Platelet‐rich fibrin and collagen membrane in the preservation of the alveolar bone: Feasibility of the elemental inorganic composition and scanning electron microscopy analysis

The success of dental implants is related to the amount, quality, and composition of the alveolar bone. The placement of platelet‐rich fibrin (PRF) clot associated with a resorbable collagen membrane (RCM) in a postextraction alveolus is a technique used for ridge preservation. This case report study analyzed the ultrastructural characteristics of cross‐sectioned alveolar bone that received PRF and RCM using scanning electron microscopy and the inorganic composition using “energy dispersive X‐ray spectrometry,” in order to explore the feasibility of this method to clinical studies. Three alveolar bone samples from two male patients (37 and 58 years old), obtained in the procedure of placing the dental implant, were analyzed. Two bone samples previously received PRF and RCM (M37 and M58), the third sample represented a physiological bone formation without treatment (M37‐control). The bone sample M37 showed irregularly shaped islets of calcified material intermingled with connective tissue. The other samples, from the 58‐year‐old patient with PRF and RCM (M58); and the other untreated bone sample from the same 37‐year‐old patient (M37‐control) showed similar ultrastructural morphology with trabecular conformation without islets agglomerations. The inorganic composition analysis showed higher concentrations of calcium and phosphorus in both samples treated with PRF and RCM in comparison to the untreated bone sample. The Ca/P ratio was higher in the M37 sample compared to the others samples. The results showed morphology and inorganic composition differences among the treatments used, suggesting that this method is feasible to analyze parameters of the alveolar bone tissue.     

Histochemical evidence of osteoclastic degradation of extracellular matrix in osteolytic metastasis originating from human lung small carcinoma (SBC-5) cells

The aim of this study was to assess the dynamics of osteoclast migration and the degradation of unmineralized extracellular matrix in an osteolytic metastasis by examining a well-standardized lung cancer metastasis model of nude mice. SBC-5 human lung small carcinoma cells were injected into the left cardiac ventricle of 6-week-old BALB/c nu/nu mice under anesthesia. At 25-30 days after injection, the animals were sacrificed and their femora and/or tibiae were removed for histochemical analyses. Metastatic lesions were shown to occupy a considerable area extending from the metaphyses to the bone marrow region. Tartrate resistant acid phosphatase (TRAPase)-positive osteoclasts were found in association with an alkaline phosphatase (ALPase)-positive osteoblastic layer lining the bone surface, but could also be localized in the ALPase-negative stromal tissues that border the tumor nodules. These stromal tissues were markedly positive for osteopontin, and contained a significant number of TRAPase-positive osteoclasts expressing immunoreactivity for CD44. We thus speculated that, mediating its affinity for CD44, osteopontin may serve to facilitate osteoclastic migration after their formation associated with ALPase-positive osteoblasts. We next examined the localization of cathepsin K and matrix metallo-proteinase-9 (MMP-9) in osteoclasts. Osteoclasts adjacent to the bone surfaces were positive for both proteins, whereas those in the stromal tissues in the tumor nests showed only MMP-9 immunoreactivity. Immunoelectron microscopy disclosed the presence of MMP-9 in the Golgi apparatus and in vesicular structures at the baso-lateral cytoplasmic region of the osteoclasts found in the stromal tissue. MMP-9-positive vesicular structures also contained fragmented extracellular materials. Thus, osteoclasts appear to either select an optimized function, namely secreting proteolytic enzymes from ruffled borders during bone resorption, or recognize the surrounding extracellular matrix by mediating osteopontin/CD44 interaction, and internalize the extracellular matrices. Microsc.     

