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1.  Study of a new bone-targeting titanium implant–bone interface 
New strategies involving bone-targeting titanium (Ti) implant–bone interface are required to enhance bone regeneration and osseointegration for orthopedic and dental implants, especially in osteoporotic subjects. In this study, a new dual-controlled, local, bone-targeting delivery system was successfully constructed by loading tetracycline-grafted simvastatin (SV)-loaded polymeric micelles in titania nanotube (TNT) arrays, and a bone-targeting Ti implant–bone interface was also successfully constructed by implanting the delivery system in vivo. The biological effects were evaluated both in vitro and in vivo. The results showed that Ti surfaces with TNT–bone-targeting micelles could promote cytoskeletal spreading, early adhesion, alkaline phosphatase activity, and extracellular osteocalcin concentrations of rat osteoblasts, with concomitant enhanced protein expression of bone morphogenetic protein (BMP)-2. A single-wall bone-defect implant model was established in normal and ovariectomized rats as postmenopausal osteoporosis models. Microcomputed tomography imaging and BMP-2 expression in vivo demonstrated that the implant with a TNT-targeting micelle surface was able to promote bone regeneration and osseointegration in both animal models. Therefore, beneficial biological effects were demonstrated both in vitro and in vivo, which indicated that the bone-targeting effects of micelles greatly enhance the bioavailability of SV on the implant–bone interface, and the provision of SV-loaded targeting micelles alone exhibits the potential for extensive application in improving local bone regeneration and osseointegration, especially in osteoporotic subjects.
doi:10.2147/IJN.S119520
PMCID: PMC5135286  PMID: 27932879
bone regeneration; titania nanotubes; targeted drug delivery; orthopedic implant; drug release; micelles
2.  Surface-mediated bone tissue morphogenesis from tunable nanolayered implant coatings* 
Science translational medicine  2013;5(191):191ra83.
The functional success of a biomedical implant critically depends on its stable bonding with the host tissue. Aseptic implant loosening accounts for over half of all joint replacement failures. Various materials, including metals and plastic, confer mechanical integrity to the device, but often these materials are not suitable for direct integration with the host tissue, which leads to implant loosening and patient morbidity. We describe a self-assembled, osteogenic, polymer-based conformal coating that promotes stable mechanical fixation of an implant in a surrogate rodent model. A single modular, polymer-based multilayered coating was deposited using a water-based layer-by-layer approach, by which each element was introduced on the surface in nanoscale layers. Osteoconductive hydroxyapatite (HAP) and osteoinductive bone morphogenetic protein 2 (BMP-2) contained within the nanostructured coating acted synergistically to induce osteoblastic differentiation of endogenous progenitor cells within the bone marrow, without indications of a foreign body response. The tuned release of BMP-2, controlled by a hydrolytically degradable poly(β-amino ester), was essential for tissue regeneration and, in the presence of HAP, the modular coating encouraged the direct deposition of highly cohesive trabecular bone on the implant surface. The bone-implant interfacial tensile strength was significantly higher than standard bone cement, did not fracture at the interface, and had long-term stability. Collectively, these results suggest that the multilayered coating system promotes biological fixation of orthopedic and dental implants to improve surgical outcomes by preventing loosening and premature failure.
doi:10.1126/scitranslmed.3005576
PMCID: PMC4001255  PMID: 23803705
3.  Bone Regeneration in Odontostomatology 
Maxillary edentulism, together with periodontal disease, is the condition that most frequently induces disruption of alveolar bone tissue. Indeed, the stimulus of the periodontal ligament is lost and the local bone tissue becomes subject to resorption processes that, in the six months following the loss of the tooth, result in alveolar defects or more extensive maxillary atrophy. In both cases, loss of vestibular cortical bone is followed by reduction in the vertical dimension of the alveolar process, producing effects that upset the morphology of the three-dimensional relations between the dental arches. Maintenance, or restoration, of sufficient bone volume to withstand prosthetic loading and the insertion of an endosseous implant, demands the implementation of operating protocols that bring about bone regeneration in the defect sites. Given the biological principles involved, this requires the implementation of osteogenesis, osteoinduction and osteoconduction protocols.
Osteogenesis is the synthesis of new bone by autologous cells that remain viable, given the capacity of the grafted material to become part of the newly forming bone tissue; osteoinduction is based on the capacity of the grafted material to induce the migration, proliferation and phenotypic conversion, into bone-producing cells, of multipotent undifferentiated cells derived from connective tissue or bone marrow; osteoconduction, meanwhile, provides three-dimensional support and guidance to osteoblast precursors within the defect. The operating procedures implemented take into account the size and morphology of the defect, for the restoration of which guided repair or an out-and-out regenerative protocol may be sufficient. Guided repair exploits the principle of resorption/replacement of the biomaterial with newly-formed bone and consists of restoring the lost bone tissue through the implantation of different, osteointegrative biomaterials. This type of repair requires the application of biocompatible osteoconductors which will gradually be absorbed and replaced by newly formed tissue. Instead, the clinical-surgical basis of bone regeneration is: guided bone regeneration (GBR), the use of growth factors and the application of grafts/osteointegrative materials. GBR, through the use of membranes (resorbable or non-resorbable) allows the filling of a defect, “guiding” the growth only of the osteogenic lines and preventing the invasion of non-osteogenic tissues that compete with the bone. This objective is achieved also thanks to the capacity of the membranes to serve as a filter, thereby strengthening the osteocompetent lines and, at the same time, keeping epithelial cells away. The clinical use of GBR, partly on account of its predictable results, is now very widespread. The growth factors used in bone regeneration are glycoproteins which exert autocrine and paracrine effects on the primordial cells in the site. One of these factors, plasma-rich protein (PRP), is an autologous source of growth factors; obtained by separating and concentrating the platelets in a small volume of plasma, it is immediately utilisable in the surgical site. As regards the osteointegrative materials we can distinguish between autologous, homologous, heterologous, and alloplastic grafts. Of these, autologous bone is the gold standard as it has osteogenic, osteoinductive, and osteoconductive properties and, being fresh, keeps osteoblasts viable. Depending on the size of the defect to be treated, harvesting is from endoral or extraoral sites (calvaria, iliac crest, tibia). The harvested material conserves the embryological characteristics of the site of origin: this principle is reflected in the bone density that develops in the regenerated site. Homologous bone supplied by tissue banks in various formulations is an osteoconductive and partially osteoinductive material that guarantees good mechanical properties even in large defects. Heterologous bone of bovine or equine origin is a carbonate-rich non-stoichiometric apatite. Despite showing low resorption, it does not withstand traction or masticatory loading. Alloplastic materials are osteoconductive materials showing different degrees of resorption; they have biomechanical properties and the speed of their resorption varies, depending on their chemical and stoichiometric formulation. The purpose of bone regeneration thus obtained is to allow the insertion of a titanium implant in the site of the regeneration. This alloplastic implant, whose rough and porous surface allows integration with the bone tissue, will support the prosthesis subsequently applied.
