Octacalcium Phosphate Crystals Research Articles

Mineralization of Octacalcium Phosphate under Magnetic Field

The mineralization of octacalcium phosphate (OCP) crystals in gel media was studied in the presence of a magnetic field. OCP crystal growth was found to be dependent on mineralization temperature, mineralization time, and the magnetic field. Higher temperatures significantly reduced the mineralization time, which is crucial for directional growth of OCP crystals. The growth of OCP crystals was accelerated by the applied magnetic field, while OCP crystals generated in the presence of a magnetic field exhibited increased length and width of oriented growth. This study provides valuable insights into the influence of mineralization factors in bioprocessing-inspired manufacturing processes.

Multifunctional Sr, Mg, Ag-substituted octacalcium phosphate/carboxymethyl chitosan scaffolds: Antibacterial activity and osteogenic differentiation of human mesenchymal stem cells

Numerous studies indicate that octacalcium phosphate (OCP) possesses promising bone regenerative properties. To further improve the regenerative properties of OCP, several ionic substitutions have been proposed. However, knowledge regarding the synergic effect of multiple ions substituted in OCP crystals and fabricated scaffolds is lacking. In this work, plate-like OCP crystals substituted with 1 and 2.5 mol.% of Mg2+, Sr2+ and Ag+ were prepared by the wet precipitation method. Rietveld refinement analysis indicated a successful substitution of Ca2+ by Mg2+, Sr2+and Ag+ ions. The prepared substituted OCP powders showed a high antibacterial effect towards Gram-negative and Gram-positive bacteria. Further, composite scaffolds based on substituted OCP and carboxymethyl chitosan were prepared by the freeze-crosslinking method. The scaffolds had a highly porous structure, with interconnected pores and homogeneously dispersed OCP within the polymer matrix, and high stability in simulated physiological conditions during 28 days. Good viability and proliferation of human bone marrow-derived mesenchymal stem/stromal cells were confirmed by Live/Dead and CyQUANT assays. Additionally, successful osteogenic differentiation was confirmed by quantification of alkaline phosphatase activity and immunocytochemical staining for type I collagen. Results indicate that ionic substitutions with 1.0 mol.% have the most beneficial effect on stem cell differentiation towards osteoblast phenotype.

Effect of Octacalcium Phosphate Crystals on the Osteogenic Differentiation of Tendon Stem/Progenitor Cells In Vitro.

Synthetic octacalcium phosphate (OCP) activates bone tissue-related cells, such as osteoblasts, osteoclasts, and vascular endothelial cells. However, the effect of OCP on tendon-related cell activation remains unknown. This study examined the response of rat tendon stem/progenitor cells (TSPCs) to OCP and related calcium phosphate crystals in vitro. TSPCs were cultured with OCP and Ca-deficient hydroxyapatite (CDHA) obtained from the original OCP hydrolysis to assess the activity of alkaline phosphatase (ALP) and the expression of osteogenesis-related genes. Compared with CDHA, the effect of OCP on promoting the osteogenic differentiation of TSPCs was apparent: the ALP activity and mRNA expression of RUNX2, Col1a1, OCN, and OPN were higher in OCP than in CDHA. To estimate the changes in the chemical environment caused by OCP and CDHA, we measured the calcium ion (Ca2+) and inorganic phosphate (Pi) ion concentrations and pH values of the TSPCs medium. The results suggest that the difference in the osteogenic differentiation of the TSPCs is related to the ionic environment induced by OCP and CDHA, which could be related to the progress of OCP hydrolysis into CDHA. These results support the previous in vivo observation that OCP has the healing function of rabbit rotator cuff tendon in vivo.

Physicochemical and cytological properties of silicon-doping octacalcium phosphate

• Si was successfully doped in the metastable OCP crystals by a precipitation method. • Si doping led to the contraction of OCP structure with smaller lattice parameters. • Si doping preferred forming a cluster-like hierarchical structure of OCP crystals. • Si-OCP notably promoted the proliferation and osteogenic differentiation of BMSCs. • Si-OCPs enhanced the responses of BMSCs by ionic release and mineral deposits. Silicon was successfully doped into an intermediate precursor of biological apatite, octacalcium phosphate (OCP), accompanied with a lattice contraction. Si-doped OCP significantly enhanced the proliferation and osteogenic capacities of bone marrow mesenchymal stem cells by the synergistical effect of bioactive ions release and mineral deposits, providing a potential bone substitute.

