Surface Functionalization of PTFE Membranes Intended for Guided Bone Regeneration Using Recombinant Spider Silk.

Alveolar bone loss is usually treated with guided bone regeneration, a dental procedure which utilizes a tissue-separation membrane. The barrier membrane prevents pathogens and epithelial cells to invade the bone augmentation site, thereby permitting osteoblasts to deposit minerals and build up bone. This study aims at adding bioactive properties to otherwise inert PTFE membranes in order to enhance cell adherence and promote proliferation. A prewetting by ethanol and stepwise hydration protocol was herein employed to overcome high surface tension of PTFE membranes and allow for a recombinant spider silk protein, functionalized with a cell-binding motif from fibronectin (FN-silk), to self-assemble into a nanofibrillar coating. HaCaT and U-2 OS cells were seeded onto soft and hard tissue sides, respectively, of membranes coated with FN-silk. The cells could firmly adhere as early as 1 h post seeding, as well as markedly grow in numbers when kept in culture for 7 days. Fluorescence and scanning electron microscopy images revealed that adherent cells could form a confluent monolayer and develop essential cell-cell contacts during 1 week of culture. Hence, functionalized PTFE membranes have a potential of better integration at the implantation site, with reduced risk of membrane displacement as well as exposure to oral pathogens.

Factors influencing marginal bone loss at a hydrophilic implant design placed with or without GBR procedures: A 5-year retrospective study.

BACKGROUND: Ongoing marginal bone loss is a threat to the longevity of implant-supported prostheses. AIM: The aim of the present study was to retrospectively evaluate the survival rate and factors affecting marginal bone levels at a hydrophilic implant design after 5 years in function. MATERIALS AND METHODS: The study group consisted of 51 consecutive patients previously treated with 159 hydrophilic implants (Neoss Straight Proactive implants) and scheduled for annual check-ups with clinical and radiographic examinations during 5 years. Data were compiled for the entire study population as well as for two subgroups: one where guided bone regeneration (GBR) was performed (91 implants) and the other where no GBR procedures (68 implants) were performed. Marginal bone levels were measured from peri-apical radiographs taken at placement and annual follow-ups. Statistical analyses were applied to evaluate the effect of different factors on marginal bone remodeling. RESULTS: Two implant failures, one from each subgroup, occurred during the first year of function resulting in an overall cumulative survival rate (CSR) of 98.7% after 5 years of loading. The mean marginal bone loss amounted to 0.7 ± 0.7 mm after 1 year and 0.8 ± 0.6 mm after 5 years. No implants showed more than 3 mm bone loss after 5 years. Age, gender, implant position, biotype, implant diameter, implant length, indication, surgical/loading protocol, and ISQ at prosthesis delivery were found to affect bone remodeling. No significant differences or correlations were seen for smoking, jaw, bone quantity, bone quality, GBR, sinus lift, and ISQ at implant placement. CONCLUSIONS: The present implant design performed well with few failures and minimal marginal bone loss after 5 years of loading. Marginal bone remodeling at implants is a complex phenomenon, which is affected by many patient-, procedure-, and implant-related factors that need to be further investigated.

Factors Influencing Resonance Frequency Analysis (RFA) Measurements and 5-Year Survival of Neoss Dental Implants.

BACKGROUND: Diagnostic instruments based on resonance frequency analysis (RFA) can be utilised to assess dental implant stability during treatment and follow-up. AIM: The aim of the present study was to investigate the influence of patient- and implant-related factors on implant stability and the 5-year implant survival. In addition, the influence of stability (ISQ value) at placement and abutment connection on implant survival was evaluated. MATERIALS AND METHODS: RFA measurements from a total of 334 consecutive patients with 745 dental implants (Neoss Ltd., Harrogate, UK) were retrospectively analysed after at least 5 years in function. Statistics were used to evaluate the influence of the different variables on implant stability and implant survival. Odds ratio calculations were performed to compare the risk for implant failure using 60, 65, 70, and 75 ISQ as threshold levels at placement and loading. RESULTS: A total of 20 implant failures in 14 patients were noted during the 5 years of follow-up, giving an overall cumulative survival rate (CSR) of 97.3% at the implant level and 95.8% at the patient level. Gender, jaw, position, bone quality, and implant diameter had an influence on implant stability at placement. Jaw, bone quality, and implant diameter had an influence on stability after 3-4 months of healing. More failures were observed in full than in partial rehabilitations. Age, gender, jaw, position, bone quantity, bone quality, implant diameter, and implant length had no influence on implant survival. Implants with ISQ values below the threshold levels showed lower survival rates compared to implants with values above these levels. CONCLUSIONS: The present study showed a significantly higher risk for implant failure, showing an ISQ value below 70 and 75 at placement or after 3-4 months of healing. The results indicate that RFA measurements can be used to identify implants with increased risk for failure.

