Dual setting α-tricalcium phosphate cements

An extension of the application of calcium phosphate cements (CPC) to load-bearing defects, e.g. in vertebroplasty, would require less brittle cements with an increased fracture toughness. Here we report the modification of CPC made of alpha-tricalcium phosphate (α-TCP) with 2-hydroxyethylmethacrylate (HEMA), which is polymerised during setting to obtain a mechanically stable polymer-ceramic composite with interpenetrating organic and inorganic networks. The cement liquid was modified by the addition of 30–70 % HEMA and ammoniumpersulfate/tetramethylethylendiamine as initiator. Modification of α-TCP cement paste with HEMA decreased the setting time from 14 min to 3–8 min depending on the initiator concentration. The 4-point bending strength was increased from 9 MPa to more than 14 MPa when using 50 % HEMA, while the bending modulus decreased from 18 GPa to approx. 4 GPa. The addition of ≥50 % HEMA reduced the brittle fracture behaviour of the cements and resulted in an increase of the work of fracture by more than an order of magnitude. X-ray diffraction analyses revealed that the degree of transformation of α-TCP to calcium deficient hydroxyapatite was lower for polymer modified cements (82 % for polymer free cement and 55 % for 70 % HEMA) after 24 h setting, while the polymerisation of HEMA in the cement liquid was quantitative according to FT-IR spectroscopy. This work demonstrated the feasibility of producing fracture resistant dual-setting calcium phosphate cements by adding water soluble polymerisable monomers to the liquid cement phase, which may be suitable for an application in load-bearing bone defects.     

Preparation of octacalcium phosphate by the hydrolysis of α-tricalcium phosphate

A new simple method of preparation for the thermodynamically unstable octacalcium phosphate [Ca₈H₂(PO₄)₆·5H₂O; OCP] has been developed using the hydrolysis of -Ca₃(PO₄)₂ instead of the conventional hydrolysis of CaHPO₄·2H₂O. The hydrolysis experiments were carried out by treating an -Ca₃(PO₄)₂(1 g)-H₂O(50 m) suspension for 3 h at temperatures in the range 40 to 80° C and at pHs in the range 3 to 7.5. The formation of OCR was limited to within a narrow region between formation regions of other phosphates. Favourable conditions for OCP preparation were, for example, 70° C, pH4.5 to 5.0 and 60° C, pH5.0. Particles of OCP were composed of tight aggregates of strip-like microcrystals growing probably along the [0 0 1] and (1 0 0) plane of the OCP structure. Nearly stoichiometric OCP was obtained under the most suitable conditions with good reproducibility. Pyrolytic processes of OCP were approximately consistent with the data published so far. However, the temperatures of the appearance and disappearance of pyrolytic crystalline phases and ionic species deviated slightly from the published data. Thermal dehydration up to 150° C without destruction of OCP and decomposition reactions above 300° C resulted in changes in surface area and average pore radius of OCP.     

Reactivity of α-tricalcium phosphate

The reactivity of -tricalcium phosphate (-TCP) in forming hydroxyapatite (HAp) at 37°C was investigated. The effects of synthesis route, HAp seeding and the presence of calcium salts on the mechanism and extent of HAp formation were examined by pH measurements and/or isothermal calorimetric analyses. A synthesis temperature at the lower end in the temperature range of 1100–1300°C and the reaction of -TCP with a high specific surface area greatly improved rate and extent of HAp formation. The time for complete reaction decreased from 18 h to 14 h, when the reaction was carried out in the presence of 1 wt% of HAp seeds; the hydrolysis mechanism did not change. At HAp seeds proportion of 5 wt% and 10 wt%, transformation occurred without a nucleation period. The calcium salt additives studied were anhydrous and dihydrate form of dicalcium phosphate (CaHPO₄ and CaHPO₄ · 2H₂O), calcium carbonate (CaCO₃), and calcium sulfate hemihydrate (CaSO₄ · 1/2H₂O). All the additives delayed HAp formation as determined by the isothermal calorimetric analyses. Their retarding effects in decreasing order are CaCO₃, CaSO₄ · 1/2H₂O, DCPD, DCP. CaCO₃ almost completely retarded HAp formation. After 24 h, hydrolysis was complete only for pure -TCP and for the -TCP-DCP blend. Reaction was complete in other formulations before 48 h except for the CaCO₃-containing blend. In all mixtures conversion to HAp occurred without forming any intermediates. However gypsum formed in the mixture containing CaSO₄ · 1/2H₂O. All the -TCP-additive mixtures, excluding -TCP-CaCO₃, reached nominally the same strength value after 24 h of reaction as governed by the transformation of -TCP to HAp. For phase-pure -TCP, the average tensile strength changed from 0.36 ± 0.03 MPa to 7.26 ± 0.6 MPa. Upon hydrolysis only the CaSO₄ · 1/2H₂O-containing mixture exhibited slightly higher strength averaging 8.36 ± 0.9 MPa.     

