Biological properties of calcium phosphate biomaterials for bone repair: a review

Bone defects are a common disease threatening the health of many people. Calcium phosphate (CaP) is an ideal bone substitutive material that is widely used for bone repair due to its excellent biological properties including osteoinductivity, osteoconductivity and biodegradability. For this reason, investigation of these properties and the effects of various influencing factors is vital for modulating calcium phosphate during the design process to maximally satisfy clinical requirements. In this study, the latest studies on the biological properties of CaP biomaterials, including hydroxyapatite (HA), tricalcium phosphate (TCP), and biphasic calcium phosphate (BCP), have been summarized. Moreover, recent advances on how these properties are altered by different factors are reviewed. Considering the limited mechanical strength of CaP materials, this study also reviews CaP composites with different materials as improvement measures. Finally, perspectives regarding future developments of CaP materials are also provided.

This article reviews the recent advances and various factors affecting the improvement of the biological properties of calcium phosphate for bone repair.     

In vitro and in vivo assessment of nanostructured porous biphasic calcium phosphate ceramics for promoting osteogenesis in an osteoporotic environment

Treatment of bone defects in osteoporotic patients with bone substitutes is difficult, due to insufficient osseointegration. The development of appropriate biomaterials to solve the problem requires the assessment of the material performance in an osteoporotic environment, which is rarely investigated. Herein, nanostructured biphasic calcium phosphate (nBCP) ceramics were prepared via the incorporation of hydroxyapatite nanoparticles (HANPs) into porous biphasic CaP (BCP) substrates, leading to an increase of over 500% in the specific surface area. Primary osteoblasts harvested from osteoporotic rats were cultured on the nBCP ceramics, and it was found that the osteoblast functions, including proliferation, alkaline phosphatase activity, osteocalcin secretion and expression of osteogenic genes, were significantly enhanced compared with osteoblasts grown on non-nanostructured BCP ceramics. To further assess the osteoinduction ability, the ceramics were implanted in the femur of osteoporotic rats. Compared to the rats implanted with non-nanostructured BCP ceramics, a higher amount of mechanically matured bone was newly formed in the rats with nBCP ceramics after 6 weeks of implantation. Such enhanced osteoinduction ability of the nBCP ceramics may be due to the incorporated HANPs, as well as the nanostructured topography induced by the HANPs. These results indicate good in vitro and in vivo osteoinductivity of the nBCP ceramics in an osteoporotic environment and offer potential benefits for treating bone defects in osteoporotic patients.

Nanostructured porous biphasic calcium phosphate ceramics are able to significantly promote bone defect healing in an osteoporotic environment.     

Sintered porous Ti6Al4V scaffolds incorporated with recombinant human bone morphogenetic protein-2 microspheres and thermosensitive hydrogels can enhance bone regeneration

A well-controlled powder sintering technique was used to fabricate porous Ti6Al4V scaffold. The thermosensitive chitosan thioglycolic acid (CS-TA) hydrogel was used as a carrier to inject recombinant human bone morphogenetic protein-2 (rhBMP-2) microspheres into pores of the Ti6Al4V scaffold at 37 °C, and then the porous Ti6Al4V/rhBMP-2 loaded hydrogel composite was obtained. The bare Ti6Al4V scaffold was used as the control. The characteristics and mechanical properties of the scaffold, rheological properties of the hydrogels and the rhBMP-2 loaded hydrogel, the release of the rhBMP-2 loaded hydrogel, and the biological properties of the two types of samples were evaluated by in vitro and in vivo tests. Results indicated that the sintered porous Ti6Al4V had high porosity, large pore size with good mechanical properties. The hydrogel and rhBMP-2 loaded hydrogel showed thermosensity. The rhBMP-2 loaded hydrogel showed a stable and extended release profile without too high burst release of rhBMP-2. Both groups showed good biocompatibility and osteogenic ability. However, according to the results of cell tests and implantation, the group with rhBMP-2 loaded hydrogel had significantly higher cell proliferation rate, faster bone growth speed, and more bone ingrowth at every time point. Therefore, the sintered porous Ti6Al4V scaffolds incorporated with rhBMP-2 microspheres and CS-TA hydrogel was effective in enhancing the bone regeneration, and prospects a good candidate for application in orthopedics.

Porous Ti6Al4V scaffolds incorporated with rhBMP-2 microspheres and CS-TA hydrogel can enhance the bone regeneration.     

