigate the effect of a surface chelation treatment on fibronectin adsorption, and MG63 cell adhesion, onto porous ceramics of hydroxyapatite (HA), beta-tricalcium phosphate (beta-TCP), and HA/TCP biphasic material (BCP). Those ceramics were immersed in an EDTA solution and the effect of this treatment on the material composition was assayed. X-ray diffraction data showed the presence of alpha- and beta-TCP phases in HA and BCP materials, which were both completely removed by the chelation treatment in the case of HA. On BCP, alpha-TCP was removed and beta-TCP partially dissolved. The TCP material, which was pure beta-TCP, underwent a mass loss, but no change in composition was observed. Adhesion of MG63 cells was overall higher on the fibronectin-coated EDTA-treated HA material, but was especially enhanced on EDTA-treated HA. Changes in surface morphologies, as compared with the use of scanning electron microscopy, did not seem to be related to the effects observed. The EDTA treatment proved to be a very efficient way of removing by-products of HA sintered materials, and thus enhancing the biocompatibility of the material.     

Osteogenicity of biphasic calcium phosphate ceramics and bone autograft in a goat model

The aim of this work was to compare the osteogenicity of calcium phosphate ceramic granules with autologous bone graft in ectopic and orthotopic sites. Biphasic calcium phosphate (BCP) granules composed of hydroxyapatite (HA) and beta-tricalcium phosphate (beta-TCP) in a 60/40 ratio were sintered at 1050, 1125 and 1200 degrees C, producing different microporosities. Either BCP ceramic granules or autologous bone chips (n=7) were implanted into paraspinal muscles. Osteoinduction was not observed in either the BCP implants or autologous bone chips after 6 or 12 weeks in the ectopic sites. Hollow and bored polytetrafluoroethylene (PTFE) cylinders were filled with autologous bone, BCP granules or left empty, then implanted into critical-sized defects in femoral epiphyses. The PTFE cylinders left empty contained marrow and blood vessels but not mineralized bone, indicating that this model prevented bone ingrowth (0.56+/-0.43% at 12 weeks). Bone formation was observed in contact with the BCP1050 and BCP1125 granules in the femoral sites after 6 weeks. The amount of bone after 12 weeks was 5.6+/-7.3 and 9.6+/-6.6% for BCP1050 and BCP1125, respectively. Very little bone formation was observed with the BCP1200 implants (1.5+/-1.3% at 12 weeks). In both the ectopic and orthotopic sites, autologous bone chips were drastically resorbed (from 19.4+/-3.7% initially to 1.7+/-1.2% at 12 weeks). This study shows that synthetic bone substitutes may have superior stability and osteogenic properties than autologous bone grafts in critical-sized bone defects.     

Comparative study of biphasic calcium phosphate ceramics impregnated with rhBMP-2 as bone substitutes

We investigated pellet-shaped implants prepared from biphasic calcium phosphate (BCP) ceramics with five different ratios of hydroxyapatite (HAP) to beta-tricalcium phosphate (beta-TCP) to evaluate these ceramics as bone substitutes. BCP ceramics impregnated with different doses of recombinant human bone morphogenetic protein 2 (rhBMP-2) (1, 5, and 10 microg) were used for experimental purposes and ceramics without rhBMP-2 were used for control. The pellets were implanted under the pericranium in adult Wistar male rats and were harvested 8 weeks after implantation. The retrieved pellets were then examined radiologically, histologically, and histomorphometrically. The results revealed that the pellets treated with rhBMP-2 exhibited new bone and bone marrow, whereas control pellets produced fibrous connective tissues. The formation of new bone induced by rhBMP-2 was dose dependent. The extent of bone and bone marrow formation and the degree of resorption of the ceramic particles were significantly higher in the pellets composed of 25% HAP-75% TCP. In this study, bioresorption of the ceramic produced favorable conditions for rhBMP-2-induced bone formation.     

Evaluation of ceramics composed of different hydroxyapatite to tricalcium phosphate ratios as carriers for rhBMP-2

We have investigated pellet-shaped implants prepared from biphasic calcium phosphate (BCP) ceramics with five different ratios of hydroxyapatite (HAP) to beta tricalcium phosphate (beta-TCP). The purpose of this study was to evaluate these BCP ceramics as carriers for rhBMP-2. BCP ceramics impregnated with the different doses of recombinant human bone morphogenetic protein 2 (rhBMP-2) (1, 5 and 10g) were used for the experimental purpose and the ceramics without rhBMP-2 were used as control. The pellets were placed into subcutaneous pockets on the dorsum of 4-week-old male Wistar rats. The animals were sacrificed 2 and 4 weeks after implantation. Bone induction was estimated by alkaline phosphatase (ALP) activity measured at 2 weeks after implantation. Pellets were also examined radiologically, histologically and histomorphometrically. The results showed that all experimental pellets exhibited new bone formation whereas the control pellets produced only fibrous connective tissue. Here, 100% HAP ceramic showed most amount of bone formation, whereas 25% HAP to 75% TCP ceramic produced the bone least in amount among different BCP ceramics at the end of 4 weeks. This study indicates that formation of new bone depends on the ceramic content with high HAP-TCP ratio and high dose of rhBMP-2.     

