Biomineralized strontium-substituted apatite/titanium dioxide coating on titanium surfaces

Bone mineral is a multi-substituted calcium phosphate. One of these ion substitutions, strontium, has been proven to increase bone strength and decrease bone resorption. Biomimetics is a potential way to prepare surfaces that provide a favorable bone tissue response, thus enhancing the fixation between bone and implants. Here we prepared double-layered strontium-substituted apatite and titanium dioxide coatings on titanium substrates via mimicking bone mineralization. Morphology, crystallinity, surface chemistry and composition of Sr-substituted coatings formed via biomimetic coating deposition on crystalline titanium oxide substrates were studied as functions of soaking temperature and time in phosphate buffer solutions with different Sr ion concentration. The morphology of the biomimetic apatite changed from plate-like for the pure HA to sphere-like for the Sr ion substituted. Surface analysis results showed that 10–33% of Ca ions in the apatite have been substituted by Sr ions, and that the Sr ions were chemically bonded with apatite and successfully incorporated into the structure of apatite.     

In vitro and in vivo degradation of biomimetic octacalcium phosphate and carbonate apatite coatings on titanium implants

Calcium phosphate (Ca-P) coatings have been applied onto titanium alloys prosthesis to combine the srength of metals with the bioactivity of Ca-P. It has been clearly shown in many publications that Ca-P coating accelerates bone formation around the implant. However, longevity of the Ca-P coating for an optimal bone apposition onto the prosthesis remains controversial. Biomimetic bone-like carbonate apatite (BCA) and Octacalcium Phosphate (OCP) coatings were deposited on Ti6Al4V samples to evaluate their in vitro and in vivo dissolution properties. The coated plates were soaked in alpha-MEM for 1, 2, and 4 weeks, and they were analyzed by Back Scattering Electron Microscopy (BSEM) and by Fourier Transform Infra Red spectroscopy (FTIR). Identical coated plates were implanted subcutaneously in Wistar rats for similar periods. BSEM, FTIR, and histomorphometry were performed on the explants. In vitro and in vivo, a carbonate apatite (CA) formed onto OCP and BCA coatings via a dissolution-precipitation process. In vitro, both coatings dissolved overtime, whereas in vivo BCA calcified and OCP partially dissolved after 1 week. Thereafter, OCP remained stable. This different in vivo behavior can be attributed to (1) different organic compounds that might prevent or enhance Ca-P dissolution, (2) a greater reactivity of OCP due to its large open structure, or (3) different thermodynamic stability between OCP and BCA phases. These structural and compositional differences promote either the progressive loss or calcification of the Ca-P coating and might lead to different osseointegration of coated implants.     

Strontium-substituted apatite coating grown on Ti6Al4V substrate through biomimetic synthesis

During the last few years Strontium has been shown to have beneficial effects when incorporated at certain doses in bone by stimulating bone formation. It is believed that its presence locally at the interface between an implant and bone will enhance osteointegration and therefore, ensure the longevity of a joint prosthesis. In this study we explore the possibility of incorporating Sr into nano-apatite coatings prepared by a solution-derived process according to an established biomimetic methodology for coating titanium based implants. The way this element is incorporated in the apatite structure and its effects on the stereochemistry and morphology of the resulting apatite layers was investigated, as well as its effect in the mineralization kinetics. By using the present methodology it was possible to incorporate increasing amounts of Sr in the apatite layers. Sr was found to incorporate in the apatite layer through a substitution mechanism by replacing Ca in the apatite lattice. The presence of Sr in solution induced an inhibitory effect on mineralization, leading to a decrease in the thickness of the mineral layers. The obtained Sr-substituted biomimetic coatings presented a bone-like structure similar to the one found in the human bone and therefore, are expected to enhance bone formation and osteointegration.     

