Calcium Phosphate Materials Research Articles

Staged implant placement after defect regeneration using biphasic calcium phosphate materials with different surface topographies in a minipig model

ObjectiveTo assess the influence of biphasic calcium phosphate materials with different surface topographies on bone formation and osseointegration of titanium implants in standardized alveolar ridge defects.Materials and methodsStandardized alveolar ridge defects (6 × 6 mm) were created in the mandible of 8 minipigs and filled with three biphasic calcium phosphate materials (BCP1–3, 90% tricalcium phosphate/10% hydroxyapatite) with different surface properties (micro- and macroporosities) as well as a bovine-derived natural bone mineral (NBM) as a control. At 12 weeks, implants were placed into the augmented defects. After further 8 weeks of healing, dissected blocks were processed for histological analysis (e.g., mineralized (MT), residual bone graft material (BS), bone-to-implant contact (BIC)).ResultsAll four biomaterials showed well-integrated graft particles and new bone formation within the defect area. MT values were comparable in all groups. BS values were highest in the NBM group (21.25 ± 13.52%) and markedly reduced in the different BCP groups, reaching statistical significance at BCP1-treated sites (9.2 ± 3.28%). All test and control groups investigated revealed comparable and statistically not significant different BIC values, ranging from 73.38 ± 20.5% (BCP2) to 84.11 ± 7.84% (BCP1), respectively.ConclusionAll bone graft materials facilitated new bone formation and osseointegration after 12 + 8 weeks of healing.

Coupling between Macrophage Phenotype, Angiogenesis and Bone Formation by Calcium Phosphates

The ability of calcium phosphate (CaP) materials to induce bone formation varies with their physicochemical properties, with surface topography as one of the most crucial triggers. In view of the natural wound healing processes initiated after surgical implantation, involving inflammation, angiogenesis, tissue formation and remodeling, we here comparatively investigated the biological cascades occurring upon ectopic implantation of a tricalcium phosphate with submicron surface topography (TCP-S, osteoinductive) and a tricalcium phosphate with micron-scale topography (TCP-B, non-osteoinductive). In vitro, TCP-S facilitated M2 polarization of macrophages derived from a human leukemic cell line (THP-1) as shown by the enhanced secretion of TGF-β and CCL18. Interestingly, the conditioned media of polarized M2 macrophages on TCP-S enhanced tube formation by human umbilical vein endothelial cells (HUVECs), while had no influence on the osteogenic differentiation of human bone marrow stromal cells (HBMSCs). Following implantation in a canine intramuscular defect, TCP-S locally increased typical M2 macrophage markers (e.g. IL-10) at week 1 and enhanced blood vessel formation after week 3 compared to TCP-B. Bone formation was observed histologically for TCP-S 6 weeks after implantation, and bone formation was reduced when an angiogenesis inhibitor (KRN633) was added to TCP-S. No bone formation was observed for TCP-B. The data here presented demonstrates strong links between surface topography, macrophage polarization, angiogenesis and bone formation in CaP materials implanted in non-osseous sites.

Current Use of Calcium Sulfate Bone Grafts

Bone graft placement is the most widely used therapeutic strategy for the surgical correction of osseous defects. In recent years, increasing attention has been given to the development of synthetic bone grafts. Of those currently available, calcium sulfate materials exhibit several unique properties that warrant discussion. These include their intrinsic osteogenic potential, their stimulatory effect on angiogenesis, the fact that they are fully biodegradable, the lack of proinflammatory responses following their placement in situ, and their lost cost of production. However, despite the attractiveness of these features, the use of calcium phosphate materials for bone grafting continues to be more widespread. This review examines the current use of calcium sulfate bone grafts in regenerative medicine. It also considers their clinical drawbacks before providing insight into the development of new calcium sulfate grafting constructs that might address these concerns.

