No evidence to indicate topographic dependency on bone formation around cp titanium implants under masticatory loading

In vitro studies have proved the topographic dependency upon osteogenesis on titanium plate by investigating the cell-adhesion, -shape, -proliferation, -differentiation, ALP activity and osteocalcin production of osteogenic stem cells, MG36, MC3T3-E1 and wild strains of bone formative cells from animal and human. However, this in vivo study on bone growth around cp titanium dental implants under masticatory loading did not demonstrate significant difference among the different surface roughness in the range of Ra 0.4–1.9 μm, Rz 2.8–11.2 μm, Rmax 3.6–28.1 μm and Sm 2.9–41.0 μm, which was estimated by measuring the bone contacts, bone occupancies and bone bonding strengths at the implant/bone marrow interface. It is revealed that the topographic dependency on the osteogenetic activity is apt to be covered with wide variation in bone healing potential under the clinical condition with functional biting load.     

Study of interface phenomena between bone and titanium and alumina surfaces in the case of monolithic and composite dental implants

The interface between mandibular bone and dental implants was examined with the in vivo dog model. Implant/bone interfaces were investigated for three types of materials: Ti–30 wt% Ta/Al2O3, titanium and Al2O3 using microscopy techniques covering a large magnification range: scanning electron microscopy, transmission electron microscopy, energy dispersive X-ray analysis and Auger spectroscopy. During the interaction of the Al2O3 ceramic with bone, an interfacial layer about 15 m thick is formed. The same phenomenon was observed at the titanium bone interface, where the thickness of the layer was about 10 m. In all cases, interface layers were sharp with well-defined borders between bone tissue and implant materials. No calcification took place inside the interface layer. A chemical analysis performed on this layer shows the presence of titanium, calcium and phosphorus in the case of titanium implants, and aluminium, calcium and phosphorus in the case of alumina implants. A rapid decrease in metal composition with increasing distances from the implant surface is correlated to a slow increase in calcium and phosphorus in the direction of the bone. Direct contact between implant and bone was observed. No biocorrosive effects were detected at the Ti–30 wt% Ta/Al2O3 metal–ceramic interface.     

Nanoscale characterization of bone–implant interface and biomechanical modulation of bone ingrowth

Bone–implant interface is characterized by an array of cells and macromolecules. This study investigated the nanomechancial properties of bone–implant interface using atomic force microscopy in vitro, and the mechanical modulation of implant bone ingrowth in vivo using bone histomorphometry. Upon harvest of screw-type titanium implants placed in vivo in the rabbit maxilla and proximal femur for 4 weeks, nanoindentation was performed in the bone–implant interface at 60-μm intervals radially from the implant surface. The average Young's Moduli (E) of the maxillary bone–implant interface was 1.13 ± 0.27 MPa, lacking significant differences at all intervals. In contrast, an increasing gradient of E was observed radially from the femur bone–implant interface: 0.87 ± 0.25 MPa to 2.24 ± 0.69 MPa, representing significant differences among several 60-μm intervals. In a separate experiment, bone healing was allowed for 6 weeks for proximal femur implants. The right femoral implant received axial cyclic loading at 200 mN and 1 Hz for 10 min/d over 12 days, whereas the left femoral implant served as control. Cyclic loading induced significantly higher bone volume, osteoblast numbers per endocortical bone surface, mineral apposition rate, and bone formation rate than controls. These data demonstrate nanoscale and microscale characterizations of bone–implant interface, and mechanical modulation of bone ingrowth surrounding titanium implants.     

Early tissue response to titanium implants inserted in rabbit cortical bone

The tissue response to screw-shaped implants of commercially pure titanium was studied 3–180 days after insertion in the rabbit tibia by means of transmission electron microscopy. Red blood cells and scattered macrophages predominated at the implant surface after 3 days. At day 7 and later intervals, multinuclear giant cells were the cell type found at the implant surface protruding into the bone marrow and in areas with no bone-titanium contact. Osteoblasts or mesenchymal cells were rarely seen at the implant surface at any time period. Two modes of mineralization could be distinguished in the interface. Firstly, the typical mineralization of osteoid seams produced by osteoblasts. Secondly, an accumulation of scattered hydroxyapatite crystals in the unmineralized collagen matrix in the interface. Mineralized tissue was observed close to the implants surface from day 14. However, the innermost 2–20 m were poorly mineralized although scattered hydroxyapatite crystals were present. The collagen fibrils did not reach the implant surface but were separated from it by an amorphous layer, being 0.3–0.5 m thick which did not decrease in width with time. An electron-dense lamina limitans-like line containing mineral was observed between the amorphous layer and the bone tissue.     

