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.     

Characterization of titanium surfaces with calcium and phosphate and osteoblast adhesion

The titanium surfaces containing calcium, phosphate ions and the carbonate apatite were characterized. The effect of surface chemistry on the initial rabbit osteoblast response on these surfaces was investigated. The cell count and alkaline phosphatase (ALP) specific activity assay were used for biochemical analyses. Scanning electron microscopy was used for morphology observation and in particular X-ray photoelectron spectroscopy (XPS) for surface chemistry characterization. The number of cells adhering to the apatite coating surface was the maximum, the number of cells on the surface containing calcium without phosphate ions was higher than that containing phosphate without calcium, and the number on the unmodified titanium surface was the least. The osteoblasts cultured on the apatite surface exhibited the highest ALP specific activity, next were the ones on the surface containing solely calcium, the lowest were on the unmodified titanium surface. On the substrate surfaces removed of adhered cells, the order of nitrogen amounts detected by XPS was consistent with ones of ALP specific activity and cell number, except for the unmodified titanium surface. For the substrate surfaces removed of adhered osteoblasts, XPS analysis showed that calcium and phosphorous amounts decreased during cell adhesion. After cell culture the Ca2p binding energy (BE) values for apatite coating and the surface containing solely calcium were similar to those of the two surfaces adsorbed bovine serum albumin (BSA). The P2p BE values for the surfaces containing phosphate ions, including the apatite coating and the surface containing solely phosphate ions, showed the same change. But after cell culture the decrease of the P2p BE value for the coating surface was larger than the one for the surface containing solely phosphate ions. Considering the bovine serum albumin adsorption on the same samples, these results indicated that calcium ions on titanium surfaces play a more important role than phosphate ions in initial interactions among culture medium, osteoblasts and titanium surfaces. On the apatite coating surface, calcium ions are active sites for osteoblast adhesion, while calcium and phosphate ions co-exist on titanium surfaces, the former promotes the osteoblast adhesion onto the phosphate sites on titanium surfaces. The cell adhesion was a complicated biological and chemical process relating to surface several elements similar to protein adsorption.     

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.     

Calcium phosphate-based coatings on titanium and its alloys

Use of titanium as biomaterial is possible because of its very favorable biocompatibility with living tissue. Titanium implants having calcium phosphate coatings on their surface show good fixation to the bone. This review covers briefly the requirements of typical biomaterials and narrowly focuses on the works on titanium. Calcium phosphate ceramics for use in implants are introduced and various methods of producing calcium phosphate coating on titanium substrates are elaborated. Advantages and disadvantages of each type of coating from the view point of process simplicity, cost-effectiveness, stability of the coatings, coating integration with the bone, cell behavior, and so forth are highlighted. Taking into account all these factors, the efficient method(s) of producing these coatings are indicated finally.     

In vitro investigation of titanium and hydroxyapatite dental implant surfaces using a rat bone marrow stromal cell culture system

In this study, rat bone marrow cells (RBM) were used to evaluate different titanium and hydroxyapatite dental implant surfaces. The implant surfaces investigated were: a titanium surface having a porous titanium plasma-sprayed coating (sample code Ti-TPS), a titanium surface with a deep profile structure (sample code Ti-DPS), an uncoated titanium substrate with a machined surface (sample code Ti-ma) and a machined titanium substrate with a porous hydroxyapatite plasma-sprayed coating (sample code Ti-HA). RBM cells were cultured on the disc-shaped test substrates for 14 days. The culture medium was changed daily and examined for calcium and phosphate concentrations. After 14 days specimens were examined by light microscopy, scanning electron microscopy, energy dispersive X-ray analysis and morphometry of the cell-covered substrate surface. All test substrates facilitated RBM growth of extracellular matrix formation. Ti-DPS and Ti-TPS to the highest degree, followed by Ti-ma and Ti-HA. Ti-DPS and Ti-TPS displayed the highest cell density and thus seem to be well suited for the endosseous portion of dental implants. RBM cells cultured on Ti-HA showed a delayed growth pattern. This may be related to its high phosphate ion release.     

