Microporous Calcium Phosphate Research Articles

Crystallization of carboplatin-loaded onto microporous calcium phosphate using high-vacuum method: Characterization and release study.

Drug delivery systems are a new approach to increase therapeutic efficacy and to reduce the side effects of traditional treatments. Calcium phosphates (CaPs) have been studied as drug delivery systems, especially in bone diseases. However, each system has some particularities that depend on the physical and chemical characteristics of the biomaterials and drug interaction. In this work, granulated CaPs were used as a matrix for loading the anticancer drug carboplatin using the high-vacuum method. Five compositions were applied: hydroxyapatite (HA), β-tricalcium phosphate (β-TCP), biphasic HAp 60%/β-TCP 40% (BCP), β-TCP/MgO nanocomposite, and β-TCP/SiO2 nanocomposite. Carboplatin drug in 50, 60, and 70 mg/g was precipitated on the surface of CaPs. Morphological, chemical and surface modifications in the carboplatin-CaPs were investigated by scanning electron microscopy (SEM), energy dispersive spectroscopy (EDS), backscattered electron microscopy (BSE), X-ray diffraction (XRD), X-ray fluorescence spectroscopy (XRF), Fourier transform infrared (FT-IR), and Raman spectroscopy. The characterization of the CaP-carboplatin biomaterials showed heterogeneous crystalline precipitation of the drug, and no morphological modifications of the CaPs biomaterials. The in vitro release profile of carboplatin from CaPs was evaluated by the ultraviolet-visible (UV-Vis) method. The curves showed a burst release of upon 60% of carboplatin loaded followed by a slow-release of the drug for the time of the study. The results were typical of a low-interaction system and physisorption mechanism. The high-vacuum method permitted to load the high amount of carboplatin drug on the surface of the biomaterials despite the low interaction between carboplatin and CaPs.

Digital Light Processing of Freeze-cast Ceramic Layers for Macroporous Calcium Phosphate Scaffolds with Tailored Microporous Frameworks.

The objective of the present study is to demonstrate the versatility of the digital light processing (DLP) technique particularly when using a freeze-cast ceramic layer as the feedstock, which can manufacture porous calcium phosphate (CaP) scaffolds with arbitrarily designed macroporous structures with tailored microporous frameworks specially designed for bone scaffold applications. For this goal, we employed camphene-camphor as the freezing vehicle and porogen for the preparation of photocurable CaP suspensions containing diurethane dimethacrylate (UDMA) monomers. After freeze-casting, the CaP suspensions could be solidified at controlled temperatures (~33–38 °C) and then be photopolymerized by DLP. All produced CaP scaffolds fairly resembled the designed macroporous structures (the gyroid structure with two interpenetrating macropore networks). In addition, numerous micropores were created in the CaP filaments, while the microporosity increased with increasing the camphene-camphor amount from 40 vol % to 60 vol %. As a consequence, compressive strength and modulus of hierarchically porous CaP scaffolds decreased due to an increase in overall porosity. However, reasonable mechanical properties could be obtained at high porosities owing to the CaP frameworks constructed in a periodic manner. In addition, excellent water penetration capability, biocompatibility, and apatite-forming ability were obtained, which were attributed to the microporous CaP frameworks with good pore interconnectivity and large surface area.

Sintering And Characterization Of A Hydroxya-patite Matrix And Hydroxyapatite Matrix Nano-composites

