Mineral Phase Of Bone Research Articles

Raman Spectroscopy of Natural Bone and Synthetic Apatites

Raman spectroscopy of natural bones and hydroxyapatites is described. In addition, how Raman spectroscopy has proved crucial in providing baseline data for the modification of synthetic apatite powders that are routinely used now as bone replacement materials is explained. It is important to understand the chemical structural properties of natural bone. Bone consists of two primary components: an inorganic or mineral phase, which is mainly a carbonated form of a nanoscale crystalline calcium phosphate, closely resembling hydroxyapatite, and an organic phase, which is composed largely of type I collagen fibers. Other constituents of bone tissue include water and organic molecules such as glycosaminoglycans, glycoproteins, lipids, and peptides. Ions such as sodium, magnesium, fluoride, and citrate are also present, as well as hydrogenophosphate. Hence, the mineral phase in bone may be characterized essentially as nonstoichiometric substituted apatite. Such a distinction is important in the development of synthetic calcium phosphates for application as skeletal implants. An understanding of bone function and its interfacial relationship to an implant clearly depends on the associated structure and composition. Therefore, it is essential to fully understand the chemical composition of bone, and Raman spectroscopy is an excellent technique for such an analysis.

Biological Evaluation of Polydimethylsiloxane Modified by Calcium Phosphate Nanoparticles for Potential Application in Spine Surgery

The biological properties of a composite made of polydimethylsiloxane matrix and two kinds of calcium phosphate nanoparticles (hydroxyapatite, beta tricalcium phosphate) were tested by means of in vitro and in vivo investigations. Chemical and phase analyses of the nanoparticles were performed. The effects of different volume fractions of nanoparticles were investigated. Human osteoblast-like cell cultures (MG 63) demonstrated that the addition of 5–15 vol.% of nanoparticles had a significantly positive effect on osteoblast viability, and that high concentration did not support cell proliferation. The highest rates of osteointegration in pig bone were attained for implants modified by 15 vol.% hydroxyapatite and more intensive formation of new bone tissue was observed. Our study indicates that hydroxyapatite probably better imitates the composition and structure of the mineral phase of real bone.

Tantalum oxide and barium sulfate as radiopacifiers in injectable calcium phosphate-poly(lactic-co-glycolic acid) cements for monitoringin vivodegradation

Monitoring the degradation of calcium phosphate-based bone substitute materials in vivo by means of noninvasive techniques (e.g., radiography) is often a problem due to the chemical resemblance of those substitutes with the mineral phase of bone. In the view of that, the present study aimed at enhancing the radiopacity of calcium phosphate cement enriched with poly(lactic-co-glycolic acid) (CPC-PLGA) microspheres, by adding tantalum oxide (Ta2O5) or the more traditional radiopacifier barium sulfate (BaSO4). The radiopacifying capacity of these radiopacifiers was first evaluated in vitro by microcomputed tomography (μCT). Thereafter, both radiopacifiers were tested in vivo using a distal femoral condyle model in rabbits, with subsequent ex vivo μCT analysis in parallel with histomorphometry. Addition of either one of the radiopacifiers proved to enhance radiopacity of CPC-PLGA in vitro. The in vivo experiment showed that both radiopacifiers did not induce alterations in biological performance compared to plain CPC-PLGA, hence both radiopacifiers can be considered safe and biocompatible. The histomorphometrical assessment of cement degradation and bone formation showed similar values for the three experimental groups. Interestingly, μCT analysis showed that monitoring cement degradation becomes feasible upon incorporation of either type of radiopacifier, albeit that BaSO4 showed more accuracy compared to Ta2O5.

Periodic ab initio bulk investigation of hydroxylapatite and type A carbonated apatite with both pseudopotential and all-electron basis sets for calcium atoms

Apatite minerals draw the attention of many researchers not only in mineralogy, but also in biology, biochemistry, and medicine because hydroxylapatite [Ca10(PO4)6(OH)2] is the main component of the mineral phase of mammalian bones. However, in nature this mineral is mostly present with various stoichiometric defects. The carbonate ion is found commonly in its structure where it can occupy different crystallographic sites; however, its configurational energy and relative orientation in the apatite lattice is still debated.

