复合磷酸钙陶瓷人工骨

磷酸钙陶瓷因具有良好的生物相容性和骨传导作用而成为人工骨的常用材料 ,但是 ,它们本身无骨诱导作用。将具有骨诱导作用的物质如 BMP、骨髓、生长因子等与磷酸钙陶瓷复合 ,可以克服磷酸钙陶瓷无骨诱导作用的缺陷     

In vivo behavior of three different injectable hydraulic calcium phosphate cements

Two dicalcium phosphate dihydrate (DCPD) hydraulic cements and one apatite hydraulic cement were implanted in epiphyseal and metaphyseal, cylindrical bone defects of sheep. The in vivo study was performed to assess the biocompatibility of the DCPD cements, using the apatite cement as control. After time periods of 2, 4 and 6 months the cement samples were clinically and histologically evaluated. Histomorphometrically the amount of new bone formation, fibrous tissue and the area of remaining cement were measured over time. In all specimens, no signs of inflammation were detectable either macroscopically or microscopically. All cements were replaced by different amounts of new bone. The two DCPD-cements showed the highest new bone formation and least cement remnants at 6 months, whereas the apatite was almost unchanged over all time periods.     

Tantalumpentoxide as a radiopacifier in injectable calcium phosphate cements for bone substitution

The chemical resemblance of calcium phosphate (CaP) cements and the mineral phase of bone is a problem in distinguishing CaP cement from bone tissue by means of common, noninvasive techniques (e.g., X-ray imaging and microcomputed tomography [μCT]). In this study, the feasibility of using tantalumpentoxide (Ta(2)O(5)) powder as radiopacifier in CaP cements was analyzed. A distal femoral condyle model in male adult Wistar rats was used. After 6 weeks of implantation time, the results were analyzed by means of μCT and histology. Unambiguous distinction of CaP cement from native bone tissue and volumetric measurements of the materials appeared to be possible by means of μCT scanning. Furthermore, there was no evidence of either inflammation or fibrous tissue around the implant materials or at the bone-material interface. In conclusion, the addition of Ta(2)O(5) as a radiopacifying additive to CaP cements allows discrimination between bone substitute and surrounding bone tissue. Consequently, Ta(2)O(5) represents an effective and biocompatible additive in CaP cements for in vivo monitoring purposes.     

In vivo behavior of a novel injectable calcium phosphate cement compared with two other commercially available calcium phosphate cements

The aim of this study was to investigate the physicochemical and biological properties of a newly developed calcium phosphate cement (CPC). The novel cement was compared with two other commercially available CPCs. After mixing the powder and liquid phase, the CPCs were injected as a paste into a rabbit distal femoral defect model. The CPCs were evaluated after 24 h, 6 weeks, 26 weeks, and 52 weeks. The novel CPC was easy to handle and was fast setting. X-ray diffraction (XRD) and Fourier Transform Infrared Spectrometry (FTIR) at the different implantation periods showed that the cement had converted to carbonated hydroxyapatite and remained stable over time. Histological evaluation showed bone apposition on the cement surface without any inflammatory response or fibrous encapsulation. At later time points, all CPCs were completely covered by a thin layer of bone. Osteoclast-like cells present at the interface resorbed parts of the cement mass. Histological and histomorphometrical analyses did not show any significant differences between the three implanted CPCs. The results indicate that the investigated CPC is biocompatible, osteoconductive, as well as osteotransductive and seems to be both biologically safe and effective as a bone void filler.     

Controlling the biological function of calcium phosphate bone substitutes with drugs

There is a growing interest in bone tissue engineering for bone repair after traumatic, surgical or pathological injury, such as osteolytic tumor or osteoporosis. In this regard, calcium phosphate (CaP) bone substitutes have been used extensively as bone-targeting drug-delivery systems. This localized approach improves the osteogenic potential of bone substitutes by delivering bone growth factors, thus extending their biofunctionality to any pathological context, including infection, irradiation, tumor and osteoporosis. This review briefly describes the physical and chemical processes implicated in the preparation of drug-delivering CaPs. It also describes the impact of these processes on the intrinsic properties of CaPs, especially in terms of the drug-release profile. In addition, this review focuses on the potential influence of drugs on the resorption rate of CaPs. Interestingly, by modulating the resorption parameters of CaP biomaterials, it should be possible to control the release of bone-stimulating ions, such as inorganic phosphate, in the vicinity of bone cells. Finally, recent in vitro and in vivo evaluations are extensively reported.     