Microporous calcium phosphate biomaterials are known for their physical and biological applications. Among the best known are the stoichiometric hydroxyapatite (HA) and tricalcium phosphate (TCP). This is because these biomaterials exhibit chemical and crystallographic compositions which are similar to that found in bones and teeth. The use of nanotechnology enables obtaining calcium phosphate nanostructured powders and calcium phosphate nanocomposite matrix formed by a second nano phase of type SiO2, TiO2, Al2O3-a, ZrO2, Mg. Different methods and techniques for the synthesis and preparation of nanostructured powders and biomaterials are noted in the literature, but it is known that not all lead to the same results. Calcium phosphates nanostructured biomaterials are a new class of biomaterials which provide new physical, morphological, nanostructural and microstructural features with interconnected microporosity which are promising to wettability, capillary action, cell adhesion and proliferation on the surface of grains and micropores. Based on research of these biomaterials, it has been found that they show potential applications in traumatology, orthopedic and dental applications in reconstruction, defects and bone tissue repairing, implants attachment and dental remineralization treatment. This study was aimed at the sintering and characterization of an HA matrix and three nanocomposite biomaterials with 5% by volume of the respective second phases: SiO2, ZrO2 and Al2O3-a in the HA matrix. The HA powder and nanocomposite HA/SiO2 were sintered at 1100 °C/2h. HA/ZrO2 nanocomposite powder followed two sintering conditions: a temperature of 1100 °C/2h and the other, at 1300 ºC/2h. HA/Al2O3-a nanocomposite powder was only sintered at 1300 ºC/2h. The biomaterials were characterized by scanning electron microscopy, X-ray diffraction and open porosity and hydrostatic density were also determined by applying the Arthur method. The results are encouraging and show for HA, HA/SiO2, HA/ZrO2 biomaterials (obtained by sintering at 1100 °C) interconnected microporous microstructures, formed by fine grains which are favorable for the expected wettability and capillarity characteristics.

Platelet-rich plasma encapsulation in hyaluronic acid/gelatin-BCP hydrogel for growth factor delivery in BCP sponge scaffold for bone regeneration.

Microporous calcium phosphate based synthetic bone substitutes are used for bone defect healing. Different growth factor loading has been investigated for enhanced bone regeneration. The platelet is a cellular component of blood which naturally contains a pool of necessary growth factors that mediate initiation, continuation, and completion of cellular mechanism of healing. In this work, we have investigated the encapsulation and immobilization of platelet-rich plasma (PRP) with natural polymers like hyaluronic acid (HA) and gelatin (Gel) and loading them in a biphasic calcium phosphate (BCP) scaffold, for a synthetic-allologous hybrid scaffold. Effect of PRP addition in small doses was evaluated for osteogenic potential invitro and invivo. BCP (10%) mixed HA-Gel hydrogel with or without PRP, was loaded into a BCP sponge scaffold. We investigated the hydrogel-induced improvement in mechanical property and PRP-mediated enhancement in biocompatibility. Invitro studies for cytotoxicity, cell attachment, and proliferation were carried out using MC3T3-E1 pre-osteoblast cells. In invitro studies, the cell count, cell proliferation, and cell survival were higher in the scaffold with PRP loading than without PRP. However, in the invivo studies using a rat model, the PRP scaffold was not superior to the scaffold without PRP. This discrepancy was investigated in terms of the interaction of PRP in the invivo environment.

Microporous calcium phosphate ceramics driving osteogenesis through surface architecture.

The presence of micropores in calcium phosphate (CaP) ceramics has shown its important role in initiating inductive bone formation in ectopic sites. To investigate how microporous CaP ceramics trigger osteoinduction, we optimized two biphasic CaP ceramics (i.e., BCP-R and BCP-S) to have the same chemical composition, equivalent surface area per volume, comparable protein adsorption, similar ion (i.e., calcium and phosphate) exchange and the same surface mineralization potential, but different surface architecture. In particular, BCP-R had a surface roughness (Ra) of 325.4 ± 58.9 nm while for BCP-S it was 231.6 ± 35.7 nm. Ceramic blocks with crossing or noncrossing channels of 250, 500, 1000, and 2000 µm were implanted in paraspinal muscle of dogs for 12 weeks. The percentage of bone volume in the channels was not affected by the type of pores (i.e., crossing vs. closed) or their size, but it was greatly influenced by the ceramic type (i.e., BCP-R vs. BCP-S). Significantly, more bone was formed in the channels of BCP-R than in those of BCP-S. Since the two CaP ceramics differed only in their surface architecture, the results hereby demonstrate that microporous CaP ceramics may induce ectopic osteogenesis through surface architecture.