Dual Contrast Agent for Computed Tomography and Magnetic Resonance Hard Tissue Imaging

Calcium phosphate cements (CPCs) are commonly used bone substitute materials, which closely resemble the composition of the mineral phase of bone. However, this high similarity to natural bone also results in difficult discrimination from the bone tissue by common imaging modalities, that is, plain X-ray radiography and three-dimensional computed tomography (CT). In addition, new imaging techniques introduced for bone tissue visualization, like magnetic resonance imaging (MRI), face a similar problem. Even at high MRI resolution, the lack of contrast between CPCs and surrounding bone is evident. Therefore, this study aimed to evaluate the feasibility of a dual contrast agent, traceable with both CT and MRI as enhancers of CPC/bone tissue contrast. Our formulation is based on the use of silica beads as vectors, which encapsulate and carry contrast-enhancing nanoparticles, in our case, colloidal Gold and Superparamagnetic Iron oxide particles (SPIO). The bead suspension was incorporated within a calcium phosphate powder. The resultant cements were then tested both in vitro and in vivo in a femoral condyle defect model in rats. Results showed that the mechanical properties of the cement were not significantly affected by the inclusion of the beads. Both in vitro and in vivo data proved the homogeneous incorporation of the contrast within the cement and its visual localization, characterized by a short-term CT contrast enhancement and a long-term MR effect recognizable by the characteristic blooming shape. Finally, no signs of adverse tissue reactions were noticed in vivo. In conclusion, this study proved the feasibility of a multimodal contrast agent as an inert and biocompatible enhancer of CaP cement versus bone tissue contrast.

Atom Probe Tomography of Apatites and Bone-Type Mineralized Tissues

Nanocrystalline biological apatites constitute the mineral phase of vertebrate bone and teeth. Beyond their central importance to the mechanical function of our skeleton, their extraordinarily large surface acts as the most important ion exchanger for essential and toxic ions in our body. However, the nanoscale structural and chemical complexity of apatite-based mineralized tissues is a formidable challenge to quantitative imaging. For example, even energy-filtered electron microscopy is not suitable for detection of small quantities of low atomic number elements typical for biological materials. Herein we show that laser-pulsed atom probe tomography, a technique that combines subnanometer spatial resolution with unbiased chemical sensitivity, is uniquely suited to the task. Common apatite end members share a number of features, but can clearly be distinguished by their spectrometric fingerprint. This fingerprint and the formation of molecular ions during field evaporation can be explained based on the chemistry of the apatite channel ion. Using end members for reference, we are able to interpret the spectra of bone and dentin samples, and generate the first three-dimensional reconstruction of 1.2 × 10(7) atoms in a dentin sample. The fibrous nature of the collagenous organic matrix in dentin is clearly recognizable in the reconstruction. Surprisingly, some fibers show selectivity in binding for sodium ions over magnesium ions, implying that an additional, chemical level of hierarchy is necessary to describe dentin structure. Furthermore, segregation of inorganic ions or small organic molecules to homophase interfaces (grain boundaries) is not apparent. This has implications for the platelet model for apatite biominerals.

Calcite as a bone substitute. Comparison with hydroxyapatite and tricalcium phosphate with regard to the osteoblastic activity

Close to the bone mineral phase, the calcic bioceramics, such as hydroxyapatite (HA) and β-tricalcium phosphate (β-TCP), are commonly used as substitutes or filling materials in bone surgery. Besides, calcium carbonate (CaCO3) is also used for their excellent biocompatibility and bioactivity. However, the problem with the animal-origin aragonite demands the new technique to synthesize pure calcite capable of forming 3D bone implant. This study aims to manufacture and evaluate a highly-pure synthetic crystalline calcite with good cytocompatibility regarding to the osteoblasts, comparing to that of HA and β-TCP. After the manufacture of macroporous bioceramic scaffolds with the identical internal architecture, their cytocompatibility is studied through MC3T3-E1 osteoblasts with the tests of cell viability, proliferation, vitality, etc. The results confirmed that the studied process is able to form a macroporous material with a controlled internal architecture, and this synthesized calcite is non-cytotoxic and facilitate the cell proliferation. Indeed requiring further improvement, the studied calcite is definitely an interesting alternative not only to coralline aragonite but also to calcium phosphate ceramics, particularly in bone sites with the large bone remodelling.

