Poly-ε-caprolactone Coated and Functionalized Porous Titanium and Magnesium Implants for Enhancing Angiogenesis in Critically Sized Bone Defects

For healing of critically sized bone defects, biocompatible and angiogenesis supporting implants are favorable. Murine osteoblasts showed equal proliferation behavior on the polymers poly-ε-caprolactone (PCL) and poly-(3-hydroxybutyrate)/poly-(4-hydroxybutyrate) (P(3HB)/P(4HB)). As vitality was significantly better for PCL, it was chosen as a suitable coating material for further experiments. Titanium implants with 600 µm pore size were evaluated and found to be a good implant material for bone, as primary osteoblasts showed a vitality and proliferation onto the implants comparable to well bottom (WB). Pure porous titanium implants and PCL coated porous titanium implants were compared using Live Cell Imaging (LCI) with Green fluorescent protein (GFP)-osteoblasts. Cell count and cell covered area did not differ between the implants after seven days. To improve ingrowth of blood vessels into porous implants, proangiogenic factors like Vascular Endothelial Growth Factor (VEGF) and High Mobility Group Box 1 (HMGB1) were incorporated into PCL coated, porous titanium and magnesium implants. An angiogenesis assay was performed to establish an in vitro method for evaluating the impact of metallic implants on angiogenesis to reduce and refine animal experiments in future. Incorporated concentrations of proangiogenic factors were probably too low, as they did not lead to any effect. Magnesium implants did not yield evaluable results, as they led to pH increase and subsequent cell death.     

Comparison of Selective Laser Melted Titanium and Magnesium Implants Coated with PCL

Degradable implant material for bone remodeling that corresponds to the physiological stability of bone has still not been developed. Promising degradable materials with good mechanical properties are magnesium and magnesium alloys. However, excessive gas production due to corrosion can lower the biocompatibility. In the present study we used the polymer coating polycaprolactone (PCL), intended to lower the corrosion rate of magnesium. Additionally, improvement of implant geometry can increase bone remodeling. Porous structures are known to support vessel ingrowth and thus increase osseointegration. With the selective laser melting (SLM) process, defined open porous structures can be created. Recently, highly reactive magnesium has also been processed by SLM. We performed studies with a flat magnesium layer and with porous magnesium implants coated with polymers. The SLM produced magnesium was compared with the titanium alloy TiAl6V4, as titanium is already established for the SLM-process. For testing the biocompatibility, we used primary murine osteoblasts. Results showed a reduced corrosion rate and good biocompatibility of the SLM produced magnesium with PCL coating.     

UV Light-Generated Superhydrophilicity of a Titanium Surface Enhances the Transfer,Diffusion and Adsorption of Osteogenic Factors from a Collagen Sponge

It is a significant challenge for a titanium implant, which is a bio-inert material, to recruit osteogenic factors, such as osteoblasts, proteins and blood effectively when these are contained in a biomaterial. The objective of this study was to examine the effect of ultraviolet (UV)-treatment of titanium on surface wettability and the recruitment of osteogenic factors when they are contained in an atelocollagen sponge. UV treatment of a dental implant made of commercially pure titanium was performed with UV-light for 12 min immediately prior to the experiments. Superhydrophilicity on dental implant surfaces was generated with UV-treatment. The collagen sponge containing blood, osteoblasts, or albumin was directly placed on the dental implant. Untreated implants absorbed only a little blood from the collagen sponge, while the UV-treated implants absorbed blood rapidly and allowed it to spread widely, almost over the entire implant surface. Blood coverage was 3.5 times greater for the UV-treated implants (p < 0.001). Only 6% of the osteoblasts transferred from the collagen sponge to the untreated implants, whereas 16% of the osteoblasts transferred to the UV-treated implants (p < 0.001). In addition, a weight ratio between transferred albumin on the implant and measured albumin adsorbed on the implant was 17.3% in untreated implants and 38.5% in UV-treated implants (p < 0.05). These results indicated that UV treatment converts a titanium surface into a superhydrophilic and bio-active material, which could recruite osteogenic factors even when they were contained in a collagen sponge. The transfer and subsequent diffusion and adsorption efficacy of UV-treated titanium surfaces could be useful for bone formation when titanium surfaces and osteogenic factors are intervened with a biomaterial.     

