Ryo Jimbo

Photo‐induced hydrophilicity enhances initial cell behavior and early bone apposition

OBJECTIVEnThe anatase form of titanium dioxide (TiO(2)) exhibits photo-induced hydrophilicity when it is irradiated with ultraviolet (UV) light. In the present study, the effect of photo-induced hydrophilicity on initial cell behavior and bone formation was evaluated.nnnMATERIALS AND METHODSnPlasma source ion implantation method and post-annealing were employed for coating the anatase form of TiO(2) to the surface of the titanium disk and implant. Half of the disks and implants were illuminated with UV for 24 h beforehand, whereas the other halves were blinded and used as controls. Photo-induced hydrophilicity was confirmed by a static wettability assay. The effects of this hydrophilicity on cell behavior were evaluated by means of cell attachment, proliferation and morphology using pluripotent mesenchymal precursor C2C12 cells. Thereafter, bone formation around the hydrophilic implant inserted in the rabbit tibia was confirmed histomorphometrically.nnnRESULTSnThe water contact angle of the photo-induced hydrophilic disk decreased markedly from 43.5 degrees to 0.5 degree. Cell attachment and proliferation on this hydrophilic disk showed significant improvement. The cell morphology on this hydrophilic disk was extremely flattened, with an elongation of the lamellipodia, whereas a round/spherical morphology was observed on the control disk. The photo-induced hydrophilic implant enhanced the bone formation with the bone-to-metal contact of 28.2% after 2 weeks of healing (control: 17.97%).nnnCONCLUSIONnThe photo-induced hydrophilic surface used in the current study improves the initial cell reactions and enhances early bone apposition to the implant.

Enhanced Initial Cell Responses to Chemically Modified Anodized Titanium

BACKGROUNDnPreviously, we reported that anodized porous titanium implants have photocatalytic hydrophilicity. However, this effect was not always sufficient for the significant improvement of bone apposition.nnnPURPOSEnThe purpose of this study was to improve the photocatalytic properties of porous titanium implants by the fluoride modification of the anodized titanium dioxide (TiO(2)), and to investigate the initial cell response to it.nnnMATERIALS AND METHODSnThe ideal concentration of ammonium hydrogen fluoride (NH(4)F-HF(2)) used in this study was determined by a static water contact angle assay. The ideal concentration of NH(4)F-HF(2) was 0.175%, and experimental disks were treated with this concentration. A pluripotent mesenchymal cell line, C2C12, was cultured on the disks in order to investigate cell attachment, morphology, and proliferation.nnnRESULTSnCell attachment after 30 minutes of culturing was significantly higher for the ultraviolet-irradiated, fluoride-modified anodized TiO(2) (p < .05), and the simultaneous scanning electron microscope observation showed a rather flattened and extended cell morphology. The proliferation rate after 24 hours was also significantly higher for the fluoride-modified anodized TiO(2).nnnCONCLUSIONnFluoride chemical modification enhances the hydrophilic property of the anodized TiO(2) and improves the initial cell response to it.

Accelerated photo-induced hydrophilicity promotes osseointegration : an animal study

BACKGROUNDnIn the previous in vitro study, fluoride-modified, anodized porous titanium was proven to have enhanced its photo-induced hydrophilicity, which induced the hyperactivation of initial cell response.nnnPURPOSEnThe purpose of the present study was to investigate in vivo bone apposition during the early stages of osseointegration in rabbit tibiae.nnnMATERIALS AND METHODSnAnodized porous titanium implants (TiU, TiUnite®, Nobel Biocare AB, Göteborg, Sweden) were modified with 0.175 wt% ammonium hydrogen fluoride solution (NH(4) F-HF(2) ). Twenty-four hours prior to the experiments, the surface-modified implants were ultraviolet-irradiated (modTiU). Blinded and unpackaged TiU implants were used as controls. Thereafter, the implants were placed in the rabbit tibial metaphyses and histomorphometrically analyzed at 2 and 6 weeks after insertion.nnnRESULTSnModTiU demonstrated a significantly greater degree of bone-to-metal contact than TiU after 2 and 6 weeks of healing.nnnCONCLUSIONnThe results proved that the enhanced photo-induced hydrophilicity of the NH(4) F-HF(2) -modified anodized implants promoted bone apposition during the early stages of osseointegration.