Preparation of microporous and macroporous biphasic calcium phosphate scaffolds modified via a biomimetic method for bone tissue engineering applications

Preparation of microporous and macroporous biphasic calcium phosphate scaffolds modified via a biomimetic method for bone tissue engineering applications

Microporous calcium phosphate ceramics as tissue engineering scaffolds for the repair of osteochondral defects: Histological results

Treatment of defects in joint cartilage aims to re-establish normal joint function. In vitro experiments have shown that the application of synthetic scaffolds is a promising alternative to existing therapeutic options. A sheep study was conducted to test the suitability of microporous pure β-tricalcium phosphate (TCP) ceramics as tissue engineering scaffolds for the repair of osteochondral defects. Cylindrical plugs of microporous β-TCP (diameter: 7mm; length: 25mm; porosity: 43.5±2.4%; pore diameter: ∼5μm) with interconnecting pores were used. Scaffolds were seeded with autologous chondrocytes in vitro and cultured for 4weeks. A drill hole (diameter 7mm) was placed in both medial femoral condyles of sheep. For the left knee the defect was filled with a TCP plug and for the right knee the defect was left empty. After 6, 12, 26 and 52weeks, seven animals from each group were killed and studied. The samples were examined employing histological, histomorphometric and immunohistological methods as well as various imaging techniques (X-ray, microcomputer tomography and scanning electron microscopy). After explantation the cartilage defects were first assessed macroscopically. There were no signs of infection or inflammation. Histological grading scales were used for assessment of bony integration and cartilage repair. An increasing degradation (81% after 52weeks) of the ceramic with concomitant bone formation was observed. The original structure of cancellous bone was almost completely restored. After 26 and 52weeks, collagen II-positive hyaline cartilage was detected in several samples. New subchondral bone had formed. The formation of cartilage began at the outer edge and proceeded to the middle. According to the O’Driscoll score, values corresponding to healthy cartilage were not reached after 1year. Integration of the newly formed cartilage tissue into the surrounding native cartilage was found. The formation of biomechanical stable cartilage began at the edge and progressed towards the centre of the defect. After 1year this process was still not completed. Microporous β-TCP scaffolds seeded with chondrocytes are suitable for the treatment of osteochondral defects.

Micro-porous calcium phosphate coatings on load-bearing zirconia substrate: Processing, property and application

This study presents the design, processing, properties and potential applications of a novel layered bio-ceramic composites consisting of three different micro-porous calcium phosphate coatings on strong zirconia cores manufactured using a recently developed slip coating-deposition and coating-substrate co-sintering technique. Detailed microstructures of the three graded micro-porous calcium phosphate coatings, and the coating/substrate interface have been investigated. Also, the flexural strength of the bio-ceramic composite and the bonding state between the coatings and zirconia substrate have been characterized. A preliminary and limited in vitro cell test indicates that the new scaffold composite has no cytotoxicity to the fibroblasts which can attach, proliferate and grow on the coating surfaces. Because of the combination of bio-function and strength, such layered load-bearing bio-ceramic composites are a potential candidate for large-scale head bone repairs.

Microporous calcium phosphate ceramics as tissue engineering scaffolds for the repair of osteochondral defects: Biomechanical results