Review paper: behavior of ceramic biomaterials derived from tricalcium phosphate in physiological condition

Various calcium phosphates are used for bone repair. Although hydroxyapatite (HA) sintered ceramics are widely used due to their osteoconductivity, its bioresorbability is so low that HA remains in the body for a long time after implantation. In contrast, tricalcium phosphate (TCP) ceramics show resorbable characters during bone regeneration, and can be completely substituted for the bone tissue after stimulation of bone formation. Therefore, much attention is paid to TCP ceramics for scaffold materials for supporting bone regeneration. This paper reviews bioresorbable properties of calcium phosphate ceramics derived from beta-TCP and alpha-TCP.     

Nanoindentation study of interfaces between calcium phosphate and bone in an animal spinal fusion model.

Intertransverse process spinal fusion is a common surgical procedure for the treatment of spinal disorders. In the present study, a porous hydroxyapatite (HA)/beta-tricalcium phosphate (beta-TCP) ceramic was tested as graft material using a rabbit lumbar transverse process (L5-L6) fusion model. The porous ceramic blocks were implanted onto the dorsal decorticated surface of the lumbar transverse processes. The specimens were harvested at the seventh week after implantation. Histomorphological observation revealed that the integration of HA/beta-TCP with the host bone of the transverse process occurred by both cancellous bone formation and cartilage formation. Scanning electron microscopy-wavelength dispersive X-ray spectrometry examinations showed significant differences in calcium, phosphorus, and sulfur contents in the newly formed tissues and the porous HA/TCP implants. Nanoindentations were used to evaluate the intrinsic mechanical properties of the implants and the newly formed tissues. The Young's moduli of the newly formed cartilage, new cancellous bone, and HA/TCP, were 0.66 +/- 0.02 GPa, 2.36 +/- 0.50 GPa, and 10.2 +/- 1.21 GPa, respectively. Nanoindentation results revealed degradation of the porous ceramics and incomplete calcification of the new cancellous bone at the seventh week after implantation. Nanoindentation appeared to be a useful technique for assessing the mechanical status of spinal fusion in animal models.     

Comparative bone growth behavior in granules of bioceramic materials of various sizes.

Various bioceramic materials were implanted into 6-mm-diameter holes made in the femoral condyles of mature Japanese white rabbits using different-sized granules to find an optimal material and granule diameter for use as a bone graft. Bioceramics include a bioinert ceramic (Alumina), surface-bioactive ceramics [hydroxyapatite (HAp) and Bioglass(R)], and resorbable bioactive ceramics [alphatricalcium phosphate (alpha-TCP), beta-TCP, tetracalcium phosphate (TeCP), Te. DCPD, Te. DCPA, and low-crystalline HAp]. Granule sizes were 100-300, 10, and 1-3 microm. Bone growth behavior varied with the kind of bioceramic and the size used. For surface-bioactive ceramics, 45S5 Bioglass(R) led to more rapid bone proliferation than synthetic HAp. In resorbable bioactive ceramics, the order of resorption was: low-crystalline HAp and OCP > TeCP, Te DCPD, Te DCPA > alpha-TCP, beta-TCP. In terms of biocompatibility, alpha-TCP was better than beta-TCP.     