Biphasic composite of calcium phosphate-based mesoporous silica as a novel bone drug delivery system

We reported the new biphasic composites of calcium phosphate and mesoporous silica material (CaP@MSi) in the form of powders and pellets as a potential bone drug delivery system for doxycycline hydrochloride (DOX). The CaP@MSi powders were synthesized by cationic surfactant-templating method. The effects of 10, 20, and 30% CaP content in the CaP@MSi powders on the molecular surface structure, the cytotoxicity against osteoblast cells in vitro, and the mineralization potential in simulated body fluid were investigated. The CaP@MSi characterized by the highest mineralization potential (30% CaP content) were used for DOX adsorption and pelletization process. The CaP which precipitated in the CaP@MSi composites was characterized as calcium-deficient with the Ca:P molar ratio between 1.0 and 1.2. The cytotoxicity assays demonstrated that the CaP content in MSi increases osteoblasts viability indicating the CaP@MSi (30% CaP content) as the most biocompatible. The combination of CaP and MSi was an effective strategy to improve the mineralization potential of parent material. Upon immersion in simulated body fluid, the CaP of composite converted into the bone-like apatite. The obtained pellets preserved the mineralization potential of CaP@MSi and provided the prolonged 5-day DOX release. The obtained biphasic CaP@MSi composites seem to have an application potential as bone-specific drug delivery system.

Experimental and numerical analysis on bending and tensile failure behavior of calcium phosphate cements

Since their discovery in the 1980s, injectable self-setting calcium phosphate cements (CPCs) are frequently used in orthopedic, oral and maxillofacial surgery due to their chemical resemblance to the mineral phase of native bone. However, these cements are very brittle, which complicates their application in load-bearing anatomical sites. Polymeric fibers can be used to transform brittle calcium phosphate cements into ductile and load-bearing biomaterials. To understand and optimize this process of fiber reinforcement, it is essential to characterize the mechanical properties of fiber-free calcium phosphate matrices in full detail. However, the mechanical performance of calcium phosphate cements is usually tested under compression only, whereas bending and tensile tests are hardly performed due to technical limitations. In addition, computational models describing failure behavior of calcium phosphate cements under these clinically more relevant loading scenarios have not yet been developed. Here, we investigate the failure behavior of calcium phosphate cements under bending and tensile loading by combining, for the first time, experimental tests and numerical modeling. To this end, a 3-D gradient-enhanced damage model is developed in a finite element framework, and numerical results are correlated to experimental three-point bending and tensile tests to characterize the mechanical properties of calcium phosphate cements in full detail. The presented computational model is successfully validated against experimental results and is able to predict the mechanical response of calcium phosphate cement under different types of loading with a unique set of parameters. This model offers a solid basis for further experimental and computational studies on the development of load-bearing bone cements.

THE EFFECT OF MINERAL TRIOXIDE AGGREGATE ALONE AND IN COMBINATION WITH BIPHASIC CALCIUM PHOSPHATE ON HEALING OF MANDIBULAR DEFECTS IN RABBIT

INTRODUCTION: Critical sized bone defects above critical size do not heal completely by themselves and thus represent major clinical challenge to reconstructive surgery. Numerous bone substitutes have already been used to promote bone regeneration. Incorporation of bone grafts such as biphasic calcium phosphate (BCP) as osteoconductive materials has been proven to be effective in promoting bone healing in oral surgery. As it is considered a potential candidate for bone regenerating biomolecules which can help in rapid bone repair because of its composition, the biocompatible material mineral trioxide aggregate(MTA) is suggested to be promising. OBJECTIVES: To compare the efficacy of MTA on the process of bone healing alone and the combination of MTA with biphasic calcium phosphate MATERIALS AND METHODS: Sixteen rabbits will be divided into two experimental groups: (1) MTA, (2) MTA+BCP. Right and left critical size bone defects will be performed in the edentulous area of rabbit mandibles (diastema). Right side bone defects will be filled with MTA, while the left side bone defects will be filled with MTA in combination with BCP. The effect of both MTA and BCP on defect healing will be assessed histologically, histomorphometrically and immunohistochemically after 6 weeks postoperatively. RESULTS: Greater amount of new bone formation was noticed in the CSDs loaded with MTA and BCP. The amount of new bone formed was significantly higher in the MTA+BCP group when compared with the MTA group. Also bone cells activities appeared greater in the former group. CONCLUSIONS: MTA induces new bone formation via an osteoinductive action and can be added to bone grafts to enhance bone regenerative capacity

Effects of argon plasma treatment on the osteoconductivity of bone grafting materials.