Evaluation of Osseointegration of Titanium Alloyed Implants Modified by Plasma Polymerization

By means of plasma polymerization, positively charged, nanometre-thin coatings can be applied to implant surfaces. The aim of the present study was to quantify the adhesion of human bone cells in vitro and to evaluate the bone ongrowth in vivo, on titanium surfaces modified by plasma polymer coatings. Different implant surface configurations were examined: titanium alloy (Ti6Al4V) coated with plasma-polymerized allylamine (PPAAm) and plasma-polymerized ethylenediamine (PPEDA) versus uncoated. Shear stress on human osteoblast-like MG-63 cells was investigated in vitro using a spinning disc device. Furthermore, bone-to-implant contact (BIC) was evaluated in vivo. Custom-made conical titanium implants were inserted at the medial tibia of female Sprague-Dawley rats. After a follow-up of six weeks, the BIC was determined by means of histomorphometry. The quantification of cell adhesion showed a significantly higher shear stress for MG-63 cells on PPAAm and PPEDA compared to uncoated Ti6Al4V. Uncoated titanium alloyed implants showed the lowest BIC (40.4%). Implants with PPAAm coating revealed a clear but not significant increase of the BIC (58.5%) and implants with PPEDA a significantly increased BIC (63.7%). In conclusion, plasma polymer coatings demonstrate enhanced cell adhesion and bone ongrowth compared to uncoated titanium surfaces.     

Morphology,Composition, and Bioactivity of Strontium-Doped Brushite Coatings Deposited on Titanium Implants via Electrochemical Deposition

Surface modification techniques have been applied to generate titanium implant surfaces that promote osseointegration for use in dental applications. In this study, strontium-doped brushite coatings were deposited on titanium by electrochemical deposition. The phase composition of the coating was investigated by energy dispersive X-ray spectroscopy and X-ray diffraction. The surface morphologies of the coatings were studied through scanning electron microscopy, and the cytocompatibility and bioactivity of the strontium-doped brushite coatings were evaluated using cultured osteoblasts. Osteoblast proliferation was enhanced by the addition of strontium, suggesting a possible mechanism by which strontium incorporation in brushite coatings increased bone formation surrounding the implants. Cell growth was also strongly influenced by the composition of the deposited coatings, with a 10% Sr-doped brushite coating inducing the greatest amount of bone formation among the tested materials.     

Surface coating with cyclic RGD peptides stimulates osteoblast adhesion and proliferation as well as bone formation

The physiological inertness of synthetic implant materials often results in insufficient implant integration and limited acceptance of implants in tissues. After implantation the implant surface is often separated from the surrounding healthy and regenerating tissue, for example by a fibrous capsule. To avoid this host-versus-graft reaction, a strong mechanical contact between tissue and implant must be ensured. An enhanced contact between graft and the surrounding tissue can be provided by coating the implant with cell-adhesive molecules. The highly active and alpha(v)beta(3)- and alpha(v)beta(5)-integrin-selective peptide c(-RGDfK-) (f=D-phenylalanine) was functionalized with various linker molecules containing an acrylamide end group by using the lysine side chain of c(-RGDfK-). The acrylamide group can be used to bind the peptide covalently to poly(methyl methacrylate) (PMMA) surfaces. The coated surfaces effectively bind to murine osteoblasts as well as human osteoblasts in vitro when a minimum distance of 3.5 nm between surface and the constrained RGD sequence is provided. In contrast to osteoblasts in cell suspension, surface-bound osteoblasts show no apoptosis but proliferate by a factor of 10 over a 22 d period. Coating of inert implant surfaces with highly active and alpha(v)-selective peptides affords a marked improvement in osteoblast binding over current technologies. In vivo studies show that peptide-coated PMMA pellets implanted into the patella groove of rabbits are integrated into the regenerating bone tissue faster and more strongly than uncoated pellets.     