A 1-7 year retrospective follow-up on consecutively placed 7-mm-long dental implants with an electrowetted surface.

BACKGROUND: This retrospective consecutive case series study was performed to determinate the survival rate and implant stability of short (7 mm length) dental implants with an electrowetted hydrophilic surface that were in function from 1 to 7 years. METHODS: A retrospective chart review identified and evaluated 86 consecutively placed 7-mm-long dental implants (ProActive, Neoss Ltd., Harrogate, England) in 75 patients. Analysis was performed for implant survival as well as implant stability, as measured by insertion torque (IT) and resonance frequency analysis (RFA). RESULTS: Clinical follow-ups were performed from 1.0 to 7.0 years after implant placement (mean 4.0 ± 2.1 years). Two implants failed prior to loading resulting in a 5-year cumulative survival rate (CSR) of 97.7%. An additional late failure occurred at 60 months post-loading for a 7-year CSR of 94.8%. Mean insertion torque was 30.1 ± 7.4 Ncm and mean RFA at insertion was 73.6 ± 8.1 ISQ. Follow-up RFA measurements suggested that the achieved primary stability was maintained throughout the healing phase. CONCLUSION: The present study demonstrates that treatment with short implants can be a predictable treatment option with high survival rate in sites with limited available bone.

In vitro evaluation of barrier function against oral bacteria of dense and expanded polytetrafluoroethylene (PTFE) membranes for guided bone regeneration.

AIM: This study evaluates biofilm formation and barrier function against Streptococcus oralis of nonresorbable polytetrafluoroethylene (PTFE) guided bone regeneration membranes having expanded (e-PTFE) and dense (d-PTFE) microstructure. MATERIALS AND METHODS: Three e-PTFE membranes of varying openness, one d-PTFE membrane, and commercially pure titanium discs were evaluated. All e-PTFE membranes consisted of PTFE nodes interconnected by fibrils. The d-PTFE membrane was fibril-free, with large evenly spaced indentations. The surfaces were challenged with S. oralis and incubated statically for 2-48h. Bacterial colonization, viability, and penetration were evaluated. RESULTS: S. oralis numbers increased over time on all surfaces, as observed using scanning electron microscopy, while cell viability decreased, as measured by colony forming unit (CFU) counting. At 24h and 48h, biofilms on d-PTFE were more mature and thicker (tower formations) than on e-PTFE, where fewer layers of cells were distributed mainly horizontally. Biofilms accumulated preferentially within d-PTFE membrane indentations. At 48h, greater biofilm biomass and number of viable S. oralis were found on d-PTFE compared to e-PTFE membranes. All membranes were impermeable to S. oralis cells. CONCLUSIONS: All PTFE membranes were effective barriers against bacterial passage in vitro. However, d-PTFE favored S. oralis biofilm formation.

A One-year Follow-up Study of a Tapered Hydrophilic Implant Design Using Various Placement Protocols in the Maxilla.