PMCID: PMC3213819
4.  Peen treatment on a titanium implant: effect of roughness, osteoblast cell functions, and bonding with bone cement 
Implant failure due to poor integration of the implant with the surrounding biomaterial is a common problem in various orthopedic and orthodontic surgeries. Implant fixation mostly depends upon the implant surface topography. Micron to nanosize circular-shaped groove architecture with adequate surface roughness can enhance the mechanical interlock and osseointegration of an implant with the host tissue and solve its poor fixation problem. Such groove architecture can be created on a titanium (Ti) alloy implant by laser peening treatment. Laser peening produces deep, residual compressive stresses in the surfaces of metal parts, delivering increased fatigue life and damage tolerance. The scientific novelty of this study is the controlled deposition of circular-shaped rough spot groove using laser peening technique and understanding the effect of the treatment techniques for improving the implant surface properties. The hypothesis of this study was that implant surface grooves created by controlled laser peen treatment can improve the mechanical and biological responses of the implant with the adjoining biomaterial. The objective of this study was to measure how the controlled laser-peened groove architecture on Ti influences its osteoblast cell functions and bonding strength with bone cement. This study determined the surface roughness and morphology of the peen-treated Ti. In addition, this study compared the osteoblast cell functions (adhesion, proliferation, and differentiation) between control and peen-treated Ti samples. Finally, this study measured the fracture strength between each kind of Ti samples and bone cement under static loading. This study found that laser peen treatment on Ti significantly changed the surface architecture of the Ti, which led to enhanced osteoblast cell adhesion and differentiation on Ti implants and fracture strength of Ti–bone cement interfaces compared with values of untreated Ti samples. Therefore, the laser peen treatment method has the potential to improve the biomechanical functions of Ti implants.
doi:10.2147/IJN.S89376
PMCID: PMC4745826  PMID: 26893563
titanium; cement; interface; PMMA; fracture strength; orthopedics; laser peen; orthodontics
5.  Techniques for dental implant nanosurface modifications 
PURPOSE
Dental implant has gained clinical success over last decade with the major drawback related to osseointegration as properties of metal (Titanium) are different from human bone. Currently implant procedures include endosseous type of dental implants with nanoscale surface characteristics. The objective of this review article is to summarize the role of nanotopography on titanium dental implant surfaces in order to improve osseointegration and various techniques that can generate nanoscale topographic features to titanium implants.
MATERIALS AND METHODS
A systematic electronic search of English language peer reviewed dental literature was performed for articles published between December 1987 to January 2012. Search was conducted in Medline, PubMed and Google scholar supplemented by hand searching of selected journals. 101 articles were assigned to full text analysis. Articles were selected according to inclusion and exclusion criterion. All articles were screened according to inclusion standard. 39 articles were included in the analysis.
RESULTS
Out of 39 studies, seven studies demonstrated that bone implant contact increases with increase in surface roughness. Five studies showed comparative evaluation of techniques producing microtopography and nanotopography. Eight studies concluded that osteoblasts preferably adhere to nano structure as compared to smooth surface. Six studies illustrated that nanotopography modify implant surface and their properties. Thirteen studies described techniques to produce nano roughness.
CONCLUSION
Modification of dental osseous implants at nanoscale level produced by various techniques can alter biological responses that may improve osseointegration and dental implant procedures.
doi:10.4047/jap.2014.6.6.498
PMCID: PMC4279049  PMID: 25558347
Intelligent surfaces; Sputtering; Superhydrophillic; Chemical vapor deposition; Osseointegration; Engineered surface
6.  Fibronectin and vitronectin promote human fetal osteoblast cell attachment and proliferation on nanoporous titanium surfaces 
Improvements in osteoconduction of implant biomaterials require focusing on the bone-implant interface, which is a complex multifactorial system. Surface topography of implants plays a crucial role at this interface. Nanostructured surfaces have been shown to promote serum protein adsorption and osteoblast adhesion when compared to microstructured surfaces for bone-implant materials. We studied the influence of the serum proteins fibronectin and vitronectin on the attachment and proliferation of osteoblasts onto nanostructured titania surfaces. Human fetal osteoblastic cells hFOB 1.19 were used as model osteoblasts and were grown on nanoporous TiO2 templates, using Ti6Al4V and commercially pure Ti substrates as controls. Results show a significant increase in cell proliferation on nanoporous TiO2 over flat substrates. Initial cell attachment data exhibited a significant effect by either fibronectin or vitronectin on cell adhesion at the surface of any of the tested materials. In addition, the extent of cell adhesion was significantly different between the nanoporous TiO2 and both Ti6Al4V and commercially pure Ti substrates, with the first showing the highest surface coverage. There was no significant difference on osteoblast attachment or proliferation between the presence of fibronectin or vitronectin using any of the material substrates. Taken together, these results suggest that the increase in osteoblast attachment and proliferation shown on the nanoporous TiO2 is due to an increase in the adsorption of fibronectin and vitronectin because of the higher surface area and to an enhanced protein unfolding, which allows access to osteoblast binding motifs within these proteins.