Local release of magnesium from mesoporous TiO2 coatings stimulates the peri-implant expression of osteogenic markers and improves osteoconductivity in vivo.

Local release of Mg ions from titanium implant surfaces has been shown to enhance implant retention and integration. To clarify the biological events that lead to this positive outcome, threaded implants coated with mesoporous TiO2 thin films were loaded with Mg-ions and placed in the tibia of rabbits for 3weeks, after surface characterization. Non-loaded mesoporous coated implants were used as controls. Peri-implant gene expression of a set of osteogenic and inflammatory assays was quantified by means of real-time quantitative polymerase chain reaction. The expression of three osteogenic markers (OC, RUNX-2 and IGF-1) was significantly more pronounced in the test specimens, suggesting that the release of Mg ions directly at the implant sites may stimulate an osteogenic environment. Furthermore, bone healing around implants was evaluated on histological slides and by diffraction-enhanced imaging (DEI), using synchrotron radiation. The histological analysis demonstrated new bone formation around all implants, without negative responses, with a significant increase in the number of threads filled with new bone for test surfaces. DEI analysis attested the high mineral content of the newly formed bone. Improved surface osteoconductivity and increased expression of genes involved in the bone regeneration were found for magnesium-incorporation of mesoporous TiO2 coatings.

Osteogenic potential of human adipose-derived stromal cells on 3-dimensional mesoporous TiO2 coating with magnesium impregnation

The aim of this study was to evaluate the osteogenic response of human adipose-derived stromal cells (ADScs) to mesoporous titania (TiO2) coatings produced with evaporation-induced self-assembly method (EISA) and loaded with magnesium. Our emphasis with the magnesium release functionality was to modulate progenitor cell osteogenic differentiation under standard culture conditions. Osteogenic properties of the coatings were assessed for stromal cells by means of scanning electron microscopy (SEM) imaging, colorimetric mitochondrial viability assay (MTT), colorimetric alkaline phosphates activity (ALP) assay and real time RT-polymerase chain reaction (PCR). Using atomic force microscopy (AFM) it was shown that the surface expansion area (Sdr) was strongly enhanced by the presence of magnesium. From MTT results it was shown that ADSc viability was significantly increased on mesoporous surfaces compared to the non-porous one at a longer cell culture time. However, no differences were observed between the magnesium impregnated and non-impregnated surfaces. The alkaline phosphatase activity confirmed that ADSc started to differentiate into the osteogenic phenotype after 2 weeks of culturing. The gene expression profile at 2 weeks of cell growth showed that such coatings were capable to incorporate specific osteogenic markers inside their interconnected nano-pores and, at 3 weeks, ADSc differentiated into osteoblasts. Interestingly, magnesium significantly promoted the osteopontin gene expression, which is an essential gene for the early biomaterial-cell osteogenic interaction.

Surface treatment with ascorbic acid and ferric chloride improves the micro-tensile bond strength of 4-META/MMA-TBB resin to dentin.

