Thin Hydroxyapatite Research Articles

Preparation and growth behaviours of low porosity hydroxyapatite with enhanced adhesion by electrochemical deposition on micro-arc oxide coatings

Electrochemical deposition (ED) has been proposed as a low temperature approach for preparing hydroxyapatite (HA) coatings. To overcome the problem of low adhesion, the preparation of thin and low-porosity coatings on Ti substrates pre-treated by micro-arc oxidation (MAO) is a good strategy. In this paper, a thin and dense HA coating was obtained by modifying parameters at pH 6.4, 45 °C, and 0.5 mA·cm−2. The growth behaviours of HA electrodeposited on MAO coatings were investigated by SEM and EDS technology and summarized that the HA was initially generated in the MAO micropores, then covered the entire MAO surface and gradually grew outwards. This growth pattern was also confirmed by ATR-FTIR, XPS, EIS, open potential, and wettability results at different ED cycles. Adjusting 1:1 power-on/off time ratio affected the morphology of HA grown in the pores in the early stage and also increased the interfacial strength, up to 10.69 MPa at duty cycle 12/24.The cross-section of the final specimen was characterised by TEM and EDS, indicating that amorphous layers, nanocrystalline HA, and submicron HA were formed sequentially from the TiO2(Ca)/HA interface outwards. The increase in Ca2+ concentration and the formation of CaO bonds at the interface may be important reasons for the adhesive enhancement. This process provides a strategy to enhance the strength of bioceramic materials on Ti substrates.

Multilayered porous hydroxyapatite coating on Ti6Al4V implant with enhanced drug delivery and antimicrobial properties

Failure of the orthopedic implant due to bacterial infections and poor osseointegration is a common problem in total hip arthroplasty. The challenge here is improving the antibacterial properties of implant surface by loading the drug, without compromising the bioactivity and its performance. Plasma sprayed porous hydroxyapatite (HA) coatings on metallic implant surface, impregnated with antibiotic is found to be a solution. In this context, sustained delivery needs more drug being incorporated in the coating, for which the coating requires to be more porous. However, increased porosity makes the brittle HA coating weaker and unsuitable for intended application. The present study introduces the interesting concept of a multilayer HA coating, with a porosity gradient from bottom to top in different layers. Thin dense HA layer, adjacent to Ti alloy substrate offers strong adhesion with the implant surface, while the gradient increase in porosity on the upper layers maintain the integrity of the coating, while allowing the loading of drug in higher quantity. Developed coating is filled with drug containing biodegradable polymer through vacuum impregnation method. This structure offered an improvement in drug loading by 145% (16 μg/mm2) and increased the drug release duration (in days) by almost 4 times. In addition to these, effective filling of the pores with polymer improved the fracture toughness of the HA coating significantly, by restricting the crack growth. Improved tribological behavior was also noted, along with excellent bioactivity and osseointegration.

Corrosion-Resistance Analysis of HA Layer Deposited through Electrophoresis on Ti4Al4Zr Metallic Substrate

An alloy surface with possible applications in the medical field, Ti4A14Zr, was improved through the deposition of a thin hydroxyapatite (HA) layer. In this paper, we analyzed the growth of a HA layer through electrophoresis and the corrosion resistance of the metallic sample covered with the ceramic layer. The substrate surface was processed via chemical procedures before the HA deposition. The state of the metallic surface and that of the layer of HA were investigated using scanning electron microscopy (SEM) and energy dispersive spectroscopy (EDS) analysis of the chemical composition. The results indicate a high increase in the corrosion resistance associated with the ceramic layer compared to the metallic basic layer. Moreover, the analysis revealed the formation of a homogeneous TiO2 layer on the surface of the metallic substrate. The titanium oxide layer identified by SEM–EDS and confirmed by EIS was very homogeneous and resistant, with a compact microstructural appearance and submicron dimension. The layer composed of TiO2 and HA provided good corrosion protection.

Influence of ZrO2 Addition on Structural and Biological Activity of Phosphate Glasses for Bone Regeneration.