Tensile force testing of optimized coin-shaped titanium implant attachment kinetics in the rabbit tibiae

In the present study, the bone response of titanium implants at early bone healing stages, was evaluated using a tensile test. Test surface of coin-shaped cp. titanium implants were standardized by grit blasting with TiO₂, grain size 180–220 m. The surface topography of the implant specimens was examined by SEM, and by a confocal laser scanner for evaluation of S _a, S _t and S _dr. The implants were placed onto the leveled site on the tibia of 12 New Zealand White rabbits, 4 implants in each animal. The rabbits were divided into three groups with different observation times i.e. 2, 4 and 6 weeks. The retention of 12 implants were tested by measuring the pull-out force needed to detach the implant from the bone. There was a significant increase in implant retention from 2 to 4 and to 6 weeks healing time (p<0.05). Four implants from each time point were randomly chosen for histological evaluation. The histological appearance of the implant–bone interface at the different healing times showed noticeable differences in the degree of bone healing and maturation, suggesting that, in rabbits, 6 weeks healing time is a suitable observation point for tensile testing of surface optimized osseointegrating implants.     

Monte Carlo simulation of trabecular bone remodelling and absorbed dose coefficients for tritium and 14C

A Monte Carlo simulation of multiple trabecular bone cavities in adult bone was developed and the absorbed radiation dose factors evaluated for 3H and 14C. The model was developed to assess the dose from radionuclide uptake in quiescent bone, but also the effects of temporal changes in bone turnover by incorporating bone-modelling units (BMU). Absorbed dose fractions were calculated for target regions that include the endosteal layer where radiation-sensitive stem cells in bone marrow are considered to reside preferentially. There were large differences in the absorbed fractions for two types of bone surface, quiescent and forming. Tritium in quiescent bone results in a dose to the endosteum about 20 times that for the same activity in forming bone surface irradiating osteoblasts. When the quiescent bone surface source was extended from an infinitely thin layer to a more realistic 1 microm thick, the tritium absorbed fractions for endosteum and red marrow targets fell by more than 2-fold.     

In vitro and in vivo evaluation of the effects of demineralized bone matrix or calcium sulfate addition to polycaprolactone–bioglass composites

The objective of this study was to improve the efficacy of polycaprolactone/bioglass (PCL/BG) bone substitute using demineralized bone matrix (DBM) or calcium sulfate (CS) as a third component. Composite discs involving either DBM or CS were prepared by compression moulding. Bioactivity of discs was evaluated by energy dispersive X-ray spectroscopy (ESCA) and scanning electron microscopy (SEM) following simulated body fluid incubation. The closest Calcium/Phosphate ratio to that of hydroxyl carbonate apatite crystals was observed for PCL/BG/DBM group (1.53) after 15 day incubation. Addition of fillers increased microhardness and compressive modulus of discs. However, after 4 and 6-week PBS incubations, PCL/BG/DBM discs showed significant decrease in modulus (from 266.23 to 54.04 and 33.45 MPa, respectively) in parallel with its highest water uptakes (36.3 and 34.7%). Discs preserved their integrity with only considerable weight loss (7.5–14.5%) in PCL/BG/DBM group. In vitro cytotoxicity tests showed that all discs were biocompatible. Composites were implanted to defects on rabbit humeri. After 7 weeks, new tissue formation and mineralization at bone-implant interface were observed for all implants. Bone mineral densities at interface were higher than that of implant site and negative controls (defects left empty) but lower than healthy bone level. However, microhardness of implant sites was higher than in vitro results indicating in vivo mineralization of implants. Addition of DBM or CS resulted with higher microhardness values at interface region (ca. 650 μm from implant) compared to PCL/BG and negative control. Histological studies revealed that addition of DBM enhanced bone formation around and into implant while CS provided cartilage tissue formation around the implant. From these results, addition of DBM or CS could be suggested to improve bone healing efficacy of PCL/BG composites.     