In vivo protein adsorption on polymers: visualization of adsorbed proteins on vascular implants in dogs

The absorption of plasma proteins is an important event at the blood-material interface, and strongly affects subsequent cellular interaction and thrombus formation. Although considerable efforts have been expended to elucidate the mechanism of protein adsorption and the role of absorbed protein layer at the blood-material interface, there has been little knowledge of how the initial adsorbed proteins are maintained or changed in a time-variant process in in vivo long-term implantation. In this study, we described detailed analyses concerning the characterization of adsorbed proteins on HEMA-styrene block copolymer surfaces (HEMA-st) and poly(ethylene oxide)(PEO) grafted Biomer® (B-PE04K) for in vivo long-term canine vascular graft implants as well as in vitro short-term experiments. Biomer vascular grafts (6 mm I.D., 7 cm in length) were fabricated by a dip coating and the luminal surface was modified with PEO grafting, HEMA-st coating, or Biomer coating (control). These surface modified grafts were recirculated for different time intervals (5, 15, 30, 60 and 120 min) using citrated canine whole blood. The grafts were then implanted in the abdominal aortas of dogs and evaluated for graft patency and protein adsorption. The adsorbed proteins (albumin, IgG and fibrinogen) were quantified using an in situ radioimmunoassay. Surface protein layer thickness was measured by transmission electron microscopy (TEM). Visualization of absorbed plasma proteins (albumin, IgG and fibrinogen) was performed with TEM using an immunoperoxidase double antibody technique. In in vitro recirculation systems, albumin and IgG showed similar Langmuir type pattern onto all test surfaces. On B-PEO4K surfaces, fibrinogen adsorption kinetics demonstrated 'Vroman effect'. The Biomer and B-PE04K grafts occluded within 1 month, while HEMA-st grafts were patent for over 3 months. Biomer and B-PE04K showed thick multilayers of adsorbed proteins, and the thickness increased with implantation periods and the composition altered with time. In contrast, HEMA-st showed a monolayer-like adsorbed protein pattern, and the composition and thickness were consistent regardless of implantation time including in vitro short-time experiments, which may attribute to less conformational change of adsorbed proteins on HEMA-st surfaces. In terms of nonthrombogenicity, the stable monolayer-like adsorbed protein layer on HEMA-st surfaces exhibited improved blood compatibility over thick multilayered adsorbed proteins on Biomer and B-PE04K surfaces.     

In vivo protein adsorption on polymers: visualization of adsorbed proteins on vascular implants in dogs.

The absorption of plasma proteins is an important event at the blood-material interface, and strongly affects subsequent cellular interaction and thrombus formation. Although considerable efforts have been expended to elucidate the mechanism of protein adsorption and the role of absorbed protein layer at the blood-material interface, there has been little knowledge of how the initial adsorbed proteins are maintained or changed in a time-variant process in in vivo long-term implantation. In this study, we described detailed analyses concerning the characterization of adsorbed proteins on HEMA--styrene block copolymer surfaces (HEMA-st) and poly(ethylene oxide) (PEO) grafted Biomer (B-PEO4K) for in vivo long-term canine vascular graft implants as well as in vitro short-term experiments. Biomer vascular grafts (6 mm I.D., 7 cm in length) were fabricated by a dip coating and the luminal surface was modified with PEO grafting, HEMA-st coating, or Biomer coating (control). These surface modified grafts were recirculated for different time intervals (5, 15, 30, 60 and 120 min) using citrated canine whole blood. The grafts were then implanted in the abdominal aortas of dogs and evaluated for graft patency and protein adsorption. The adsorbed proteins (albumin, IgG and fibrinogen) were quantified using an in situ radioimmunoassay. Surface protein layer thickness was measured by transmission electron microscopy (TEM). Visualization of absorbed plasma proteins (albumin, IgG and fibrinogen) was performed with TEM using an immunoperoxidase double antibody technique. In in vitro recirculation systems, albumin and IgG showed similar Langmuir type pattern onto all test surfaces. On B-PEO4K surfaces, fibrinogen adsorption kinetics demonstrated 'Vroman effect'. The Biomer and B-PEO4K grafts occluded within 1 month, while HEMA-st grafts were patent for over 3 months. Biomer and B-PEO4K showed thick multilayers of adsorbed proteins, and the thickness increased with implantation periods and the composition altered with time. In contrast, HEMA-st showed a monolayer-like adsorbed protein pattern, and the composition and thickness were consistent regardless of implantation time including in vitro short-time experiments, which may attribute to less conformational change of adsorbed proteins on HEMA-st surfaces. In terms of nonthrombogenicity, the stable monolayer-like adsorbed protein layer on HEMA-st surfaces exhibited improved blood compatibility over thick multilayered adsorbed proteins on Biomer and B-PEO4K surfaces.     