The osteoconductive properties of bone grafting materials represent one area of research for the management of bony defects found in the fields of periodontology and oral surgery. From a physico-chemical aspect, the wettability of the graft has been demonstrated to be one of the most important factors for new bone formation. It is also well-known that argon plasma treatment (PAT) and ultraviolet irradiation (UV) may increase the surface wettability and, consequently, improve the regenerative potential of the bone grafts. Therefore, the aim of the present in vitro study was to evaluate the effect of PAT and UV treatment on the osteoconductive potential of various bone grafts. The following four frequently used bone grafts were selected for this study: synthetic hydroxyapatite (Mg-HA), biphasic calcium phosphate (BCP), cancellous and cortical xenogenic bone matrices (CaBM, CoBM). Sixty-six serially numbered disks 10 mm in diameter were used for each graft material and randomly assigned to the following three groups: test 1 (PAT), test 2 (UV), and control (no treatment). Six samples underwent topographic analysis using SEM pre- and post-treatments to evaluate changes in surface topography/characteristics. Additionally, cell adhesion and cell proliferation were evaluated at 2 and 72 h respectively following incubation in a three-dimensional culture system utilizing a bioreactor. Furthermore, the effects of PAT and UV on immune cells were assessed by measuring the viability of human macrophages at 24 h. The topographic analysis showed different initial morphologies of the commercial biomaterials (e.g., Mg-HA and BCP showed flat morphology; BM samples were extremely porous with high roughness). The surface analysis following experimental treatments did not demonstrate topographical difference when compared with controls. Investigation of cells demonstrated that PAT treatment significantly increased cell adhesion of all 4 evaluated bone substitutes, whereas UV failed to show any statistically significant differences. The viability test revealed no differences in terms of macrophage adhesion on any of the tested surfaces. Within their limitations, the present results suggest that treatment of various bone grafting materials with PAT appears to enhance the osteoconductivity of bone substitutes in the early stage by improving osteoblast adhesion without concomitantly affecting macrophage viability. Treatment of bone grafts with PAT appears to result in faster osseointegration of the bone grafting materials and may thus favorably influence bone regeneration.

APPLICATION OF BIOGLASS IN THE TREATMENT OF FRACTURES AND DEFECTS OF LONG BONES

One of the most promising trends in contemporary traumatology and orthopaedics towards the treatment of fractures and bone defects is the use of synthetic calcium-phosphate biomaterials. The purpose of this study is to identify the features of bone defect healing in rats during the implantation of calcium phosphate glass crystalline materials, and to substantiate the feasibility of the use of bioglass in the clinical treatment of fractures and defects of long bones. Materials and methods. In this study we used bioactive calcium-phosphate glass crystal material BS-11. The experimental part was performed on 36 white rats. The animals were subjected to defect modelling in the femur with following bioglass filling. Results. We observed the formation of osteogenic fibroreticular tissue and bone tissue of varying degrees of maturity around the implanted filling samples through all the healing periods. The relative content of bone tissue in the surrounding regenerated tissue around the studied samples BS-11 increased in parallel with the increase of the observation period and reached 100% in 30 days after the surgical operation and at the end of the study we registered the formation of bone tissue of the lamellar structure around the samples inserted. We observed no signs of inflammation or any cellular reaction in any of the cases studies that points out the biocompatibility of the material. Conclusions. Morphological study has demonstrated that the insertion of cylindrical blocks made of glass-crystalline calcium phosphate materials BC-11 in the distal metaphysis of the femur of rats and healing is accompanied with osteoregeneration around them in accordance with the conventional stages of the bone tissue formation. This material can be recommended for treating long bone defects of different aetiology.