This work investigated the suitability of microporous β-tricalcium phosphate (TCP) scaffolds pre-seeded with autologous chondrocytes for treatment of osteochondral defects in a large animal model. Microporous β-TCP cylinders (Ø 7mm; length 25mm) were seeded with autologous chondrocytes and cultured for 4weeks in vitro. Only the upper end of the cylinder was seeded with chondrocytes. Chondrocytes formed a multilayer on the top. The implants were then implanted in defects (diameter 7mm) created in the left medial femoral condyle of ovine knees. The implants were covered with synovial membrane from the superior recess of the same joint. For the right knees, an empty defect with the same dimensions served as control. Twenty-eight sheep were split into 6-, 12-, 26- and 52week groups of seven animals. Indentation tests with a spherical (Ø 3mm) indenter were used to determine the biomechanical properties of regenerated tissue. A software-based limit switch was implemented to ensure a maximal penetration depth of 200μm and maximal load of 1.5N. The achieved load, the absorbed energy and the contact stiffness were measured. Newly formed cartilage was assessed with the International Cartilage Repair Society Visual Assessment Scale (ICRS score) and histomorphometric analysis. Results were analysed statistically using the t-test, Mann–Whitney U-test and Wilcoxon test. Statistical significance was set at p<0.05. After 6weeks of implantation, the transplanted area tolerated an indentation load of 0.05±0.20N. This value increased to 0.10±0.06N after 12weeks, to 0.27±0.18N after 26weeks, and 0.27±0.11N after 52weeks. The increase in the tolerated load was highly significant (p<0.0001), but the final value was not significantly different from that of intact cartilage (0.30±0.12N). Similarly, the increase in contact stiffness from 0.87±0.29Nmm−1 after 6weeks to 3.14±0.86Nmm−1 after 52weeks was highly significant (p<0.0001). The absorbed energy increased significantly (p=0.02) from 0.74×10−6±0.38×10−6Nm after 6weeks to 2.83×10−6±1.35×10−6Nm after 52weeks. At 52weeks, the International Cartilage Repair Society (ICRS) scores for the central area of the transplanted area and untreated defects were comparable. In contrast, the score for the area from the edge to the centre of the transplanted area was significantly higher (p=0.001) than the score for the unfilled defects. A biomechanically stable cartilage was built outside the centre of defect. After 52weeks, all but one empty control defect were covered by bone and a very thin layer of cartilage (ICRS 7 points). The empty hole could still be demonstrated beneath the bone. The histomorphometric evaluation revealed that 81.0±10.6% of TCP was resorbed after 52weeks. The increase in TCP resorption and replacement by spongy bone during the observation period was highly significant (p<0.0001). In this sheep trial, the mechanical properties of microporous TCP scaffolds seeded with transplanted autologous chondrocytes were similar to those of natural cartilage after 52weeks of implantation. However, the central area of the implants had a lower ICRS score than healthy cartilage. Microporous TCP was almost fully resorbed at 52weeks and replaced by bone.

Antimicrobial peptides on calcium phosphate-coated titanium for the prevention of implant-associated infections

Prevention of implant-associated infections has been one of the main challenges in orthopaedic surgery. This challenge is further complicated by the concern over the development of antibiotic resistance as a result of using traditional antibiotics for infection prophylaxis. The objective of this study was to develop a technique that enables the loading and local delivery of a unique group of cationic antimicrobial peptides (AMP) through implant surfaces. A thin layer of micro-porous calcium phosphate (CaP) coating was processed by electrolytic deposition onto the surface of titanium as the drug carrier. The broad spectrum AMP Tet213 (KRWWKWWRRC) was selected and loaded onto the CaP coating. SEM, XRD and FTIR analyses confirmed the CaP coating to be micro-porous octacalcium phosphate. By using a luminescence spectrometer technique, it was demonstrated that a 7 μm thick porous CaP coating could load up to 9 μg of AMP/cm2 using a simple soaking technique. The drug-loaded CaP coating (CaP-Tet213) was not cytotoxic for MG-63 osteoblast-like cells. The CaP-Tet213 implants had antimicrobial activity against both Gram-positive (Staphylococcus aureus) and Gram-negative (Pseudomonas aeruginosa) bacteria with 106-fold reductions of both bacterial strains within 30 min as assessed by measuring colony-forming units (CFU). Repeated CFU assays on the same CaP-Tet213 specimen demonstrated retention of antimicrobial activity by the CaP-Tet213 surfaces through four test cycles. The susceptibility of bacteria to the CaP-Tet213 surfaces was also evaluated by assessing the inhibition of luminescence of P. aeruginosa containing a luxCDABE cassette at 4 h and 24 h with ∼92% and ∼77% inhibition of luminescence, respectively. It was demonstrated that CaP-Tet213 was a more efficient antimicrobial coating than CaP-MX226, CaP-hLF1-11 or CaP-tobramycin following incubation of CaP implants with equimolar concentrations of Tet213, the commercially developed antimicrobial peptide MX-226, hLF1-11 or tobramycin. A device coated with CaP-Tet213 could be a potential solution for the prevention of the peri-implant infection in orthopaedics.