Individual plasma proteins detected on rough biomaterials by phase imaging AFM

In the past several years, atomic force microscopy (AFM) has provided topographic images of adsorbed plasma proteins in situ at unprecedented resolution. Imaging has been limited to adsorbed protein on relatively smooth model substrates such as mica, graphite, or self-assembled monolayers on which the small height of the protein can be observed from the background. The inherent roughness of biomaterial surfaces has prevented observation of adsorbed proteins in topographic images. We report imaging isolated fibrinogen molecules adsorbed on National Heart Lung and Blood Institute (NHLBI) reference materials polydimethylsiloxane and low-density polyethylene in situ using phase imaging AFM. Fibrinogen, a plasma protein important for blood coagulation and platelet aggregation, was adsorbed from dilute solution onto reference biomaterial surfaces at sub-monolayer coverage. Tapping mode AFM was used to image the samples. For polydimethylsiloxane, the lateral size of the surface features is much greater than the dimensions of proteins. This allowed adsorbed proteins to be observed in topographic images. The phase imaging signal of tapping mode AFM provides information on differences in material properties of the surface, and was used to distinguish individual protein molecules from the underlying polymer surface. On the low-density polyethylene surface, characteristic topographical features are of the same magnitude as the protein molecules, so that protein cannot be distinguished from the surface using topographic images. However, phase images were used to successfully locate and characterize the distribution of the protein. Phase imaging was not able to distinguish fibrinogen adsorbed onto expanded polytetrafluoroethylene. The utility and limitations of the phase imaging technique for characterizing protein adsorption to rough surfaces is discussed.     

Synthèse et greffage de polymères bioactifs sur des surfaces en titane pour favoriser l’ostéointégration

Titanium is widely used in orthopedic and dental implants for its excellent resistance to corrosion and its biocompatibility. In order to improve the long-term osteointegration of titanium, bioactive polymers bearing ionics groups such as sulfonates (sodium polysytrene sulfonate, polyNaSS) are grafted by a covalent way onto titanium surface. The surface is chemically modified and then bioactive polymers are grafted by radical polymerization. The chemical composition of grafted surfaces is given by ATR/FTIR and XPS which certified the presence of sulfonate groups at the surface of grafted titanium. Quantitative grafting of polyNaSS is determined by a colorimetric method and evaluated at 5 μg/cm².

In vitro study is performed in order to see the effect of these bioactive polymers on the mineralization of human osteoblast (line MG63). After 28 days of cultured cells on grafted titanium surfaces and non-grafted ones, the amount of calcium onto surfaces is quantified. The results show that the mineralization of these cells is improved with the presence of polyNaSS. The amount of calcium is increased on grafted surfaces compared to non-grafted ones. Cell adhesion was evaluated. Cells were seeded onto grafted and non-grafted titanium and then subjected to detachment forces. The results show that the attachment of human osteoblasts-like cells is increased for grafted titanium with polyNaSS. A study on titanium surface grafted by polymers bearing ionics groups such as carboxylate and phosphate is in progress.     

Vitronectin adsorption on surfaces visualized by tapping mode atomic force microscopy