Nurse's A-Phase-Silicocarnotite Ceramic-Bone Tissue Interaction in a Rabbit Tibia Defect Model.

Calcium phosphate materials are widely used as bone substitutes due to their bioactive and biodegradable properties. Also, the presence of silicon in their composition seems to improve the bioactivity of the implant and promote bone tissue repair. The aim of this study was to develop a novel ceramic scaffold by partial solid-state sintering method with a composition lying in the field of the Nurse’s A-phase–silicocarnotite, in the tricalcium phosphate–dicalcium silicate (TCP–C2S) binary system. Also, we evaluated its osteogenic and osteoconductive properties after being implanted into tibia defects in New Zealand rabbits. X-ray, microcomputer tomography, and histomorphometry studies demonstrated that this porous ceramic is highly biocompatible and it has excellent osteointegration. The material was being progressively reabsorbed throughout the study and there was no unspecified local or systemic inflammatory response observed. These results suggest that ceramic imitates the physicochemical characteristics of bone substitutes used in bone reconstruction.

Insights into structural characteristics and the mechanism of silicate-based calcium phosphates with phosphoric acid modulation

Si-containing calcium phosphate materials recently have attracted attention owning to their biomineral capability and benefit to the proliferation and differentiation of osteoblast-like cells. In this study, tri-calcium silicate (C3S), which can induce apatite layer deposition on its surface, and phosphoric acid are used to prepare silicate-based calcium phosphates via aqueous precipitation method. It's known that calcium phosphate undergoes phase transformation when the pH value changes. So, the different ratios of C3S and phosphoric acid are designed to study the phase transformation of calcium phosphate. The pH, ions release of solution, composition and structure are detected. The results show the phase transformation of calcium phosphate from hydroxyapatite to brushite with the increased content of phosphoric acid. The structure of calcium silicate hydrate (C–S–H) also undergoes a series of changes, including protonation and polymerization. Briefly, this study focuses primarily on the interaction between C–S–H and calcium phosphates. In addition, it may provide a new sight for the sustained delivery of bisphosphonates achieved by depositing on the surface of C3S for the treatment of osteoporosis.

Structural and metabolic features of the femur of rats after the implantation of glass crystalline material

In vivo research is an important link in the development of osteoplastic materials.Aim: to study the structural features and markers of bone metabolism after an implantation into the distal metaphysis of the rat femur of glass crystalline calcium phosphate material (GCM). Methods: hole defects in the distal metaphysis of the femur were performed in 28 white laboratory male rats (age 4 months, weight 200–260 g). GCM cylindrical samples with a diameter of 1 mm and a height of 3 mm were placed in defects. A comparison group for biochemical studies was 5 intact rats. Animals were sacrificed 7, 14, 28, and 90 days after surgery. The histological studies of the bone structure around the implant and assessment of osseointegration were performed. In the blood serum the activity of alkaline phosphatase, the content of chondroitin sulfates, urea, the activity of alanine aminotransferase were determined.Results: 7 days after surgery the relative content of bone tissue around implants was (28.59 ± 1.33) %, after 14 days it increased by 2.71 times (p < 0.001). After 30 days bone tissue was located around the entire perimeter of the implant. Bioresorption of the material continued throughout the observation period. A significant increase in the activity of the alkaline phosphatase in blood serum compared with the rate of intact rats was shown from the 7th to the 30th day of observation. The maximum increase of chondroitin sulfates was recorded on the 7th day. The content of urea in the blood serum and the activity of alanine aminotransferase did not statistically significantly differ in terms of the experiment or from indices of intact animals.Conclusions: the studied material is biocompatible, has osteoinductive and osteoconductive qualities. After implanting it into the bone, bone repair is not impaired, and lamellar bone tissue is formed for the final study period (90 days). A characteristic feature of the material is its gradual resorption until the 90th day of observation.