In vivo evaluation of plasma-sprayed titanium coating after alkali modification

In this paper, plasma-sprayed titanium coatings were modified by alkali treatment. The changes in chemical composition and structure of coatings were examined by SEM and AES. The results obtained indicated that a net-like microscopic texture feature, which was full of the interconnected fine porosity, appeared on the surface of alkali-modified titanium coatings. The surface chemical composition was also altered by alkali modification. A sodium titanate compound was formed on the surface of the titanium coating and replaced the native passivating oxide layer. Its thickness was measured as about 150 nm which was about 10 times of that of the as-sprayed coating. The bone bonding ability of titanium coatings were investigated using a canine model. The histological examination and SEM observation demonstrated that more new bone was formed on the surface of alkali-modified implants and grew more rapidly into the porosity. The alkali-modified implants were found to appose directly to the surrounding bone. In contrast, a gap was observed at the interface between the as-sprayed implants and bone. The push-out test showed that alkali-modified implants had a higher shear strength than as-sprayed implants after 1 month of implantation. An interfacial layer, containing Ti, Ca and P, was found to form at the interface between bone and the alkali-modified implant by EDS analysis.     

A hybrid coating of biomimetic apatite and osteocalcin

A novel hybrid coating of biomimetic apatite(BAp) and osteocalcin (OC) was prepared by incubating BAp-coated Ti6A14V coupons in an osteocalcin-containing medium. A significant amount (up to 1.0 wt %) of OC was adsorbed by the BAp coating within 3 h of incubation as demonstrated by high-performance liquid chromatography. Characterizations of the hybrid coating with environmental scanning microscopy and X-ray diffraction indicated that protein adsorption does not alter the microstructure of the coating. The presence of OC in the hybrid coating was visualized with fluorescence microscopy using an immuno-labeling procedure. The affinity of OC to the BAp coating was examined using a 20-h elution test in phosphate-buffered saline and only a minimal amount (<10%) of the loaded OC was eluted out. When the coating was fully dissolved in hydrochloric acid solution after elution, about 78% of the loaded OC could be recovered. Enzyme-linked immunosorbent assay and peptide sodium dodecylsulfate-polyacrylamide gel electrophoresis confirmed the integrity and activity of OC molecules throughout the tests. The preliminary cell culture tests showed a significant effect of OC on the attachment and proliferation of osteoblasts. The quick loading profile and high affinity of OC to the BAp coating make it an ideal candidate for the hybrid coating preparation in clinical environment.     

Gene delivery via DNA incorporation within a biomimetic apatite coating

Integrating inductivity with conductivity in a material may advance tissue engineering. An organic/inorganic hybrid was developed by incorporating plasmid DNA encoding for the β-gal gene complexed with Lipofectamine 2000^® (DNA–Lipoplex) within apatite via coprecipitation. It was hypothesized that this system will result in enhanced transfection efficiency compared to DNA–Lipoplexes adsorbed to the mineral surface and DNA coprecipitated without Lipofectamine 2000^®. PLGA films were cast onto glass slips and apatite and DNA were coprecipitated in modified simulated body fluid (mSBF). DNA–Lipoplex presence in mineral, DNA–Lipoplex stability (vs. coprecipitation time), and transfection efficiency (determined with C3H10T1/2 cells) as a function of coprecipitation time, DNA–Lipoplex concentration, and DNA incorporation method were studied. DNA–Lipoplex presence and spatial distribution on apatite were confirmed through fluorescence. Transfection efficiency was highest for 6 h of DNA–Lipoplex coprecipitation. Differences in transfection efficiency were found between the DNA concentrations, with the highest efficiency for coprecipitation being 40 μg/ml (p ≤ 0.009 relative to other coprecipitation concentrations). Significant differences in transfection efficiency existed between incorporation methods (p < 0.05) with the highest efficiency for DNA–Lipoplex coprecipitation. This hybrid material system not only integrates inductivity provided by the DNA and conductivity provided by the apatite, but it also has significant implications in non-viral gene delivery due to its ability to increase transfection efficiency.     