Evaluation of calcium phosphates and experimental calcium phosphate bone cements using osteogenic cultures

In this study, rat bone marrow cells (RBM) were used to evaluate two biodegradable calcium phosphate bone cements and bioactive calcium phosphate ceramics. The substances investigated were: two novel calcium phosphate cements, Biocement F and Biocement H, tricalcium phosphate (TCP), surface-modified alpha-tricalcium phosphate [TCP (s)] and a rapid resorbable calcium phosphate ceramic consisting of CaKPO(4) (sample code R5). RBM cells were cultured on disc-shaped test substrates for 14 days. The culture medium was changed daily and also examined for calcium, phosphate, and potassium concentrations. Specimens were evaluated using light microscopy, and morphometry of the cell-covered substrate surface, scanning electron microscopy, and energy dispersive X-ray analysis and morphometry of the cell-covered substrate surface. Areas of mineralization were identified by tetracyline labeling. Except for R 5, rat bone-marrow cells attached and grew on all substrate surfaces. Of the different calcium phosphate materials tested, TCP and TCP (s) facilitated osteoblast growth and extracellular matrix elaboration to the highest degree, followed by Biocements H and F. The inhibition of cell growth encountered with R 5 seems to be related to its high phosphate and potassium ion release.     

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.     

Processing and in vivo evaluation of multiphasic calcium phosphate cements with dual tricalcium phosphate phases

Calcium phosphate cements (CPCs) use the simultaneous presence of several calcium phosphates phases. This is done to generate specific bulk and in vivo properties. This work has processed and evaluated novel multiphasic CPCs containing dual tricalcium phosphate (TCPs) phases. Dual TCPs containing α- and β-TCP phases were obtained by thermal treatment. Standard CPC (S-CPC) was composed of α-TCP, anhydrous dicalcium phosphate and precipitated hydroxyapatite, while modified CPC (DT-CPC) included both α- and β-TCP. Physicochemical characterization of these CPCs was based on scanning electron microscopy, X-ray diffraction, specific surface area (SSA) and particle size (PS) analysis and mechanical properties. This characterization allowed the selection of one DT-CPC for setting time, cohesion and biological assessment compared with S-CPC. Biological assessment was carried out using a tibial intramedullary cavity model and subcutaneous pouches in guinea pigs. Differences in the surface morphology and crystalline phases of the treated TCPs were detected, although PS analysis of the milled CPC powders produced similar results. SSA analysis was significantly higher for DT-CPC with α-TCP treated at 1100°C for 5h. Poorer mechanical properties were found for DT-CPC with α-TCP treated at 1000°C. Setting time and cohesion, as well as the in vivo performance, were similar in the selected DT-CPC and the S-CPC. Both CPCs created the desired host reactions in vivo.     

Heparin modification of calcium phosphate bone cements for VEGF functionalization

A promising strategy to promote angiogenesis within an engineered tissue is the local and sustained delivery of an angiogenic factor by the substitute itself. Recently, we reported on functionalization of Biocement D (BioD) and several modifications of this calcium phosphate bone cement with vascular endothelial growth factor (VEGF). Maintenance of biological activity of VEGF after release from the cement was improved by modification of BioD with mineralized collagen type I (BioD/coll). However, BioD/coll composites showed a higher initial burst of VEGF release than do the unmodified BioD. In the present study, VEGF release from BioD/coll composites modified with different amounts of heparin was investigated. We found a distinct reduction of the initial burst of release by adding heparin in a concentration-dependent manner. Moreover, the heparin modification had a positive impact on the biological activity of released VEGF. An advancement of biological properties of BioD/coll by addition of heparin was further shown by improved adhesion of endothelial cells on the cement surface. Characterization of material properties of the heparin-modified BioD/coll composites revealed a finer microstructure with smaller HA-particles and a higher specific surface area than heparin-free BioD/coll. However, higher amounts of heparin resulted in a reduced compressive strength. The rheological properties of these cement pastes have been found to be favorable for good handling particularly with regard to their clinical application.     