Zirconium doped calcium phosphate-based bioglasses are the most prominent bioactive materials for bone and dental repair and regeneration implants. In the present study, a 8ZnO–22Na2O–(24 − x)CaO–46P2O5–xZrO2 (0.1 ≤ x ≤ 0.7, all are in mol%) bioglass system was synthesized by the conventional melt-quenching process at 1100 °C. The glass-forming ability and thermal stability of the glasses were determined by measuring the glass transition temperature (Tg), crystallization temperature (Tc), and melting temperature (Tm), using differential thermal analysis (DTA). The biological activity of the prepared samples was identified by analyzing X-ray diffraction (XRD), Fourier transform infrared spectroscopy (FTIR) and scanning electron microscopy-energy dispersive spectra (SEM-EDS), before and after immersion in simulated body fluid (SBF) for various intervals of 0, 1 and 5 days, along with the magnitude of pH and the degradation of glasses also evaluated. The obtained results revealed that the glass-forming ability and thermal stability of glasses increased with the increase in zirconia mol%. The XRD, FTIR, and SEM-EDS data confirmed a thin hydroxyapatite (HAp) layer over the sample surface after incubation in SBF for 1 and 5 days. Furthermore, the development of layer found to be increased with the increase of incubation time. The degradation of the glasses in SBF increased with incubation time and decreased gradually with the increase content of ZrO2 mol% in the host glass matrix. A sudden rise in initial pH values of residual SBF for 1 day owing to ion leaching and increase of Ca2+ and PO43− ions and then decreased. These findings confirmed the suitability of choosing material for bone-related applications.

Electron Irradiation Effect in Surface Modification of Hydroxyapatite Ceramics

The structure of ultrathin sections of dense submicrocrystalline hydroxyapatite (HA) (Ca10(PO4)6(OH)2) ceramics has been studied by X-ray diffraction and high-resolution transmission electron microscopy (TEM). The results demonstrate that high-energy electron irradiation can lead to surface modification with thin CaO, α-Ca3(PO4)2, and HA nanoparticles. Two-level interference contrast and double Fourier transformation of TEM images have been shown to be effective tools for revealing and identifying nanoparticles, respectively.

Fabrication and In vitro Bioactivity of Robust Hydroxyapatite Coating on Porous Titanium Implant

Compared with the traditional dental implant, TixOs® manufactured by direct laser metal forming(DLMF) technology exhibits improved capability for bone osteointegration due to its porous surface structure, and has achieved remarkable clinical effect. However, like the traditional titanium and other alloyed implants, the porous titanium implant TixOs® also has relatively weak bioactivity. To address this issue, a proper surface modification method may be needed. Hydroxyapatite(HA) has been widely used in implant surface coating for its similar chemical composition to bone tissue and its osteoconductive properties. Thus, combining TixOs® implants with hydroxyapatite can be an efficient way to enhance their bioactivity. We herewith reported a competent pulsed laser deposition(PLD) method of coating nano-sized HA thin film onto the porous TixOs® implant. The HA coatings were characterized by means of scanning electron microscopy(SEM), energy dispersive X-ray spectroscopy(EDS), X-ray photoelectron spectroscopy(XPS) and focused ion beam(FIB) method, and nanocrystal sized thin HA films were identified on the surface of TixOs® implants. The low cytotoxicity and improved cell proliferation ability of HA coated implants were further tested and verified using MC 3T3 E1 cells with the consideration of the controlling group. Our results show that a stable and bioactive HA thin film is able to form on the surface of the porous titanium implant by PLD method. This may benefit the further clinical application of TixOs® implants.

In vitro cytotoxicity of calcium phosphate cement reinforced with multiwalled carbon nanotubes.

The in vitro cytotoxicity of both the multiwalled carbon nanotubes (MWCNT) in suspension with culture medium and the tetracalcium phosphate/monetite cement with addition of 0.8 wt% of MWCNTs on fibroblasts and osteoblasts were studied. The cytotoxicity was evaluated by MTS test (formazan) and live/dead staining. No cytotoxicity of MWCNT extract was measured contrary to about 60% reduction in proliferation of fibroblasts in MWCNT suspension as compared with negative control. The several contact cytotoxicity of MWCNT composite cement surfaces on seeded cells was demonstrated by MTS test and live/dead staining of damaged fibroblasts and dead osteoblasts after 72 h of culture. The detailed microstructure analysis showed a significant refinement of the surface texture due to the formation of thin needle-like hydroxyapatite particles on MWCNTs and this effect could be responsible for cytotoxicity of composites.