Bonelike apatite coatings on plasma-sprayed porous titanium by biomimetic processing

INTRODUCTION　　 Hydroxyapatite(HA) has many biological benefits,such as direct bonding tobone and enhances new bone formation around it.It has been demonstrated thatdental and orthopaedic implants coated with HA show superiorhistological resultstothe uncoated ones.Various methodsaswell as plasma spraying,which is commonlyused,have been developed to coat HA on metals.However,Plasma-sprayed HAcoatings are limited by specific drawbacks such as low crystallinity,weak bondstrength to the s…     

The effect of temperature and initial pH on biomimetic apatite coating

Bone-like apatite coatings were prepared using a biomimetic method in a simulated body fluid (SBF). The effect of initial pH values and immersing temperatures on biomimetic apatite coating formation was studied. Three different temperatures were used in this study: 24 (room temperature), 40, and 60 degrees C. At each temperature, SBF solutions with three different initial pHs were chosen: low, medium, and high. The total inorganic carbon (TIC) content and pH-time profile of each coating system were recorded during the coating formation. The apatite coatings were characterized using X-ray diffraction (XRD), field emission scanning electron microscope (FESEM), and Fourier transform infra-red (FTIR). It has been found that SBF temperature has a great effect on the bicarbonate decomposition rate. The bicarbonate ions tend to decompose faster as the temperature increases. The decomposition of bicarbonate ions results in a pH increase in the SBF. With different initial SBF pHs, the decomposition of different amounts of bicarbonate ions is required to reach the critical pH range of apatite formation. With different amounts of bicarbonate ions in the SBF, the surface morphology of the biomimetic apatite coating formed is different. Therefore, the initial pH of the SBF solution plays a vital role in controlling the surface morphology of the biomimetic apatite coating. Also, it was found that as the SBF temperature increased, the critical pH range at which biomimetic apatite coating forms decreased. The critical pH range for the SBF prepared at 24, 40, and 60 degrees C was 6.65-6.71, 6.55-6.65, and 6.24-6.42, respectively.     

In vivo evaluation of defined polished titanium surfaces to prevent soft tissue adhesion

Soft tissue-implant adhesion is often required for implant integration into the body; however, in some situations, the tissue is required to glide freely over an implant. In the case of distal radius fracture treatment, current literature describes how titanium and its alloys tend to lead to more intra-tendon inflammatory reactions compared with stainless steel. This leads to tendon-implant adhesion and damage possibly causing limited palmar flexion and even tendon rupture. The goal of this study was to analyze the effect of different surface polishings of titanium and titanium molybdenum implants on soft tissue reactions in vivo, with the aim to prevent direct soft tissue adhesion. Using a nonfracture model, to allow for study of the soft-tissue-implant surface interactions only, six surface variants of the same plate design were implanted onto the tibia of 24 New Zealand white rabbits and left in situ for 12 weeks. Results indicate that paste polished commercially pure titanium and titanium molybdenum alloy had the least soft tissue adhesion, with the concomitant development of a soft tissue capsule. Surface topography did not appear influence the thickness of the connective tissue surrounding the plate. Therefore, suitable surface polishing could be applied to plates for clinical use, where free gliding of tissues is required.     

Incorporation of tobramycin into biomimetic hydroxyapatite coating on titanium

Calcium phosphate coatings containing an antibiotic were produced on titanium alloy (Ti6Al4V) implants using a biomimetic approach. Thin, amorphous calcium phosphate (ACP) coatings were first deposited onto Ti6Al4V plates by immersion in 5 times concentrated simulated body fluid (SBF), for 24h at 37 degrees C. The ACP-coated implants were then immersed in a supersaturated calcium phosphate (SCP) solution containing 0, 100, 200, 400, 600 or 800 mg/l of tobramycin for 48 h at 37 degrees C. A carbonated hydroxyapatite (CHA) layer, approximately 40 microm thick, was formed. Approximately 3 microg/mg of tobramycin was co-precipitated with the CHA crystals onto titanium alloy plates, using 800mg/l tobramycin in the coating solution. For comparison, plasma-sprayed calcium phosphate coatings were also immersed in solutions containing 100, 200, 400 or 1,000 mg/l of tobramycin for 10, 40 min, or 48 h. A maximum of about 0.3 microg/mg could be adsorbed onto the plasma-sprayed calcium phosphate coating with the comparable concentration of 800 mg/l in solution. The dissolution of coating and release of tobramycin were also measured in vitro using saline solution buffered at pH 5.0 or 7.3 at 37 degrees C. The release rate of tobramycin was faster at pH 7.3 than at pH 5, with 50 and 4 microg/ml/min, respectively. Tobramycin released from the biomimetic-coated plates could inhibit growth of Staphylococcus aureus bacteria. The result of this study, therefore, indicates that the biomimetic CHA coatings containing antibiotics could be used to prevent post-surgical infections in orthopaedic or trauma.     