Mechanical and histological evaluations of cobalt-chromium alloy and hydroxyapatite plasma-sprayed coatings in bone

This study evaluated the mechanical and histological behavior of cobalt-chromium (CoCr) alloy and hydroxyapatite (HA) plasma-sprayed coatings in canine cortical bone after 6 and 12 weeks of implantation, using CoCr alloy as the substrate. the substrate was bond-coated with microtextured CoCr alloy coating to ensure adherence between the substrate and top coats. A macrotextured CoCr alloy top coat with surface roughness R _a=34.25±5.50 m was produced to create suitable pores ranging from 25 m to 200 m for bone ingrowth. For HA top coat, a relatively smooth surface (R _a=15.14±3.21 m) was prepared for bone apposition. Shear testing of bone/implant interfaces showed that the CoCr alloy top coat exhibited significantly lower (p<0.01) mean shear strength than the HA top coat at each time interval. The maximum shear strength was 10.88±0.38 MPa for HA-coated implants 12 weeks post-implantation. After histological evaluations, substantial differences in the extent of new bone formation and the types of implant/bone contact were found between two kinds of implants. Direct bone-to-HA coating contact was consistently observed, while a layer of fibrous tissue intervening at the bone-CoCr alloy coating interface was found. Occasionally, partial dissolution of HA coating was seen after 12 weeks of implantation. The results of this study suggested that plasma-sprayed macrotextured CoCr coatings may not be an effective alternative for biological fixation.     

Voxel-based models representing the male and female ICRP reference adult--the skeleton

For the forthcoming update of organ dose conversion coefficients, the International Commission on Radiological Protection (ICRP) will use voxel-based computational phantoms due to their improved anatomical realism compared with the class of mathematical or stylized phantoms used previously. According to the ICRP philosophy, these phantoms should be representative of the male and female reference adults with respect to their external dimensions, their organ topology and their organ masses. To meet these requirements, reference models of an adult male and adult female have been constructed at the GSF, based on existing voxel models segmented from tomographic images of two individuals whose body height and weight closely resemble the ICRP Publication 89 reference values. The skeleton is a highly complex structure of the body, composed of cortical bone, trabecular bone, red and yellow bone marrow and endosteum ('bone surfaces' in their older terminology). The skeleton of the reference phantoms consists of 19 individually segmented bones and bone groups. Sub-division of these bones into the above-mentioned constituents would be necessary in order to allow a direct calculation of dose to red bone marrow and endosteum. However, the dimensions of the trabeculae, the cavities containing bone marrow and the endosteum layer lining these cavities are clearly smaller than the resolution of a normal CT scan and, thus, these volumes could not be segmented in the tomographic images. As an attempt to represent the gross spatial distribution of these regions as realistically as possible at the given voxel resolution, 48 individual organ identification numbers were assigned to various parts of the skeleton: every segmented bone was subdivided into an outer shell of cortical bone and a spongious core; in the shafts of the long bones, a medullary cavity was additionally segmented. Using the data from ICRP Publication 89 on elemental tissue composition, from ICRU Report 46 on material mass densities, and from ICRP Publication 70 on the distribution of the red bone marrow among and marrow cellularity in individual bones, individual elemental compositions for these segmented bone regions were derived. Thus, most of the relevant source and target regions of the skeleton were provided. Dose calculations using these regions will be based on fluence-to-dose response functions that are multiplied with the particle fluence inside specific bone regions to give the dose quantities of interest to the target tissues.     