High dose glucocorticoid hampers bone formation and resorption after bone marrow ablation in rat

We analyzed the effect of glucocorticoid on bone regeneration after bone marrow ablation in tibiae of 8-week-old rats. Methylprednisolone sodium succinate (MPSS) was injected intramuscularly at a dose of 100 mg/kg/day for 3 days. Tibiae on days 1, 3, 5, 7, 10, 12, and 14 after ablation were subjected to tartrate-resistant acid phosphatase staining, immunohistochemistry, in situ hybridization, and transmission electron microscopy (TEM), and measurement of the volume of newly-formed bone and the osteoclast number. MPSS significantly decreased the newly-formed bone volume on day 7, and immature bone still remained on day 10 in the MPSS-treated group. The volume of this bone was significantly higher than that in the control group. However, there were no differences between the groups in the osteoclast number, the expression of mRNAs for osteoblast differentiation markers, and alkaline phosphatase and cathepsin K judged by immunohistochemistry. TEM findings showed no difference in the form of osteoblasts, whereas osteoclasts in the MPSS-treated group had less developed ruffled borders, compared to those in the control group. These results suggest that MPSS treatment affects neither the differentiation nor the shape of osteoblasts, and does not change the osteoclast number or the cathepsin K level. However, high dose MPSS inhibits both bone formation and resorption during bone regeneration after rat tibial bone marrow ablation, and inhibits ruffled border formation in osteoclasts. These data will be useful to develop bone regenerative therapies for bone diseases due to high dose steroid administration.     

A novel closed-loop electromechanical stimulator to enhance osseointegration with immediate loading of dental implant restorations

The degree of osseomechanical integration of dental implants is acutely sensitive to their mechanical environment. Bone, both as a tissue and structure, adapts its mass and architecture in response to loading conditions. Therefore, application of predefined controlled loads may be considered as a treatment option to promote early maturation of bone/implant interface prior to or in conjunction with crown/prosthesis attachment. Although many studies have established that the magnitude, rate of the applied strain, and frequency have significant effects on the osteogenic response, the actual specific relationships between strain parameters and frequency have not yet been fully defined. The purpose of this study was to develop a stimulator to apply defined mechanical stimuli to individual dental implants in vivo immediately after implantation, exploring the hypothesis that immediate controlled loading could enhance implant integration. An electromechanical device was developed, based on load values obtained using a two-dimensional finite element analysis of the bone/implant interface generating 1000 to 4000 pe and operated at 30 and 3 Hz respectively. The device was then tested in a cadaveric pig mandible, and periosteal bone surface strains were recorded for potential future comparison with a three-dimensional finite element model to determine loading regimens to optimize interface strains and iterate the device for clinical use.     

Finite element analysis of bone stress and strain around a distal osseointegrated implant for prosthetic limb attachment

Direct skeletal attachment techniques have recently been identified as an alternative method for percutaneous attachment of prosthetic limbs. Osseointegrated implants for prosthetic attachments are subjected to a complex load condition. This finite element study investigates the effect of varying geometries of the implant on the stress and strain distribution in the area of the bone/implant interface. Simplified three-dimensional axisymmetric models of the femur and seven implants with different core diameters, external diameters, implant lengths and thread pitches were derived. The resulting stress and strain distributions were compared. The significance of each implant geometry was identified for improving implant design in the light of benefit to the bone/implant osseointegration and the bone remodelling of the femur.     

In vivo response to HA-polyhydroxybutyrate/polyhydroxyvalerate composite

This study examined the morphological and compositional structure of bone–implant interfaces after in vivo implantation into the tibias of rabbits. The implants were composed of biodegradable polyhydroxybutyrate/polyhydroxyvalerate copolymer reinforced with synthetic hydroxyapatite (HA) particles. Optical and scanning electron microscopy techniques were used, including energy‐dispersive X‐ray analysis. The interface was found to be morphologically, biologically and chemically active throughout the period of study. There was a strong tendency to rebuild the bone structure at the interface after implantation, independent of the composition of the implant, but direct bone bonding with the implant depended on the bioactive nature of the interface, as represented by the HA particles. At all implantation times, lamellar bone formed at the interface and replaced degrading polymer matrix, while engulfing HA filler particles. In regions about 50–100 from the interface, the bone region displayed an osteon organization. Osteoblasts and osteocytes were identified throughout the interface region. The thickness of the newly formed bone significantly increased over the period of the experiment from about 130 µm at 1 month to about 770 µm at 6 months. Materials that behave in this manner may be useful in some bone replacement therapies.     