Effect of iron on the setting properties of α-TCP bone cements

New ceramic materials with the ability to set like cement, after mixing a powder phase made of one and/or several of these new reactants and a liquid phase, have been obtained within the ternary system “CaO-P₂O₅-FeO”. These new reactants have magnetic properties, i.e. cement made from them maintains its magnetic property during the whole setting and hardening. These new materials can be of use, for example, in dental applications, in the treatment of certain types of bone cancer and, in general, in the fields of Biomaterials and Bone Tissue Engineering. In this article, we report on the effect of iron-modified α -tricalcium phosphate, which is the main reactant of commercial calcium phosphate bone cements, on their new setting and hardening properties.     

Calcium phosphate cement: in vitro and in vivo studies of the α-tricalcium phosphate-dicalcium phosphate dibasic-tetracalcium phosphate monoxide system

In this paper, calcium phosphate cement consisting of -tricalcium phosphate (-TCP), dicalcium phosphate dibasic (DCPD) and tetracalcium phosphate monoxide (TeCP) was investigated in vitro and in vivo. Measurements of compressive strength against soaking time in simulated body fluid (SBF) showed a rapid increase of the hardness for the first 7 days. The gained strength was retained up to 1 year and the maximal mean value was 94.7 (±14.4) MPa. X-ray diffraction (XRD) and scanning electron microscopy (SEM) presented precipitates of hydroxyapatite (HA) after mixing, also after soaking in SBF and after implantation in rat subcutaneous tissues. However, the conversion to HA happened in different ways between in vitro and in vivo exposures. Histologic examinations showed that the cement causes the same reactions at the interface with surrounding soft tissues as HA. The authors consider the cement to be a promising material as a bone substitute, bone cement or dental material, however, further studies in a paste form and in bone tissue environments are necessary.     

Composite bone substitute materials based on β-tricalcium phosphate and magnesium-containing sol–gel derived bioactive glass

In the present study, bioceramic composites with improved mechanical and biological properties were synthesized by sintering mixtures of β-tricalcium phosphate and SiO₂–CaO–MgO–P₂O₅ sol–gel derived bioactive glass at 1000–1200°C. The physical, mechanical, structural and biological properties of the composites were evaluated by appropriate experiments such as microhardness, bending strength, XRD, SEM and MTT. The results showed that 1000 and 1100°C were not appropriate temperatures for sintering the composites and in contrast, the microhardness, bending strength and bulk density significantly increased by increasing in quantity of bioglass phase when the samples were sintered at 1200°C. No significant difference was found between the fracture toughness of the composites and pure β-tricalcium phosphate. β-tricalcium phosphate was structurally stable up to 1200°C and did not transform to its alpha form even in the presence of the bioglass phase but migration of magnesium cations from the glass composition into its lattice structure was found by right-shift in XRD patterns, especially when the composite contained higher amount of bioglass component. Calcium silicate was also crystallized in the composition of the composites, which was more detectable in higher sintering temperatures. The results of the MTT test showed that proliferation of human osteosarcoma cells on the composites was considerably better than that of pure β-TCP.     