Enhanced Gap Filling and Osteoconduction Associated with Alendronate-Calcium Phosphate-Coated Porous Tantalum

Porous tantalum has been shown to be effective in achieving bone ingrowth. However, in some circumstances, bone quality or quantity may be insufficient to allow adequate bone ingrowth. We hypothesized that local delivery of alendronate from porous tantalum would enhance the ability of the tantalum to achieve bone ingrowth when there is a gap between the implant and bone. We evaluated the effect of alendronate-coated porous tantalum on new bone formation in an animal model incorporating a gap between the implant and bone. A cylindrical porous tantalum implant was implanted in the distal part of each femur in eighteen rabbits (a total of thirty-six implants) and left in situ for four weeks. Three types of porous tantalum implants were inserted: those with no coating (the control group), those with microporous calcium phosphate coating, and those coated with microporous calcium phosphate and alendronate. Subcutaneous fluorescent labeling was used to track new bone formation. Bone formation was analyzed with backscattered electron microscopy and fluorescent microscopy of undecalcified samples. The relative increases in the mean volume of gap filling, bone ingrowth, and total bone formation in the group treated with the porous tantalum implants coated with calcium phosphate and alendronate were 143% (p < 0.001), 259% (p < 0.001), and 193% (p < 0.001), respectively, compared with the values in the control group treated with the uncoated porous tantalum implants. The percentage of the length of the implant that was in contact with new bone in the group treated with the calcium phosphate and alendronate coating was increased by an average of 804% compared with the percentage in the group treated with the uncoated implants. The study demonstrated significantly enhanced filling of the bone-implant gap and bone ingrowth in association with the porous tantalum implants coated with calcium phosphate and alendronate.

Modeling vancomycin release kinetics from microporous calcium phosphate ceramics comparing static and dynamic immersion conditions

The release kinetics of vancomycin from calcium phosphate dihydrate (brushite) matrices and polymer/brushite composites were compared using different fluid replacement regimes, a regular replacement (static conditions) and a continuous flow technique (dynamic conditions). The use of a constantly refreshed flowing resulted in a faster drug release due to a constantly high diffusion gradient between drug loaded matrix and the eluting medium. Drug release was modeled using the Weibull, Peppas and Higuchi equations. The results showed that drug liberation was diffusion controlled for the ceramics matrices, whereas ceramics/polymer composites led to a mixed diffusion and degradation controlled release mechanism. The continuous flow technique was for these materials responsible for a faster release due to an accelerated polymer degradation rate compared with the regular fluid replacement technique.

Ectopic bone formation by microporous calcium phosphate ceramic particles in sheep muscles

Calcium phosphate ceramics are widely used in bone reconstructive surgery because of their osteconductive properties. However, these materials generally lack osteoinductive properties required to support bone healing in large defects. In this article, we study the osteoinductive potential of calcium phosphate ceramic particles implanted for 6 months into the dorsal muscles of eight adult female sheep. Microporous biphasic calcium phosphate (MBCP) granules of 1–2 mm composed of hydroxyapatite and beta-tricalcium phosphate (60/40) had macropores of 450 μm, micropores of 0.43 μm, and a specific surface area of 1.8 m 2/g. After 6 months in the back muscles of sheep, the explants composed of MBCP granules were hard and encapsulated by normal muscle tissue. Ectopic bone formation with Haversian structures was observed in close contact with the MBCP granules in histological sections. Back-scattered electron microscopy and micro-computed tomography indicated that approximately 10% of well-mineralized bone with mature osteocytes had formed between or upon the granules. The ectopic bone showed trabeculae bridging the MBCP granules. Both the number and thickness of the trabeculae formed between the MBCP particles were comparable to those measured in spongious bone. The overall results therefore confirmed the presence of mature bone after intramuscular implantation of MBCP granules. The different hypotheses explaining ectopic bone formation induced by MBCP granules are discussed. Synthetic bone substitutes with osteoinductive properties could be used in bone reconstructive surgery.