Severe Cortical and Trabecular Osteopenia in Secondary Hyperparathyroidism

Background: Peripheral quantitative computed tomography (pQCT) provides real volumetric bone density values, not only of the total, but also of trabecular and cortical bone, separately. In addition, it provides data on bone geometry that can be related to the risk of fracture. Methods: Total, cortical, and trabecular volumetric bone mineral densities (BMD), as well as the main geometric parameters (cross‐sectional area, cortical area, trabecular area, and cortical thickness) were assessed by pQCT at the distal radius in 24 hemodialysis patients affected by severe secondary hyperparathyroidism (PTH, mean ± SD: 1444 ± 695 pg/mL). The strength‐strain index (SSI), a biomechanical parameter describing bone fragility, was also determined. Results: Compared with a control group of 64 healthy age‐matched subjects, volumetric BMD (mg/cm³) was significantly reduced in all patients (total BMD: 243 ± 87 vs. 405 ± 138, cortical BMD: 605 ± 218 vs. 856 ± 204, trabecular BMD: 95 ± 51 vs. 182 ± 75). Cortical area and cortical thickness showed significant modifications, while cross‐sectional area did not. SSI was significantly reduced (547 ± 125 vs. 927 ± 306 mm³). PTH levels showed a significant inverse correlation with cortical BMD (r = ?0.56), cortical thickness (r = ?0.46), cortical area (r = ?0.61), and SSI (r = ?0.54). Quantitative analysis of bone demonstrated cortical porosity. Conclusions: In dialysis patients with severe secondary hyperparathyroidism, pQCT showed a significant cortical osteopenia, associated with geometric and mechanical bone impairment. Interestingly, we also found a comparable deficit of trabecular bone, which may be related to the very high PTH levels. Generalized cortical thinning, intracortical porosity and cortical‐endosteal resorption (“trabecularization” of the cortical bone) are major determinants of reduced bone strength, which may be quantitated by pQCT.     

The effect of bone ingrowth depth on the tensile and shear strength of the implant–bone e-beam produced interface

New technologies, such as selective electron beam melting, allow to create complex interface structures to enhance bone ingrowth in cementless implants. The efficacy of such structures can be tested in animal experiments. Although animal studies provide insight into the biological response of new structures, it remains unclear how ingrowth depth is related to interface strength. Theoretically, there could be a threshold of ingrowth, above which the interface strength does not further increase. To test the relationship between depth and strength we performed a finite element study on micro models with simulated uncoated and hydroxyapatite (HA) coated surfaces. We examined whether complete ingrowth is necessary to obtain a maximal interface strength. An increase in bone ingrowth depth did not always enhance the bone–implant interface strength. For the uncoated specimens a plateau was reached at 1,500 μm of ingrowth depth. For the specimens with a simulated HA coating, a bone ingrowth depth of 500 μm already yielded a substantial interface strength, and deeper ingrowth did not enhance the interface strength considerably. These findings may assist in optimizing interface morphology (its depth) and in judging the effect of bone ingrowth depth on interface strength.     

<Emphasis Type="Italic">In vitro</Emphasis> study on bone formation and surface topography from the standpoint of biomechanics

Effect of surface topography upon cell-adhesion, -orientation and -differentiation was investigated by in vitro study on cellular responses to titanium substratum with different surface roughness. Cell-shape, -function and -differentiation depending upon the surface topography were clarified by use of bone formative group cells (BFGCs) derived from bone marrow of beagles femur. BFGCs consisted of hematopoietic stem cells (HSC) and osteogenetic stem cells (OSC). Cell differentiation of BFGCs was expressed and promoted by structural changes of cytoskeleton, and cell-organella, which was caused by mechanical stress with cytoplasmic stretching of cell adhesions to the substratum. Phagocytic monocytes of HSC differentiated to osteomediator cells (OMC) by cytoplasmic stretching with cell adhesion to the substratum. The OMC mediated and promoted cell differentiation from OSC to osteoblast through osteoblastic phenotype cell (OBC) by cell-aggregation of nodules with pile up phenomenon of OBC onto OMC. The osteogenesis might be performed by coupling work of both cells, OMC originated from monocyte of HSC and OBC originated from OSC, which were explained by SEM, TEM and fluorescent probe investigation on BFGCs on the test plate of cp titanium plates with different topographies. This osteogenetic process was proved by investigating cell proliferation, DNA contents, cell-adhesion, alkaline phosphatase activity and osteocalcine productivity for cells on the titanium plates with different topographies. The study showed increased osteogenic effects for cells cultured on Ti with increased surface roughness. Possible mechanisms were discussed from a biomechanical perspective.     