Titania Nanotubes/Hydroxyapatite Nanocomposites Produced with the Use of the Atomic Layer Deposition Technique: Estimation of Bioactivity and Nanomechanical Properties.

Titanium dioxide nanotubes/hydroxyapatite nanocomposites were produced on a titanium alloy (Ti6Al4V/TNT/HA) and studied as a biocompatible coating for an implant surface modification. As a novel approach for this type of nanocomposite fabrication, the atomic layer deposition (ALD) method with an extremely low number of cycles was used to enrich titania nanotubes (TNT) with a very thin hydroxyapatite coating. X-ray diffraction (XRD) and scanning electron microscopy (SEM) were used for determination of the structure and the surface morphology of the fabricated nanocoatings. The biointegration activity of the layers was estimated based on fibroblasts’ proliferation on the TNT/HA surface. The antibacterial activity was determined by analyzing the ability of the layers to inhibit bacterial colonization and biofilm formation. Mechanical properties of the Ti6Al4V/TNT/HA samples were estimated by measuring the hardness, Young’s module, and susceptibility to scratching. The results revealed that the nanoporous titanium alloy coatings enriched with a very thin hydroxyapatite layer may be a promising way to achieve the desired balance between biofunctional and biomechanical properties of modern implants.

Application of Solution Plasma Surface Modification Technology to the Formation of Thin Hydroxyapatite Film on Titanium Implants

Hydroxyapatite (HA) coatings on titanium implants enhance rapid bone formation around the implant due to their osteoconductive property. The present study aimed to achieve a thin and uniform HA film coating on titanium implants by solution plasma treatment (SPT). Commercially pure titanium and porous titanium disks were employed. A pulse plasma generator was used on the disks for 30 min. Morphologic and crystallographic features of the deposited films were examined by scanning electron microscopy (SEM) and X-ray diffractometry (XRD). To evaluate the wettability of the disks, water droplet (20 µL) surfaces were measured using a contact angle analyzer. The initial attachment of osteoblast-like cells (MC3T3E1) on the titanium substrates before and after solution plasma treatment was evaluated by counting the number of attached cells after incubation for 4 h. After immersion in the mineralizing solution for up to seven days, no crystals were observed on the polished-Ti surface. A more uniform and dense precipitation of round and grown crystals with diameters of approximately 1–5 µm was observed on Ti-SPT. XRD clearly showed that the precipitated crystals on titanium disks were HA. The contact angle of the polished-Ti increased with time (θ = 37°–51°). The surface of the Ti-SPT remained hydrophilic (θ ˂ 5°) after up to 30 days of aging. The number of attached cells on the Ti-SPT after aging for 30 days remained above 85% of that on the Ti-SPT without aging. SPT in a mineralizing solution can be used to acquire a homogenous precipitation of HA on porous-surfaced titanium implants.

A doxycycline-treated hydroxyapatite implant surface attenuates the progression of peri-implantitis: A radiographic and histological study in mice.

Oral rehabilitation with dental implants has become increasingly common; however, the increase of peri-implantitis is a great concern. Doxycycline (DOX) is a widely used antibiotic that inhibits bacteria growth, inflammation, and bone resorption. To evaluate the progression of peri-implantitis of hydroxyapatite (HA)-coated implants with (5 mg/mL, DOX group) or without (HA group) DOX treatment on the surface. The maxillary first molars of 20 male mice were extracted. Eight weeks later, small titanium screw implants coated with thin HA and treated with or without DOX were placed at the extracted sites. Four weeks after implant placement, half of the animals in both groups were sacrificed, and ligatures were placed around the implant necks in the other half. These mice were sacrificed 4 weeks later. The bone around the implants was examined radiologically and histologically. Four weeks after the ligature placement, the radiographic measurements revealed that peri-implant bone levels of palatal and mesial sites, and histological measurements showed that bone levels of mesial and distal sites in the DOX group were significantly higher than those in the HA group. The present results indicating that the DOX-treated HA implant surface attenuates the progression of peri-implantitis.