On-Growth and In-Growth Osseointegration Enhancement in PM Porous Ti-Scaffolds by Two Different Bioactivation Strategies: Alkali Thermochemical Treatment and RGD Peptide Coating

A lack of primary stability and osteointegration in metallic implants may result in implant loosening and failure. Adding porosity to metallic implants reduces the stress shielding effect and improves implant performance, allowing the surrounding bone tissue to grow into the scaffold. However, a bioactive surface is needed to stimulate implant osteointegration and improve mechanical stability. In this study, porous titanium implants were produced via powder sintering to create different porous diameters and open interconnectivity. Two strategies were used to generate a bioactive surface on the metallic foams: (1) an inorganic alkali thermochemical treatment, (2) grafting a cell adhesive tripeptide (RGD). RGD peptides exhibit an affinity for integrins expressed by osteoblasts, and have been reported to improve osteoblast adhesion, whereas the thermochemical treatment is known to improve titanium implant osseointegration upon implantation. Bioactivated scaffolds and control samples were implanted into the tibiae of rabbits to analyze the effect of these two strategies in vivo regarding bone tissue regeneration through interconnected porosity. Histomorphometric evaluation was performed at 4 and 12 weeks after implantation. Bone-to-implant contact (BIC) and bone in-growth and on-growth were evaluated in different regions of interest (ROIs) inside and outside the implant. The results of this study show that after a long-term postoperative period, the RGD-coated samples presented higher quantification values of quantified newly formed bone tissue in the implant’s outer area. However, the total analyzed bone in-growth was observed to be slightly greater in the scaffolds treated with alkali thermochemical treatment. These results suggest that both strategies contribute to enhancing porous metallic implant stability and osteointegration, and a combination of both strategies might be worth pursuing.     

Titanium implant materials with improved biocompatibility through coating with phosphonate-anchored cyclic RGD peptides

One key point for improving osseous integration of implants is to render them osteopromotive by specifically favoring the adhesion of osteoblasts. Mimicking the physiological adhesion process of osteoblasts to the extracellular matrix improves cell adhesion in vitro and results in improved and earlier osseous integration of implants in vivo. Our approach involves coating titanium implants with a tailor-made cyclic-RGD peptide, thus allowing them to bind to specific integrin receptors on the cell surface through multimeric phosphonates. The advantages of this very stable, new type of anchoring for practical application are presented.     

Enhanced mechanical properties of porous titanium implants via in-situ synthesized titanium carbide in lamellar pore walls

Directional lamellar porous titanium scaffolds are widely used as bone implant bearing materials because of their anisotropic pore structure. Their mechanical properties can be effectively improved by enhancing the strength of pore walls through the introduction of ceramics. In this work, porous titanium implants were prepared by freeze casting combined with TiH₂ decomposition. The graphene was introduced into the pore walls of porous titanium, which could transform into titanium carbide (TiC) in situ upon sintering. TiC was evenly distributed in the lamellar pore walls, and the interface was well bonded. The compression strength of the fabricated implants was up to 389.94 MPa when the graphene content was 3 wt%, which was 377.8% times as high as the porous titanium. The crack propagation was resisted by TiC because of the “pinning” effect on the pore wall. Some of TiC were pulled out from the matrix, and others were fractured. The strength of the fabricated implants was improved significantly by the large consumption of fracture energy. Also, fabricated porous titanium implants with TiC are suitable for bone implantation.     

Early Fixation of Cobalt-Chromium Based Alloy Surgical Implants to Bone Using a Tissue-engineering Approach

To establish the methods of demonstrating early fixation of metal implants to bone, one side of a Cobalt-Chromium (CoCr) based alloy implant surface was seeded with rabbit marrow mesenchymal cells and the other side was left unseeded. The mesenchymal cells were further cultured in the presence of ascorbic acid, β-glycerophosphate and dexamethasone, resulting in the appearance of osteoblasts and bone matrix on the implant surface. Thus, we succeeded in generating tissue-engineered bone on one side of the CoCr implant. The CoCr implants were then implanted in rabbit bone defects. Three weeks after the implantation, evaluations of mechanical test, undecalcified histological section and electron microscope analysis were performed. Histological and electron microscope images of the tissue engineered surface exhibited abundant new bone formation. However, newly formed bone tissue was difficult to detect on the side without cell seeding. In the mechanical test, the mean values of pull-out forces were 77.15 N and 44.94 N for the tissue-engineered and non-cell-seeded surfaces, respectively. These findings indicate early bone fixation of the tissue-engineered CoCr surface just three weeks after implantation.     