生物陶瓷微颗粒引发的细胞和组织损害

为论证生物陶瓷烧结不完全形成的微颗粒（〈5μm）引发细胞和组织损害的假设，对该类颗粒在体内和体外的细胞和组织损害进行了研究：（1）对4种双相生物陶瓷（BCP）进行细胞毒性试验。试验发现所有的浸出液出现细胞毒性，但是浸出液经离心后，毒性消失；（2）对羟基磷灰石（HA）、p磷酸三钙（pTCP）和40％pTCP／60％HA混合物微颗粒进行细胞抑制实验。结果显示随着微颗粒的浓度增加，成纤维细胞活力下降；而当微颗粒浓度达到一万个／细胞时，细胞活力和增殖能力完全消失；（3）HA，pTCP和BCP陶瓷颗粒（500-1500um）被植入到兔子股骨远端，种植12周后β-TCP的降解率为40％，BCP为5％，但是HA接近不降解。新骨形成在β-TCP（21％）和HA（18％）比BCP（12％）更为明显。同时BCP颗粒的周围有很多的微颗粒形成，可见吞噬细胞吞噬微颗粒，形成吞噬体。以上结果提示，微颗粒可能是局部炎症和细胞损害的首要原因，而且有可能影响骨形成。因此，我们必须注意生物陶瓷烧结的重要性，它们的烧结不良就可形成微颗粒，引发细胞和组织的损害。特别是BCP陶瓷含有两种需要不同烧结温度的粉体，它的烧结难度较高，很易形成微颗粒。     

Sol-gel synthesis and characterization of macroporous calcium phosphate bioceramics containing microporosity

Amorphous calcium phosphate powders were precipitated from calcium metal and phosphoric acid in ethanol. Depending on the quantity of reagent, the CaP powders had different chemical compositions and, after heating, formed beta-tricalcium phosphate (beta-TCP), hydroxyapatite (HA) or BCP mixtures. Dilatometric measurements indicated that shrinkage of compacted CaP powders occurred first at around 650 degrees C and continued up to 1200 degrees C. The amorphous CaP powders were mixed with urea beads, compacted under isostatic pressure at 140 MPa and sintered at 1100 degrees C for 5 h. Scanning electron microscopy indicated that macro-microporous ceramics were produced. The ceramics had spherical macropores of 700-1200 microm in diameter, with limited interconnections and a macroporosity of 42% as determined by microcomputed tomography. The micropores ranged from 0.1 to 1 microm in diameter. These ceramics made of HA, beta-TCP or BCP exhibiting both macroporosity and microporosity can be used as bone fillers.     

α-磷酸三钙/HA晶须/羧甲基壳聚糖-明胶复合增强多孔骨水泥的制备研究

为研究羟基磷灰石(HA)晶须和羧甲基壳聚糖-明胶(CMC-Gel)对多孔磷酸钙骨水泥(CPC)力学性能的影响,将α-磷酸三钙(α-TCP)粉、HA晶须和致孔剂L-谷氨酸钠按一定的质量比进行混合,加入调和液制备成α-TCP/HA晶须复合多孔骨水泥,然后将其浸润到一系列不同CMC和Gel质量比的溶液中以制备α-TCP/HA晶须/CMC-Gel复合增强多孔骨水泥,对其进行抗压强度测试和扫描电镜观察。结果显示,当HA晶须含量为4%,未添加CMC和Gel时,α-TCP/HA晶须复合多孔骨水泥的抗压强度达到2.57MPa,与未复合HA晶须的骨水泥相比提高了81%;当CMC和Gel的质量比为50∶50时,α-TCP/HA晶须/CMC-Gel复合多孔骨水泥的抗压强度达到最大值3.34MPa,与单纯的多孔α-TCP骨水泥相比提高了135%,同时韧性也有较大改善。     

The fabrication and biochemical evaluation of alumina reinforced calcium phosphate porous implants

Alumina reinforced calcium phosphate porous implants were manufactured to improve the mechanical strength while maintaining the bioactivity of calcium phosphate ceramics. The alumina porous bodies, which provided the mechanical strength, were fabricated by a polyurethane sponge method and multiple coating techniques resulted in the porous bodies with a 90-75% porosity and a compressive strength of up to approximately 6MPa. The coating of hydroxyapatite (HAp) or tricalcium phosphate (beta-TCP) was performed by dipping the alumina porous bodies into calcium phosphate ceramic slurries and sintering the specimens. The fairly strong bonding between the HAp or TCP coating layer and the alumina substrate was obtained by repeating the coating and sintering processes. The biochemical evaluations of the porous implants were conducted by in vitro and in vivo tests. For in vitro test, the implants were immersed in Ringer's solution and the release of Ca and P ions were detected and compared with those of calcium phosphate powders. For in vivo test, the porous bodies were implanted into mixed breed dogs and bone mineral density measurements and histological studies were conducted. The alumina reinforced HAp porous implants had a higher strength than the HAp porous implants and exhibited a similar bioactivity and osteoconduction property to the HAp porous implants.     