Biodegradable β-tricalcium phosphate cement with anti-washout property based on chelate-setting mechanism of inositol phosphate

Novel biodegradable β-tricalcium phosphate (β-TCP) cements with anti-washout properties were created on the basis of chelate-setting mechanism of inositol phosphate (IP6) using β-TCP powders. The β-TCP powders were ball-milled using ZrO₂ beads for 0–6 h in the IP6 solutions with concentrations from 0 to 10,000 ppm. The chelate-setting β-TCP cement with anti-washout property was successfully fabricated by mixing the β-TCP powder ball-milled in 3,000 ppm IP6 solution for 3 h and 2.5 mass% Na₂HPO₄ solution, and compressive strength of the cement was 13.4 ± 0.8 MPa. An in vivo study revealed that the above cement was directly in contact with host and newly formed bones without fibrous tissue layers, and was resorbed by osteoclast-like cells on the surface of the cement. The chelate-setting β-TCP cement with anti-washout property is promising for application as a novel injectable artificial bone with both biodegradability and osteoconductivity.     

The controlled resorption of porous α-tricalcium phosphate using a hydroxypropylcellulose coating

Tricalcium phosphate (TCP) ceramic is known in orthopedics to be a bioresorbable bone substitute. A porous TCP ceramic body also has high potential as a drug delivery system in bony defects. Porous alpha-TCP ceramic can be easily fabricated using conventional sintering of beta-TCP, since alpha-TCP is the thermodynamically stable phase at temperatures above 1 100 degrees C. However, the solubility of alpha-TCP is much higher than that of beta-TCP. Therefore, the dissolution of porous alpha-TCP progresses at a higher rate than bone repair. In the present study, we attempted to reduce the dissolution rate of porous alpha-TCP by employing an organic polymer coating. We fabricated porous alpha-TCP ceramic with a continuous 10-50 microm diameter pore structure by sintering a body made from a beta-TCP and potato starch slurry. The porous body obtained was coated with hydroxypropylcellulose (HPC), and then subjected to heat treatment. The chemical durability and mechanical properties of the body were examined before and after coating with the HPC. The dissolution of porous alpha-TCP in buffered solutions was reduced by coating with HPC and drying at 60 degrees C. The compressive strength of the porous alpha-TCP was also improved by coating with HPC. The results of in vivo experiments showed that some parts of the porous alpha-TCP ceramic coated with HPC remained in the canal of the tibia of a rabbit four weeks after implantation, whereas no residual was observed in a non-coated alpha-TCP ceramic. Coating with HPC was found to be effective for controlling bioresorption and improving the workability of porous alpha-TCP ceramic. The prepared porous alpha-TCP ceramic is expected to be useful as a novel material for bone fillers by incorporating it with drugs or osteoinductive factors.     

Preparation of hydroxyapatite/α-tricalcium phosphate composites by colloidal process

Synthesised hydroxyapatite (HAp)/α-tricalcium phosphate (α-TCP) composites were prepared by colloidal process. Mixed slurries composed of 30 vol% HAp/α-TCP powder and 70 vol% aqueous solution containing a small amount of polymer dispersant were produced. HAp/α-TCP powder was blended with 0, 0.2, 0.4 or 1.0 mass fraction of α-TCP. Rheological behaviour of the HAp/α-TCP mixed slurries depended on the quantity of polymer dispersant and pH. Addition of suitable quantity of polymer dispersant at higher pH resulted in an increase in the stability of colloidal state due to electrostatic stabilisation. The optimum quantity of dispersant for the colloidal process was found to be proportional to the mass fraction of α-TCP. In the dehydration process of the slurries, pressure filtration technique was effective in preventing the gravity segregation of α-TCP. Thus, HAp/α-TCP composites with homogeneous microstructure and high relative density could be prepared by optimising the colloidal process.     