Measurement of temperature change during the implant site preparation to determine influence of tool characteristics

Implant site preparation procedure is the most important factor that affects early osseointegration performance of a dental implant. During the side preparation procedure increase in the bone temperature above critic limit causes irreversible osteonecrosis. This heat rise compromises implant area around implants thus ending with unsuccessful osseointegration outcomes.In this experimental study drill tip geometry, drill tip angle and drill sharpness affects on procedure temperature were investigated. Experiments were carried on fresh bovine bones and implant sites were prepared individually for each experimental set.     

Numerical simulation of bone remodelling around dental implants

Crestal bone loss can result in the failure of dental implants and can be caused, by among other factors, the development of non-physiological mechanical conditions. Bone remodelling (BR) is the physiological process through which bone adapts itself to the mechanical environment. A previously published mathematical model of BR is used in this work to study the homogenized structural evolution of peri-implant bone. This model is used to study the influence of the diameter and length of a dental implant of pure titanium on its long-term stability. The temporal evolution of porosity and microstructural damage of the peri-implant bone are the variables analysed in this study. The results show that damage and porosity increase as the implant length decreases and, more pronouncedly, as its diameter decreases. The increase in damage and porosity levels is localized, as many other studies confirm, at the implant neck due to the stress concentration that is created in that area. The main conclusion of this study is that in implants with a diameter equal to or greater than 3 mm the damage is under control and there is no mechanical failure of the peri-implant bone in the long term.     

Feasibility of detecting trabecular bone around percutaneous titanium implants in rabbits by in vivo microfocus computed tomography

Objectives: The goal of this study was to examine the feasibility of in vivo imaging of trabecular bone around titanium implants by means of microfocus computed tomography (micro‐CT) and the use of rabbits for this purpose. Materials and Methods: Ten male rabbits type Hollander, received a titanium implant (1.7 mm diameter and 10 mm length) in the trabecular bone of the left tibia. Seven weeks later a micro‐CT scan was taken. Four rabbits were used to monitor potential harmful effects from X‐ray absorption until 4 weeks after scanning. A second group of six rabbits was used for testing the hypothesis that a good correlation exists between in vivo micro‐CT images and histological images of trabecular bone around titanium implants. The six rabbits were scanned and sacrificed immediately. The tibias were extracted and submitted to standard histological procedures. This resulted in a total of 12 histological sections and their corresponding 12 micro‐CT images. Bone area measurements were performed at the left and right side of the implant in three regions: 0–500, 500–1000 and 1000–1500 μm distance from the implant interface. Intra‐class correlations (ICC) were calculated between both techniques. Results: The four rabbits did not show any sign of radiodermatitis 4 weeks after scanning. In the micro‐CT images of the group of six rabbits, trabeculae are visible, but not well defined, due to the presence of noise in the image. The ICC for the right implant side were 0.44 for zone 0–500 μm, 0.48 for zone 500–1000 μm and 0.40 for zone 1000–1500 μm. The ICC for the left implant side could not be calculated. Conclusion: A low agreement was found between the bone measurements from histology and in vivo micro‐CT images. The use of the in vivo micro‐CT for trabecular bone imaging around metallic implants should be restricted to track tendencies in follow‐up studies.     