Pedicle screws with a thin hydroxyapatite coating for improving fixation at the bone-implant interface in the osteoporotic spine: experimental study in a porcine model.

OBJECTIVE Instrumentation failure caused by the loosening of pedicle screws (PSs) in patients with osteoporosis is a serious problem after spinal surgery. The addition of a thin hydroxyapatite (HA) surface coating applied by using a sputtering process was reported recently to be a promising method for providing bone conduction around an implant without a significant risk of coating-layer breakage. In this study, the authors evaluated the biomechanical and histological features of the bone-implant interface (BII) of PSs with a thin HA coating in an in vivo porcine osteoporotic spine model. METHODS Three types of PSs (untreated/standard [STPS], sandblasted [BLPS], and HA-coated [HAPS] PSs) were implanted into the thoracic and lumbar spine (T9-L6) of 8 mature Clawn miniature pigs (6 ovariectomized [osteoporosis group] and 2 sham-operated [control group] pigs). The spines were harvested from the osteoporosis group at 0, 2, 4, 8, 12, or 24 weeks after PS placement and from the control group at 0 or 24 weeks. Their bone mineral density (BMD) was measured by peripheral quantitative CT. Histological evaluation of the BIIs was conducted by performing bone volume/tissue volume and bone surface/implant surface measurements. The strength of the BII was evaluated with extraction torque testing. RESULTS The BMD decreased significantly in the osteoporosis group (p < 0.01). HAPSs exhibited the greatest mean extraction peak torque at 8 weeks, and HAPSs and BLPSs exhibited significantly greater mean torque than the STPSs at 12 weeks (p < 0.05). The bone surface/implant surface ratio was significantly higher for HAPSs than for STPSs after 2 weeks (p < 0.05), and bonding between bone and the implant surface was maintained until 24 weeks with no detachment of the coating layer. In contrast, the bone volume/tissue volume ratio was significantly higher for HAPSs than for BLPSs or STPSs only at 4 weeks. CONCLUSIONS Using PSs with a thin HA coating applied using a sputtering process strengthens bonding at the BII, which might improve early implant fixation after spinal surgery for osteoporosis. However, the absence of increased bone mass around the screw remains a concern; prescribing osteoporosis treatment to improve bone quality might be necessary to prevent fractures around the screws.

Functionalized coatings by cold spray: An in vitro study of micro- and nanocrystalline hydroxyapatite compared to porous titanium

Three different surface treatments on a Ti6Al4V alloy have been in vitro tested for possible application in cementless joint prosthesis. All of them involve the novelty of using the Cold Spray technology for their deposition: (i) an as-sprayed highly rough titanium and, followed by the deposition of a thin hydroxyapatite layer with (ii) microcrystalline or (iii) nanocrystalline structure. Primary human osteoblasts were extracted from knee and seeded onto the three different surfaces. Cell viability was tested by MTS and LIVE/DEAD assays, cell differentiation by alkaline phosphatase (ALP) quantification and cell morphology by Phalloidin staining. All tests were carried out at 1, 7 and 14 days of cell culture.Different cell morphologies between titanium and hydroxyapatite surfaces were exhibited. At 1 day of cell culture, cells on the titanium coating were spread and flattened, expanding the filopodia actin filaments in all directions, while cells on the hydroxyapatite coatings showed round like-shape morphology due to slower attachment. Higher cell viability was detected at all times of cell culture on titanium coating due to a better attachment at 1 day. However, from 7 days of cell culture, cells on hydroxyapatite showed good attachment onto surfaces and highly increased their proliferation, mostly on nanocrystalline, achieving similar cell viability levels than titanium coatings. ALP levels were significantly higher in titanium, in part, because of greatest cell number. Overall, the best cell functional results were obtained on titanium coatings whereas microcrystalline hydroxyapatite presented the worst cellular parameters. However, results indicate that nanocrystalline hydroxyapatite coatings may achieve promising results for the faster cell proliferation once cells are attached on the surface.