Correlation between properties and microstructure of laser sintered porous β-tricalcium phosphate bone scaffolds

Abstract

A porous β-tricalcium phosphate (β-TCP) bioceramic scaffold was successfully prepared with our homemade selective laser sintering system. Microstructure observation by a scanning electron microscope showed that the grains grew from 0.21 to 1.32 μm with the decrease of laser scanning speed from 250 to 50 mm min^?1. The mechanical properties increased mainly due to the improved apparent density when the laser scanning speed decreased to 150 mm min^?1. When the scanning speed was further decreased, the grain size became larger and the mechanical properties severely decreased. The highest Vickers hardness and fracture toughness of the scaffold were 3.59 GPa and 1.16 MPa m^1/2, respectively, when laser power was 11 W, spot size was 1 mm in diameter, layer thickness was 0.1–0.2 mm and laser scanning speed was 150 mm min^?1. The biocompatibility of these scaffolds was assessed in vitro with MG63 osteoblast-like cells and human bone marrow mesenchymal stem cells. The results showed that all the prepared scaffolds are suitable for cell attachment and differentiation. Moreover, the smaller the grain size, the better the cell biocompatibility. The porous scaffold with a grain size of 0.71 μm was immersed in a simulated body fluid for different days to assess the bioactivity. The surface of the scaffold was covered by a bone-like apatite layer, which indicated that the β-TCP scaffold possesses good bioactivity. These discoveries demonstrated the evolution rule between grain microstructure and the properties that give a useful reference for the fabrication of β-TCP bone scaffolds.     

Repair of goat tibial defects with bone marrow stromal cells and β-tricalcium phosphate

Tissue engineering techniques have been proven effective in bone regeneration and repairing load-bearing bone defects. Previous studies, however, have heretofore been limited to the use of slowdegradable or natural biomaterials as scaffolds. There are, however, no reports on using biodegradable, synthetic beta-tricalcium phosphate (β-TCP) as scaffolds to repair weight-bearing bone defects in large animals. In the present study, highly porous β-TCP scaffolds prepared by the polymeric sponge method were used to repair goat tibial defects. Fifteen goats were randomly assigned to one of three groups, and a 26 mm-long defect at the middle part of the right tibia in each goat was created. In Group A (six goats), a porous β-TCP ceramic cylinder that had been loaded with osteogenically induced autologous bone marrow stromal cells (BMSCs) was implanted in the defect of each animal. In Group B (six goats), the same β-TCP ceramic cylinder without any cells loaded was placed in the defect. In Group C (three goats), the defect was left untreated. In Group A, bony union can be observed by gross view, X-ray and micro-computed tomography (Micro-CT) detection, and histological observation at 32 weeks post-implantation. The implanted β-TCP scaffolds were almost completely replaced by tissue-engineered bone. Bone mineral density in the repaired area of Group A was significantly higher (p < 0.05) than that of Group B, in which scant new bone was formed in each defect and the β-TCP hadn’t been completely resorbed at 32 weeks. Moreover, the tissue-engineered bone of Group A had similar biomechanical properties as that of the normal left tibia in terms of bending strength and Young’s modulus (p > 0.05). In Group C, little or no new bone was formed, and non-union occurred, showing that the 26 mm segmental defect of the goat tibia was critical sized at 32 weeks. Thus, it can be concluded that the mechanical properties of the BMSCs/β-TCP composites could be much improved via tissue engineering approach and β-TCP might be used to repair the weight-bearing segmental defects of goat tibias. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Effect of silicate incorporation on in vivo responses of α-tricalcium phosphate ceramics

In addition to calcium phosphate-based ceramics, glass-based materials have been utilized as bone substitutes, and silicate in these materials has been suggested to contribute to their ability to stimulate bone repair. In this study, a silicate-containing α-tricalcium phosphate (α-TCP) ceramic was prepared using a wet chemical process. Porous granules composed of silicate-containing α-TCP, for which the starting composition had a molar ratio of 0.05 for Si/(P + Si), and silicate-free α-TCP were prepared and evaluated in vivo. When implanted into bone defects that were created in rat femurs, α-TCP ceramics either with or without silicate were biodegraded, generating a hybrid tissue composed of residual ceramic granules and newly formed bone, which had a tissue architecture similar to physiological trabecular structures, and aided regeneration of the bone defects. Supplementation with silicate significantly promoted osteogenesis and delayed biodegradation of α-TCP. These results suggest that silicate-containing α-TCP is advantageous for initial skeletal fixation and wound regeneration in bone repair.     