Vitronectin (Vn) is an abundant glycoprotein present in plasma and the extracellular matrix of most tissue and is an important medium required for bone cells to attach and spread on biomaterial surfaces. Hence, Vn adsorption is an initial and key step in implantation. In this study, adsorption of Vn molecules on mica and oxidized titanium substrates in Milli-Q water (pH 5.6) and in simulated body fluid (SBF, pH 7.4) has been investigated using tapping mode atomic force microscopy. Conformation of the adsorbed proteins was determined from the images. The effect of adsorption time was investigated on mica surfaces. The protein first attaches to the surface, then immediately spreads out with different lateral dimensions of adsorbed protein and becomes aggregated. After 15 min of adsorption, globular Vn molecules form clusters of aggregates in extended chains on the mica surface. Globular Vn molecules appear flatter (i.e., oblate ellipsoids) and coalesce on the ridges of the uneven oxidized titanium surface. From cross-sectional analyses of the Vn images, it was found that the contact areas of Vn molecules associated with Ti surfaces are larger than those with mica surfaces. This suggests that the different surface properties of substrates contribute to this different conformation of adsorbed Vn molecules. The larger contact areas of Vn associated with Ti substrates indicates higher affinity for the biomaterial oxidized titanium surface than for mica. In SBF, Vn interacted weakly with the mica surfaces and Vn molecules were easily desorbed during the imaging process. On oxidized titanium substrates, after 3 h of incubation time in Vn-SBF solution, images can be obtained showing adsorbed Vn aggregates on the oxidized titanium surface. In this case, the protein colloids are strongly adherent, with increased lateral dimensions compared with the adsorption on mica.     

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.     

Local induction of calcium phosphate formation on TiO2 coatings on titanium via surface treatment with a CO2 laser

Sol-gel-derived TiO(2) coatings are known to promote bonelike hydroxyapatite formation on their surfaces in vitro and in vivo. Hydroxyapatite integrates into bone tissue. In some clinical applications, the surface of an implant is simultaneously interfaced with soft and hard tissues, so it should match the properties of both. A new method is introduced for treating the coatings locally in a controlled manner. The local densification of sol-gel-derived titania coatings on titanium substrates with a CO(2) laser was studied in terms of the in vitro calcium phosphate-inducting properties. CO(2)-laser-treated multilayer coatings were compared with furnace-fired coatings prepared with the same recipe and previously shown to be bioactive. Additionally, local areas of furnace-fired multilayer coatings (previously shown to be bioactive in vitro) were further laser-treated to achieve various properties in the same implant. Topological surface properties were examined with atomic force microscopy. The formation of hydroxyapatite was studied with Fourier transform infrared and scanning electron microscopy energy-dispersive X-ray analysis. The results show that calcium phosphate formation can be adjusted locally by laser treatment. Calcium phosphate is a bonelike hydroxyapatite. The local treatment of sol-gel-derived coatings with a CO(2) laser is a promising technique for creating implants with various properties to interface different tissues and a possible way of coating implants that do not tolerate furnace firing.     

Mechanisms of fibrinogen domains : biomaterial interactions

Spontaneous adsorption of fibrinogen is critical to the pathogenesis of biomaterial-mediated inflammatory responses. However, the mechanism by which adsorbed fibrinogen affects phagocyte responses is still not clear. To investigate the molecular interaction between fibrinogen and biomaterials, fibrinogen fragments (D 100 and E50) were generated and used in the present study. The results indicate that biomaterial : D100 interaction is essential to fibrinogen-mediated inflammatory responses, because biomaterials precoated with D100, but not E50, prompt strong inflammatory responses. Furthermore, the results from in vitro studies show that whole molecule fibrinogen and D100 exhibit very similar protein:surface interactions. Specifically: (1) both D100 and fibrinogen have high affinity for biomaterial surfaces; and (2) the retention rates of adsorbed D 100 in both in vivo and in vitro environments are as high as that for adsorbed fibrinogen. On the other hand, E50 does bind to biomaterials but with low affinity because, once bound, it is not tightly adherent to the biomaterial surfaces. Taken together, the results suggest that the mechanism of fibrinogen-mediated inflammatory responses may involve the following three consecutive events: (1) after contact with blood or tissue fluid, the D domain tends to interact with biomaterial surfaces and is important in the tight binding of fibrinogen to implant surfaces; (2) the biomaterial surface then promotes conformational changes within the D domain, exposing P1 epitope (y 190-202, which interacts with phagocyte Mac-1 integrin); and (3) the engagement of Mac-1 integrin with P1 epitope then triggers subsequent phagocyte adherence and reactions.     