Comparative Study between Laser Light Stereo-Lithography 3D-Printed and Traditionally Sintered Biphasic Calcium Phosphate Scaffolds by an Integrated Morphological, Morphometric and Mechanical Analysis.

In dental districts, successful bone regeneration using biphasic calcium phosphate materials was recently explored. The present study aimed to perform a comparative study between 3D-printed scaffolds produced by laser light stereo-lithography (SLA) and traditionally sintered biphasic calcium phosphate scaffolds by an integrated morphological, morphometric and mechanical analysis. Methods: Biphasic calcium phosphate (30% HA/70% β-TCP) samples, produced by SLA-3D-printing or by traditional sintering methods, were tested. The experimental sequence included: (1) Microtomography (microCT) analyses, to serve as control-references for the 3D morphometric analysis; (2) loading tests in continuous mode, with compression up to fracture, to reconstruct their mechanical characteristics; and (3) microCT of the same samples after the loading tests, for the prediction of the morphometric changes induced by compressive loading of the selected materials. All the biomaterials were also studied by complementary scanning electron microscopy to evaluate fracture regions and surfaces. Results: The characterization of the 3D mineralized microarchitecture showed that the SLA-3D-printed biomaterials offer performances comparable to and in some cases better than the traditionally sintered ones, with higher mean thickness of struts and pores. Interestingly, the SLA-3D-printed samples had a higher ultimate strength than the sintered ones, with a smaller plastic region. Moreover, by SEM observation, it was observed that fractures in the SLA-3D-printed samples were localized in the structure nodes or on the external shells of the rods, while all the traditionally sintered samples revealed a ductile fracture surface. Conclusions: The reduction of the region of plastic deformation in the SLA-3D-printed samples with respect to traditionally sintered biomaterials is expected to positively influence, in vivo, the cell adhesion. Both microCT and SEM imaging revealed that the studied biomaterials exhibit a structure more similar to human jaw than the sintered biomaterials.

Effects of Fluoride and Calcium Phosphate Materials on Remineralization of Mild and Severe White Spot Lesions.

Fixed orthodontic treatments often lead to enamel demineralization and cause white spot lesions (WSLs). The aim of this study was to evaluate the mineralization degree of 2 types of WSLs based on ICDAS index and compare the remineralizing efficacy of 3 oral hygiene practices after 1 month and 3 months. 80 mild demineralized and 80 severe demineralized enamel specimens were randomized into three treatments: fluoride toothpaste (FT), fluoride varnish plus fluoride toothpaste (FV+FT), and CPP-ACP plus fluoride toothpaste (CPP-ACP+FT). Microhardness tester, DIAGNODent Pen 2190, and scanning electron microscope were used to evaluate the changes of mineralization degree. Both qualitative and quantitative indicators suggested that the mild and severe white spot lesions were different in the degree of mineralization. Severe WSLs demineralized much more seriously than mild lesions even after 3 months of treatment. Despite the variation in severity, both lesions had the same variation trend after each measure was applied: FT had weak therapeutic effect, FV + FT and CPP-ACP + FT were effective for remineralization. Their remineralizing efficacy was similar after 1 month, and combined use of CPP-ACP plus F toothpaste was more effective after 3 months. In order to fight WSLs, early diagnosis was of great importance, and examination of the tooth surface after air-dry for 5 seconds was recommended. Also, when WSLs were found, added remineralizing treatments were required.