Bone-Bonding Ability of Ce-TZP/Al&lt;sub&gt;2&lt;/sub&gt;O&lt;sub&gt;3&lt;/sub&gt; Nanocomposite with a Microporous Surface and Calcium Phosphate Coating

The nano-composite of a ceria-stabilized tetragonal zirconia polycrystals (Ce-TZP) and alumina (Al2O3) polycrystals (Ce-TZP/Al2O3) is attractive as a load-bearing bone substitute because of its mechanical properties and phase stability. We have developed a new method of hydrofluoric acid and heat treatment (HFT) to give a microporous structure to the surface of this Ce-TZP/Al2O3 nanocomposite ceramic. Bone-bonding ability of a microporous surface and calcium phosphate coating on Ce-TZP/Al2O3 composite has been investigated through in vivo detaching model. Thin calcium phosphate coating layer was added by alternate soaking process, and thick CaP layer was produced by soaking in simulated body fluid for 5 days. HFT treated Ce-TZP/Al2O3 composite showed high bone-bonding ability compared with the control group. Thick and thin CaP coating accelerated bone-bonding ability in early post-implantation period. The submicron microporous surface was beneficial for achieving mechanical interlocking between the ceramic and surrounding bone. These results suggest the possibility of using a Ce-TZP/Al2O3 nanocomposite ceramic with microporous surface and calcium phosphate coating as the bearing material for uncemented total joint replacements.

Microneedles coated with porous calcium phosphate ceramics: Effective vehicles for transdermal delivery of solid trehalose

Trehalose (alpha-D-glucopyranosyl-alpha-D-glucopyranoside) is recognized as a promising fast-dissolving solid reservoir capable of stabilizing the native structure of proteins and suitable for loading with a wide variety of bioactive substances. Currently, there is a growing interest in developing cost-effective methods for immobilizing solid trehalose on arrays of microneedles for delivering protein-based and DNA-based vaccine to the epidermis. In the present work, micro-porous calcium phosphate coatings were used to provide a biocompatible interface with a large surface area for the effective immobilization of trehalose on microneedles. Calcium phosphate coatings with varying degrees of porosity were electrochemically synthesized on the tips of stainless steel acupuncture needles and loaded with solid trehalose. Skin experiments were designed to determine the ability of micro-porous calcium phosphate coatings to deliver solid trehalose into epidermis without breaking during insertion. The mechanical performance of the coatings was assessed by inserting the tips of the coated needles into human skin to an average depth of 100-300 microm and then removing them for analysis by scanning electron microscopy. Microporous calcium phosphate coatings loaded with trehalose effectively breached the stratum corneum and allowed direct access to the epidermis without breaking and without stimulating nerves in deeper tissues.

A Hybrid Laser/Aerosol Method for the Synthesis of Porous Nanostructured Calcium Phosphate Materials for Bone Tissue Engineering Applications

ABSTRACTWe present a new synthesis method based on laser generation and processing of aerosol particles that can produce calcium phosphate coatings in a porous nanostructured configuration. The process uses laser ablation of crystalline hydroxyapatite targets to produce a calcium phosphate aerosol comprising micro- and nanoparticles that are processed and deposited on metallic substrates under well-controlled temperature and ambient conditions, creating a microporous calcium phosphate network suitable for growth of biogenic calcium phosphate materials. Laser ablation is carried out using a KrF excimer laser at fluences between 0.4 J/cm2 and 2.8 J/cm2 and temperatures ranging from 500°C to 760°C. X-ray diffraction and scanning electron microscopy measurements on samples deposited above 750°C show that the obtained material is crystalline hydroxyapatite with good mechanical stability. Its microstructure features a porous framework of partially sintered microparticles surrounded by nanoparticulate material.