Improvement of bonding strength between biomimetic apatite coating and substrate

Bone-like apatite coatings were prepared using a biomimetic method in a modified simulated body fluid (m-SBF). The effect of the m-SBF volume on the apatite coating quality was studied. Three m-SBF volumes, 50, 100, and 200 mL, were employed to immerse titanium substrates in a sealed container so as to produce apatite coatings with different properties, namely types I, type II, and type III apatite coatings, respectively. The coatings were characterized using X-ray diffraction and environmental scanning electron microscope. The bonding between the coating and the Ti substrate was evaluated using an adhesive strength test. All three apatite coatings demonstrated a poorly crystallized structure, and the coatings formed exhibited a uniformed surface morphology. Further increasing the m-SBF volume, small globules of apatite started to form on the surface of the coating. The bonding strength for the three coating systems were 8.52 +/- 2.41, 10.36 +/- 2.78, and 17.23 +/- 2.55 MPa for types I, II, and III apatite coatings, respectively. The failure analyses suggested that type III coating failed mostly at the interface between the coating and the substrate, while type I and II coatings failed mostly within the apatite coating. Our study revealed that a dense, thick, well-adhered apatite coating could be achieved by carefully controlling the volume of m-SBF.     

Adherent apatite coating on titanium substrate using chemical deposition

Plasma-sprayed "HA" coatings on commercial orthopedic and dental implants consist of mixtures of calcium phosphate phases, predominantly a crystalline calcium phosphate phase, hydroxyapatite (HA) and an amorphous calcium phosphate (ACP) with varying HA/ACP ratios. Alternatives to the plasma-spray method are being explored because of some of its disadvantages. The purpose of this study was to deposit an adherent apatite coating on titanium substrate using a two-step method. First, titanium substrates were immersed in acidic solution of calcium phosphate resulting in the deposition of a monetite (CaHPO4) coating. Second, the monetite crystals were transformed to apatite by hydrolysis in NaOH solution. Composition and morphology of the initial and final coatings were identified using X-ray diffraction (XRD), Scanning Electron Microscopy, and Energy Dispersive Spectroscopy (EDS). The final coating was porous and the apatite crystals were agglomerated and followed the outline of the large monetite crystals. EDS revealed the presence of calcium and phosphorous elements on the titanium substrate after removing the coating using tensile or scratching tests. The average tensile bond of the coating was 5.2 MPa and cohesion failures were observed more frequently than adhesion failures. The coating adhesion measured using scratch test with a 200-microm-radius stylus was 13.1N. Images from the scratch tracks demonstrated that the coating materials were squashed without fracturing inside and/or at the border of the tracks until the failure point of the coating. In conclusion, this study showed the potential of a chemical deposition method for depositing a coating consisting of either monetite or apatite. This method has the advantage of producing a coating with homogenous composition on even implants of complex geometry or porosity. This method involves low temperatures and, therefore, can allow the incorporation of growth factors or biogenic molecules.     

In vivo evaluation of plasma-sprayed wollastonite coating

Wollastonite coatings were prepared by plasma spraying. The bioactivity of wollastonite coatings was investigated in vivo by implanting in dog's muscle, cortical bone and marrow, respectively. The behaviour of bone tissue around wollastonite coatings were examined by histological and SEM observation. After 1 month in the muscle, a bone-like apatite layer was found to form on the surface of the wollastonite coating. When implanted in cortical bone, histological observation demonstrated that bone tissue could extend and grow along the surface of the wollastonite coating. The coating bonded directly to the bone without any fibrous tissue, indicating good biocompatibility and bone conductivity. SEM and EDS analysis revealed that bone did not bond to wollastonite coating directly, but through a Ca/P layer. This suggested that the formation of bone-like apatite layer was very important for bonding to the bone tissue. The amount of bone-implant contact was also measured. Wollastonite coating was shown to stimulate more bone formation on its surface than titanium coating after implantation for 1 month, enhancing the short-term osseointegration properties of implant. The test in marrow indicated that wollastonite coatings could induce new bone formation on their surface showing good bone inductivity property.     

Carbonate apatite coating on titanium induced rapidly by precalcification.