Fabrication and in vivo evaluation of Ti6Al4V implants with controlled porous structure and complex shape

Electron beam melting process was used to fabricate porous Ti6Al4V implants. The porous structure and surface topography of the implants were characterized by scanning electron microscopy (SEM) and digital microscopy (DM). The results showed that the pore size was around 600 and the porosity approximated to 57%. There was about±50 μm of undulation on implants surfaces. Standard implants and a custom implant coupled with porous sections were designed and fabricated to validate the versatility of the electron beam melting (EBM) technique. After coated with bone-like apatite, samples with fully porous structures were implanted into cranial defects in rabbits to investigate the in vivo performance. The animals were sacrificed at 8 and 12 weeks after implantation. Bone ingrowth into porous structure was examined by histological analysis. The histological sections indicated that a large amount of new bone formation was observed in porous structure. The newly formed bone grew from the calvarial margins toward the center of the bone defect and was in close contact with implant surfaces. The results of the study showed that the EBM produced Ti6Al4V implants with well-controlled porous structure, rough surface topography and bone-like apatite layer are beneficial for bone ingrowth and apposition.     

In vitro assessment of bone-implant interface using an acoustic emission transmission test

The aim of this in vitro study was to determine the feasibility of monitoring the primary stability of dental implants using a simple transmission test with acoustic emission. Forty screw-shaped titanium dental implants were installed in the middle of 10 fresh bovine ribs obtained from different animals. The implants were divided into two size groups, 8.5 mm x 3.5 mm and 13 mm x 4.5 mm, and were inserted in either tight- or loose-fitting conditions. For each implant, pulses of acoustic energy were injected at the centre of a customised gold abutment 3 mm in height using a standard pencil lead break source (Hsu-Nielsen source). A total of 30 acoustic emission recordings were made for each implant in which the transmitted energy was measured on the surface of the bone using an acoustic sensor mounted at the middle of the rib. The transmitted acoustic energy for the implants under tight-fitting conditions was significantly higher than for the loose-fitting for both sizes of implant. The acoustic emission energy values for the 13 mm implants were also higher than for the 8.5 mm implants. The results indicate that implants with good primary stability (tight-fitting) had higher acoustic emission energy than implants where primary stability was poor (loose-fitting). The longer and wider implants produced higher acoustic emission energy than shorter and narrower implants. Together, the findings suggest that a simple transmission test, properly calibrated, should be able to assess the quality of the contact between the implant and the bone in the clinical situation.     

Effects of material properties and hole designs of the jig on the fatigue life of dental implants

A purpose of this paper is to analyze the effects of material properties and hole designs of the jig on fatigue test results of dental implants. An implant fatigue test method is described in ISO14801, which requires that the jig should firmly hold the implant and the elastic modulus of the jig should be more than 3 GPa. However, these requirements are insufficient to represent a dental implant in the jawbone as the fixture is osseointegrated in the jawbone that comprises of the cortical bone and the cancellous bone. In this paper, three different materials for the jig and two different hole designs of the jig for holding the implant were examined in FE simulations and fatigue tests. From the simulation and test results, the effects of material properties and the hole designs of the jig were evaluated in the light of fatigue life of dental implants.     

Design and fabrication of custom-made dental implants

Dental implant has been attracting more and more attention due to their advantages of reliability and comfort. It is estimated that 10% of people will need dental implants in their life time. Especially, companies offer a system for customer to choice. But, no one research on custom-made dental implants. Traditional implants have their limitation and they are not better fit due to the difference of patient??s oral condition. The advantages of custom implant are accuracy fit and esthetic emergence profile. So, custom-made implant is desirable. Another key problem of custom-made dental implant is manufacturing. Dental implant is difficult to machine due to its complex features and its material (titanium). With the ever increasing demand for tight tolerance and increased complexity and accuracy, traditional machine tools have become ineffective for machining them. So, we design and built a PC-based CNC Turn-Mill-Hob professional machining center for machining our custom-made dental implants accurately and efficiently. This paper introduces our method: firstly, the custom-made dental implant in various oral conditions is designed by using our implant database. And then, FEA results indicate the stress distribution and magnitude of implant-bone interface for dentist referring. Finally, samples are automatic machined by our machining center.     