In vitro and in vivo evaluations of mechanical properties,biocompatibility and osteogenic ability of sintered porous titanium alloy implant

A new sintering technique using Ti₆Al₄V powder suspension was performed to prepare porous Ti₆Al₄V alloys. The porous alloys could be fabricated with different porosities and pore sizes by controlling the quantity and size of spacer particles added to the Ti₆Al₄V powder. The characteristics and biological properties of the porous Ti₆Al₄V with two different porosities were evaluated by mechanical tests, cell tests and implantation. Dense Ti₆Al₄V was used as the control. Compared with the control group, the porous Ti₆Al₄V showed good biocompatibility and osteogenic ability, which makes this type of porous alloy a good prospective material for biomedical application. And compared with 50% porosity, the alloy with 75% porosity had the optimal mechanical properties, and suitable pore size and porosity, which allowed more bone ingrowth.

The sintered porous Ti₆Al₄V with 75% porosity has optimal mechanical properties, good biocompatibility and osteogenic ability for more bone ingrowth.     

Effect of biodegradable magnesium-copper coatings on bone integration based on the porous structures in a rabbit model

Biodegradable magnesium (Mg)–copper (Cu) coatings are promising in orthopedic implants to enhance osteogenesis. The present study aimed to compare the osteogenic effect of Mg–Cu coating and non-coating implants using a rabbit model. Mg/Mg–Cu coating of porous Ti6Al4V alloys was performed by the arc ion plating method. Five porous implants (smooth, porous, Mg coated, Mg–0.1Cu coated, and Mg–0.7Cu coated) were implanted into the distal femurs of the rabbits. The rabbits were sacrificed after one and two months, respectively, and the distal femurs with the implants were analyzed via micro-computed tomography (CT), double fluorescent labeling, and hard tissue biopsy to evaluate their osteogenic effect. During the two months of observation, the Mg/Mg–Cu coatings exhibited no advantages when compared with the bare porous structures in terms of bone integration; however, the porous structures were more conducive for bone ingrowth than the smooth implants. The osteogenic application of Mg–Cu coated orthopedic implants is worth further investigation. Furthermore, due to its long-term antibacterial ability, the biodegradable Mg–Cu coatings are promising in orthopedic applications.

The results of the present study suggest that the application of Mg–Cu coating in orthopedic implants is mainly due to its long-term antibacterial ability rather than its osteogenesis ability.     

Fabrication of a Cu/Zn co-incorporated calcium phosphate scaffold-derived GDF-5 sustained release system with enhanced angiogenesis and osteogenesis properties

Synthetic scaffolds with multifunctional properties, including angiogenesis and osteogenesis capacities, play an essential role in accelerating bone regeneration. In this study, various concentrations of Cu/Zn ions were incorporated into biphasic calcium phosphate (BCP) scaffolds, and then growth differentiation factor-5 (GDF-5)-loaded poly(lactide-co-glycolide) (PLGA) microspheres were attached onto the ion-doped scaffold. The results demonstrated that with increasing concentration of dopants, the scaffold surface gradually changed from smooth grain crystalline to rough microparticles, and further to a nanoflake film. Additionally, the mass ratio of β-tricalcium phosphate/hydroxyapatite increased with the dopant concentration. Furthermore, GDF-5-loaded PLGA microspheres attached onto the BCP scaffold surface exhibited a sustained release. In vitro co-culture of bone mesenchymal stem cells and vascular endothelial cells showed that the addition of Cu/Zn ions and GDF-5 in the BCP scaffold not only accelerated cell proliferation, but also promoted cell differentiation by enhancing the alkaline phosphatase activity and bone-related gene expression. Moreover, the vascular endothelial growth factor secretion level increased with the dopant concentration, and attained a maximum when GDF-5 was added into the ions-doped scaffold. These findings indicated that BCP scaffold co-doped with Cu/Zn ions exhibited a combined effect of both metal ions, including angiogenic and osteogenic capacities. Moreover, GDF-5 addition further enhanced both the angiogenic and osteogenic capacities of the BCP scaffold. The Cu/Zn co-incorporated BCP scaffold-derived GDF-5 sustained release system produced multifunctional scaffolds with improved angiogenesis and osteogenesis properties.

A Cu/Zn co-incorporated BCP scaffold-derived GDF-5 sustained release system was successfully prepared and exhibited improved angiogenic and osteogenic capacities.     

Icariin controlled release on a silk fibroin/mesoporous bioactive glass nanoparticles scaffold for promoting stem cell osteogenic differentiation

The treatment of bone defects caused by various reasons is still a major problem in orthopedic clinical work. Many studies on osteogenic implant materials have used various biologically active factors such as osteogenic inducers, but these biologically active factors have various side effects. Therefore, in this study, silk fibroin (SF) was used as a scaffold material, mesoporous bioactive glass nanoparticles (MBGNs) as a sustained release carrier, and the traditional Chinese drug icariin (ICA) was loaded to promote bone formation. The experiments in this study have proven that SF/MBGNs-ICA scaffolds can successfully load and release ICA for a long time, and the sustained-release ICA can promote the proliferation and differentiation of BMSCs for a long time. This controlled-release ICA organic/inorganic two-component scaffold material is expected to become a new bone grafting solution.