Histological and histomorphometrical comparative study of the degradation and osteoconductive characteristics of alpha- and beta-tricalcium phosphate in block grafts

The purpose of the present study was to compare alpha- and beta-tricalcium phosphate (TCP) as bone graft material for augmenting highly resorbed alveolar ridges. The cranial bones of 15 rabbits were used. Three titanium chambers filled with porous blocks of alpha-TCP, beta-TCP, or blood clots were placed in each slit. The two TCP blocks had similar inner/outer structures and purities. Animals were sacrificed after 2, 4, and 8 weeks. Specimens were embedded in polyester resin as nondecalcified specimens, and evaluated both histologically and histomorphometrically. In both TCP groups, blocks had hardly degraded at 2 weeks while in the alpha-TCP group, the block had notably started degrading after 4 weeks. In the beta-TCP group, degradation began at 4 weeks and this degradation had increased just slightly after 8 weeks. The alpha-TCP block degraded significantly more than the beta-TCP block. Residual alpha-TCP particles surrounded by newly formed bone decreased over time, and both particles and newly formed bone were simultaneously absorbed by osteoclast-like cells. These observations suggest that residual alpha-TCP particles surrounded by newly formed bone may disappear progressively from bone and could be incorporated into the bone remodeling cycle in combination with newly formed bone.     

Phase formation and evolution in the silicon substituted tricalcium phosphate/apatite system

The sintering of silicon doped calcium phosphate ceramics prepared from a basic colloidal hydroxyapatite (Ca5(PO4)3OH or HA) precipitate mixed with silica over 800 degrees C yields a phase mixture of tricalcium phosphate phases (TCP) designated Si-TCP, beta-TCP and a silicon substituted dehydrated apatite (Si-Ap). The Si-TCP phase is defined as a combination of a silicon stabilized TCP in which the silicon content attains a saturated value (Ca3(P0.9Si0.1O3.95)2 or Si-TCP(sat)) and alpha-TCP (Ca3(PO4)2). Si-TCP(sat) has the same crystalline space group (P2(1)/a) as alpha-TCP, but with characteristically different lattice parameters due to the substitution of silicon in tetrahedral phosphorus sites. The nucleation and growth kinetics of Si-TCP in samples of composition 0.2 mol SiO2:mol HA (0.2:1) and 1 mol SiO2:mol HA (1:1) can be understood in terms of the initial growth of alpha-TCP at a silica-HA interface followed by a transformation to Si-TCP(sat) or beta-TCP. A thermodynamic model for the formation of Si-TCP(sat) predicts a nucleation temperature of 795 degrees C, in close agreement with experiment. If sufficient silicon is available, the alpha-TCP transforms to Si-TCP(sat) during extended sintering. In the absence of sufficient silicon, the alpha-TCP transforms to beta-TCP.     

A preliminary study on osteoinduction of two kinds of calcium phosphate ceramics.

With respect to the effect of material factors on calcium phosphate biomaterial-induced osteogenesis, the osteoinductive property of two kinds of porous hydroxyapatite ceramics, which were made by different producers, was investigated in dorsal muscles of dogs. One hydroxyapatite ceramic (S-HA), macroporous implants with rough pore walls containing abundant micropores, was made by Sichuan Union University (Chengdu, China); the other hydroxyapatite ceramic (J-HA), porous implants with smooth macropore walls composed of regularly aligned crystal grains, was provided by Mitsubishi Ceramic Int. (Japan). Different tissue response was detected histologically and microradiographically after the ceramic samples had been implanted in dorsal muscles of dogs for 3 and 6 months. Bone formation was found in S-HA at 3 months, which increased at 6 months. In contrast, no bone formation was detected in J-HA at both 3 and 6 months. These results indicate that with the special architecture, calcium phosphate ceramic can induce bone formation in soft tissue. As both materials were very similar in their chemical and crystallographic structures, but varied in their microstructures, the latter seem to be an important factor affecting the osteoinductive capacity of calcium phosphate ceramics. These data suggest that, by controlling the preparation of calcium phosphate ceramic, bone substitutes with intrinsic osteoinductive property can be developed from calcium phosphates.     