Kappa-carrageenan-Functionalization of octacalcium phosphate-coated titanium Discs enhances pre-osteoblast behavior and osteogenic differentiation.

Bioactive coatings are promising for improving osseointegration and the long-term success of titanium dental or orthopaedic implants. Biomimetic octacalcium phosphate (OCP) coating can be used as a carrier for osteoinductive agents. κ-Carrageenan, a highly hydrophilic and biocompatible seaweed-derived sulfated-polysaccharide, promotes pre-osteoblast activity required for bone regeneration. Whether κ-carrageenan can functionalize OCP-coating to enhance osseointegration of titanium implants is unclear. This study aimed to analyze carrageenan-functionalized biomimetic OCP-coated titanium structure, and effects of carrageenan functionalization on pre-osteoblast behavior and osteogenic differentiation. Titanium discs were coated with OCP/κ-carrageenan at 0.125-2mg/ml OCP solution, and physicochemical and biological properties were investigated. κ-Carrageenan (2mg/ml) in the OCP coating of titanium discs decreased the pore size in the sheet-like OCP crystal by 41.32%. None of the κ-carrageenan concentrations tested in the OCP-coating did affect hydrophilicity. However, κ-carrageenan (2mg/ml) increased (1.26-fold) MC3T3-E1 pre-osteoblast spreading at 1h i.e., κ-Carrageenan in the OCP-coating increased pre-osteoblast proliferation (max. 1.92-fold at 2mg/ml, day 1), metabolic activity (max. 1.50-fold at 2mg/ml, day 3), and alkaline phosphatase protein (max. 4.21-fold at 2mg/ml, day 3), as well as matrix mineralization (max. 5.45-fold at 2mg/ml, day 21). κ-Carrageenan (2mg/ml) in the OCP-coating increased gene expression of Mepe (4.93-fold) at day 14, and Runx2 (2.94-fold), Opn (3.59-fold), Fgf2 (3.47-fold), Ocn (3.88-fold), and Dmp1 (4.59-fold) at day 21 in pre-osteoblasts. In conclusion, κ-carrageenan modified the morphology and microstructure of OCP-coating on titanium discs, and enhanced pre-osteoblast metabolic activity, proliferation, and osteogenic differentiation. This suggests that κ-carrageenan-functionalized OCP coating may be promising for in vivo improvement of titanium implant osseointegration.

Intercalated molecule releasing process of thiomalate substituted octacalcium phosphate crystals during phase conversion

Octacalcium phosphate (OCP) is a layered calcium phosphate compound that has attracted significant attention as a new biomaterial due to its excellent biocompatibility. Its interlayer structure, known as HPO4-OH layer, can be easily replaced by dicarboxylic acid. However, the kinetical ionic and/or molecular exchange process of substituted dicarboxylic acid into OCP interlayer is still unclear. Therefore, in this study, we evaluate the molecular exchange process for thiomalate (SH-malate)-substituted OCP (OCP-SH-malate) during the phase conversion process to other phases in disodium hydrogen phosphate solution (Na2HPO4). When the OCP-SH-malate is immersed in 1 mol/L Na2HPO4 at 40 °C while shaking at 200 rpm, SH-malate, first desorbed from the interlayer of OCP-SH-malate and the HPO4-OH layer structure, is formed at the interlayer where the SH-malate was dissolved. The HPO4-OH layer structure is unstable, making it easily convert to hydroxyapatite (HAp) via dehydration upon further immersion. During this reaction, besides the dissolution of marginal edges of the crystals, crystals morphology was no obvious changes, neither were there evidences of dissolution and collapsing of OCP crystals nor nucleation of newly created crystals.

Multiple proliferation signaling pathways are modulated by octacalcium phosphate in osteoblasts.