Effect of X-ray irradiation on the elastic strain evolution in the mineral phase of bovine bone under creep and load-free conditions

Both the load partitioning between hydroxyapatite (HAP) and collagen during compressive creep deformation of bone and the HAP residual strain in unloaded bone have been shown in previous synchrotron X-ray diffraction studies to be affected by the X-ray irradiation dose. Here, through detailed analysis of the X-ray diffraction patterns of bovine bone, the effect of X-ray dose on (i) the rate of HAP elastic strain accumulation/shedding under creep conditions and (ii) the HAP lattice spacing and average root mean square (RMS) strain under load-free conditions are examined. These strain measurements exhibit three stages in response to increasing X-ray dose. Up to ∼75kGy (stage I) no effect of dose is observed, indicating a threshold behavior. Between ∼75 and ∼300kGy (stage II) in unloaded bone the HAP d-spacing increases and the RMS strain decreases with dose, indicating strain relaxation of HAP. Furthermore, under constant compressive load creep conditions, the rate of compressive elastic strain accumulation in HAP decreases with increasing dose until, at ∼115kGy, it changes sign, indicating that the HAP phase is shedding load during creep deformation. These stage II behaviors are consistent with HAP–collagen interfacial damage, which allows the HAP elastic strain to relax within both the loaded and unloaded samples. Finally, for doses in excess of ∼300kGy (stage III, measured up to 7771kGy) the HAP lattice spacing and RMS strain for load-free samples and the rate of HAP elastic strain shedding for crept samples remain independent of dose, suggesting a saturation of damage and/or stiffening of the collagen matrix due to intermolecular cross-linking.

Evaluation of α-Tricalcium Phosphate Cement Obtained at Different Temperatures

The calcium phosphate cements (CPCs) based on α-tricalcium phosphate (α-TCP) are highly attractive for use in medicine and odontology, since they have similar chemical and phase composition of mineral phase of bones (calcium deficient hydroxyapatite (CDHA)). However, one of the biggest difficulties for use of this type of cement is its low mechanical strength due to the presence of undesirable phases, such as β-tricalcium phosphate. The route for obtaining α-TCP is at high temperature by solid state reaction, mixing calcium carbonate and calcium pyrophosphate. The aim of this work was to obtain calcium phosphate cements with improved strength, by studying the obtaining of α-TCP at temperatures of 1300, 1400 and 1500°C. The samples were analyzed by crystalline phases, pH, setting time, particle size, in vitro test (Simulated Body Fluid), porosity, density and compressive strength. The results show that the synthesis temperatures influence strongly the phases of powders obtained and the mechanical properties of cement, being unnecessary quenching for obtaining pure α-TCP.

Nanonstructural Analysis of Hydroxyapatite Thin Films Using HRTEM/FIB Techniques

Hydroxyapatite, with a composition that resembles the mineral phase of bone, is often used as a coating of metallic prostheses to improve the adhesion and osseointegration of implants in different parts of the skeleton. Among the available techniques that allow the deposition of this material on metallic substrates, Pulsed Laser Deposition (PLD) is capable of producing high quality and highly biocompatible coatings with excellent final attachment.

Effect of the composition of hydroxyapatite/bioactive glass nanocomposite foams on their bioactivity and mechanical properties

Nanocomposite foams were prepared on addition of 63S bioactive glass (BG) to pure hydroxyapatite (HA), in proportion of 0, 25, 50, 75, and 100wt.%, and their mechanical properties and bioactivity were compared. The prepared nanocomposite foams have a grain size in the range 24–38nm and pore size in the range 100–400μm. The compressive strength and elastic modulus increased with increasing the amount of BG addition up to 25wt.% and then decreased by more addition of BG. The maximum values of compressive strength and elastic modulus were found to be about 2.78MPa and 219MPa, respectively. The mean values of total and interconnected porosity were calculated in the range 84–88% and 57–76%, respectively. The obtained composite foams have chemical composition similar to the mineral phase of bone and by changing the ratio of HA/BG it can reach the appropriate bioactivity and biodegradability level needed for different biomedical applications.

Effect of strontium-substituted biphasic calcium phosphate on inflammatory mediators production by human monocytes

Calcium phosphate materials are widely used as bone substitutes because of their properties close to those of the mineral phase of bones. Nevertheless, after several months, calcium phosphate-based materials release particles that may be phagocytosed by monocytes, leading to an inflammatory reaction. Strontium is well known to counteract the osteoporosis process, but little is known about its effect on inflammatory processes. The purpose of this work was to study the effect of biphasic calcium phosphate (BCP) particles substituted with strontium on the inflammatory reaction. Human primary monocytes stimulated or not by lipopolysaccharide (LPS) were exposed to BCP particles containing strontium for 6 and 24h. Inflammatory mediators (cytokines and matrix metalloproteinases (MMPs)) production was then quantified by ELISA and zymography. We observed that the presence of strontium had few effects on unstimulated cells, but it decreased the production of pro-inflammatory cytokines and the chemokine interleukin 8 in LPS-stimulated cell-conditioned medium. This work suggests for the first time that strontium may be involved in the control of inflammatory processes following BCP phagocytosis by human monocytes.