Long-term promotion of osteogenic differentiation through silk fibroin/mesoporous bioactive glass-loaded sustained release of icariin.     

Cell-tailored calcium carbonate particles with different crystal forms from nanoparticle to nano/microsphere

Inspired by biomineralization, the first synthesis of size-tunable calcium carbonates from nanoparticles (YC-CaCO₃ NPs) to nano/microspheres (YC-CaCO₃ N/MSs) with a porous structure was accomplished using a facile method under the mediation of the secretion from yeast cells (YCs). The biomolecules derived from the secretion of YCs were used as conditioning and stabilizing agents to control the biosynthesis of the YC-CaCO₃ materials. The morphology and crystal forms of YC-CaCO₃ materials can be affected by the biomolecules from the secretion of YCs. With increasing concentrations of biomolecules, the morphologies of the obtained CaCO₃ materials changed from nanoparticles to nano/microspheres with a porous structure, while the crystal forms transformed from amorphous to calcite. Functional investigations showed that YC-CaCO₃ NSs with a porous structure effectively acted as anticancer drug carriers with accurate and selective drug release in tumor tissue, which suggests that they have great potential to function as a therapeutic delivery system. These application features are mainly attributed to the satisfactory biocompatibility and biodegradability, high drug-loading capacity, and pH-dependent sustained drug release performance of the porous YC-CaCO₃ NSs. The biomimetic synthesis strategy of YC-CaCO₃ materials mediated by YC secretion not only helps to shed light on the biomineralization mechanism in organisms, but may also lead to a new means of biosynthesizing organic–inorganic nanocomposites.

The synthesis of cell-tailored calcium carbonate with different crystal forms can be controlled from nanoparticle to nano/microsphere by a bio-inspired strategy.     

Reduced graphene oxide-incorporated calcium phosphate cements with pulsed electromagnetic fields for bone regeneration

Natural calcium phosphate cements (CPCs) derived from sintered animal bone have been investigated to treat bone defects, but their low mechanical strength remains a critical limitation. Graphene improves the mechanical properties of scaffolds and promotes higher osteoinduction. To this end, reduced graphene oxide-incorporated natural calcium phosphate cements (RGO-CPCs) are fabricated for reinforcement of CPCs'' characteristics. Pulsed electromagnetic fields (PEMFs) were additionally applied to RGO-CPCs to promote osteogenic differentiation ability. The fabricated RGO-CPCs show distinct surface properties and chemical properties according to the RGO concentration. The RGO-CPCs’ mechanical properties are significantly increased compared to CPCs owing to chemical bonding between RGO and CPCs. In in vitro studies using a mouse osteoblast cell line and rat-derived adipose stem cells, RGO-CPCs are not severely toxic to either cell type. Cell migration study, western blotting, immunocytochemistry, and alizarin red staining assay reveal that osteoinductivity as well as osteoconductivity of RGO-CPCs was highly increased. In in vivo study, RGO-CPCs not only promoted bone ingrowth but also enhanced osteogenic differentiation of stem cells. Application of PEMFs enhanced the osteogenic differentiation of stem cells. RGO-CPCs with PEMFs can overcome the flaws of previously developed natural CPCs and are anticipated to open the gate to clinical application for bone repair and regeneration.

Natural calcium phosphate cements (CPCs) derived from sintered animal bone have been investigated to treat bone defects, but their low mechanical strength remains a critical limitation.     

Fe/Zn-modified tricalcium phosphate (TCP) biomaterials: preparation and biological properties

Bone repairing materials play an essential role in the repair treatment of bone defects. The presence of calcium phosphate invertebrates is of significance for bone repairing processes. However, the mechanical properties and osteogenic activities of many current calcium phosphate materials are not ideal, which limit their biological applications. Therefore, it is an effective alternative strategy to study the modification of calcium phosphate biomaterials to address these limitations. In this research, in order to enhance the biological performance of tricalcium phosphate (β-TCP), metal species (Fe and Zn) modified β-TCP materials through the co-precipitation method were successfully developed. The physical, chemical and biological properties of the binary composites were carefully studied for the first time. The bioactivities of the Fe-TCP and Zn-TCP were evaluated by simulating body fluid (SBF) immersion experiments, blood compatibility, and cytotoxicity tests. The findings demonstrated that the metal-TCP with excellent cytocompatibility and osteogenic properties shows good potential in medical applications.