Comparative Effect of Calcium Phosphate Biocomposite Materials on the Healing of Experimental Defect of Compact Bone Tissue

Introduction For the treatment of bone defects, a considerably large number of biocomposite calcium-phosphate materials has been developed and used. However, in the scientific literature, there is no information about the comparative effect of biocomposite materials based on β-tricalcium phosphate, hydroxyapatite, and collagen on the dynamics of healing of the defect of compact bone tissue. Materials and Methods The experiment was performed on 48 white Wistar rats. In the middle third of the femoral shaft, a perforated defect 2.5 mm in diameter was reproduced in the bone marrow canal, which was filled with the calcium phosphate material Collapan (Intermedapatit, Moscow, Russia) (hydroxyapatite/collagen/antibiotics) in the animals of the first group, and with Guidor easy-graft Crystal (Sunstar S.A., Etoy, Switzerland) (hydroxyapatite/β-tricalcium phosphate) in the animals of the second group. Fragments of the injured bones were examined on the 15th and 30th days by light microscopy with morphometry and scanning electron microscopy. Results It was found that, in the area of implantation of Collapan and of Guidor easy-graft Crystal, signs of inflammation were not detected, and osteogenic cells exhibited high topism to biocomposite materials. The biomaterials during the entire period of the experiment are subjected to resorption and replacement by the bone tissue of the regenerate. On the 15th and 30th days of the experiment, the predominance in the rate of biomaterial resorption (of 35.04% and 53.47%, respectively) and the formation of regenerate bone tissue (of 58.67% and 50.47%, respectively) was in the area of implantation of Collapan. Conclusion The biocomposite materials tested exhibit a high biocompatibility, osteoconductive properties, and good integration with the tissues of the regenerate. However, the biocomposite material Collapan undergoes resorption and replacement by the bone tissue of the regenerate much faster, and Guidor easy-graft Crystal provides stability of the defect volume of compact bone tissue due to full resorption and good integration with the tissues of the regenerate.

Surfactant-free electrochemical synthesis of fluoridated hydroxyapatite nanorods for biomedical applications

Abstract Fluoridated hydroxyapatite (FHA) [Ca10(PO4)6Fx(OH)2−x, x = 0–2] is believed to be a promising calcium phosphate (CaP) to replace pure hydroxyapatite (HA) for next-generation implants, owing to its better biocompatibility, higher antibacterial activity, and lower solubility. Notably, the shape and size of the CaP crystals play key roles in their performance and can influence their applications. One-dimensional (1D) FHA nanorods are important CaP materials which have been widely used in regenerative medicine applications such as restorative dentistry. Unfortunately, the traditional synthesis methods for FHA nanorods either employ surfactants or take a relatively long time. In this study, we aimed to propose a facile synthesis route to fabricate FHA nanorods without any surfactants using an electrochemical deposition method for the first time. This study focused on preparing FHA nanorods without the assistance of any surfactant, unlike the traditional synthesis methods, to avoid chemical impurities. FHA nanorods with lengths of 124–2606 nm, diameters of 28–211 nm, and aspect ratios of 4.4–21.8 were synthesized using the electrochemical method, followed by a heat treatment. For the as-synthesized FHA nanorods, the Ca/P ratio was 1.60 and the atomic concentration of F was 2.06 at.%. An ultrastructure examination revealed that each FHA nanorod possessed long-range order, good crystallinity, and a defect-free lattice with a certain crystallographic plane orientation along the whole rod. In short, we propose a novel, surfactant-free, cost-saving, and more efficient route to synthesize FHA nanorods which can be widely applied in multiple biomedical applications, including drug delivery, bone repair, and restorative dentistry.

In situ calcium isotopic ratio determination in calcium carbonate materials and calcium phosphate materials using laser ablation-multiple collector-inductively coupled plasma mass spectrometry