PMCID: PMC3718483  PMID: 23858975
nanoporous TiO2; hFOB 1.19; protein adsorption; vitronectin; fibronectin
7.  Strontium Ranelate: The Pathophysiological Rationale 
Skeletal metabolism and the replacement of damaged tissue with the same amount of intact bone depends on the correct balance between bone formation and bone resorption.
The existence of an imbalance between bone formation and resorption is a concept central to understanding of the pathophysiology of osteoporosis and the reduction of fracture risk.
With aging, the volume of bone that is formed during the bone remodelling process and after injury is less than the volume absorbed during the bone resorption phase; this results in bone loss and increased bone fragility. In addition to bone mineral density, many other properties of bone are determined by the balance between bone formation and bone resorption. A bone that is biomechanically more fragile is also a bone that consolidates more slowly after a fracture event. Although the fracture healing stages are the same even in the presence of osteoporosis, recent studies have shown a slowdown in the process of consolidation when osteoporosis is present. In particular, strategies to reduce fracture risk and facilitate the process of consolidation of the fracture may be a primary criterion for selection.
The ability to modulate anabolic and catabolic phenomena in the skeleton, both locally and systemically, opens up a new horizon for the reduction of fracture risk and the enhancement of bone healing, particularly when the bone is qualitatively and/or quantitatively compromised.
Clinical research has recently allowed the development of therapies, such as treatment with strontium ranelate, able to increase production of bone matrix by osteoblasts and to act positively on the distribution of the skeletal microarchitecture. Strontium ranelate is able to rebalance bone turnover in favour of the formation of more resistant and elastic bone, by stimulating osteoblasts and inhibiting the resorptive activity of osteoclasts, thereby ensuring rapid and lasting protection against the risk of fractures. In vitro studies have shown that the drug is able to promote replication of the first pre-osteoblasts and their differentiation into mature osteoblasts and osteocytes interacting with the receptor CaSR and through the increased synthesis of OPG. Thanks, again, to the participation of the CaSR receptor, but also by reducing the production of RANKL, strontium ranelate decreases the resorptive activity of osteoclasts. The anabolic action of strontium ranelate in terms of mineral apposition rate in both cortical and trabecular bone was demonstrated on bone biopsies analysed by three-dimensional micro-CT. The drug was shown to increase the number of trabeculae, the cortical thickness, and the total bone volume. The bone-forming activity of strontium ranelate was also demonstrated in comparative studies versus teriparatide and antiresorptive agents. In experimental studies the bone-forming effect of strontium ranelate leads to an increase in the bone callus volume and its maturation and, in turn, to an acceleration of the consolidation of the fracture and better implant osteointegration.
In conclusion, the mechanism of action of strontium ranelate, which inhibits bone resorption in favour of new bone formation, is able to counteract, in a physiological manner, the bone loss associated with advancing age. The net effect is an increase in bone mass, trabecular and cortical bone, which explains its anti-fracture efficacy. The drug’s ability to stimulate bone formation seems to unfold at the level of the callus allowing improved fracture healing and in the case of implants potential improvement of implant osteointegration.
PMCID: PMC3213810
8.  A novel bionic design of dental implant for promoting its long-term success using nerve growth factor (NGF): Utilizing nano-springs to construct a stress-cushioning structure inside the implant 
Summary
The absence of periodontium causes masticatory load in excess of the self-repairing potential of peri-implant bone; peri-implant bone loss caused by occlusal overload is not uncommon in patients and greatly diminishes chances of long-term success. Regenerative treatments may be useful in inducing peri-implant bone regeneration, but are only stopgap solutions to the aftermaths caused by the imperfect biomechanical compatibility of the dental implant. Despite promising success, the tissue-engineered periodontal ligament still needs a period of time to be perfected before being clinically applied. Hence, we propose a novel design of dental implant that utilizes nano-springs to construct a stress-cushioning structure inside the implant. Many studies have shown that NGF, a neurotrophin, is effective for nerve regeneration in both animal and clinical studies. Moreover, NGF has the potential to accelerate bone healing in patients with fracture and fracture nonunion and improve osseointegration of the implant. The key point of the design is to reduce stress concentrated around peri-implant bone by cushioning masticatory forces and distributing them to all the peri-implant bone through nano-springs, and promote osseoperception and osseointegration by NGF-induced nerve regeneration and new bone formation. This design, which transfers the main biomechanical interface of the implant from outside to inside, if proven to be valid, may to some extent compensate for the functions of lost periodontium in stress cushioning and proprioception.
doi:10.12659/MSM.883253
PMCID: PMC3560710  PMID: 22847209
bionic design; dental implant; nano-springs; nerve growth factor; osseointegration; osseoperception
9.  Behavior of osteoblastic cells cultured on titanium and structured zirconia surfaces 
Head & Face Medicine  2008;4:29.
Background
Osseointegration is crucial for the long-term success of dental implants and depends on the tissue reaction at the tissue-implant interface. Mechanical properties and biocompatibility make zirconia a suitable material for dental implants, although surface processings are still problematic. The aim of the present study was to compare osteoblast behavior on structured zirconia and titanium surfaces under standardized conditions.