Insights into Surface Treatment Methods of Titanium Dental Implants

Titanium is the most widely used material for dental implants, due to its desirable properties, e.g., high biocompatibility, low density, high stiffness and strength, etc. More importantly, titanium implants may osseointegrate with living bone, meaning that new bone grows directly onto the surface of the implant, without any intermediate soft tissue layer. A successfully osseointegrated implant generally has a strong bonding to the adjacent bone; consequently, it usually functions well and remains stable for long service period. It also has been clinically proven that surface treatment methods can improve the rate and quality of titanium implants' osseointegration. This article focuses on two such methods, i.e., surface roughening and hydroxyapatite (HA) coating. In addition, we discuss a promising new methodology, which attempts to modify the surface charge of titanium materials. This paper focuses on the current best surface treatment methods for titanium dental implants developed and improved in the past two decades, i.e., 1990–2010.     

Nanoroughening of sandblasted 3Y-TZP surface by alumina coating deposition for improved osseointegration and bacteria reduction

Zirconia-based (3Y-TZP) dental implants are being introduced as an alternative to established titanium counterparts. As 3Y-TZP’s bioinertness poses a challenge to osseointegration, modifications of the implants’ surface are necessary. Roughening is a common method to improve the cell response and should ideally combine micro- and nano-scale features. Here we show a simple synthesis procedure for the nano-roughening of sandblasted 3Y-TZP by precipitation of a nanostructured alumina coating. As-modified surfaces were characterised regarding their morphology and phase composition, roughness, wetting behaviour and zeta potential, and correlated to the behaviour of human osteoblasts and Staphylococcus aureus bacteria. Osteoblasts were successfully cultured on all surfaces regardless of their modification. Surfaces exhibiting a combination of micro- and nano-roughening performed best regarding initial attachment and proliferation of osteoblasts. Alumina-coated surfaces also limited the bacterial adhesion.     

Antimicrobial Brushes on Titanium via “Grafting to” Using Phosphonic Acid/Pyridinium Containing Block Copolymers

Coating medical implants with antibacterial polymers may prevent postoperative infections which are a common issue for conventional titanium implants and can even lead to implant failure. Easily applicable diblock copolymers are presented that form polymer brushes via “grafting to” mechanism on titanium and equip the modified material with antibacterial properties. The polymers carry quaternized pyridinium units to combat bacteria and phosphonic acid groups which allow the linear chains to be anchored to metal surfaces in a convenient coating process. The polymers are synthesized via reversible-addition-fragmentation-chain-transfer (RAFT) polymerization and postmodifications and are characterized using NMR spectroscopy and SEC. Low grafting densities are a major drawback of the “grafting to” approach compared to “grafting from”. Thus, the number of phosphonic acid groups in the anchor block are varied to investigate and optimize the surface binding. Modified titanium surfaces are examined regarding their composition, wetting behavior, streaming potential, and coating stability. Evaluation of the antimicrobial properties revealed reduced bacterial adhesion and biofilm formation for certain polymers, albeit the cell biocompatibility against human gingival fibroblasts is also impaired. The presented findings show the potential of easy-to-apply polymer coatings and aid in designing next-generation implant surface modifications.     

Osteoinduction properties of graphite-like amorphous carbon films evaluated in-vitro