OBJECTIVEnThe purpose of this study was to investigate the efficacy of an experimental dentin conditioner consisting of ascorbic acid (AA) and ferric chloride (Fe), in terms of micro-tensile bond strength (microTBS).nnnMETHODSnFive experimental solutions were prepared and evaluated: 10% AA and 5% Fe (10AA-5Fe); 10% AA and 0% Fe (10AA-0Fe); 0% AA and 5% Fe (0AA-5Fe); 10% citric acid and 3% ferric chloride (10-3); and, 0% AA and 0% Fe (0AA-0Fe). Flattened dentin surfaces were treated with each of the experimental solutions. A composite material rod was bonded to the dentin surface with a self curing luting agent (4-META/MMA-TBB resin). The specimens were stressed to failure under tension after 24h of immersion in water.nnnRESULTSnThe 10AA-5Fe group showed significantly higher bond strength than the 10AA-0Fe, 0AA-5Fe, 10-3 and AA-0Fe groups.nnnCONCLUSIONnIt was found that an experimental conditioner consisting of 10% ascorbic acid and 5% ferric chloride significantly improved the micro-tensile bond strength between 4-META/MMA-TBB resin and dentin.

The physicochemical characterization and in vivo response of micro/nanoporous bioactive ceramic particulate bone graft materials

In this study, the physicochemical characteristics of calcium phosphate based bioactive ceramics of different compositions and blends presenting similar micro/nanoporosity and micrometer scale surface texture were characterized and evaluated in an in vivo model. Prior to the animal experiment, the porosity, surface area, particle size distribution, phase quantification, and dissolution of the materials tested were evaluated. The bone regenerative properties of the materials were evaluated using a rabbit calvaria model. After 2, 4, and 8 weeks, the animals were sacrificed and all samples were subjected to histologic observation and histomorphometric analysis. The material characterization showed that all materials tested presented variation in particle size, porosity and composition with different degrees of HA/TCP/lower stoichiometry phase ratios. Histologically, the calvarial defects presented temporal bone filling suggesting that all material groups were biocompatible and osteoconductive. Among the different materials tested, there were significant differences found in the amount of bone formation as a function of time. At 8 weeks, the micro/nanoporous material presenting ~55%TCP:45%HA composition ratio presented higher amounts of new bone regeneration relative to other blends and a decrease in the amount of soft tissue infiltration.

Modulation of the nanometer pore size improves magnesium adsorption into mesoporous titania coatings and promotes bone morphogenic protein 4 expression in adhering osteoblasts

OBJECTIVEnMesoporous (MP) titania films used as implant coatings have recently been considered as release systems for controlled administration of magnesium to enhance initial osteoblast proliferation in vitro. Tuning of the pore size in such titania films is aimed at increasing the osteogenic potential through effects on the total loading capacity and the release profile of magnesium.nnnMETHODSnIn this study, evaporation-induced self-assembly (EISA) was used with different structure-directing agents to form three mesoporous films with average pore sizes of 2nm (MP1), 6nm (MP2) and 7nm (MP3). Mg adsorption and release was monitored using quartz crystal microbalance with dissipation (QCM-D). The film surfaces were characterized with atomic force microscopy (AFM), scanning electron microscopy (SEM) and X-ray photoelectron spectroscopy (XPS). The effect of different Mg release on osteogenesis was investigated in human fetal osteoblasts (hFOB) using pre-designed osteogenesis arrays and real-time polymerase chain reaction (RT-PCR).nnnRESULTSnResults showed a sustained release from all the films investigated, with higher magnesium adsorption into MP1 and MP3 films. No significant differences were observed in the surface nanotopography of the films, either with or without the presence of magnesium. MP3 films (7nm pore size) had the greatest effect on osteogenesis, up-regulating 15 bone-related genes after 1 week of hFOB growth and significantly promoting bone morphogenic protein (BMP4) expression after 3 weeks of growth.nnnSIGNIFICANCEnThe findings indicate that the increase in pore width on the nano scale significantly enhanced the bioactivity of the mesoporous coating, thus accelerating osteogenesis without creating differences in surface roughness.