This study presents a highly precise and accurate Ca isotope analysis method using laser ablation multiple collector inductively coupled plasma mass spectrometry (LA-MC-ICP-MS) in calcium carbonates and calcium phosphates. High-sensitivity cones and nitrogen gas were used to improve the performance of the LA-MC-ICP-MS. The experiment results show that at the N2 mode, the signal intensity of 44Ca+ in the high-sensitivity cones (Jet + X cones) was 5-fold higher than that in the normal cones (S + H cones), while the polyatomic interferences of 40Ar1H1H+ and 40Ar16O+ in the high-sensitivity cones were suppressed by factors of 6.1 and 10.6, respectively, after the addition of 8 ml min−1 N2. The combination of high sensitivity cones and N2 addition did not increase the production of double charged ions. Additionally, the interference correction of doubly charged Sr ions was investigated in detail. The experiment results demonstrate that the concentration ratio of Sr to Ca (represented as I87Sr++/I44Ca+), the 87Sr/86Sr ratio and the mass fractionation factor of Sr (fSr) during the measurement process were key parameters for in situ Ca isotope ratios analysis. The threshold values of these key parameters were evaluated and are recommended to ensure an accurate correction of doubly charged Sr ions. Using the optimized instrumental conditions and correction strategies, an excellent within-run precision (<0.06‰, 2SE) and external reproducibility (<0.15‰, 2SD) for the microanalysis of δ44/42Ca915b were obtained in natural calcium carbonates and calcium phosphates. These precisions are better than the values previously reported in the literature regarding using LA-MC-ICP-MS. Matrix effects were observed during the in situ Ca isotope analysis among calcium carbonates, hydroxy calcium phosphates and igneous apatites. Using the matrix-matching standards, accurate δ44/42Ca915b values in natural materials with various matrices were achieved, confirming the availability of the proposed method to resolve the spatial variation of δ44/42Ca in natural materials.

Molecular dynamics exploration of the amorphous surface structures and properties of the biomimetic β-tricalcium phosphate

β-Tricalcium phosphate (β-TCP) is one of the main components in bone matrix. Its synthetic analogue has been widely applied as bioceramics or catalysts. Experimental evidences have shown that the surface area of the bioactive calcium phosphate materials is in amorphous state rather than perfect crystal phase. In this work, based on molecular dynamics annealing simulation, we have constructed the amorphous surface structures for β-TCP (100) plane. INTERFACE force field (IFF) shows very good performance for simulating the properties of r β-TCP such as lattice parameters, infrared spectroscopy, and melting points. Our calculations show that the amorphous surface structures of β-TCP have obvious nano-grooves, which might be beneficial to the adsorption of biomolecules and the enhancement of biological activity. The mechanical properties of annealed structure of β-TCP were also calculated and discussed. Our results show that the annealing temperatures have obvious effects on the surface energy but not mechanical properties. As a results, in designing or optimizing the biomaterials, the sintering should not be the only way to improve the mechanical properties of β-TCP. The present amorphous surface models for the β-TCP show interesting properties and are useful in further investigations on the interactions between biomaterial surface and proteins.

Immune Modulation by Transplanted Calcium Phosphate Biomaterials and Human Mesenchymal Stromal Cells in Bone Regeneration.

A wide variety of biomaterials have been developed as both stabilizing structures for the injured bone and inducers of bone neoformation. They differ in chemical composition, shape, porosity, and mechanical properties. The most extensively employed and studied subset of bioceramics are calcium phosphate materials (CaPs). These materials, when transplanted alongside mesenchymal stem cells (MSCs), lead to ectopic (intramuscular and subcutaneous) and orthotopic bone formation in preclinical studies, and effective fracture healing in clinical trials. Human MSC transplantation in pre-clinical and clinical trials reveals very low engraftment in spite of successful clinical outcomes and their therapeutic actions are thought to be primarily through paracrine mechanisms. The beneficial role of transplanted MSC could rely on their strong immunomodulatory effect since, even without long-term engraftment, they have the ability to alter both the innate and adaptive immune response which is critical to facilitate new bone formation. This study presents the current knowledge of the immune response to the implantation of CaP biomaterials alone or in combination with MSC. In particular the central role of monocyte-derived cells, both macrophages and osteoclasts, in MSC-CaP mediated bone formation is emphasized. Biomaterial properties, such as macroporosity and surface microstructure, dictate the host response, and the ultimate bone healing cascade. Understanding intercellular communications throughout the inflammation, its resolution and the bone regeneration phase, is crucial to improve the current therapeutic strategies or develop new approaches.