Characterization of the transformation from calcium-deficient apatite to β-tricalcium phosphate

The structural changes that occur during the transformation of a Ca-deficient apatite, prepared by a wet chemical method, to -TCp were investigated. X-ray diffraction (XRD) analysis of as-prepared samples and samples calcined at temperatures between 500 and 1100 °C showed that the transformation occurs over the temperature range 710–740 °C, under non-equilibrium conditions. The change in crystallite size with increasing calcination/sintering temperature was studied by XRD using the Scherrer formula. Fourier transform infra-red (FTIR) analysis indicated considerable structural change in samples above and below this temperature range. Changes were observed in the hydroxyl, carbonate and phosphate bands as the calcination temperature was increased from 500 to 1100 °C. Even once a single -TCP phase is obtained at 740 °C there remains a considerable amount of structural change at temperatures between 740 and 1100 °C. This effect was illustrated by an unusual change in the lattice parameters of the -TCP structure and significant changes in the phosphate bands of FTIR spectra as the calcination temperature was increased. The results obtained in this study show that the combined experimental techniques of XRD and FTIR are excellent complimentary methods for characterizing structural changes that occur during phase transformations.     

Characterization of the transformation from calcium-deficient apatite to β-tricalcium phosphate

The structural changes that occur during the transformation of a Ca-deficient apatite, prepared by a wet chemical method, to -TCP were investigated. X-ray diffraction (XRD) analysis of as-prepared samples and samples calcined at temperatures between 500 and 1100 °C showed that the transformation occurs over the temperature range 710–740 °C, under non-equilibrium conditions. The change in crystallite size with increasing calcination/sintering temperature was studied by XRD using the Scherrer formula. Fourier transform infra-red (FTIR) analysis indicated considerable structural change in samples above and below this temperature range. Changes were observed in the hydroxyl, carbonate and phosphate bands as the calcination temperature was increased from 500 to 1100 °C. Even once a single -TCP phase is obtained at 740 °C there remains a considerable amount of structural change at temperatures between 740 and 1100 °C. This effect was illustrated by an unusual change in the lattice parameters of the -TCP structure and significant changes in the phosphate bands of the FTIR spectra as the calcination temperature was increased. The results obtained in this study show that the combined experimental techniques of XRD and FTIR are excellent complimentary methods for characterizing structural changes that occur during phase transformations.     

Hydrothermal growth of whitlockite coating on β-tricalcium phosphate surfaces for enhancing bone repair potential

In this study, we developed a simple approach for the controllable growth of whitlockite(WH) on a β-tricalcium phosphate surface and investigated its cell viability via CCK-8, its live-dead staining and its alkaline phosphatase activity. Herein, WH with controllable morphologies was prepared by regulating the hydrothermal reaction conditions. The results of scanning electron microscopy, X-ray diffraction and Xray photoelectron spectroscopy indicated that pure hexagonal plates of WH were prepared successfully.In vitro cell experiments showed that WH possessed excellent biocompatibility and effectively promoted the adhesion and proliferation of mouse bone mesenchymal stem cells. The osteogenesis of the WH was also enhanced. The obtained WH was expected to be utilized for promising applications as implantable block materials for bone repair.     