Graphite-like amorphous carbon (a-C) films were deposited by magnetron sputtering on medical grade stainless steel (SS) substrates. The purpose of the study was the evaluation of amorphous carbon as a coating for medical implants in contact with bone, in which the formation of bone on the surface is of special interest to improve the implant performance. The initial step consisted of the evaluation of the cytocompatibility of the a-C films. Cytocompatibility was evaluated by seeding human osteoblast cells on the surfaces and measuring their adhesion, proliferation and viability up to seven days. These data are also of interest for the osteoinduction process because the initial phase of the bone-forming activity of osteoblasts entails the attachment, proliferation, cell differentiation and extracellular matrix synthesis, followed by a second phase of bone matrix mineralization. The evaluation in-vitro of the osteoinduction properties of the surfaces consisted on the study of cell differentiation by measuring the alkaline phosphatase activity, the morphological examination of the mineralized extracellular matrix by electron microscopy, and the evaluation of a selection of proteins involved in the bone-formation process.The cytocompatibility (cell attachment, proliferation and viability) was determined in comparison with SS, titanium coatings and the plastic tissue control. We observed a superior cellular adhesion and proliferation in the a-C surfaces, while the viability was slightly better on the Ti coatings, but nevertheless, the a-C values were much higher than in the plastic control, indicating that the a-C films are no toxic and biocompatible. On the other hand, all the results of the osteoinduction; ALP specific activity and expression of bone-growth specific proteins indicated that the a-C surface significantly promotes the cell differentiation, leading to the formation of the extracellular matrix and its mineralization.     

Antibacterial Titanium Implants Biofunctionalized by Plasma Electrolytic Oxidation with Silver,Zinc, and Copper: A Systematic Review

Patients receiving orthopedic implants are at risk of implant-associated infections (IAI). A growing number of antibiotic-resistant bacteria threaten to hamper the treatment of IAI. The focus has, therefore, shifted towards the development of implants with intrinsic antibacterial activity to prevent the occurrence of infection. The use of Ag, Cu, and Zn has gained momentum as these elements display strong antibacterial behavior and target a wide spectrum of bacteria. In order to incorporate these elements into the surface of titanium-based bone implants, plasma electrolytic oxidation (PEO) has been widely investigated as a single-step process that can biofunctionalize these (highly porous) implant surfaces. Here, we present a systematic review of the studies published between 2009 until 2020 on the biomaterial properties, antibacterial behavior, and biocompatibility of titanium implants biofunctionalized by PEO using Ag, Cu, and Zn. We observed that 100% of surfaces bearing Ag (Ag-surfaces), 93% of surfaces bearing Cu (Cu-surfaces), 73% of surfaces bearing Zn (Zn-surfaces), and 100% of surfaces combining Ag, Cu, and Zn resulted in a significant (i.e., >50%) reduction of bacterial load, while 13% of Ag-surfaces, 10% of Cu-surfaces, and none of Zn or combined Ag, Cu, and Zn surfaces reported cytotoxicity against osteoblasts, stem cells, and immune cells. A majority of the studies investigated the antibacterial activity against S. aureus. Important areas for future research include the biofunctionalization of additively manufactured porous implants and surfaces combining Ag, Cu, and Zn. Furthermore, the antibacterial activity of such implants should be determined in assays focused on prevention, rather than the treatment of IAIs. These implants should be tested using appropriate in vivo bone infection models capable of assessing whether titanium implants biofunctionalized by PEO with Ag, Cu, and Zn can contribute to protect patients against IAI.     

Porous titanium and nitinol implants synthesized by SHS/SLS: Microstructural and histomorphological analyses of tissue reactions

The comparative microstructural analyses and histomorphological studies of tissue reactions to porous titanium and nitinol implants synthesized by Selective Laser Sintering (SLS) are presented for a rat model for bone implants. It was discovered that the surface of porous pegs of titanium and nitinol made by SHS/SLS has a significantly favorable structure to the mechanical interlocking with bone and soft tissues. Histological analysis of decalcified paraffin sections after implant removal could only show that trabecular bone structures and marrow cavities were observed around the porous implants. In the connective tissue of the remaining implant beds the following cells: macrophages, fibroblasts, adipocytes and lymphocytes are discernible. It was shown that the nitinol synthesized by combined SHS/SLS technique has a developed and ordered microstructure.     