Bioactive bone cements containing nano-sized titania particles for use as bone substitutes

Three types of bioactive polymethylmethacrylate (PMMA)-based bone cement containing nano-sized titania (TiO2) particles were prepared, and their mechanical properties and osteoconductivity are evaluated. The three types of bioactive bone cement were T50c, ST50c, and ST60c, which contained 50 wt% TiO2, and 50 and 60 wt% silanized TiO2, respectively. Commercially available PMMA cement (PMMAc) was used as a control. The cements were inserted into rat tibiae and allowed to solidify in situ. After 6 and 12 weeks, tibiae were removed for evaluation of osteoconductivity using scanning electron microscopy (SEM), contact microradiography (CMR), and Giemsa surface staining. SEM revealed that ST60c and ST50c were directly apposed to bone while T50c and PMMAc were not. The osteoconduction of ST60c was significantly better than that of the other cements at each time interval, and the osteoconduction of T50c was no better than that of PMMAc. The compressive strength of ST60c was equivalent to that of PMMAc. These results show that ST60c is a promising material for use as a bone substitute.     

Kinetic study of citric acid influence on calcium phosphate bone cements as water-reducing agent

Most of the research performed on calcium phosphate bone cements (CPBCs) has dealt with the improvement of bone cement formulations for new, demanding bone-filling applications. In particular, the development of injectable bone cements is of real interest for the biomedical community. The aim of this work was to study the effect of citric acid on the injectability and the setting properties of alpha-tricalcium phosphate-based cements. A comparative kinetic study was performed on cements with and without citric acid relating the hardening curves and the hydration rates using a mathematical approach. Citric acid behaved as a fluidificant during the first stages of the cement mixing. The dissolution-precipitation reactions of the alpha-tricalcium phosphate were retarded with the addition of citric acid and the compressive strength at saturation increased. In conclusion, citric acid can behave as a water-reducing admixture.     

Conversion of octacalcium phosphate in calcium phosphate cements

This study investigated the in vitro conversion reaction in calcium phosphate cements (CPCs) containing octacalcium phosphate (OCP) as one of the reagents. OCP is known to be a precursor for apatite formation in vivo. The reaction products were characterized using infrared spectroscopy and X-ray diffraction. Although the conversion of OCP into hydroxyapatite is thermodynamically favorable, OCP only yields apatite formation in CPC provided it is combined with a highly soluble Ca(2+) and OH(-) releasing reaction partner. In this respect, tetracalcium phosphate is a promising compound. Adding small amounts of monocalcium phosphate monohydrate can stimulate the setting through intermediate brushite formation. The preparation method of OCP might drastically affect the performance of the cement. The reaction path of the setting of these CPC probably does not conform to the singular point principle described in the literature, and an in situ hydrolysis of OCP to apatite is conceivable. Simulation of apatite formation using OCP as the precursor and/or seed in CPC might be beneficial for some biomedical applications.     

New depowdering-friendly designs for three-dimensional printing of calcium phosphate bone substitutes

Powder-based three-dimensional printing (3DP) is a versatile method that allows creating synthetic calcium phosphate (CaP) scaffolds of complex shapes and structures. However, one major drawback is the difficulty of removing all remnants of loose powder from the printed scaffolds, the so-called depowdering step. In this study, a new design approach was proposed to solve this problem. Specifically, the design of the printed scaffolds consisted of a cage with windows large enough to enable depowdering while still trapping loose fillers placed inside the cage. To demonstrate the potential of this new approach, two filler geometries were used: sandglass and cheese segment. The distance between the fillers was varied and they were either glued to the cage or free to move after successful depowdering. Depowdering efficiency was quantified by microstructural morphometry. The results showed that the use of mobile fillers significantly improved depowdering. Based on this study, large 3DP scaffolds can be realized, which might be a step towards a broader clinical use of 3D printed CaP scaffolds.     