Vital confocal microscopy in bone

We wished to exploit confocal microscopy for high spatial and temporal resolution vital microscopy in bone. To this end, we evolved implants with glass windows supported in titanium, which were placed in the medial proximal tibial plateau of the rabbit, and special small, self-focussing objectives (dry 10/0.25, water immersion 20/0.45, and oil immersion 45/0.65 and 120/1.0) which mated and matched to the conical window entrance section of the metal components. At intervals of up to 21 months after implant healing, these lenses were used to study live tissue using two genera of confocal microscope: multiple aperture disc, tandem scanning, microscopes for observation in reflection, and video rate confocal laser scanning microscopes for recording, mainly in the fluorescence mode. The latter allowed the study of a variety of intravenously administered substances, including fluorescein, fluorescein-dextrans, fluorescent microspheres, acridine orange, DASPMI, calcein, and tetracycline. We were able to remove blood, stain cells with fluorescent markers, and replace them into the circulation. Calcein and tetracycline bind to the mineral front in bone: this labelling was studied in progress. We observed that both substances partition and remain for long periods (at least days) in adipocytes. Further characterisation of the system used both confocal fluorescence and scanning electron microscopy methods in the study of retrieved implants. These studies showed that the subimplant cortical bone remodelled to a less compact structure with a rich microvasculature extremely close to bone. The points of attachment of bone to glass were found to involve coarse fibres, with the matrix containing large numbers of large cells: some of this tissue was cartilage and some immature bone. An amorphous, mineralised matrix was in immediate contact with glass. The results provide further confirmation of the general utility of high-scan speed confocal methodology in physiology.     

Morphological and structural study of pseudowollastonite implants in bone

In vitro experiments show that pseudowollastonite (α-CaSiO₃) is a highly bioactive material that forms a hydroxyapatite surface layer on exposure to simulated body fluid and also to human parotid saliva. This finding is very significant, as it indicates that the pseudowollastonite can be physically and chemically integrated into the structure of living bone tissue, and therefore could be suitable for repair or replacement of living bone.   The physical and chemical nature of the remodelled interface between the pseudowollastonite implants and the surrounding bone has been studied after in vivo implantation of 20 pseudowollastonite cylinders into rat tibias. The interfaces formed after 3, 6, 8 and 12 weeks of implantation were examined histologically using an optical microscope and also by analytical scanning electron microscopy.   SEM and X-ray elemental analysis showed that the new bone was growing in direct contact with the implants. Other examinations found that the bone was fully mineralized. The ionic exchange taking place at the implant interface with the body fluids was essential in the process of the implant integration through a dissolution–precipitation–transformation mechanism. The study found the interface biologically and chemically active over the 12-week implantation period. The rate of new bone formation decreased after the first 3 weeks and reached constant value over the following 9 weeks. The osteoblastic cells migrated towards the interface and colonized the surface at the contact areas with the cortical regions and also bone marrow.     

Confocal laser scanning microscopy for the study of the morphological changes of the postextraction sites

A better understanding of the remodeling process of postextraction sockets is essential in dental treatment planning. The aim of this study was to evaluate whether confocal laser scanning microscopy (CLSM) can be applied to imaging contour changes of postextraction sites, as well as to its quantification with image analysis of obtained three‐dimensional images. This work describes a new application of the CLSM technique. The system used was the OLS3100‐USS, LEXT model (Olympus®). CLSM was used for the surface analysis of the extraction site. The measurements taken with CLSM were: (1) mesio‐distal distance, (2) alveolar ridge thickness, and (3) vestibular and lingual alveolar ridge height. Results of study cast scanning at baseline, 1 and 3 months after tooth extraction, with CLSM are well‐detailed images of postextraction areas. The CLSM technique used in study casts is a valid method to measure the dimensional changes that happen in the edentulous area after tooth extraction. This technique allows the evaluation of changes in mesio‐distal distance, thickness of the alveolar ridge and alveolar ridge height based on the measurements on the alveolar contours. Microsc. Res. Tech., 2011. © 2011 Wiley‐Liss, Inc.     