Methods
The surface characteristics were determined by scanning electron microscopy (SEM). In primary bovine osteoblasts attachment kinetics, proliferation rate and synthesis of bone-associated proteins were tested on different surfaces.
Results
The results demonstrated that the proliferation rate of cells was significantly higher on zirconia surfaces than on titanium surfaces (p < 0.05; Student's t-test). In contrast, attachment and adhesion strength of the primary cells was significant higher on titanium surfaces (p < 0.05; U test). No significant differences were found in the synthesis of bone-specific proteins. Ultrastructural analysis revealed phenotypic features of osteoblast-like cells on both zirconia and titanium surfaces.
Conclusion
The study demonstrates distinct effects of the surface composition on osteoblasts in culture. Zirconia improves cell proliferation significantly during the first days of culture, but it does not improve attachment and adhesion strength. Both materials do not differ with respect to protein synthesis or ultrastructural appearance of osteoblasts. Zirconium oxide may therefore be a suitable material for dental implants.
doi:10.1186/1746-160X-4-29
PMCID: PMC2614982  PMID: 19063728
10.  In situ dental implant installation after decontamination in a previously peri-implant diseased site: a pilot study 
Purpose
The aim of this study was to examine whether a previous peri-implantitis site can affect osseointegration, by comparing implant placement at a site where peri-implantitis was present and at a normal bone site. A second aim of this study was to identify the tissue and bone reaction after treating the contaminated implant surface to determine the optimal treatment for peri-implant diseases.
Methods
A peri-implant mucositis model for dogs was prepared to determine the optimal treatment option for peri-implant mucositis or peri-implantitis. The implants were inserted partially to a length of 6 mm. The upper 4 mm part of the dental implants was exposed to the oral environment. Simple exposure for 2 weeks contaminated the implant surface. After 2 weeks, the implants were divided into three groups: untreated, swabbed with saline, and swabbed with H2O2. Three implants from each group were placed to the full length in the same spot. The other three implants were placed fully into newly prepared bone. After eight weeks of healing, the animals were sacrificed. Ground sections, representing the mid-buccal-lingual plane, were prepared for histological analysis. The analysis was evaluated clinically and histometrically.
Results
The untreated implants and H2O2-swabbed implants showed gingival inflammation. Only the saline-swabbed implant group showed re-osseointegration and no gingival inflammation. There was no difference in regeneration height or bone-to-implant contact between in situ implant placement and implant placement in the new bone site.
Conclusions
It can be concluded that cleaning with saline may be effective in implant decontamination. After implant surface decontamination, implant installation in a previous peri-implant diseased site may not interfere with osseointegration.
doi:10.5051/jpis.2012.42.1.13
PMCID: PMC3296930  PMID: 22413069
Dental implants; Decontamination; Hydrogen peroxide; Peri-implantitis
11.  Ti nanorod arrays with a medium density significantly promote osteogenesis and osteointegration 
Scientific Reports  2016;6:19047.
Ti implants are good candidates in bone repair. However, how to promote bone formation on their surface and their consequent perfect integration with the surrounding tissue is still a challenge. To overcome such challenge, we propose to form Ti nanorods on their surface to promote the new bone formation around the implants. Here Ti nanorod arrays (TNrs) with different densities were produced on pure Ti surfaces using an anodizing method. The influence of TNr density on the protein adsorption as well as on the adhesion, proliferation, and osteogenic differentiation of MC3T3-E1 pre-osteoblastic cells were assessed. The TNrs were also implanted into the bone defects in rabbits to test their application in promoting bone formation and osteointegration at the implant-bone interface. TNrs with the medium density were found to show the best capability in promoting the protein adsorption from surrounding medium, which in turn efficiently enhanced osteogenic differentiation in vitro and osteointegration in vivo. Our work suggests that growing TNrs with a medium density on the surface of traditional Ti implants is an efficient and facile method for promoting bone formation and osteointegration in bone repair.
doi:10.1038/srep19047
PMCID: PMC4705471  PMID: 26743328
12.  Nanostructured Surfaces of Dental Implants 
The structural and functional fusion of the surface of the dental implant with the surrounding bone (osseointegration) is crucial for the short and long term outcome of the device. In recent years, the enhancement of bone formation at the bone-implant interface has been achieved through the modulation of osteoblasts adhesion and spreading, induced by structural modifications of the implant surface, particularly at the nanoscale level. In this context, traditional chemical and physical processes find new applications to achieve the best dental implant technology. This review provides an overview of the most common manufacture techniques and the related cells-surface interactions and modulation. A Medline and a hand search were conducted to identify studies concerning nanostructuration of implant surface and their related biological interaction. In this paper, we stressed the importance of the modifications on dental implant surfaces at the nanometric level. Nowadays, there is still little evidence of the long-term benefits of nanofeatures, as the promising results achieved in vitro and in animals have still to be confirmed in humans. However, the increasing interest in nanotechnology is undoubted and more research is going to be published in the coming years.
doi:10.3390/ijms14011918
PMCID: PMC3565355  PMID: 23344062
adult stem cells; nanotechnologies; differentiation; osteogenesis; surfaces; dental implant
13.  A novel in vivo method for quantifying the interfacial biochemical bond strength of bone implants 
Quantifying the in vivo interfacial biochemical bond strength of bone implants is a biological challenge. We have developed a new and novel in vivo method to identify an interfacial biochemical bond in bone implants and to measure its bonding strength. This method, named biochemical bond measurement (BBM), involves a combination of the implant devices to measure true interfacial bond strength and surface property controls, and thus enables the contributions of mechanical interlocking and biochemical bonding to be distinguished from the measured strength values. We applied the BBM method to a rabbit model, and observed great differences in bone integration between the oxygen (control group) and magnesium (test group) plasma immersion ion-implanted titanium implants (0.046 versus 0.086 MPa, n=10, p=0.005). The biochemical bond in the test implants resulted in superior interfacial behaviour of the implants to bone: (i) close contact to approximately 2 μm thin amorphous interfacial tissue, (ii) pronounced mineralization of the interfacial tissue, (iii) rapid bone healing in contact, and (iv) strong integration to bone. The BBM method can be applied to in vivo experimental models not only to validate the presence of a biochemical bond at the bone–implant interface but also to measure the relative quantity of biochemical bond strength. The present study may provide new avenues for better understanding the role of a biochemical bond involved in the integration of bone implants.