Development and characterization of rhVEGF-loaded poly(HEMA–MOEP) coatings electrosynthesized on titanium to enhance bone mineralization and angiogenesis

Osteointegration of titanium implants could be significantly improved by coatings capable of promoting both mineralization and angiogenesis. In the present study, a copolymeric hydrogel coating, poly-2-hydroxyethyl methacrylate-2-methacryloyloxyethyl phosphate (P(HEMA–MOEP)), devised to enhance calcification in body fluids and to entrap and release growth factors, was electrosynthesized for the first time on titanium substrates and compared to poly-2-hydroxyethyl methacrylate (PHEMA), used as a blank reference. Polymers exhibiting negatively charged groups, such as P(HEMA–MOEP), help to enhance implant calcification. The electrosynthesized coatings were characterized by X-ray photoelectron spectroscopy and atomic force microscopy. MG-63 human osteoblast-like cell behaviour on the coated specimens was investigated by scanning electron microscopy, MTT viability test and osteocalcin mRNA detection. The ability of negatively charged phosphate groups to promote hydroxyapatite-like calcium phosphate deposition on the implants was explored by immersing them in simulated body fluid. Similar biological responses were observed in both coated specimens, while calcium-phosphorus globules were detected only on P(HEMA–MOEP) surfaces pretreated with alkaline solution. Testing of the ability of P(HEMA–MOEP) hydrogels to entrap and release human recombinant vascular endothelial growth factor, to tackle the problem of insufficient oxygen and nutrient delivery, suggested that P(HEMA–MOEP)-coated titanium prostheses could represent a multifunctional material suitable for bone restoration applications.     

Nano-scale study of the nucleation and growth of calcium phosphate coating on titanium implants

The nucleation and growth of a calcium phosphate (Ca-P) coating deposited on titanium implants from simulated body fluid was investigated by using atomic force microscopy (AFM) and environmental scanning electron microscopy (ESEM). Forty titanium alloy plates were assigned into two groups. One group with a smooth surface having a maximum roughness R(max) < 0.10 microm (s-Ti6Al4V) and a group with a rough surface with an R(max) < 0.25 microm (r-Ti6Al4V) were used. Titanium samples were immersed in SBF concentrated by five (SBF x 5) from 10 min to 5 h and examined by AFM and ESEM. Scattered Ca-P deposits of approximately 15 nm in diameter appeared after only 10 min of immersion in SBF x 5. These Ca-P deposits grew up to 60-100 nm after 4 h on both s- and r-Ti6Al4V substrates. With increasing immersion time, the packing of Ca-P deposits with size of tens of nanometers in diameter formed larger globules and then a continuous Ca-P film on titanium substrates. A direct contact between the Ca-P coating and the Ti6Al4V surface was observed. The Ca-P coating was composed of nanosized deposits and of an interfacial glassy matrix. This interfacial glassy matrix might ensure the adhesion between the Ca-P coating and the Ti6Al4V substrate. In the case of s-Ti6Al4V substrate, failures within this interfacial glassy matrix were observed overtime. Part of the glassy matrix remained on s-Ti6Al4V while part detached with the Ca-P film. The Ca-P coating detached from the smooth substrate, whereas the Ca-P film extended onto the whole rough titanium surface over time. In the case of r-Ti6Al4V, the Ca-P coating covered evenly the substrate after immersion in SBF x 5 for 5 h. The present study suggested that the heterogeneous nucleation of Ca-P on titanium was immediate and did not depend on the Ti6Al4V surface topography. The further growth and mechanical attachment of the final Ca-P coating strongly depended on the surface, for which a rough topography was beneficial.     

Interactions between calcium, phosphate, and albumin on the surface of titanium.

The deposition of calcium phosphate on chemically polished commercially pure titanium immersed in Hank's balanced salt solution (HBSS) with bovine serum albumin (BSA) (concentrations 0 and 4 mg/mL) has been investigated. Electrochemical techniques, 125I labeling of albumin, scanning electron microscopy, energy dispersive spectroscopy, X-ray diffraction, and X-ray photoelectron spectroscopy were used. A tricalcium phosphate layer with a thickness of ca. 1 microm was formed for periods of immersion in HBSS ranging between 1 and 2 weeks. A concentration of 4 mg/mL of BSA prevented its formation, even for periods as long as 1 month. In the absence of BSA, the electrochemical behavior of titanium specimens was significantly affected by the length of immersion time, reflecting the changes that slowly occur on their surface. In the presence of BSA, the surfaces maintained most of their original electrochemical activity. Surface studies have shown that calcium and phosphate become incorporated in the surface at very early stages of immersion. Albumin, which was rapidly adsorbed on titanium, was slowly desorbed when titanium was placed in HBSS. Protein and phosphate may coexist on the same surface, but initially adsorbed albumin molecules prevent the precipitation of a thick layer of tricalcium phosphate.     