Promotion of bone regeneration on titanium implants through a chemical treatment process using calcium phosphate slurry: Microscopic analysis, cellular response, and animal experiment.

The present study provides scientific evidence that a new chemical treatment process using calcium phosphate slurry promotes bone regeneration on titanium (Ti) implants. The material's surface modified by the treatment was analyzed using microscopic observation and the bone regeneration efficacy was evaluated both in vitro and in vivo. Formation of a thin hydroxyapatite layer with a thickness of about 50 nm and an increase of surface roughness were confirmed by microscopic observations. Histological evaluation of rat femora implanted with the specimens showed that the areas of the specimens directly attached to bone tissue were significantly more extensive than those implanted with control Ti at 2 and 8 weeks. Likewise, on the treated Ti, ALP activity, osteopontin, osteocalcin, and calcium contents of rat bone marrow stromal cells were significantly higher than on the control Ti. Furthermore, reverse transcription polymerase chain reaction showed greater expression of messenger ribonucleic acid encoding Cbfa1 and collagen type1 on the treated Ti at 2 weeks. Based on these results, we concluded that the new process was effective to enhance the osteoconductivity of Ti. © 2018 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 106B: 2716-2724, 2018.

Influence of Heating Conditions for Formation of a Thin Apatite Film on Zirconia Using a Molecular Precursor Method

The influence of heating conditions, heating temperature, and heating time on the formation of a thin carbonate-containing hydroxyapatite (CA) film onto partially stabilized zirconia using a molecular precursor method was evaluated. The molecular precursor solution was prepared from a mixture of calcium-ethylenediaminetetraacetic acid complex and phosphate compounds at Ca/P ratio of 1.67. After the application of molecular precursor solution onto zirconia, four different heating conditions—namely, 600 °C-2 h, 800 °C-2 h, 1000 °C-2 h, and 600 °C-4 h—were applied. No distinct difference of surface appearance of CA coating was observed between 600 and 800 °C-2 h. Fusion of apatite crystals was observed at 1000 °C-2 h. Surface roughness of CA film at 1000 °C-2 h was significantly higher than those under other heating conditions. Heating at 800 °C produced a significantly more hydrophilic surface and higher degree of crystallization. No significant differences were recognized in the critical load at the first crack in the coating among the four samples by scratch tests. After 30 days’ immersion in phosphate buffered saline, the four different CA coating films were still present. Simulated body fluid immersion experiments were performed as in vitro biocompatibility tests. After 48 h immersion, the CA film at 800 °C-2 h showed a greater amount of spherical crystal precipitation. It was suggested that properties of CA coating on partially stabilized zirconia using a molecular precursor method were influenced by the heating temperature and time.

Improvement of the Mechanical Properties of AZ91D Magnesium Alloys by Deposition of Thin Hydroxyapatite Film

Structural and mechanical behavior of thin hydroxyapatite (HA) films deposited via radio-frequency magnetron sputtering on AZ91D magnesium alloy was investigated. Nanoindentationwas employed to evaluate nanohardness and Young’s modulus of the uncoated and HA-coated AZ91 magnesium alloy. The HA-coated AZ91D magnesium alloy exhibited a higher hardness of 7.1 GPa and a higher modulus of 86 GPa compared withthe uncoated substrate revealing a strong load-bearing capacity.

Inflammatory cell response to ultra-thin amorphous and crystalline hydroxyapatite surfaces

It has been suggested that surface modification with a thin hydroxyapatite (HA) coating enhances the osseointegration of titanium implants. However, there is insufficient information about the biological processes involved in the HA-induced response. This study aimed to investigate the inflammatory cell response to titanium implants with either amorphous or crystalline thin HA. Human mononuclear cells were cultured on titanium discs with a machined surface or with a thin, 0.1 μm, amorphous or crystalline HA coating. Cells were cultured for 24 and 96 h, with and without lipopolysaccharide (LPS) stimulation. The surfaces were characterized with respect to chemistry, phase composition, wettability and topography. Biological analyses included the percentage of implant-adherent cells and the secretion of pro-inflammatory cytokine (TNF-α) and growth factors (BMP-2 and TGF-β1). Crystalline HA revealed a smooth surface, whereas the amorphous HA displayed a porous structure, at nano-scale, and a hydrophobic surface. Higher TNF-α secretion and a higher ratio of adherent cells were demonstrated for the amorphous HA compared with the crystalline HA. TGF-β1 secretion was detected in all groups, but without any difference. No BMP-2 secretion was detected in any of the groups. The addition of LPS resulted in a significant increase in TNF-α in all groups, whereas TGF-β1 was not affected. Taken together, the results show that thin HA coatings with similar micro-roughness but a different phase composition, nano-scale roughness and wettability are associated with different monocyte responses. In the absence of strong inflammatory stimuli, crystalline hydroxyapatite elicits a lower inflammatory response compared with amorphous hydroxyapatite.