Laser ablation in titanium implants followed by biomimetic hydroxyapatite coating: Histomorphometric study in rabbits

Titanium surface texture and chemistry modification successfully improves the host response and consequently the bone-to-implant contact surrounding dental implants. The aim of the present study was to investigate, using histomorphometrical-analysis, the effects of titanium surface modification by laser-ablation (Nd:YAG) followed by thin chemical deposition of HA. Forty-eight rabbits received one implant by tibiae of AS-machined (MS), laser-modified (LMS), or biomimetic hydroxyapatite-coated (HA) surface. Bone-to-implant contact (BIC) and bone area (BBT) were evaluated after 4, 8, and 12 weeks, at cortical and cancellous regions. Average BIC in the cortical region was higher (P < 0.001) on the LMS and HA implants for all periods, with no differences between LMS and HA. For the cancellous area, the LMS and HA implants showed higher (P < 0.01) BIC than MS at the initial periods. The LMS and HA showed similar values in the cortical region, but a tendency of higher values for HA in the cancellous region was observed in all periods. For the BBT, the differences were found only between HA and MS after 4 weeks in the cortical region (P < 0.05), and after 12 weeks in the cancellous area (P < 0.05). Our results showed that HA biomimetic coating preceded by laser treatment induced the contact osteogenesis and allowed the formation of a more stable bone-implant interface, even in earlier periods.     

Transmission electron microscopy of the interface between bone and pseudowollastonite implant

This paper reports on the structural morphology of the interface in vivo between implants composed of bioactive synthetic pseudowollastonite ceramic and bone in rat tibias. Thin sections of the interfaces were examined after 6 and 8 weeks of implantation period in a high resolution transmission electron microscope up to the lattice plane resolution level.
The interfaces developed normal biological and chemical activities and remained reactive over the 8-week period. The regions showing direct bone tissue bonding to the implant contained nanocrystals of hydroxyapatite-like phase growing epitaxially across the interface in the [002] direction. The nanocrystals were also identified in the bone tissue formed in the interfacial area.
The reactivity of the implant caused in the first instance formation of an amorphous woven type of bone, which transformed into a crystalline lamellar type containing collagen fibres. The Ca/P ratio of the interfacial region was found to be between 1.67 in the mature bone tissue formed about 5 µm from the interface, and 2.06 in the regions right at the interface.     

Primary and long-term stability of a short-stem hip implant

The new generation short-stem hip implants are designed to encourage physiological-like loading, to minimize stress-strain shielding and therefore implant loosening in the long term. As yet there are no long-term clinical studies available to prove the benefits of these short-stem implants. Owing to this lack of clinical data, numerical simulation may be used as a predictor of longer term behaviour. This finite element study predicted both the primary stability and long-term stability of a short-stem implant. The primary implant stability was evaluated in terms of interface micromotion. This study found primary stability to fall within the critical threshold for osseointegration to occur. Longer term stability was evaluated using a strain-adaptive bone remodelling algorithm to predict the long-term behaviour of the bone in terms of bone mineral density (BMD) changes. No BMD loss was observed in the classical Gruen zones 1 and 7 and bone remodelling patterns were comparable with hip resurfacing results in the literature.     

The signaling pathway in modulating bone metabolism after dental implant in diabetes

Diabetes Mellitus is a systematic disease with complications in multi-organs, including decreased implant osseointegration and a high failure rate of dental transplants. Accumulating evidence indicates that the signaling pathway directly impacts the process of bone metabolism and inflammatory response implicated with dental implants in diabetic patients. This review summarizes the recent advance in signaling pathways regulate osseointegration and inflammatory response in dental transplantation, aiming to identify the potential therapeutic target to reduce the dental transplant failure in diabetes patients, with emphasis on the surface characteristics of the implant, inflammatory signaling, AMPK, PPARγ, WNT, ROS, and adiponectin signaling.