Bisphosphonate coating on titanium screws increases mechanical fixation in rat tibia after two weeks

Recently published data indicate that immobilized N-bisphosphonate enhances the pullout force and energy uptake of implanted stainless steel screws at 2 weeks in rat tibia. This study compares titanium screws with and without a bisphosphonate coating in the same animal model. The screws were first coated with an approximately 100-nm thick crosslinked fibrinogen film. Pamidronate was subsequently immobilized into this film via EDC/NHS-activated carboxyl groups within the fibrinogen matrix, and finally another N-bisphosphonate, ibandronate, was physically adsorbed. The release kinetics of immobilized (14)C-alendronate was measured in buffer up to 724 h and showed a 60% release within 8 h. Mechanical tests demonstrated a 32% (p = 0.04) and 48% (p = 0.02) larger pullout force and energy until failure after 2 weeks of implantation, compared to uncoated titanium screws. A control study with physically adsorbed pamidronate showed no effect on mechanical fixation, probably due to a too small adsorbed amount. We conclude that the fixation of titanium implants in bone can be improved by fibrinogen matrix-bound bisphosphonates.     

Osteoinductive Implants: The Mise-en-scène for Drug-Bearing Biomimetic Coatings

In orthopaedic and dental implantology, novel tools and techniques are being sought to improve the regeneration of bone tissue. Numerous attempts have been made to enhance the osteoconductivity of titanium prostheses, including modifications in their surface properties and coating with layers of calcium phosphate. The technique whereby such layers are produced has recently undergone a revolutionary change, which has had profound consequences for their potential to serve as drug-carrier systems. Hitherto, calcium phosphate layers were deposited upon the surfaces of metal implants under highly unphysiological physical conditions, which precluded the incorporation of proteinaceous osteoinductive drugs. These agents could only be adsorbed, superficially, upon preformed layers. Such superficially adsorbed molecules are released too rapidly within a biological milieu to be effective in their osteoinductive capacity. Now, it is possible to deposit calcium phosphate layers under physiological conditions of temperature and pH by the so-called biomimetic process, during which bioactive agents can be coprecipitated. Since these molecules are integrated into the inorganic latticework, they are released gradually in vivo as the layer undergoes degradation. This feature enhances the capacity of these coatings to act as a carrier system for osteogenic agents.     

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.     

Biological performance of biomimetic calcium phosphate coating of titanium implants in the dog mandible

The aim of the present study was to analyze the in vivo effect of biomimetic calcium phosphate coating of titanium implants on periimplant bone formation and bone-/implant contact. Five types of implants were used: 1) Ti6Al4V implants with a polished surface; 2) Ti6Al4V implants with collagen coating; 3) Ti6Al4V implants with a mineralized collagen layer; 4) Ti6Al4V implants with sequential coating of hydroxyapatite (HA) and collagen; and 5) Ti6Al4V implants with HA coating only. All implants had square cross sections with an oblique diameter of 4.6 mm and were inserted press fit into trephine burr holes of 4.6 mm in the mandibles of ten beagle dogs. The implants of five animals each were evaluated after a healing period of 1 month and 3 months, respectively, during which time sequential fluorochrome labeling of bone formation had been performed. Bone formation was evaluated by morphometric measurement of the newly formed bone around the implants and the percentage of implant bone contact. After 1 month, there was a significantly higher percentage of mean bone/implant contact in the HA-coated implants compared to those with polished surface and those with the collagen-coated surface. After 3 months, these differences were not present anymore. Bone apposition was significantly higher next to implants with sequential HA/collagen coating compared to polished surfaces and mineralized collagen layer. It is concluded that biomimetic coating of titanium implants with HA has shown the clearest trend to increase bone-implant contact in the early ingrowth period. The addition of collagen to an HA coating layer may hold some promise when used as sequential HA/collagen coating with mineralized collagen as the surface layer.