Chemical treatments have been thought to be promised methods for improving bioactivity of titanium. In this work, the effect of precalcification with boiling saturated Ca(OH)2 solution on bioactivation of titanium was investigated. After precalcification and soaking in supersaturated Ca-P solution (SCP), calcium phosphate rapidly precipitated onto the surfaces of titanium, and after only three days an uniform apatite layer was found up to thickness of a few micrometers. The observation using scanning electron microscopy (SEM) showed that the coating was composed of a number of small crystal grains. The investigation by X-ray energy dispersion spectroscopy (EDS), Fourier transform infrared spectroscopy (FTIR) and X-ray diffraction (XRD) indicated that the coating was Ca-deficient carbonate apatite. Based on the analyses for the surfaces and SCP, a mechanism of precipitation of apatite was proposed in thermal dynamics and kinetics.     

Electrochemical activation of titanium for biomimetic coating of calcium phosphate

This article reports an electrochemical method to activate titanium surface for biomimetic calcium phosphate (Ca-P) coatings. Titanium serving as cathode was treated in an electrochemical cell with a supersaturated calcium and phosphate solution serving as electrolyte. This treatment generated a gel-like film with thickness of about 100 nm on the titanium surface. The amorphous film was composed by calcium and phosphate ions and contained a large number of crystal nuclei of octacalcium phosphate (OCP). The effectiveness of this novel treatment was demonstrated by comparing the behavior of treated and untreated titanium when used for biomimetic coating. A uniform Ca-P coating was formed on the treated titanium after immersion for several hours in aqueous solution. This work explored a new method to activate surfaces of metal implants for osseointegration, which is considerably faster than treatments currently in use, such as alkaline treatment.     

In vitro bone formation on a bone-like apatite layer prepared by a biomimetic process on a bioactive glass-ceramic

In this study we have investigated the behavior of fetal rat osteoblasts, cultured up to 23 days, on a bioactive apatite-wollastonite (AW) glass-ceramic and on the same material on which a carbonated apatite layer had been formed by a biomimetic process (AWa). At the last day of culture, the specific activity of alkaline phosphatase activity, as determined biochemically, was about 30% greater on AWa compared with AW disks. After the cell layers had been scraped off, scanning electron microscopic (SEM) observations of the materials' surfaces revealed that mineralized bone nodules remained attached to both surfaces but in larger amounts on AWa. X-ray microanalysis indicated the presence of calcium (Ca) and phosphorus (P) in the bone tissue throughout the AWa surface and Ca, P, and silicon (Si) on the AW surface. The AW/ and AWa/bone interfaces also were analyzed after fracturing of the disks. The interfacial analysis showed firm bone bonding to the AW and AWa surfaces, confirmed by the X-ray microanalytic mappings. These results indicate the importance of surface composition in supporting differentiation of osteogenic cells and the subsequent apposition of bone matrix, which allows a strong bond of the bioactive materials to the bone. Furthermore, prefabrication of a biologic apatite layer by a method that mimics biomineralization could find application to bone-repairing materials.     

Porous titanium and silicon-substituted hydroxyapatite biomodification prepared by a biomimetic process: Characterization and in vivo evaluation

Porous titanium with a pore size of 150–600 μm and a porosity of 67% was prepared by fiber sintering. The porous titanium had a complete three-dimensional (3D) interconnected structure and a high yield strength of 100 MPa. Si-substituted hydroxyapatite (Si-HA) was coated on the surface by a biomimetic process to improve the surface bioactivity. X-ray diffraction results showed that Si-HA coating was not well crystallized. New bone tissue was found in the uncoated porous titanium after 2 weeks of implantation and a significant increase (p < 0.05) in the bone ingrowth rate (BIR) was found after 4 weeks of implantation, indicating the good osteoconductivity of the porous structure. The HA-coated and Si-HA-coated porous titanium exhibited a significantly higher BIR than the uncoated titanium at all intervals, highlighting the better surface bioactivity and osteoconductivity of the HA- and Si-HA coatings. Also, the Si-HA-coated porous titanium demonstrated a significantly higher BIR than the HA-coated porous titanium, showing that silicon plays an active role in the surface bioactivity. For Si-HA-coated porous titanium, up to 90% pore area was covered by new bone tissue after 4 weeks of implantation in cortical bone. In the bone marrow cavity, the pore spaces were filled with bone marrow, displaying that the interconnected pore structure could provide a channel for body fluid. It was concluded that both the 3D interconnected pore structure and the Si-HA coating contributed to the high BIR.     