Fabrication and characterization of triple nanobioceramic composite foam

In this study, triple nanobioceramic composite foam consisting of bioactive glass (BG), forsterite, and hydroxyapatite (HA) with highly porous structure and high mechanical strength was successfully fabricated via gelcasting method. The BG and forsterite were added to HA as second-phase ceramics to enhance simultaneously its bioactivity, biodegradability, and mechanical properties. The prepared foams have a crystallite size in the range 30–52 nm (while the crystallite size of initial HA and forsterite powders is about 22 nm), pore size in the range 100–300 µm, and total and interconnected porosity upto 87% and 81%, respectively. The compressive strength measurements showed that the foams have an excellent compressive strength (2.19–3.68 MPa), which is appropriate for load-bearing tissue engineering applications and is close to the compressive strength of spongy bone. In vitro tests proved the good bioactivity and resorbability of the foams. The resulting composite foams have similar chemical composition to the mineral phase of bone and it is possible to optimize the ratio of BG phase to ceramic phases for achieving appropriate mechanical strength, bioactivity, and biodegradability level needed for different applications.

Polymeric additives to enhance the functional properties of calcium phosphate cements

The vast majority of materials used in bone tissue engineering and regenerative medicine are based on calcium phosphates due to their similarity with the mineral phase of natural bone. Among them, calcium phosphate cements, which are composed of a powder and a liquid that are mixed to obtain a moldable paste, are widely used. These calcium phosphate cement pastes can be injected using minimally invasive surgery and adapt to the shape of the defect, resulting in an entangled network of calcium phosphate crystals. Adding an organic phase to the calcium phosphate cement formulation is a very powerful strategy to enhance some of the properties of these materials. Adding some water-soluble biocompatible polymers in the calcium phosphate cement liquid or powder phase improves physicochemical and mechanical properties, such as injectability, cohesion, and toughness. Moreover, adding specific polymers can enhance the biological response and the resorption rate of the material. The goal of this study is to overview the most relevant advances in this field, focusing on the different types of polymers that have been used to enhance specific calcium phosphate cement properties.

Synthesis and Characterization of Hydroxyapatite-Based Nanostructures: Nanoparticles, Nanoplates, Nanofibers and Nanoribbons

Material science is taking an increasing important role in bioengineering and biomedical sciences, aiming to develop new systems and materials capable of adapting to the highly demanding environment of a living organism. One of those materials, Hydroxyapatite (HAp), is the principal calcium orthophosphate present in the mineral phase of bone.

Biologic Response to Carbonated Hydroxyapatite Associated with Orthopedic Device: Experimental Study in a Rabbit Model

BackgroundCarbonated hydroxyapatite (CHA) and related calcium phosphates have been studied for many years as implant materials due to their similarity with the mineral phase of bone. The main limitation of CHA ceramics as well as other bioactive materials is that they have poor mechanical proprieties. It is thought that the mechanical device can cause an increase in metabolic activity and bone healing. In this study we investigated the reactivity and tissue behaviour of implanted CHA biomaterial reinforced by mini external fixator.MethodsThe evaluation of biomaterial biocompatibility and osteogenesis was performed on a rabbit model over a period of 6 weeks by radiological, histological and scanning electron microscopy (SEM) coupled with energy dispersive X-ray SEM-energy-dispersive X-ray (EDX) analysis.ResultsWhile rabbits treated with CHA exhibited more bone formation, and fibrous tissue was observed when empty bone defects were observed. EDX analysis detected little calcium and phosphorus on the surface of the bone that was not implanted, while high content of calcium (62.7%) and phosphorus (38%) was found on the interface bone cement.ConclusionsBone repairing showed that the mini external fixator stimulated the ossification which was pushed when grafted by CHA. This effect may play an important role in the prevention of implant loosening.

Determination of Collagen Fibers Arrangement in Bone Tissue by Using Transformations of Raman Spectra Maps

The goal of this work was to evaluate the ability of Raman spectroscopy to identify molecular organization and chemical composition of extracellular matrix such as the collagen fibers arrangement, the level of mineralization, and the carbonate accumulation in mineral phase in spongy bone of the human head of the femur. Changes in composition and structure of the spongy bone tissue were illustrated using maps of polarized Raman spectra. In particular, the purpose of the present study was determination of arrangement of mineralized collagen on surface of trabecula by using transformations of Raman spectra maps. Transformations of Raman spectra maps were needed in order to remove impact of chemical composition on images of Raman spectra map, which display the collagen fibers orientation. These transformations allow to obtain simultaneously the distribution of constituents of bone and arrangement of collagen fibers on tissue surface. A method to indicate the collagen orientations is developed to understand the molecular organization in healthy and unhealthy bone at the microstructural level.