Glancing Angle Deposition of Zn-Doped Calcium Phosphate Coatings by RF Magnetron Sputtering

Zn-substituted hydroxyapatite with antibacterial effect was used in radiofrequency (RF) magnetron deposition of calcium phosphate coating onto Ti- and Si-inclined substrates. The development of surface nanopatterns for direct bacteria killing is a growing area of research. Here, we combined two approaches for possible synergetic antibacterial effect by manufacturing a patterned surface of Zn-doped calcium phosphate using glancing angle deposition (GLAD) technique. A significant change in the coating morphology was revealed with a substrate tilt angle of 80°. It was shown that an increase in the coating crystallinity for samples deposited at a tilt angle of 80° corresponds to the formation of crystallites in the bulk structure of the thin film. The variation in the coating thickness, uniformity, and influence of sputtered species energy on Si substrates was analyzed. Coatings deposited on tilted samples exhibit higher scratch resistance. The coating micro- and nano-roughness and overall morphology depended on the tilt angle and differently affected the rough Ti and smooth Si surfaces. GLAD of complex calcium phosphate material can lead to the growth of thin films with significantly changed morphological features and can be utilized to create self-organized nanostructures on various types of surfaces.

Culture of hybrid spheroids composed of calcium phosphate materials and mesenchymal stem cells on an oxygen-permeable culture device to predict in vivo bone forming capability

Three-dimensional (3-D) cell culture can better mimic physiological conditions in which cells interact with adjacent cells and the extracellular matrix than monolayer culture. We have developed a 3-D cell culture device, the Oxy chip, which can be used to generate and supply oxygen to cell spheroids to prevent hypoxia. Here, we used the Oxy chip to generate hybrid spheroids comprising calcium phosphate (CaP) particles (hydroxyapatite (HA), β-tricalcium phosphate (β-TCP) or octacalcium phosphate (OCP)) and mesenchymal stem cells (MSCs, C3H10T1/2 cells or D1 cells) that can be used to analyze cell differentiation mechanisms. We showed that the 3-D cell-cell and cell-material interactions and oxygenation offered by the Oxy chip promoted osteoblastic differentiation of MSCs. We also used histomorphometric analysis of hematoxylin and eosin staining, quality analyses by μCT and collagen orientation observation with picrosirius red staining in bone regeneration following implantation of three CaPs in a critical-sized defect in mouse calvaria. The in vivo bone formation capacity of the three tested CaP materials was OCP ≥ β-TCP > HA: the newly formed bone by OCP had a structure relatively close to that of the calvaria intact bone. When MSCs were 3-D cultured with the CaP materials using the Oxy chip, the in vitro osteogenic capacity of these materials was highly similar to trends observed in vivo. The in vitro alkaline phosphatase activity of D1 cells had the highest correlation with in vivo bone volume (R = 0.900). Chemical and FTIR spectroscopic analyses confirmed that differentiation of D1 cells could be associated with amorphous calcium phosphate (ACP) precipitation concomitant with OCP hydrolysis. Taken together, hybrid spheroid cultures using the Oxy chip can be used to screen and predict bone forming potential of bone substitute materials. Statement of significanceAn oxygen permeable-culture chip (Oxy chip) can be used to induce formation of cell spheroids by mesenchymal stem cells (MSCs). Use of the Oxy chip avoids hypoxia in the spheroid core and enhances MSC osteoblastic differentiation relative to conventional spheroid culture methods. The present study showed that the Oxy chip mimics the in vivo environment associated with bone formation and can be used to generate hybrid spheroids consisting of calcium phosphates and MSCs that are useful for analyzing cell differentiation mechanisms. Bone formation analysis following implantation of calcium phosphate materials in mouse calvaria defects showed positive correlation with the in vitro results. We propose that hybrid spheroids cultured on the Oxy chip can be used to screen and predict the bone forming potential of bone substitute materials.