doi:10.1098/rsif.2009.0060
PMCID: PMC2839374  PMID: 19369221
bone–implant interface; interfacial biochemical bond; bonding strength measurement; titanium; metal plasma source ion implantation; surface property
14.  Direct communication between osteocytes and acid-etched titanium implants with a sub-micron topography 
The osteocyte network, through the numerous dendritic processes of osteocytes, is responsible for sensing mechanical loading and orchestrates adaptive bone remodelling by communicating with both the osteoclasts and the osteoblasts. The osteocyte network in the vicinity of implant surfaces provides insight into the bone healing process around metallic implants. Here, we investigate whether osteocytes are able to make an intimate contact with topologically modified, but micrometre smooth (S a < 0.5 µm) implant surfaces, and if sub-micron topography alters the composition of the interfacial tissue. Screw shaped, commercially pure (cp-Ti) titanium implants with (i) machined (S a = ~0.2 µm), and (ii) two-step acid-etched (HF/HNO3 and H2SO4/HCl; S a = ~0.5 µm) surfaces were inserted in Sprague Dawley rat tibia and followed for 28 days. Both surfaces showed similar bone area, while the bone-implant contact was 73 % higher for the acid-etched surface. By resin cast etching, osteocytes were observed to maintain a direct intimate contact with the acid-etched surface. Although well mineralised, the interfacial tissue showed lower Ca/P and apatite-to-collagen ratios at the acid-etched surface, while mineral crystallinity and the carbonate-to-phosphate ratios were comparable for both implant surfaces. The interfacial tissue composition may therefore vary with changes in implant surface topography, independently of the amount of bone formed. Implant surfaces that influence bone to have higher amounts of organic matrix without affecting the crystallinity or the carbonate content of the mineral phase presumably result in a more resilient interfacial tissue, better able to resist crack development during functional loading than densely mineralised bone.
doi:10.1007/s10856-016-5779-1
PMCID: PMC5047930  PMID: 27699573
15.  Effects of nanoporous anodic titanium oxide on human adipose derived stem cells 
The aim of current bone biomaterials research is to design implants that induce controlled, guided, successful, and rapid healing. Titanium implants are widely used in dental, orthopedic, and reconstructive surgery. A series of studies has indicated that cells can respond not only to the chemical properties of the biomaterial, but also, in particular, to the changes in surface topography. Nanoporous materials remain in focus of scientific queries due to their exclusive properties and broad applications. One such material is nanostructured titanium oxide with highly ordered, mutually perpendicular nanopores. Nanoporous anodic titanium dioxide (TiO2) films were fabricated by a three-step anodization process in propan-1,2,3-triol-based electrolyte containing fluoride ions. Adipose-derived stem cells offer many interesting opportunities for regenerative medicine. The important goal of tissue engineering is to direct stem cell differentiation into a desired cell lineage. The influence of nanoporous TiO2 with pore diameters of 80 and 108 nm on cell response, growth, viability, and ability to differentiate into osteoblastic lineage of human adipose-derived progenitors was explored. Cells were harvested from the subcutaneous abdominal fat tissue by a simple, minimally invasive, and inexpensive method. Our results indicate that anodic nanostructured TiO2 is a safe and nontoxic biomaterial. In vitro studies demonstrated that the nanotopography induced and enhanced osteodifferentiation of human adipose-derived stem cells from the abdominal subcutaneous fat tissue.
doi:10.2147/IJN.S116263
PMCID: PMC5072627  PMID: 27789947
adipose-derived stem cells; anodic titanium oxide; nanotopography; osteogenic differentiation; biomaterials
16.  Numerical simulation of in vivo intraosseous torsional failure of a hollow-screw oral implant 
Head & Face Medicine  2006;2:36.
Background
Owing to the complexity and magnitude of functional forces transferred to the bone-implant interface, the mechanical strength of the interface is of great importance. The purpose of this study was to determine the intraosseous torsional shear strength of an osseointegrated oral implant using 3-D finite element (FE) stress analysis implemented by in vivo failure torque data of an implant.
Methods
A Ø 3.5 mm × 12 mm ITI® hollow screw dental implant in a patient was subjected to torque failure test using a custom-made strain-gauged manual torque wrench connected to a data acquisition system. The 3-D FE model of the implant and peri-implant circumstances was constructed. The in vivo strain data was converted to torque units (N.cm) to involve in loading definition of FE analysis. Upon processing of the FE analysis, the shear stress of peri-implant bone was evaluated to assume torsional shear stress strength of the bone-implant interface.
Results
The in vivo torque failure test yielded 5952 μstrains at custom-made manual torque wrench level and conversion of the strain data resulted in 750 N.cm. FE revealed that highest shear stress value in the trabecular bone, 121 MPa, was located at the first intimate contact with implant. Trabecular bone in contact with external surface of hollow implant body participated shear stress distribution, but not the bone resting inside of the hollow.