Ultrastructure of the bone-titanium interface in rabbits

The interface zone between cortical bone and threaded non-alloyed titanium implants inserted in the rabbit tibia for 12 months was examined by light and electron microscopy. The implants were removeden bloc with the perfusion-fixed surrounding bone and the undecalcified specimens were, after osmification, dehydrated and embedded in plastic resin (LR White). In ground sections (about 10 µm thick) cortical bone appeared to be in direct contact with the implant surface and the implants were thus osseointegrated. Sections for light microscopy (1 µm thick) and electron microscopy (40 nm to 0.5 µm) were prepared by using an electropolishing technique by which the bulk part of the metal was electrochemically removed and a fracture technique by which the implant was separated from the embedded tissue before sectioning. In the electropolished specimens an unmineralized zone, 2–10 µm wide, was observed at the interface. The interface zone contained osteoid-like tissue (densely packed collagen fibrils, osteocyte canaliculi) but in general no deposits of calcium mineral. This feature of the interface could not be observed in specimens prepared by the fracture technique, indicating that the electropolishing technique had induced serious artefacts, including decalcification of the interface bone. In sections prepared by the fracture technique, mineralized bone was present very close to the implant surface. No gradient of mineral was observed. A thin layer of amorphous material (100–200 nm wide) was present peripheral to the mineralized bone. An electron dense line about 100 nm wide was formed at the border between the mineralized bone and the amorphous layer. The dense layer had the same characteristics as the lamina limitans observed around osteocyte lacunae and canaliculi or the zone between areas of bone with different degree of mineralization.Our observations suggest that mineralized bone reached close to the surface of titanium implants inserted in the rabbit tibia for 12 months but that a direct contact is not established.     

Tissue response to hafnium

The aim of the present experimental study was to evaluate the tissue response to hafnium (Hf) a reactive metal closely related to titanium (Ti) and zirconium (Zr). Hf has not been previously evaluated as implant material in a biologic environment. In a first experiment, 21 machined Hf non-threaded implants (test) and 21 similar Ti implants (control) were inserted in the abdominal wall of 21 rats. Animals were sacrificed after 8 days (6 rats), 6 (7 rats) and 12 weeks (8 rats). In a second experiment, 18 rabbits received 18 Hf and 18 Ti threaded implants in their tibiae, one implant in each tibia. The rabbits were sacrificed after 6, 12 and 24 weeks (6 animals/time interval). The bulk metal of the abdominal wall implants, embedded together with the surrounding tissue, was electrolytically dissolved and semithin (1 m) sections of the intact tissue–implant interface were evaluated by light microscopy (morphometry). Bone-implant contact and bone area within threads were evaluated in ground sections. In soft tissues, a fluid space containing predominantly monocytes/macrophages surrounded the abdominal implants at 8 days. At 6 and 12 weeks, a fibrous capsule, consisting of layers of macrophages and fibroblasts, surrounded the implants. Macrophages, including multinuclear giant cells, always formed the innermost layer in contact with the implant surface. No quantitative or qualitative difference in the tissue organization was detected between Ti and Hf implants. In rabbits, 6 weeks after insertion, the proximal two threads located within the cortical bone were filled with bone in contact with Hf and Ti. The distal threads contained bone marrow. After 12 and 24 weeks, mature bone was present in the proximal 3–4 implant threads. No statistically significant difference was found between Hf and Ti implants at any time periods. It is concluded that Hf is an interesting metal for biomedical applications in bone and soft tissue. © 2001 Kluwer Academic Publishers     

Early tissue response to titanium implants inserted in rabbit cortical bone

The tissue response to screw-shaped implants of commercially pure titanium was studied by light microscopy 3–180 days after insertion in the rabbit tibia. The implant site in the tibial metaphysis consisted mainly of cortical bone. Three days after implantation, osteoblasts, producing osteoid, were observed at the endosteal surface and elongated mesenchymal cells were present in the injury area. Some macrophages but rather few other inflammatory cells were identified. Multinuclear giant cells were in direct contact with the implant and formed an almost continuous layer along the surface from the 7th day. The number of giant cells decreased with time and with increased bone-titanium contact. Bone formation was never seen direct on the implant surface but was first observed at day 7 as a woven trabecular bone formed at the endosteal surface and extending towards the implant and as a solitary formation of woven bone close to the implant. The solitary bone matrix served as a base for surface osteoblasts which produced osteoid in a lamellar arrangement. With time the two types of newly formed bone fused and more bone filled the threads and became remodelled by bone remodelling units. Light microscopic morphometry in ground sections demonstrated that the bone/titanium contact and bone area in the threads increased with time up to 6 months after implantation     

Physicochemical characterisation of novel ultra-thin biodegradable scaffolds for peripheral nerve repair