Development of a novel smart carrier for drug delivery: Ciprofloxacin loaded vaterite/reduced graphene oxide/PCL composite coating on TiO2 nanotube coated titanium

In the field of orthopaedic implants, post-surgery infections and biocompatibility are the most challenging obstacles. Sustained and controlled antibiotic release is a key factor in novel drug delivery systems. A novel drug delivery system combined with vaterite microsphere, graphite oxide (GO), reduced graphene oxide (rGO) incorporated in a polycaprolactone (PCL) matrix on TiO₂ nanotube coated Ti (TNT-Ti) is established. Anodization was employed to develop TiO₂ nanotubular arrays on Ti. Ciprofloxacin hydrochloride (CPF–HCl) loaded vaterite microspheres were synthesized by in situ precipitation method. Deposition of vaterite/PCL, vaterite-GO/PCL and vaterite-rGO/PCL composite coating on TNT-Ti was carried out by dip coating method. The composite coatings were characterized for their phase content, morphological features and functional groups. Among the three types of composite coatings, vaterite-rGO/PCL composite coating is found to be capable of encapsulating CPF-HCl to a level of 75.14 μg. The drug release profile of CPF-HCl from the vaterite-rGO/PCL composite coating exhibits a controlled release amounting to only 35.02 % of release at the end of 120 h. The vaterite-rGO/PCL composite coating exhibits a low dissolution rate and possesses adequate bioactivity in HBSS and SBF solutions at 37 °C for 14 and 10 days, respectively. The in situ loaded CPF-HCL drug on vaterite microspheres, PCL polymer matrix and GO/rGO nanofillers does not affect the cytocompatibility and all the composite coatings supported cell viability and proliferation. The ability of vaterite-rGO/PCL composite coating to provide a slow and steady release of antibiotics with sufficient bioactivity and biocompatibility at the tissue implant interface makes it a promising for osteomyelitis infection of bone tissue implant materials.     

Preparation of Calcium Phosphate Compounds on Zirconia Surfaces for Dental Implant Applications

Titanium is widely used in medical implants despite the release of heavy metal ions over long-term use. Zirconia is very close to the color of teeth; however, its biological inertness hinders bonding with bone tissue. Alkaline treatment and coatings of calcium phosphate can be used to enhance bone regeneration adjacent to dental implants. This study examined the effects of alkaline treatment, calcium phosphate coatings, and sintering, on the physical properties of implant material. Our analysis confirmed that the calcium phosphate species were octacalcium phosphate (OCP). The sintering of calcium phosphate was shown to create B-type HAP, which is highly conducive toward the differentiation of mesenchymal stem cells (MSCs) into osteoblasts for the facilitation of bone integration. Conclusions: This study demonstrated the room-temperature fabrication of dental implants with superhydrophilic surfaces to enhance biocompatibility.     

Plasma‐Sprayed Ceramic Coatings for Osseointegration

Osseointegration of an orthopedic implant involves a cascade of cellular and extracellular biological events that take place at bone/implant interface. The process can be enhanced by coating implants with materials of various compositions, chemistry, topographies. The aim of this article was to review the literatures on development and application of numerous plasma‐sprayed ceramic coatings (including hydroxyapatite, wollastonite, dicalcium silicate, titanium oxide, and zirconia coatings) for establishing and maintaining the osseointegration based on some major biological requirements, such as good cytocompatibility, advantageous cell response, superior tissue response, effective anti‐infection activities.     

Biomimetic Hydroxyapatite Coating on Metal Implants

The combination of the high mechanical strength of metals with the osteoconductive properties of calcium phosphates make hydroxyapatite coatings on titanium implants widely used in orthopedic surgery. However, the most popular coating method, plasma spraying, exhibits some important drawbacks: the inability to cover porous implants and to incorporate biologically active agents, delamination, and particle release. The aim of this study was to elaborate a dense, strong, and thick calcium-phosphate coating on titanium and porous-tantalum implants using a two-step biomimetic procedure. In the first step, the implants were soaked in a solution that was 5 times more concentrated than regular simulated body fluid (SBF-A solution). A thin but uniform amorphous calcium-phosphate coating was deposited on the metal. Then, the implants were immersed in the SBF-B solution, which had a similar composition as the SBF-A solution, but with decreased contents of crystal growth inhibitors (i.e., Mg²⁺ and HCO₃⁻). This resulted in the fast precipitation of a 30 μm thick crystalline calcium-phosphate coating. The pH of the SBF-B solution and the thickness of the crystalline coating layer were studied as a function of time. The Fourier transform infrared spectra and X-ray diffraction patterns showed that this new coating closely resembles bone mineral. Our biomimetic coating should facilitate rapid bone formation around the implant, reducing therewith the patient's recovery time after surgery.