Osteoconductive surfaces generated from peptide functionalized poly(2-hydroxyethyl methacrylate-co-2-(methacryloyloxy)ethyl phosphate) brushes

Modification of bone implant materials with biofunctional surfaces that facilitate osteoblast adhesion and promote matrix mineralization is an attractive strategy to improve osseointegration. This manuscript discusses the preparation and properties of RGD peptide modified poly(2-hydroxyethyl methacrylate-co-2-(methacryloyloxy)ethyl phosphate) (P(HEMA-co-MEP)) brushes. Whereas the RGD peptide serves to mediate osteoblast adhesion, the MEP phosphate groups were incorporated to mimic the function of natural bone extracellular matrix phosphorylated proteins and stabilize the bone mineral phase. The polymer brush coatings were obtained via surface-initiated atom transfer radical copolymerization of HEMA and MEP followed by peptide post-polymerization modification. The functional properties of these polymer brushes were evaluated in vitro using MC3T3-E1 pre-osteoblast cells. Whereas cell proliferation and differentiation were not significantly influenced by variations in brush composition (MEP, peptide content), incorporation of MEP was found to promote matrix mineralization. Matrix mineralization was most pronounced on substrates that presented both phosphate groups and RGD peptide sequences, indicating that a simultaneous presentation of both of these cues is most favorable.

Setting properties, mechanical strength and in vivo evaluation of calcium phosphate-based bone cements

Calcium phosphate cement (CPC) is a promising material for use in minimally invasive surgery for bone defect repair due to its similarity to the mineral phase of bone, biocompatibility, bioactivity, self-setting characteristics, low setting temperature, adequate stiffness and ease of shaping in complicated geometrics. In this study, we systematically investigate the influence of preparation variables on the final properties of CPCs. We determined the effects of CPC composition, accelerators, seed hydroxyapatite and reaction temperatures on the setting times and compressive strength of CPCs based on tetracalcium phosphate (TTCP), dicalcium phosphate dehydrate (DCPD), dicalcium phosphate anhydrous (DCPA), and α-tricalcium phosphate (α-TCP). The three types of CPCs (TTCP/DCPD, TTCP/DCPA, and TTCP/α-TCP-based bone cements) were prepared by varying the amounts of seed hydroxyapatite and citric acid used as a hardening accelerator. After 24 h of incubation, all three types of bone cements exhibited the characteristic peaks attributable to hydroxyapatite (HA) without characteristic peaks of unreacted raw materials. These results indicated that the bone cements were completely converted to HA. TTCP/DCPD-based bone cements showed faster setting times than TTCP/DCPA and TTCP/α-TCP-based bone cements. As citric acid concentrations in the liquid phase increased, the setting times of all three types of bone cements gradually decreased. However, the concentrations of seed HA in the cements were not related to significant changes in setting time. The compressive strengths of CPCs were significantly influenced by composition and reaction temperature. We also studied the effects of immersion time in physiological solution on the properties of the various CPCs. In the results of in vivo tests, subjects with bone defects implanted with CPCs exhibited more bone formation than control subjects that did not receive implantations of CPCs.

Calcium Phosphate Materials for Bone Repair Application

Calcium phosphates, with a composition and structure similar to the mineral phase of bone, have been recognized as satisfactory materials for bone repair because of their excellent biocompatibility, osteoconductive, and bioactivity. In this paper, the recent related patents are reviewed, including four parts, such as calcium phosphate bioceramics, calcium phosphate coatings, calcium phosphate based particles and calcium phosphate with electromagnetic stimulation for bone repair, which might indicate that calcium phosphate materials have been widely used to repair bone. However, there are still some other interesting points, which are expected to be included in the coming patents, four points of which, such as the calcium phosphates with novel specific microstructures, composites that include calcium phosphates and carbon nanotubes, composites that include calcium phosphates and collagen and calcium phosphates based system that could be combined with many different drugs, are described in the conclusion and perspectives part. Keywords: Calcium phosphate bioceramics, calcium phosphate coatings, calcium phosphate based particles, electromagnetic stimulation, bone repair, BIOCERAMICS, hydroxyapatite, CaPs