In vivo evaluation of resorbable supercritical CO2-treated collagen membranes for class III furcation-guided tissue regeneration: IN VIVO EVALUATION OF RESORBABLE scCO2-TREATED COLLAGEN MEMBRANES

The study evaluated the effects of a Supercritical CO2 (scCO2 ) on a commercially available decellularized/delipidized naturally derived porcine pericardium collagen membrane, Vitala®. The Vitala® and scCO2 treated experimental membranes were evaluated for guided tissue regeneration (GTR) of periodontal tissue in class III furcation defects utilizing a dog model. Physical material characterization was performed by scanning electron microscopy (SEM), thermogravimetric analysis (TGA), and differential scanning calorimetry (DSC). The in vivo portion of the study was allocated to three-time points (6, 12, and 24-weeks) using standardized class III furcation defects created in the upper second and third premolars. The experimental defects (n = 5) were covered with either a collagen membrane (positive control), scCO2 -treated collagen membrane (experimental) or no membrane (negative control). Following sacrifice, histologic serial sections were performed from cervical to apical for morphologic/morphometric evaluation. Morphometric evaluation was carried out by ranking the presence of collagen membrane, amount of bone formation within the defect site and inflammatory cell infiltrate content. SEM showed the experimental scCO2 -treated membrane to have a similar gross fibrous appearance and chemical structure in comparison to the Vitala® Collagen membrane. A significant increase in membrane thickness was noted in the scCO2 -treated membranes (366 ± 54 μm) vs non-treated membranes (265 ± 75 μm). TGA and DSC spectra indicated no significant qualitative differences between the two membranes. For the in vivo results, both membranes indicated significantly greater amounts of newly formed bone (scCO2 : 2.85 ± 1.1; Vitala®: 2.80 ± 1.0) within the covered defects relative to uncovered controls (0.8 ± 0.27) at 24 weeks. Both membrane types gradually degraded as time elapsed in vivo from 6 to 12 weeks, and presented nearly complete resorption at 24 weeks. The inflammatory infiltrate at regions in proximity with the membranes was commensurate with healthy tissue levels from 6 weeks in vivo on, and periodontal ligament regeneration onset was detected at 12 weeks in vivo. The effect of the supplementary scCO2 treatment step on the collagen membrane was demonstrated to be biocompatible, allowing for the infiltration of cells and degradation over time. The treated membranes presented similar performance in GTR to non-treated samples in Class III furcation lesions. Defects treated without membranes failed to achieve regeneration of the native periodontium.

Influence of different drilling preparation on cortical bone: A biomechanical, histological, and micro-CT study on sheep

OBJECTIVEnThe aim of this study was to investigate the extent of cortical bone remodeling between two different drilling protocols by means of histomorphometric, µ-CT, and biomechanical analyses.nnnMATERIAL AND METHODSnA total of 48 implants were inserted into the mandible of six sheep following two drilling protocols: Group A (Test, nxa0=xa024), undersized preparation; Group B (Control, nxa0=xa024), non-undersized preparation. The animals were euthanatized to obtain 5 and 10xa0weeks of implantation time. Removal torque (RTQ) was measured on 12 implants of each group and the peri-implant bone was µ-CT scanned. Bone volume density (BV/TV) was calculated in pre-determined cylindrical volumes, up to 1.5xa0mm from implant surface. Non-decalcified histology was prepared on the remaining 12 implants from each group, where total bone-to-implant contact (totBIC) and newly-formed BIC (newBIC) was measured. Bone Area Fraction Occupancy (BAFO) was determined in pre-determined areas up to 1.5xa0mm from implant surface. Paired sample t test or Wilcoxon signed-rank test was used to investigate differences between the groups.nnnRESULTSnGroup A presented significantly increased RTQ value at 5xa0weeks, while no difference was observed at 10xa0weeks. Group B presented increased BV/TV value at 5xa0weeks. Both groups showed comparable values for totBIC at both time-points. However, Group A presented significantly lower newBIC at 5xa0weeks. Higher BAFO was observed in Group B at 5xa0weeks.nnnCONCLUSIONSnImplants inserted into undersized sites has an increased biomechanical performance, but provoked major remodeling of the cortical bone during the early healing period compared to non-undersized preparations. After 10xa0weeks, no difference was observed.