Osteoinduction of hydroxyapatite/β-tricalcium phosphate bioceramics in mice with a fractured fibula

Many studies have shown that calcium phosphate ceramics can induce bone formation in non-osseous sites without the application of any osteoinductive biomolecules, but the mechanisms of this phenomenon (intrinsic osteoinduction of bioceramics) remain unclear. In this study, we compared the intrinsic osteoinduction of porous hydroxyapatite/β-tricalcium phosphate (HA/β-TCP) implanted in mice at different sites. In 30 mice the left fibula was fractured and the right fibula was kept intact. A porous HA/β-TCP cylinder was implanted into both the left (group 1) and right (group 2) leg muscles of each animal. In addition, two HA/β-TCP cylinders were bilaterally implanted into leg subcutaneous pockets (group 3) in each of the remaining 15 mice. New bone formation was studied in the three groups by histology, histomorphometry and immunostaining. In group 1 new bone was observed at week 6 and bone marrow appeared at week 12. In group 2 new bone was observed at week 8 and bone marrow appeared at week 12. The new bone area percentage in group 1 was significantly higher than in group 2 at both weeks 8 and 12. In contrast, group 3 did not show any new bone within the period studied. These differences were explained based on the location of the implants and thus their proximity to the osteogenic environment of fracture healing. The results support the hypothesis that intrinsic osteoinduction by calcium phosphate ceramics is the result of adsorption of osteoinductive substances on the surface.     

Marrow cell induced osteogenesis in porous hydroxyapatite and tricalcium phosphate: a comparative histomorphometric study of ectopic bone formation

To investigate the bone formation ability of porous hydroxyapatite (HA) and tricalcium phosphate (TCP), ceramic discs were implanted with or without rat marrow cells into subcutaneous sites in syngeneic rats. The discs of HA and TCP had identical microstructures: pore size was 190-230 microns, porosity was 50-60%, and they were fully interconnected. Implants without marrow cells (discs themselves) did not show bone formation, whereas implants with marrow cells showed bone formation in the pores of the ceramics. The bone formation of both HA and TCP occurred initially on the surface of the ceramic and progressed towards the center of the pore. The de novo bone was quantitated from decalcified serial sections of the implants. One month after implantation with marrow cells, the percentage fractions of the pore area filled with bone for implanted HA and TCP were 16.9 and 15.1, respectively. At 2 months after implantation with marrow cells, the fractions of bone were 34.3 and 30.9, respectively. These results indicate that both HA and TCP ceramics can show comparable osteogenic ability in the presence of marrow cells.     

Bone growth on and resorption of calcium phosphate coatings obtained by pulsed laser deposition

Three different calcium phosphate coatings of crystalline hydroxyapatite (HA), alpha- and beta-tricalcium phosphate (alpha+beta-TCP), or amorphous calcium phosphate (ACP) obtained by pulsed laser deposition on Ti-6Al-4V were incubated in a potentially osteogenic primary cell culture (rat bone marrow) in order to evaluate the amount and mode of mineralized bone matrix formation after 2 weeks with special emphasis on the type of interfacial structure that was created. Evaluation techniques included fluorescence labeling and scanning electron microscopy. The resistance to cellular resorption by osteoclasts was also studied. Bone matrix delaminated from the ACP coatings, while it remained on the HA and the alpha+beta-TCP coatings even after fracturing. A cementlike line was seen as the immediate contiguous interface with the nondegrading dense HA surface and with the surface of the remaining porous beta-TCP coating. Highly dense and crystalline HA coatings do not dissolve but are capable of establishing a strong bond with the bone matrix grown on top. Chemical and mechanical bonding were considered in this case. Cellular resorption was practically not observed on the HA coatings, but it was observed on the alpha+beta-TCP coatings. Resorption took place as dissolution that was due to the acidic microenvironment.     

Biological performance of uncoated and octacalcium phosphate-coated Ti6Al4V

The in vivo behavior of a porous Ti6Al4V material that was produced by a positive replica technique, with and without an octacalcium phosphate (OCP) coating, has been studied both in the back muscle and femur of goats. Macro- and microporous biphasic calcium phosphate (BCP) ceramic, known to be both osteoconductive and able to induce ectopic bone formation, was used for comparison purpose. The three groups of materials (Ti6Al4V, OCP Ti6Al4V and BCP) were implanted transcortically and intramuscularly for 6 and 12 weeks in 10 adult Dutch milk goats in order to study their osteointegration and osteoinductive potential. In femoral defects, both OCP Ti6Al4V and BCP were performing better than the uncoated Ti6Al4V, at both time points. BCP showed a higher bone amount than OCP Ti6Al4V after 6 weeks of implantation, while after 12 weeks, this difference was no longer significant. Ectopic bone formation was found in both OCP Ti6Al4V and BCP implants after 6 and 12 weeks. The quantity of ectopically formed bone was limited as was the amount of animals in which the bone was observed. Ectopic bone formation was not found in uncoated titanium alloy implants, suggesting that the presence of calcium phosphate (CaP) is important for bone induction. This study showed that CaPs in the form of coating on metal implants or in the form of bulk ceramic have a significantly positive effect on the bone healing process.