Effect of ball milling on the processing of bone substitutes with calcium phosphate powders

Decreasing the microscale morphology of synthetic bone substitutes is of primary importance in order to enhance the morphology of the surface of the material, which is directly in contact with osteoconductive cells when it is implanted in bone. The aim of this study was to investigate the influence of ball milling of slurries on the microscale morphology of hydroxyapatite and tricalcium phosphate bone substitutes and the influence on their processing. Ball milling appeared to be a successful method in order to raise the sintering reactivity of the powders, that is, to decrease the sintering temperature and microstructural morphology of the material. However, it was demonstrated that ball milling had a great influence on dispersion, which became very difficult under long milling times because of dissolution of the calcium phosphate powders. Due to dissolution, ionic species were generated in the slurry and interfered with the dispersing agent. Moreover a reprecipitation process occurred simultaneously, and large particles of the most stable phase (HAP) formed. The presence of such large particles generated stress gradients and cracks in the material during the sintering stage.     

Characterization of calcium phosphate cements modified by addition of amorphous calcium phosphate

In this study the influence of amorphous calcium phosphate (ACP) on the setting of, and the formed apatite crystallite size in, a calcium phosphate cement (CPC) based on α-tricalcium phosphate (α-TCP) or tetracalcium phosphate (TTCP)/monocalcium phosphate monohydrate (MCPM) was investigated. Setting times at 22 °C were measured in air atmosphere; those at 37 °C were measured at 100% relative humidity. The phase composition of the set cements was investigated after 1 week using X-ray diffractometry and infrared spectroscopy and the morphology was investigated using scanning electron microscopy. The compressive strength (CS) of the set CPCs was measured after 1 day. Viability of MC3T3-E1 cells on the CPCs was analyzed after 7, 14 and 21 days of incubation using the CellTiter 96^® Aqueous Non-Radioactive Cell Proliferation Assay. The α-TCP-based cement exhibited long setting times, a high CS and was converted to a calcium-deficient hydroxyapatite (CDHAp). The TTCP/MCPM-based CPC was only partly converted to CDHAp, produced acceptable setting times and had a low CS. Addition of ACP to these two CPCs resulted in cements that exhibited good setting times, CS suitable for non-load-bearing applications and a full conversion to nanocrystalline CDHAp. Moreover, the ACP containing CPCs demonstrated good cell viability, making them suitable candidates for bone substitute materials.     

Biodegradation behavior of various calcium phosphate materials in bone tissue

In order to study the biodegradation behavior of calcium phosphate materials, cylinders of standard size were implanted in the tibiae of rabbits. Material parameters were stoichiometry (hydroxyapatite with a Ca/P ratio of 1.67 versus tricalcium phosphate with a Ca/P ratio of 1.50), crystallographic structure (apatite versus beta-whitlockite), microporosity, and macroporosity. The extent of biodegradation was evaluated by radiography, light and fluorescence microscopy, microradiography, and porosity measurements. All calcium phosphate materials were biocompatible in bone tissue. Hydroxyapatite ceramics had a higher osteogenic potential than beta-whitlockite materials. Depending on their porosities, sintered tricalciumphosphate (beta-whitlockite) materials were more or less biodegradable, in contrast to sintered hydroxyapatite materials, which showed no detectable resorption over a period of 9 months of implantation.     

In vivo bone response to porous calcium phosphate cement

We conducted an in vivo experiment to evaluate the resorption rate of a calcium phosphate cement (CPC) with macropores larger than 100 microm, using the CPC called Biocement D (Merck Biomaterial, Darmstadt, Germany), which after setting only shows pores smaller than 1 microm. The gas bubble method used during the setting process created macroporosity. Preset nonporous and porous cement implants were inserted into the trabecular bone of the tibial metaphysis of goats. The size of the preset implants was 6 mm and the diameter of the drill hole was 6.3 mm, leaving a gap of 0.3 mm between implant surface and drill wall. After 2 and 10 weeks, the animals were euthanized and cement implants with surrounding bone were retrieved for histologic evaluation. Light microscopy at 2 weeks revealed that the nonporous implants were surrounded by connective tissue. On the cement surface, we observed a monolayer of multinucleated cells. Ten weeks after implantation, the nonporous implants were still surrounded by connective tissue. However, a thin layer of bone now covered the implant surface. No sign of cement resorption was observed. In contrast, the porous cement evoked a completely different bone response. At 2 weeks, bone formation had already occurred inside the implant porosity. Bone formation even appeared to occur as a result of osteoinduction. Also, at their outer surface, the porous implants were completely surrounded by bone. At 2 weeks, about 31% of the initial cement was resorbed. After 10 weeks, 81% of the initial phosphate cement was resorbed and new bone was deposited. On the basis of these observations, we conclude that the creation of macropores can significantly improve the resorption rate of CPC. This increased degradation is associated with almost complete bone replacement.     