&lt;i&gt;In Vitro&lt;/i&gt; Assessment of Hydroxyapatite Coating on the Surface of Additive Manufactured Ti6Al4V Scaffolds

Custom orthopedic and dental implants may be fabricated by additive manufacturing (AM), for example using electron beam melting technology. This study is focused on the modification of the surface of Ti6Al4V alloy coin-like scaffolds fabricated via AM technology (EBM®) by radio frequency (RF) magnetron sputter deposition of hydroxyapatite (HA) coating. The scaffolds with HA coating were characterized by Scanning Electron microscopy, X-ray diffraction. HA coating showed a nanocrystalline structure with the crystallites of an average size of 32±9 nm. The ability of the surface to support adhesion and the proliferation of human mesenchymal stem cells was studied using biological short-term tests in vitro. In according to in vitro assessment, thin HA coating stimulated the attachment and proliferation of cells. Human mesenchymal stem cells cultured on the HA-coated scaffold also formed mineralized nodules.

Electrophoretic deposition and characterization of thin hydroxyapatite coatings formed on the surface of NiTi shape memory alloy

The NiTi surface was modified so as to improve its biocompatibility by forming multifunctional protective layers of TiO2 and hydroxyapatite (HAp). A special focus was placed on obtaining relatively thin (~2.4µm), crack-free and homogenous HAp coatings using the electrophoretic deposition method (EPD). A well-dispersed and stable colloidal suspension with the predominant particles size of ca 100nm was used for the deposition process. The coatings were obtained at the following deposition parameters: 40V /120s and after vacuum-sintering at 800°C by 2h providing to an increase of the surface roughness. It was revealed that the composite coatings had adhesion features of (15.1±1.4) MPa and crack resistance to deformation of 2.48%. Moreover, no layer spalling was found, even at the maximum deformation value ε=3.53%. The newly manufactured coatings were also observed to have no impact on the martensitic transformation responsible for shape memory effect. Additionally, a two-step martensitic transformation below room temperature was observed.

Successful Bone Healing of a Critical-Sized and Immediately Loaded Rabbit Mandibular Segmental Defect Using a Thin Hydroxyapatite Coated Titanium Fiber Mesh Scaffolds

Segmental bone defect in the mandible is a most difficult defect to repair in bone reconstructive/regenerative therapy because the mandibular bone constantly moves and is loaded immediately after surgery. Vascularized free bone flap is the most reliable treatment option; however, the procedure involves a long operation time and significant morbidity related to bone harvesting. Coating biomaterials with a thin hydroxyapatite (HA) was proven effective in enhancing bone compatibility. We developed submicron-thin HA-coated titanium fiber mesh scaffolds to reconstruct immediately loaded segmental mandibular defects and evaluated their bone compatibility in vivo.

Formation and Characterization of Crystalline Hydroxyapatite Coating with the (002) Texture

This study reports the effect of titanium (Ti) microstructure on the mechanical properties and surface wettability of thin (<800 nm) hydroxyapatite (HA) coating deposited via radio-frequency (RF) magnetron sputtering. It was revealed that the sand-blasting (SB) and acid etching (AE) of Ti prior deposition led to a wide range of surface roughness in nano/micro scale. After nanostructured HA coating deposition such physico-mechanical characteristics as nanohardness H, Young's modulus E, H/E ratio and H3/E2 were significantly improved. Moreover, HA coatings exhibited improved wear resistance, lower friction coefficient and ability of the coating to wetting.