Octacalcium phosphate (OCP), a type of bioactive ceramics, may be associated with dentine, tooth apatite, and especially bone generation, and promotes wound healing after fracture. Recently, commercial bone grafting products containing a large amount of OCP material have been released because OCP can be synthesized in large quantities. It is reported to increase cell proliferation, but the interaction between OCP and cell signaling pathways is still unclear. In this study, first, we demonstrated OCP mediated cell signaling pathways with only purified OCP materials. OCP regulated P38, JNK (c-Jun N-terminal kinase), Src, and AKT (protein kinase B) signaling pathways. OCP crystals appeared in the characteristic ribbon shape but varied by several tens of micrometers in size. The X-ray diffraction pattern was the same as previously reported. We studied two concentrations of OCP (10 mg/ml and 20 mg/ml) to understand whether the effect of OCP on cell signaling pathways is dose dependent. We confirmed that OCP treatment affected cell proliferation and alkaline phosphatase and disrupted Src phosphorylation but did not change the total protein level. P38 phosphorylation was activated with OCP treatment and inhibited by SB203580, but P38 total protein level did not change. OCP inhibited JNK phosphorylation signaling, whereas PD98509 inhibited JNK phosphorylation with or without OCP. Interestingly, the AKT total level decreased after OCP treatment, but AKT phosphorylation increased considerably. Our results demonstrate that OCP materials modulate cell signaling pathways and increase cell proliferation.

Octacalcium phosphate crystals including a higher density dislocation improve its materials osteogenecity

Herein, we show that the enhanced osteogenecity of octacalcium phosphate (OCP) biomaterial, recently identified as an important element in hybrid organic–inorganic nanocomposites involved in the initial hydroxyapatite crystal expansion in mammal bones, results from an enhanced chemical property, stemming from the presence of lattice strain and dislocations. Two types of OCPs were synthesized by wet-chemical processing in the presence (c-OCP) and absence (w-OCP) of gelatin, respectively, and subjected to structural, chemical, and biological analyses. High-resolution transmission electron microscopy (HRTEM) and fast Fourier transform (FFT) analyses revealed that c-OCP crystals contained approximately six times higher edge dislocations with Burgers vectors perpendicular to a-axis than that in the case of w-OCP. The dissolution of c-OCP crystal in tris-HCl buffer occurred toward the long axis of the crystal, most likely, toward the lattice strain along the c-axis direction, while w-OCP crystal dissolved toward the a-axis direction. The study suggested that the increment of internal energy by the higher dislocation density contributed promoting c-OCP dissolution and hydrolysis through decreasing the activation energy. c-OCP crystal displayed enhanced in vitro mesenchymal stem 2D cell and 3D spheroid differentiation, in vivo bone formation, and apatite crystallographic orientation in critical-sized rat calvarial defect model as compared to w-OCP crystal, at the same time, converting to apatite structure earlier than w-OCP. The present study demonstrates that dislocation-related dissolution along with enhanced conversion of OCP is a determinant in bone induction, which may be relevant to normal bone development utilizing OCP biomaterials.

Transformation from calcium sulfate to calcium phosphate in biological environment

The formation of a nano-apatite surface layer is frequently considered a measure of bioactivity, especially for non-phosphate bioceramics. In the present study, strontium-doped calcium sulfate, (Ca,Sr)SO4, was used to verify the feasibility of this measure. The (Ca,Sr)SO4 specimen was prepared by mixing 10% SrSO4 by weight with 90% CaSO4·½H2O powder by weight. A solid solution of (Ca,7.6%Sr)SO4 was then produced by heating the powder mixture at 1100 °C for 1 h. The resulting (Ca,Sr)SO4 specimen was readily degradable in phosphate solution. A newly formed surface layer in the form of flakes was formed within one day of specimen immersion in phosphate solution. Structural and microstructure–compositional analyses indicated that the flakes were composed of octacalcium phosphate (OCP) crystals. An amorphous interface containing OCP nanocrystals was found between the newly formed surface layer and the remaining (Ca,Sr)SO4 specimen. The specimen was also implanted into a rat distal femur bone defect. In addition to new bone, fibrous tissue and inflammatory cells were found to interlace the (Ca,Sr)SO4 specimen. The present study indicated that a more comprehensive evaluation is needed to assess the bioactivity of non-phosphate bioceramics.The newly formed surface layer on the (Ca,Sr)SO4 specimen after soaking in phosphate solution for 28 days.