Interaction of calcium and phosphate in apatite coating on titanium with serum albumin

A Ca-deficient carbonate apatite coating on titanium was prepared by pre-calcifying titanium in a saturated Ca(OH)2 solution and then immersing in a supersaturated calcium phosphate solution. The interaction of the protein with the apatite coating on titanium was investigated by scanning electron microscopy with X-ray energy dispersion spectroscopy. X-ray photoelectron spectroscopy, X-ray diffraction and Fourier transform infrared spectroscopy. During immersion of the coating in bovine serum albumin (BSA) solution, accompanied by an adsorption of BSA onto the coating, calcium and phosphate ions dissolved and reprecipitated, resulting in the formation of the coating containing BSA from the surface to subsurface layers. The adsorption modified the structure and morphology of the apatite coating on titanium and changed the protein configuration. It was also found that the protein chemically adsorbed onto surfaces containing calcium or phosphorus, showed that both Ca and P on the apatite coating were the binding sites with protein. The BSA adsorption onto the coating involved several elements and groups. In this process. Ca played an essential role, and the interaction of Ca on the apatite coating with the protein stimulated the bond of the protein at P sites.     

氟涂层镁铝合金动物体内生物相容性实验

目的研究氟涂层镁铝合金动物体内生物相容性。方法普通级新西兰大白兔14只,雄性,4月龄,体质量约2.0~2.5 kg。同一品系健康、SPF级昆明小白鼠15只,雌雄不限,6周龄,雌鼠无孕,体质量17~23 g。应用镁合金材料在小白鼠体内进行急性全身毒性实验,分3组(n=5)。新西兰大白兔股骨髁外侧植入氟涂层或无涂层镁铝合金圆销,分2组。术后观察兔子一般情况,术后当天、12周采用Micro CT检查。术前、术后抽兔血进行镁离子浓度检测。数据应用SPSS 18.0进行统计学分析。结果急性全身毒性实验显示,各组动物体质量变化百分比未超过10%。术后所有兔子一般情况良好,创口愈合良好,均无皮下气肿。植入12周后,镁铝合金组圆销与骨组织间出现轻微空白影;两组圆销周围均未出现骨质吸收溶解情况。氟涂层组植入物边缘略不规则,与周围骨质间存在细小空隙。植入镁铝合金后,各时间段血清镁离子浓度差异无统计学意义(F=1.695,P=0.136)。结论氟涂层镁铝合金体内无毒性反应。     

Differentiation of human bone-derived cells grown on GRGDSP-peptide bound titanium surfaces

Various surface modifications have been applied to titanium alloy (Ti-6Al-4V) implants, in an attempt to enhance osseointegration; crucial for ideal prosthetic fixation. Despite the numerous studies demonstrating that peptide-modified surfaces influence in vitro cellular behavior, there is relatively little data reporting their effects on bone remodeling. The objective of this article was to examine the effects of chemically modifying Ti-6Al-4V surfaces with a common RGD sequence, a 15-residue peptide containing GRGDSP (glycine-arginine-glycine-aspartate-serine-proline), on the modulation of bone remodeling. The expression of proteins known to be associated with osseous matrix and bone resorption were studied during the growth of human bone-derived cells (HBDC) on these peptide-modified surfaces. HBDC grown for 7 days on RGD surfaces displayed significantly increased levels of osteocalcin, and pro-collagen Ialpha1 mRNAs, compared with the production by HBDC grown on the native Ti-6Al-4V. A pattern that was also reflected at the protein levels for osteocalcin, type I collagen, and bone sialoprotein. Moreover, HBDC grown for 7 and 14 days on RGD-modified Ti-6Al-4V expressed significantly higher level of osteoclast differentiation factors and lower levels of osteoprotegerin and IL-6 proteins compared with other surfaces tested. These results suggest that different chemical treatments of implant material (Ti-6Al-4V) surface result in differential bone responses, not only their ability to form bone but also to stimulate osteoclastic formation.