Fe/Zn-TCP biomaterials were prepared and their bioactivities to enhance the synthetic bone-repair materials were studied in comparison.     

Repair of segmental rabbit radial defects with Cu/Zn co-doped calcium phosphate scaffolds incorporating GDF-5 carrier

Repair of segmental bone defects is a challenge in orthopaedics. A bone substitute is a potential solution for this challenge, and angiogenesis and osteogenesis are critical to the performance of scaffold materials. For enhancing angiogenesis and osteogenesis activities of implanted scaffolds, Cu/Zn co-doped calcium phosphate scaffolds carrying GDF-5-release microspheres were prepared and implanted into surgically created critical-sized rabbit radial defects. Radiological examination, histological analysis and biomechanical tests were used to evaluate the bone healing-union. Results showed that, with increasing Cu/Zn concentrations, new bone area, new blood vessel density, and bending failure load all increased significantly. Furthermore, Cu/Zn co-doped scaffolds incorporating GDF-5-release microspheres exhibited further increased angiogenesis and osteogenesis (vs. Cu/Zn co-doped alone), as well as a superior bending failure load. These show that, simultaneous incorporation of trace essential ions and GDF-5 combines pro-angiogenic and pro-osteogenic actions of these bioactive substances, potentially offering an effective approach to assist the healing of critical-sized bone defects.

Cu/Zn co-doped calcium phosphate scaffolds carrying GDF-5-release microspheres were prepared and implanted into surgically created critical-sized rabbit radial defects.     

Self-assembled hierarchical porous NiMn2O4 microspheres as high performance Li-ion battery anodes

Hierarchical structured porous NiMn₂O₄ microspheres assembled with nanorods are synthesized through a simple hydrothermal method followed by calcination in air. As anode materials for lithium ion batteries (LIBs), the NiMn₂O₄ microspheres exhibit a high specific capacity. The initial discharge capacity is 1126 mA h g⁻¹. After 1000 cycles, the NiMn₂O₄ demonstrates a reversible capacity of 900 mA h g⁻¹ at a current density of 500 mA g⁻¹. In particular, the porous NiMn₂O₄ microspheres still could deliver a remarkable discharge capacity of 490 mA h g⁻¹ even at a high current density of 2 A g⁻¹, indicating their potential application in Li-ion batteries. This excellent electrochemical performance is ascribed to the unique hierarchical porous structure which can provide sufficient contact for the transfer of Li⁺ ion and area for the volume change of the electrolyte leading to enhanced Li⁺ mobility.

Hierarchical structured porous NiMn₂O₄ microspheres assembled with nanorods are synthesized through a simple hydrothermal method followed by calcination in air.     

A three-dimensional hydroxyapatite/polyacrylonitrile composite scaffold designed for bone tissue engineering

In recent years, various composite scaffolds based on hydroxyapatite have been developed for bone tissue engineering. However, the poor cell survival micro-environment is still the major problem limiting their practical applications in bone repairing and regeneration. In this study, we fabricated a class of fluffy and porous three-dimensional composite fibrous scaffolds consisting of hydroxyapatite and polyacrylonitrile by employing an improved electrospinning technique combined with a bio-mineralization process. The fluffy structure of the hydroxyapatite/polyacrylonitrile composite scaffold ensured the cells would enter the interior of the scaffold and achieve a three-dimensional cell culture. Bone marrow mesenchymal stem cells were seeded into the scaffolds and cultured for 21 days in vitro to evaluate the response of cellular morphology and biochemical activities. The results indicated that the bone marrow mesenchymal stem cells showed higher degrees of growth, osteogenic differentiation and mineralization than those cultured on the two-dimensional hydroxyapatite/polyacrylonitrile composite membranes. The obtained results strongly supported the fact that the novel three-dimensional fluffy hydroxyapatite/polyacrylonitrile composite scaffold had potential application in the field of bone tissue engineering.

A fluffy and porous (3D) HA composite fibrous scaffold was fabricated by employing an improved electrospinning technique combined with a bio-mineralization process.     

Hybrid catalysts based on N-heterocyclic carbene anchored on hierarchical zeolites

Hybrid materials have been synthesized by anchoring a N-heterocyclic carbene (NHC) precursor on different inorganic zeolitic supports with hierarchical porosity, in particular hierarchical HZSM-5 and SAPO-5. Hierarchical porous inorganic supports have been obtained both by top-down and bottom-up approaches and the role of hierarchical porosity has been evaluated. A detailed physico-chemical characterization has been performed on the organic–inorganic hybrids using a multi-technique approach (XRD, volumetric and thermogravimetric analysis, ssNMR and FTIR) in order to establish a structure–property relationship. The hybrids were tested in the benzoin condensation reaction of furfural, a base catalyzed reaction.