Conclusion
The torsional strength of hollow-screw implants is basically provided by the marginal bone and the hollow part has negligible effect on interfacial shear strength.
doi:10.1186/1746-160X-2-36
PMCID: PMC1636028  PMID: 17083739
17.  Silicon-Doped Titanium Dioxide Nanotubes Promoted Bone Formation on Titanium Implants 
While titanium (Ti) implants have been extensively used in orthopaedic and dental applications, the intrinsic bioinertness of untreated Ti surface usually results in insufficient osseointegration irrespective of the excellent biocompatibility and mechanical properties of it. In this study, we prepared surface modified Ti substrates in which silicon (Si) was doped into the titanium dioxide (TiO2) nanotubes on Ti surface using plasma immersion ion implantation (PIII) technology. Compared to TiO2 nanotubes and Ti alone, Si-doped TiO2 nanotubes significantly enhanced the expression of genes related to osteogenic differentiation, including Col-I, ALP, Runx2, OCN, and OPN, in mouse pre-osteoblastic MC3T3-E1 cells and deposition of mineral matrix. In vivo, the pull-out mechanical tests after two weeks of implantation in rat femur showed that Si-doped TiO2 nanotubes improved implant fixation strength by 18% and 54% compared to TiO2-NT and Ti implants, respectively. Together, findings from this study indicate that Si-doped TiO2 nanotubes promoted the osteogenic differentiation of osteoblastic cells and improved bone-Ti integration. Therefore, they may have considerable potential for the bioactive surface modification of Ti implants.
doi:10.3390/ijms17030292
PMCID: PMC4813156  PMID: 26927080
silicon doping; titanium dioxide nanotubes; titanium; osteogenic differentiation; MC3T3-E1 cells; osseointegration
18.  A patient with meningeal melanomatosis treated for periodontal disease with a bone regeneration procedure and dental implants: clinical and behavioral management to support medical compliance 
Oral & Implantology  2014;6(3):75-81.
SUMMARY
Background.
During the last three decades dental implants have become increasingly used in partially edentulous periodontally compromised patients. The type of bacteria in the peri-implant sulcus is influenced by the periodontal bacteria present on the surfaces of the remaining teeth. Peri-implant sulci of partially edentulous individuals harbour more motile rods and spirochetes than those of fully edentulous individuals. If Peri-implantitis arises, it may lead to implant failure. This complication occurs more frequently in patients with poor oral hygiene. This is a site-specific bacterial infection similar to that caused by periodontal bacteria around teeth and it should be prevented.
Aim.
This study was conducted to radiographically evaluate hard tissue response around 6 implants, over a 2-year period, in a previously surgically treated patient affected by severe chronic periodontitis. Psychological considerations and behavioral management of the patient are described.
Materials and methods.
A complex implant-perio-prosthodontic case of a 54-year-old man affected by meningeal melanomatosis with a history of generalized severe chronic periodontitis was recruited. A comprehensive periodontal examination around teeth was accomplished before periodontal and implant treatment. After diagnostic work-up, compromised teeth from 1.3 to 2.3 and from 3.2 to 4.2 were extracted. Tooth 1.7 was also extracted. Afterwards fixed provisional restoration rehabilitated all the natural dentition and the missing teeth. Endodonthic therapies were conducted on all the teeth due to high dentinal sensitivity and pre-prosthodontic crown reconstructions performed. Periodontal surgery with modified Widman flaps were then accomplished on all the teeth. Three months later four maxillary implants in position 1.3,1.1,2.1,2.3 and two mandibular implants in position 4.2,3.2 were inserted. During mandibular implants positioning, the mental mussels were isolated and detached to achieve proper guided bone regeneration.
During implant surgery, due to systemic conditions concern, the patient underwent intravenous sedation. Five months later the implants and the teeth were rehabilitated with fixed metal-ceramic bridges. Regarding the upper prosthetic rehabilitation, the incisors marginal edges were kept vertical to the nasal spine, due to lack of previous reference points. According to the reference points previously determined, the difference in bone level between radiographs taken at implants insertion and at the maintenance appointments was calculated.
Results.
The health of the periodontally treated teeth resulted greatly enhanced. The mean alveolar bore loss was 0,30 mm after a 2-year observation period.
Conclusions.
The control of the periodontal disease before implant insertion in patients with severe chronic periodontitis is of paramount importance, as well as a regular maintenance program is essential for the health of the periodontal and peri-implant tissues. The management of patients with complex needs requires a multidisciplinary team designed to meet all the patient’s needs on various levels.
PMCID: PMC3982304  PMID: 24772265
dental implants; bacterial colonization; bone loss; periodontally compromised patients; patient compliance
19.  Promotion of osteogenic differentiation of stem cells and increase of bone-bonding ability in vivo using urease-treated titanium coated with calcium phosphate and gelatin 
Because of its excellent biocompatibility and low allergenicity, titanium has been widely used for bone replacement and tissue engineering. To produce a desirable composite with enhanced bone response and mechanical strength, in this study bioactive calcium phosphate (CaP) and gelatin composites were coated onto titanium (Ti) via a novel urease technique. The cellular responses to the CaP/gelatin/Ti (CaP/gel/Ti) and bone bonding ability were evaluated with proliferation and osteogenic differentiation of mesenchymal stem cells (MSCs) on CaP/gel/Ti and CaP/Ti in vitro. The results showed that the optical density values, alkaline phosphatase expression and genes expression of MSCs on CaP/gel/Ti were similar to those on CaP/Ti, yet significantly higher than those on pure Ti (p < 0.05). CaP/gel/Ti and CaP/Ti rods (2 mm in diameter, 10 mm in length) were also implanted into femoral shaft of rabbits and pure Ti rods served as control (n = 10). Histological examination, scanning electron microscopy (SEM) and energy dispersive spectroscopy (EDS) measurements were performed at 4 and 8 weeks after the operation. The histological and SEM observations demonstrated clearly that more new bone formed on the surface of CaP/gel/Ti than in the other two groups at each time point. The CaP/gel/Ti bonded to the surrounding bone directly with no intervening soft tissue layer. An interfacial layer, containing Ti, Ca and P, was found to form at the interface between bone and the implant on all three groups by EDS analysis. However, the content of Ca, P in the surface of CaP/gel/Ti implants was more than in the other two groups at each time point. The CaP/gel/Ti modified by the urease method was not only beneficial for MSCs proliferation and osteogenic differentiation, but also favorable for bone bonding ability on Ti implants in vivo, suggesting that Ti functionalized with CaP and gelatin might have a great potential in clinical joint replacement or dental implants.