In this study, the physicochemical properties of microporous poly (ε-caprolactone) (PCL) films and a composite material made of PCL and polylactic acid (PLA) blend were tested. Fabricated by solvent casting using dichloromethane, these ultra-thin films (60 ± 5 μm in thickness) have a novel double-sided surface topography, i.e. a porous surface with pores 1–10 μm in diameter and a relatively smooth surface with nano-scaled texture. Porous surfaces were found to be associated with increased protein adsorption and the treatment of these polyester scaffolds with NaOH rendered them more hydrophilic. Differential Scanning Calorimetry (DSC) showed that the incorporation of PLA reduced the crystallinity of the original homopolymer. Chemical changes were investigated by means of Fourier Transform Infrared Spectroscopy (FTIR) and X-ray photoelectron spectroscopy (XPS). Average surface roughness (Ra), hydrophilicity/hydrophobicity and mechanical properties of these materials were also assessed for the suitability of these materials as nerve conduits.     

Effect of biological implant surface coatings on bone formation, applying collagen, proteoglycans, glycosaminoglycans and growth factors

Objectives The aim of the present study was to evaluate six different implant surface coatings with respect to bone formation. Being major structural components of the extracellular matrix, collagen, the non-collagenous components decorin/chondroitin sulphate (CS) and the growth factors TGF-β1/BMP-4 served in different combinations as coatings of experimental titanium implants. Materials and methods Eight miniature pigs received each six implants in the mandible. The implant design showed two circular recesses along the length axis. Three, four, five and six weeks after implant placement, the animals were sacrificed in groups of two. Bone-implant contact (BIC) was evaluated along the outer implant surface and within the recesses. Bone volume was determined by synchrotron radiation micro computed tomography (SRμCT) for one implant of each surface state, 6 weeks after placement. Results At each week of observation, collagen/CS or collagen/CS/BMP-4 coated implants showed the highest BIC of all surface states. This was statistically significant at week five (p = 0.030, p = 0.040) and six (p = 0.025, p = 0.005). SRμCT measurements determined the highest bone volume for a collagen/CS coated implant. Conclusion The results indicate that collagen/CS and collagen/CS/BMP-4 lead to a higher degree of bone formation compared to other ECM components.     

Characterization of microblasted and reactive ion etched surfaces on the commercially pure metals niobium, tantalum and titanium

In surface-roughened metallic implant materials, the topography, chemistry and energy of the surfaces play an important role for the cell and tissue attachment. The highly reactive commercially pure metals niobium, tantalum and titanium were analysed after microblasting (with Al2O3 powder and consecutive shot-peening with ZrSiO2), and after additional reactive ion etching (RIE, with CF4). Scanning electron microscopy in combination with energy-dispersive X-ray analysis and surface roughness measurements showed, for all microblasted surfaces, a heterogeneous roughening (Ra about 0.7 m), and a contamination with blasting particles. RIE resulted in a further roughening (Ra about 1.1 m), and a total cleaning from contaminations, except for traces of aluminium. Determination of surface energy by dynamic contact angle measurements showed an increase in surface energy after microblasting, which further increased after RIE, most pronounced for commercially pure niobium. In conjunction with superior electrochemical properties, this makes niobium and tantalum promising candidates for implant purposes, at least equal to the generally used titanium.     

Applying a calcium phosphate layer on PEO/PBT copolymers affects bone formation in vivo

Recently a copolymer (Polyactive^R) has been introduced that combines elastomeric and bone-bonding properties. Since calcification of the copolymer is a prerequisite for bone bonding. Polyactive was precalcified in vitro in order to increase the bone-bonding rate. Precalcification was performed by subsequent incubation in Ca and P solutions and resulted in formation of a hydroxyapatite layer on the surface of the implant. Within one week after implantation this layer had disappeared from the surface and a new calcification zone was formed under the surface of the copolymer. Longer implantation periods showed that in precalcified implants bone was apposited along the walls of the pores, while in control implants new bone was first formed in the centre of the pores. Consequently, the percentage of bone contact was increased in precalcified implants, however, the amount of bone ingrowth was equal in both control and precalcified implants. Transmission electron microscopy showed the presence of an electron-dense layer at the bone implant interface, which was indicative for bone-bonding. It is concluded from these experiments that precalcification of PEO/PBT copolymers affected the direction of bone apposition and increased the bone-bonding rate.