Development of some calcium phosphate cements from combinations of α-TCP,MCPM and CaO

From previous studies it is known that alpha-tertiary calcium phosphate (-TCP), monocalcium phosphate monohydrate (MCPM) and calcium oxide form cements upon mixing combinations of them with water. In this study some formulations were optimized with respect to the particle size of the constituents, their molar ratio, amounts of hydroxyapatite or beta-tertiary calcium phosphate (-TCP) added and the water/powder ratio. Three suitable products were obtained. Product 1 had a relatively short setting time and might be suitable as a dental cavity liner or for filling parodontological pockets or for filling alveolar cavities to prevent alveolar ridge resorption. Products 2 and 3 may be more suitable for orthopaedic purposes. Their compressive strength being 35 and 12 MPa, respectively, after soaking for 1 day in Ringer's solution at 37°C. Product 1 reaches its full strength within 4 h, whereas products 2 and 3 take about 12 h and 10 h, respectively.     

Radiation synthesis of gelatin/CM-chitosan/β-tricalcium phosphate composite scaffold for bone tissue engineering

A series of biodegradable composite scaffolds was fabricated from an aqueous solution of gelatin, carboxymethyl chitosan (CM-chitosan) and β-tricalcium phosphate (β-TCP) by radiation-induced crosslinking at ambient temperature. Ultrasonic treatment on the polymer solutions significantly influenced the distribution of β-TCP particles. An ultrasonic time of 20 min, followed by 30 kGy irradiation induced a crosslinked scaffold with homogeneous distribution of β-TCP particles, interconnected porous structure, sound swelling capacity and mechanical strength. Fourier Transform Infrared Spectroscopy and X-ray Diffraction analysis indicated that β-TCP successfully incorporated with the network of gelatin and CM-chitosan. In vivo implantation of the scaffold into the mandible of beagle dog revealed that the scaffolds had excellent biocompatibility and the presence of β-TCP can accelerate bone regeneration. The comprehensive results of this study paved way for the application of gelatin/CM-chitosan/β-TCP composite scaffolds as candidate of bone tissue engineering material.     

Characterization and distribution of mechanically competent mineralized tissue in micropores of β-tricalcium phosphate bone substitutes

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Osteoblast cell response to β-tricalcium phosphate scaffolds with controlled architecture in flow perfusion culture system

Porous β-tricalcium phosphate (β-TCP) scaffolds with controlled architecture and improved mechanical properties were fabricated by combining the gel-casting and rapid prototyping techniques. The pore morphology, size, and distribution of the β-TCP scaffolds were characterized using a scanning electron microscope. The porosity of the resulting scaffolds with pore size range from 300 to 500 μm was 46%. The average compressive strength was 16.1 MPa. X-ray diffraction was used to determine the crystal structure and chemical composition of scaffolds. The result indicated that the sintering process has not changed the composition of β-TCP. Flow perfusion culture system was developed in our lab to improve mass transfer for seeded cells. For scaffold/cell constructs cultured under flow perfusion for 4, 8, and 16 days, there was greater scaffold cellularity and alkaline phosphatase activity compared with static culture condition. These results indicated that flow perfusion culture system had evident effects on osteoblast viability and functions in vitro.     

The effect of rhBMP-2 and PRP delivery by biodegradable β-tricalcium phosphate scaffolds on new bone formation in a non-through rabbit cranial defect model

This study evaluated whether the combination of biodegradable β-tricalcium phosphate (β-TCP) scaffolds with recombinant human bone morphogenetic protein-2 (rhBMP-2) or platelet-rich plasma (PRP) could accelerate bone formation and increase bone height using a rabbit non-through cranial bone defect model. Four non-through cylindrical bone defects with a diameter of 8-mm were surgically created on the cranium of rabbits. β-TCP scaffolds in the presence and absence of impregnated rhBMP-2 or PRP were placed into the defects. At 8 and 16 weeks after implantation, samples were dissected and fixed for analysis by microcomputed tomography and histology. Only defects with rhBMP-2 impregnated β-TCP scaffolds showed significantly enhanced bone formation compared to non-impregnated β-TCP scaffolds (P < 0.05). Although new bone was higher than adjacent bone at 8 weeks after implantation, vertical bone augmentation was not observed at 16 weeks after implantation, probably due to scaffold resorption occurring concurrently with new bone formation.