doi:10.1088/1468-6996/14/5/055001
PMCID: PMC5090371  PMID: 27877608
mesenchymal stem cells; hydroxyapatite (HAp); osteogenic differentiation; bone bonding
20.  Specific Biomimetic Hydroxyapatite Nanotopographies Enhance Osteoblastic Differentiation and Bone Graft Osteointegration 
Tissue Engineering. Part A  2013;19(15-16):1704-1712.
Impaired healing of cortical bone grafts represents a significant clinical problem. Cadaveric bone grafts undergo extensive chemical processing to decrease the risk of disease transmission; however, these processing techniques alter the bone surface and decrease the osteogenic potential of cells at the healing site. Extensive work has been done to optimize the surface of bone grafts, and hydroxyapatite (HAP) and nanotopography both increase osteoblastic differentiation. HAP is the main mineral component of bone and can enhance osteoblastic differentiation and bone implant healing in vivo, while nanotopography can enhance osteoblastic differentiation, adhesion, and proliferation. This is the first study to test the combined effects of HAP and nanotopographies on bone graft healing. With the goal of identifying the optimized surface features to improve bone graft healing, we tested the hypothesis that HAP-based nanotopographic resurfacing of bone grafts improves integration of cortical bone grafts by enhancing osteoblastic differentiation. Here we show that osteoblastic cells cultured on processed bones coated with specific-scale (50–60 nm) HAP nanotopographies display increased osteoblastic differentiation compared to cells on uncoated bone, bones coated with poly-l-lactic acid nanotopographies, or other HAP nanotopographies. Further, bone grafts coated with 50–60-nm HAP exhibited increased formation of new bone and improved healing, with mechanical properties equivalent to live autografts. These data indicate the potential for specific HAP nanotopographies to not only increase osteoblastic differentiation but also improve bone graft incorporation, which could significantly increase patient quality of life after traumatic bone injuries or resection of an osteosarcoma.
doi:10.1089/ten.tea.2012.0560
PMCID: PMC3700012  PMID: 23510012
21.  Decreased Staphylococcus aureus and increased osteoblast density on nanostructured electrophoretic-deposited hydroxyapatite on titanium without the use of pharmaceuticals 
Background
Plasma-spray deposition of hydroxyapatite on titanium (Ti) has proven to be a suboptimal solution to improve orthopedic-implant success rates, as demonstrated by the increasing number of orthopedic revision surgeries due to infection, implant loosening, and a myriad of other reasons. This could be in part due to the high heat involved during plasma-spray deposition, which significantly increases hydroxyapatite crystal growth into the nonbiologically inspired micron regime. There has been a push to create nanotopographies on implant surfaces to mimic the physiological nanostructure of native bone and, thus, improve osteoblast (bone-forming cell) functions and inhibit bacteria functions. Among the several techniques that have been adopted to develop nanocoatings, electrophoretic deposition (EPD) is an attractive, versatile, and effective material-processing technique.
Objective
The in vitro study reported here aimed to determine for the first time bacteria responses to hydroxyapatite coated on Ti via EPD.
Results
There were six and three times more osteoblasts on the electrophoretic-deposited hydroxyapatite on Ti compared with Ti (control) and plasma-spray-deposited hydroxyapatite on Ti after 5 days of culture, respectively. Impressively, there were 2.9 and 31.7 times less Staphylococcus aureus on electrophoretic-deposited hydroxyapatite on Ti compared with Ti (control) and plasma-spray-deposited hydroxyapatite on Ti after 18 hours of culture, respectively.
Conclusion
Compared with uncoated Ti and plasma-sprayed hydroxyapatite coated on Ti, the results provided significant promise for the use of EPD to improve bone-cell density and be used as an antibacterial coating without resorting to the use of antibiotics.
doi:10.2147/IJN.S55733
PMCID: PMC3986289  PMID: 24748789
bacteria; nanotechnology; electrophoretic deposition; inhibition
22.  Success rate of dental implants inserted in autologous bone graft regenerated areas: a systematic review 
Oral & Implantology  2012;4(3-4):3-10.
SUMMARY
Background and Aim
To assess the success rate of implants placed in atrophic ridges, regenerated by means of block bone grafts harvested from iliac crest, calvaria or intraoral donor sites (mandibular ramus, chin).
Methods and Materials
A systematic review of all prospective and retrospective studies analyzing the success rate of implants placed simultaneously or as a second surgery following ridge augmentation by means of onlay graft technique, compared with implants placed in pristine bone, was performed. To be included, studies had to involve at least five consecutively treated patients and to report clearly specified success criteria. It was also necessary a minimum follow-up period of six months, to allow the observation of potential biological complications during function, rather than early implant failures. In order to assess the success rate of implants in terms of health of periimplant tissues, implant stability, osteointegration and bone resorption, studies reporting only the survival rate of implants, were excluded.
Results
From 323 potentially relevant studies, 65 full-text publications were screened and eight were identified as fulfilling the inclusion criteria. The success rate of implants placed in onlay graft regenerated ridges ranged from 72,8% to 97% after follow-up periods ranging from 6 months to 10 years, with all the studies but two, reporting a success rate higher than 84% (range 84–97%).