A CRYSTAL CHEMISTRY OF HYDROXYAPATITE : A REVIEW

In chemistry of inorganic crystals, the octacalcium phosphate (OCP) is an apatite based crystals and having a hydrated layers which used in producing of needle or plate-shaped hydroxyapatite (HAP) nanocrystals. Although, the crystals is prepared by a dissolution precipitation reaction. These reaction led to a hexagonal HAP nanocrystals formation under hydrothermal condition from OCP at 180 for 3 hours with pH of solution adjusted to 5.5 and incorporating dicarboxylate e.g. succinate (OOC.(CH2)2.COO)2- ions having Ca/P molar ratio is expected to be 1.56±0.02, where the morphology of OCP are retained. During incorporating of succinate ions in OCP crystals, the hydrogen phosphate (HPO42-) ions in the hydrated layers of OCP are being substituted by succinate ions. Since the crystal system of HAP is hexagonal and its crystalline size in the longitudinal direction of various (a,b,c) axes depending on the thickness of the laminated plate-shaped HAP crystals. Here, their size as perpendicular to the (100) plane which is calculated by introducing of Scherrers equation, D100 = Kλ/(β cos ). The organically modified OCP which generated to HAP have unique nanostructure with micrometer thickness are characterized by using of SEM, FTIR and X-ray diffraction analysis.

Biomimetic Octacalcium Phosphate Bone Has Superior Bone Regeneration Ability Compared to Xenogeneic or Synthetic Bone.

Octacalcium phosphate (OCP) is a precursor of biological apatite crystals that has attracted attention as a possible bone substitute. On the other hand, few studies have examined this material at the experimental level due to the limitations on OCP mass production. Recently, mass production technology of OCP was developed, and the launch of OCP bone substitutes is occurring. In this study, the bone regeneration capacity of OCP products was compared with two of the most clinically used materials: heat-treated bovine bone (BHA) and sintered biphasic calcium phosphate (BCP). Twelve rabbits were used, and defects in each tibia were filled with OCP, BHA, BCP, and left unfilled as control (CON). The tibias were harvested at 4 and 12 weeks, and 15 μm slides were prepared using the diamond grinding method after being embedded in resin. Histological and histomorphometric analyses were performed to evaluate the bone regeneration ability and mechanism. The OCP showed significantly higher resorption and new bone formation in both periods analysed (p < 0.05). Overall, OCP bone substitutes can enhance bone regeneration significantly by activating osteoblasts and a rapid phase transition of OCP crystals to biological apatite crystals (mineralization), as well as providing additional space for new bone formation by rapid resorption.

Bone Tissue Response to Different Grown Crystal Batches of Octacalcium Phosphate in Rat Long Bone Intramedullary Canal Area.

The microstructure of biomaterials influences the cellular and biological responses in the bone. Octacalcium phosphate (OCP) exhibits higher biodegradability and osteoconductivity than hydroxyapatite (HA) during the conversion process from OCP to HA. However, the effect of the microstructure of OCP crystals on long tubular bones has not been clarified. In this study, two types of OCPs with different microstructures, fine-OCP (F-OCP) and coarse-OCP (C-OCP), were implanted in rat tibia for 4 weeks. F-OCP promoted cortical bone regeneration compared with C-OCP. The osteoclasts appearance was significantly higher in the C-OCP group than in the control group (defect only) at 1-week post-implantation. To investigate whether the solubility equilibrium depends on the different particle sizes of OCPs, Nano-OCP, which consisted of nanometer-sized OCPs, was prepared. The degree of supersaturation (DS) tended to decrease modestly in the order of C-OCP, F-OCP, and Nano-OCP with respect to HA and OCP in Tris-HCl buffer. F-OCP showed a higher phosphate ion concentration and lower calcium ion concentration after immersion in the buffer than C-OCP. The crystal structures of both OCPs tended to be converted to HA by rat abdominal implantation. These results suggest that differences in the microstructure of OCPs may affect osteoclastogenesis and result in osteoconductivity of this material in long tubular bone by altering dissolution behavior.

Understanding the Steric Structures of Dicarboxylate Ions Incorporated in Octacalcium Phosphate Crystals.