N-heterocyclic carbene has been grafted on hierarchical HZSM-5 and SAPO-5 to obtain hybrid catalysts, that are active in the benzoin condensation reaction.     

SiO2 thin film growth through a pure atomic layer deposition technique at room temperature

In this study, less contaminated and porous SiO₂ films were grown via ALD at room temperature. In addition to the well-known catalytic effect of ammonia, the self-limitation of the reaction was demonstrated by tuning the exposure of SiCl₄, NH₃ and H₂O. This pure ALD approach generated porous oxide layers with very low chloride contamination in films. This optimized RT-ALD process could be applied to a wide range of substrates that need to be 3D-coated, similar to mesoporous structured membranes.

In this study, less contaminated and porous SiO₂ films were grown via ALD at room temperature.     

Synthesis and plugging behavior of fluorescent polymer microspheres as a kind of conformance control agent in reservoirs

The fluorescent polymer microsphere is a newly developed chemical agent for conformance control in reservoirs. In this paper, one kind of fluorescent polymer microspheres P(AM-BA-RhB) was synthesized via the inverse suspension polymerization method with Rhodamine B as a fluorescence functional monomer. Laboratory experiments were performed to characterize the morphology, fluorescent property, swelling property and plugging behavior of fluorescent polymer microspheres. The experimental results showed that the polymer microspheres P(AM-BA-RhB) displayed stable fluorescence performance in solutions containing metal ions at pH values between 3.0 and 10.0. The swelling property was not dramatically affected by the Rhodamine B embedded in the polymer microspheres by grafting. Both a visual micromodel test and sand-pack tubes experiment demonstrated that the fluorescent polymer microspheres could pass directly or by deformation through porous media and get into the in-depth formation. The injection pressure showed the phenomenon of “Wave-type Variation”. Three plugging behaviors such as piston plugging, protruding plugging and fingering plugging were put forward. The introduction of fluorescent polymer microspheres could provide one method to research the conformance control and EOR mechanism of polymer microspheres in the reservoirs.

Three plugging behavior such as piston plugging, protruding plugging and fingering plugging were put forward.     

Nanoporous materials with predicted zeolite topologies

An increasing number of newly synthesized materials have been found to be previously present in databases of predicted porous materials. This has been observed not only for zeolites, but also for other inorganic materials and for MOFs. We here quantify the number of synthesized zeolites that are present in a large database of predicted zeolite structures as well as the number of other inorganic crystals and MOFs present in this same database. We find a significant number of real materials are in this predicted database of zeolite-like structures. These results suggest that many other predicted structures in this database may be suitable targets for designer materials synthesis.

Topological exploration of crystal structures demonstrates the presence of known zeolites, inorganics, and MOFs in a database of predicted materials.     

Heavy oil oxidation in the nano-porous medium of synthetic opal

Increasing interest to study hydrocarbon behavior in fine porous media, awakened by the shale revolution, requires the application of suitable model porous media. In the current study we prepared nano-porous synthetic opal, profoundly investigated its morphological and textural properties, and studied the kinetics of combustion of heavy oil impregnated into nanopores. Comparison of kinetic parameters of the oil oxidation process for nano-porous and coarse-porous media revealed that nanoconfinement affects the reactivity of oil.

In the current study we synthesized nano-porous opal, investigated its morphological and textural properties, and showed that nanoconfinement affects reactivity of oil.     

Inorganic process for wet silica-doping of calcium phosphate

Silica is not only a biocompatible trace element but also an essential element for bone formation and metabolism. Therefore, it is often doped into bioceramics such as calcium phosphate and calcium carbonate for enhancing biomaterial ability. Heretofore, organic silica materials are employed as silica sources, but the residual organic matter is a significant drawback in biomaterial applications. Therefore, in this study, we introduce a one-pot inorganic synthesis method for the formation of silica-doped octacalcium phosphate (OCP) using Na₂SiO₃ as the silica source. Silica was intercalated into the OCP unit lattice, replacing its hydrous layer structure, and then a layer-by-layer structure of apatite and silica was formed. Furthermore, by immersing the fabricated silica-doped OCP into suitable solutions, both silica-doped hydroxyapatite and carbonate apatite were fabricated through a one-step inorganic processes.

We introduced a one-pot synthesis method for silica doping of calcium phosphate. Silica easily incorporated into OCP interlayer optimizing Na₂SiO₃ concentrations.