Conclusion
The obtained data demonstrated that the success rate of implants placed in regenerated areas are very similar to those obtained in case of implants placed in pristine bone, and suggested that onlay graft augmentation is a quite predictable technique to allow the placement of implants in severely atrophic areas. Despite that, the current review revealed that there are not many studies providing data on the success rate of dental implants placed in onlay graft augmented ridges and demonstrated, on average, a poor methodological quality. So randomized controlled studies adopting standardized criteria to define success and failure of implants are required and data from this review must be considered indicative.
PMCID: PMC3530967  PMID: 23277867
dental implants; success rate; onlay graft; bone block; harvesting; ridge augmentation; delayed placement; immediate placement; human
23.  Development of Implant/Interconnected Porous Hydroxyapatite Complex as New Concept Graft Material 
PLoS ONE  2012;7(11):e49051.
Background
Dental implant has been successfully used to replace missing teeth. However, in some clinical situations, implant placement may be difficult because of a large bone defect. We designed novel complex biomaterial to simultaneously restore bone and place implant. This complex was incorporated implant into interconnected porous calcium hydroxyapatite (IP-CHA). We then tested this Implant/IP-CHA complex and evaluated its effect on subsequent bone regeneration and implant stability in vivo.
Methodology/Principal Findings
A cylinder-type IP-CHA was used in this study. After forming inside of the cylinder, an implant was placed inside to fabricate the Implant/IP-CHA complex. This complex was then placed into the prepared bone socket in the femur of four beagle-Labrador hybrid dogs. As a control, implants were placed directly into the femur without any bone substrate. Bone sockets were allowed to heal for 2, 3 and 6 months and implant stability quotients (ISQ) were measured. Finally, tissue blocks containing the Implant/IP-CHA complexes were harvested. Specimens were processed for histology and stained with toluidine blue and bone implant contact (BIC) was measured. The ISQs of complex groups was 77.8±2.9 in the 6-month, 72.0±5.7 in the 3-month and 47.4±11.0 in the 2-month. There was no significant difference between the 3- or 6-month complex groups and implant control groups. In the 2-month group, connective tissue, including capillary angiogenesis, was predominant around the implants, although newly formed bone could also be observed. While, in the 3 and 6-month groups, newly formed bone could be seen in contact to most of the implant surface. The BICs of complex groups was 2.18±3.77 in the 2-month, 44.03±29.58 in the 3-month, and 51.23±8.25 in the 6-month. Significant difference was detected between the 2 and 6-month.
Conclusions/Significance
Within the results of this study, the IP-CHA/implant complex might be able to achieve both bone reconstruction and implant stability.
doi:10.1371/journal.pone.0049051
PMCID: PMC3494665  PMID: 23152848
24.  Biomolecular surface coating to enhance orthopaedic tissue healing and integration 
Biomaterials  2007;28(21):3228-3235.
Implant osseointegration is a prerequisite for clinical success in orthopaedic and dental applications, many of which are restricted by loosening. Biomaterial surface modification approaches, including calcium-phosphate ceramic coatings and macro/microporosity, have had limited success in promoting integration. To improve osseointegration, titanium surfaces were coated with the GFOGER collagen-mimetic peptide, selectively promoting α2β1 integrin binding, a crucial event for osteoblastic differentiation. Titanium surfaces presenting GFOGER triggered osteoblastic differentiation and mineral deposition in bone marrow stromal cells, leading to enhanced osteoblastic function compared to unmodified titanium. Furthermore, this integrin-targeted coating significantly improved in vivo peri-implant bone regeneration and osseointegration, as characterized by bone-implant contact and mechanical fixation, compared to untreated titanium in a rat cortical bone-implant model. GFOGER-modified implants also significantly enhanced osseointegration compared to surfaces modified with full-length type I collagen, highlighting the importance of presenting specific biofunctional domains within the native ligand. In addition, this biomimetic implant coating is generated using a simple, single-step procedure that readily translates to a clinical environment with minimal processing and cytotoxicity concerns. Therefore, this study establishes a biologically active and clinically relevant implant coating strategy that enhances bone repair and orthopaedic implant integration.
doi:10.1016/j.biomaterials.2007.04.003
PMCID: PMC2034748  PMID: 17448533
biomimetic material; cell adhesion; collagen; osseointegration; integrin
25.  Is Graphene a Promising Nano-Material for Promoting Surface Modification of Implants or Scaffold Materials in Bone Tissue Engineering? 
Bone tissue engineering promises to restore bone defects that are caused by severe trauma, congenital malformations, tumors, and nonunion fractures. How to effectively promote the proliferation and osteogenic differentiation of mesenchymal stem cells (MSCs) or seed cells has become a hot topic in this field. Many researchers are studying the ways of conferring a pro-osteodifferentiation or osteoinductive capability on implants or scaffold materials, where osteogenesis of seed cells is promoted. Graphene (G) provides a new kind of coating material that may confer the pro-osteodifferentiation capability on implants and scaffold materials by surface modification. Here, we review recent studies on the effects of graphene on surface modifications of implants or scaffold materials. The ability of graphene to improve the mechanical and biological properties of implants or scaffold materials, such as nitinol and carbon nanotubes, and its ability to promote the adhesion, proliferation, and osteogenic differentiation of MSCs or osteoblasts have been demonstrated in several studies. Most previous studies were performed in vitro, but further studies will explore the mechanisms of graphene's effects on bone regeneration, its in vivo biocompatibility, its ability to promote osteodifferentiation, and its potential applications in bone tissue engineering.
doi:10.1089/ten.teb.2013.0638
PMCID: PMC4186769  PMID: 24447041

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