Octacalcium phosphate (OCP) can incorporate various dicarboxylate ions in the interlayer spaces of its layered structure. Although not proven, these incorporated ions are believed to have a linear structure. In this study, the steric structures of twelve different dicarboxylate ions incorporated into OCP were investigated by comparing the experimentally determined interlayer distance of the OCP with the distance estimated using the molecular sizes of dicarboxylic acids calculated by considering their steric structures. The results revealed that the incorporated succinate, glutarate, adipate, pimelate, suberate, and aspartate ions possessed linear structures, whereas the incorporated azelate, sebacate, methylsuccinate, and malate ions exhibited bent structures. Further, the incorporated mercaptosuccinate ions featured linear, bent, other types of structures. Moreover, the steric structure of the incorporated malonate ion significantly differed from those of other dicarboxylate ions. The computational approach employed in this study is expected to deepen our understanding of the steric structures of dicarboxylate ions incorporated in the OCP interlayer spaces.

Succinic acid and apatite world in producing thin plate-shaped Nanocrystals

Octacalcium phosphate (OCP) crystal is composed of apatite having general formula Ca5(PO4)3X, where X=F, Cl or OH and they are key component of bone and teeth, where its hydrated layers producing thin plate-shaped hydroxyapatite (HAP) nanocrystals. Here, we have been reported the synthesis of sample of HAP nanocystals as CALPHOS or CONTROL and SUC or Suc-20 under hydrothermal condition from succinic acid’s ion based OCP at 180 0C for 3hours. During incorporating of succinate ions in OCP crystals the hydrogen phosphate (HPO42-) ions in the hydrated layers of OCP are being substituted by succinate ions. Since, HAP crystal system is hexagonal and it characterized by using of SEM, XRD and FTIR technique. The crystallite size of HAP crystals are as perpendicular to the (100) plane which can calculate by using as D100 = Kλ/(β cos θ) equation of Scherrer.

Cemp1-p3 Peptide Promotes the Transformation of Octacalcium Phosphate into Hydroxyapatite Crystals

Dental cementum contains unique molecules that regulate the mineralization process in vitro and in vivo, such as cementum protein 1 (CEMP1). This protein possesses amino acid sequence motifs like the human recombinant CEMP1 with biological activity. This novel cementum protein 1-derived peptide (CEMP1-p3, from the CEMP1’s N-terminal domain: (QPLPKGCAAVKAEVGIPAPH), consists of 20 amino acids. Hydroxyapatite (HA) crystals could be obtained through the combination of the amorphous precursor phase and macromolecules such as proteins and peptides. We used a simple method to synthesize peptide/hydroxyapatite nanocomposites using OCP and CEMP1-p3. The characterization of the crystals through scanning electron microscopy (SEM), powder X-ray diffraction (XRD), high--resolution transmission electron microscopy (HRTEM), and Raman spectroscopy revealed that CEMP1-p3 transformed OCP into hydroxyapatite (HA) under constant ionic strength and in a buffered solution. CEMP1-p3 binds and highly adsorbs to OCP and is a potent growth stimulator of OCP crystals. CEMP1-p3 fosters the transformation of OCP into HA crystals with crystalline planes (300) and (004) that correspond to the cell of hexagonal HA. Octacalcium phosphate crystals treated with CEMP1-p3 grown in simulated physiological buffer acquired hexagonal arrangement corresponding to HA. These findings provide new insights into the potential application of CEMP1-p3 on possible biomimetic approaches to generate materials for the repair and regeneration of mineralized tissues, or restorative materials in the orthopedic field.

A thermostability perspective on enhancing physicochemical and cytological characteristics of octacalcium phosphate by doping iron and strontium

Investigation of thermostability will lead the groundbreaking of unraveling the mechanism of influence of ion-doping on the properties of calcium phosphates. In this work, octacalcium phosphate (OCP), a metastable precursor of biological apatite, was used as a stability model for doping ions (Fe3+ and Sr2+) with different ionic charges and radii. After treated under hot air at different temperatures (110–200 °C), the phase, morphology, structure, physicochemical properties, protein affinity, ions release, and cytological responses of the ion-doped OCPs were investigated comparatively. The results showed that the collapse of OCP crystals gradually occurred, accompanying with the dehydration of hydrated layers and the disintegration of plate-like crystals as the temperature increased. The collapsed crystals still retained the typical properties of OCP and the potential of conversion into hydroxyapatite. Compared to the undoped OCP, Fe-OCP, and Sr-OCP had lower and higher thermostability respectively, leading to different material surface properties and ions release. The adjusted thermostability of Fe-OCP and Sr-OCP significantly enhanced the adsorption of proteins (BSA and LSZ) and the cytological behavior (adhesion, spreading, proliferation, and osteogenic differentiation) of bone marrow mesenchymal stem cells to a varying extent under the synergistic effects of corresponding surface characteristics and early active ions release. This work paves the way for understanding the modification mechanism of calcium phosphates utilizing ion doping strategy and developing bioactive OCP-based materials for tissue repair.