Frozen delivery of brushite calcium phosphate cements

Calcium phosphate cements typically harden following the combination of a calcium phosphate powder component with an aqueous solution to form a matrix consisting of hydroxyapatite or brushite. The mixing process can be very important to the mechanical properties exhibited by cement materials and consequently when used clinically, since they are usually hand-mixed their mechanical properties are prone to operator-induced variability. It is possible to reduce this variability by pre-mixing the cement, e.g. by replacing the aqueous liquid component with non-reactive glycerol. Here, for the first time, we report the formation of three different pre-mixed brushite cement formulations formed by freezing the cement pastes following combination of the powder and liquid components. When frozen and stored at -80 degrees C or less, significant degradation in compression strength did not occur for the duration of the study (28 days). Interestingly, in the case of the brushite cement formed from the combination of beta-tricalcium phosphate with 2 M orthophosphoric acid solution, freezing the cement paste had the effect of increasing mean compressive strength fivefold (from 4 to 20 MPa). The increase in compression strength was accompanied by a reduction in the setting rate of the cement. As no differences in porosity or degree of reaction were observed, strength improvement was attributed to a modification of crystal morphology and a reduction in damage caused to the cement matrix during manipulation.     

Comparison of hydroxyapatite and beta tricalcium phosphate as bone substitutes after excision of bone tumors

Long-term results are reported in 23 patients and short-term results in 30 patients presenting with bone tumors treated by curettage or resection followed by implantation of hydroxyapatite (HA) or highly purified beta-tricalcium phosphate (beta-TCP), respectively. Mean follow-up was 97 and 26 months in cases involving HA implantation and beta-TCP implantation, respectively. Radiographs revealed HA incorporation into host bone in all but two cases; moreover, no obvious evidence of HA biodegradation was observed. A single patient exhibited late deformity following implantation of HA. All grafted beta-TCP was, at least partially, absorbed and replaced by newly formed bone. The mean period required for the disappearance of radiolucent zones between the ceramics and host bone was 17 weeks in HA and 9.7 weeks in beta-TCP. Highly purified beta-TCP appears to be advantageous relative to HA for surgical intervention in bone tumors consequent to the nature of remodeling and superior osteoconductivity.     

Histological and mechanical evaluation of self-setting calcium phosphate cements in a sheep vertebral bone void model

We investigated the histological and compressive properties of three different calcium phosphate cements (CPCs) using a sheep vertebral bone void model. One of the CPCs contained barium sulfate to enhance its radiopacity. Bone voids were surgically created in the lumbar region of 23 ovine spines - L3, L4, and L5 (n = 69 total vertebral bodies) - and the voids were filled with one of the three CPCs. A fourth group consisted of whole intact vertebrae. Histologic evaluation was performed for 30 of the 69 vertebrae 2 or 4 months after surgery along with radiographic evaluation. Compressive testing was performed on 39 vertebrae 4 months after surgery along with micro-CT analysis. All three CPCs were biocompatible and extremely osteoconductive. Osteoclasts associated with adjacent bone formation suggest that each cement can undergo slow resorption and replacement by bone and bone marrow. Compressive testing did not reveal a significant difference in the ultimate strength, ultimate strain, and structural modulus, among the three CPCs and intact whole vertebrae. Micro-CT analysis revealed good osseointegration between all three CPCs and adjacent bone. The barium sulfate did not affect the CPCs biocompatibility or mechanical properties. These results suggest that CPC might be a good alternative to polymethylmethacrylate for selected indications.