Introductory of Scherrer’s Equation in Laminated Plate-Shaped Hexagonal Hydroxyapatite Nanocrystals System

Since, hydroxyapatite (HAP) crystal system is hexagonal and its crystalline size in the longitudinal directions of various (a,b,c) axes, which are depending on the thickness of the laminated plate-shaped HAP crystals. Hence, by applying of Scherrer’s equation D100 = Kλ / (β cos ) we calculated their crystallite size as perpendicular to the (100) plane as well as to comparing the thickness of synthesized needle or laminated thin plate shaped HAP nanocrystals of CALPHOS or CONTROL and SUC or Suc-20 samples under hydrothermal condition via organically modified apatite based octacalcium phosphate (OCP) at 180 for 3h. The pH of solution adjusted to 5.5 with incorporating dicarboxylate or succinate ions having Ca/P molar ratio is expected to be 1.56±0.02, where the morphology of OCP are retained. During incorporating of succinate ions in OCP crystals, the hydrogen phosphate (HPO42-) ions in the hydrated layers of OCP are being substituted by succinate ions. These organically modified OCP which generated to HAP with unique nanocrystallite structure have been characterized by using of SEM, FTIR and X-ray diffraction analysis.

Hydrogen Phosphate (HPO42-) ions Substitution in Hydrated Layers of OCP by Dicarboxylate ions to Hexagonal HAP Nanocrystals System

The octacalcium phosphate (OCP) crystals are apatite bases with hydrated layers in producing of needle or plate-shaped hydroxyapatite (HAP) nanocrystals, which prepared by a dissolution precipitation reaction. The reaction led to a hexagonal HAP nanocrystals formation under hydrothermal condition via OCP at 180? for 3h with pH of solution adjusted to 5.5 with incorporating dicarboxylate or succinate ions having Ca/P molar ratio is expected to be 1.56±0.02, where the morphology of OCP are retained. During incorporating of succinate ions in OCP crystals, the hydrogen phosphate (HPO42-) ions in the hydrated layers of OCP are being substituted by succinate ions. These organically modified OCP which generated to HAP with unique nanostructures have been characterized by using of SEM, FTIR and X-ray diffraction analysis. Since the crystal system of HAP is hexagonally and its crystalline size in the direction of various (a,b,c) axes depending on the thickness of the plate-shaped HAP crystals where their size as perpendicular to the (100) plane is calculated by using of Scherrer equation D100 = K?/(? cos?). The formed hexagonal HAP are used in soft and hard tissue engineering system for biomedical application.

Comparative analysis of bovine serum albumin adsorption onto octacalcium phosphate crystals prepared using different methods.

This study compared bovine serum albumin (BSA) adsorption onto octacalcium phosphate (OCP) materials prepared from two wet preparations in the absence (w-OCP) and presence (c-OCP) of gelatin. Raman spectroscopy was used to analyze the BSA adsorption onto OCPs in a 150 mM Tris-HCl buffer containing 0.5 mM calcium and inorganic phosphate (Pi) ions at pH 7.4 and at 37°C. The degree of supersaturation of the supernatants after the adsorption was determined by measuring the ion composition. The results showed that BSA adsorption onto w-OCP was higher than that for c-OCP. The calcium ion concentration of the supernatant decreased for both w-OCP and c-OCP, whereas the Pi ion concentration increased, approaching OCP equilibria at different saturation levels. BSA adsorbed even onto c-OCP, which included a small amount of gelatin during c-OCP preparation. These results indicate that the biodegradability of w-OCP and c-OCP may be modulated through interactions with serum proteins.

Effect of Surrounding Chemical Environment on Adsorption and Accumulation of Serum Protein onto Octacalcium Phosphate Crystals

Effect of Surrounding Chemical Environment on Adsorption and Accumulation of Serum Protein onto Octacalcium Phosphate Crystals