PURPOSE: To study the clinical/radiographic outcomes and stability of a tapered implant design with a hydrophilic surface when placed in the maxilla using various protocols and followed for one year. METHODS: Ninety-seven consecutive patients treated as part of daily routine in two clinics with 163 tapered implants in healed sites, in extraction sockets and together with bone augmentation procedures in the maxilla were evaluated after one year in function. Individual healing periods varying from 0 to 6 months had been used. Insertion torque (IT) and resonance frequency analysis (RFA) measurements were made at baseline. Follow-up RFA registrations were made after 6 and 12 months of loading. The marginal bone levels were measured in intraoral radiographs from baseline and after 12 months. A reference group consisting of 163 consecutive straight maxillary implants was used for the comparison of baseline IT and RFA measurements. RESULTS: Five implants failed before loading, giving an implant survival rate of 96.9% and a prosthesis survival rate of 99.4% after one year. The mean marginal bone loss after one year was 0.5 mm (SD 0.4). The mean IT was statistically significantly higher for tapered than for straight reference implants (41.3 ± 12.0 Ncm vs 33.6 ± 12.5 Ncm, p < 0.001). The tapered implants showed a statistically insignificantly higher mean ISQ value than the straight references implants (73.7 ± 6.4 ISQ vs 72.2 ± 8.0 ISQ, p=0.119). There was no correlation between IT and marginal bone loss. There was a correlation between IT and RFA measurements (p < 0.001). CONCLUSION: The tapered implant showed a high survival rate and minimal marginal bone loss after one year in function when using various protocols for placement. The tapered implant showed significantly higher insertion torque values than straight reference implants.

One-piece zirconia oral implants: one-year results from a prospective case series. 2. Three-unit fixed dental prosthesis (FDP) reconstruction.

AIM: To evaluate the clinical and radiological outcome of one-piece zirconia oral implants for three-unit fixed dental prosthesis (FDP) replacement after 1 year. MATERIALS: Twenty eight patients were recruited for the investigation and signed an informed consent. All patients were treated with a one-stage implant surgery and a three-unit immediate temporary restoration on two one-piece zirconia implants. The implants were fabricated of yttria-stabilized tetragonal zirconia (y-TZP). The endosseous part of the implants was tapered with a porous surface. A total of 56 implants were inserted in the 28 patients. A total of 12 implants were placed in the upper jaws (six in the anterior area and six in the posterior area) and 44 in mandibles (all in the posterior area). At implant insertion and after 1 year, standardized radiographs were taken to evaluate the peri-implant bone loss. To evaluate any influences from different baseline parameters on the marginal bone loss a univariate analysis was performed. Clinical soft tissue parameters probing depth (PD), clinical attachment level (CAL), modified bleeding index (mBI) and modified plaque index (mPI) were recorded. Implant cumulative survival rates were calculated using actuarial life table analysis. Changes in the clinical variables were assessed using the Wilcoxon Signed Ranks test (PD, CAL) and the Sign test (mBl, mPl). All significance tests were conducted at a 5% level of significance. RESULTS: After 1 year, one implant was lost resulting in a survival rate of 98.2%. The patient was excluded from further analysis. The marginal bone loss after 1 year amounted to 1.95 mm. In 40% of the patients a bone loss of at least 2 mm and in 28% of the patients a loss of more than 3 mm were observed. The PD decreased for implant and tooth sites over time, the values being significantly higher for implants than for teeth. Over 1 year, the CAL increased slightly around the implants and decreased around the teeth. At the 1-year follow-up, the CAL at the implant sites was statistically significantly higher than at the reference teeth. The mBI was significantly lower at implants than at teeth. The same result was found for the plaque index. CONCLUSIONS: A high frequency of increased radiographic bone loss (>2 mm) after 1 year around the presented one-piece zirconia implant system was found. The bone loss seems to be higher compared to the very limited availability of zirconia implant data. Therefore, within the limits of the present investigation, it may be concluded that the presented zirconia implant system possibly performs inferior to conventional titanium implants and to other zirconia implants regarding peri-implant bone loss.

One-piece zirconia oral implants: one-year results from a prospective cohort study. 1. Single tooth replacement.

AIM: To investigate the clinical and radiographic outcome of a one-piece zirconia oral implant for single tooth replacement after 1 year. MATERIALS AND METHODS: A total of 65 patients received a one-stage implant surgery with immediate temporization. Standardized radiographs were taken at implant insertion and after 1 year to monitor peri-implant bone loss. A univariate analysis of the influence of different baseline parameters on marginal bone loss from implant insertion to 12 months was performed. Soft tissue parameters were evaluated at prosthesis insertion and at the 1-year follow-up. RESULTS: After 1 year, three implants were lost, giving a cumulative survival rate of 95.4%. The marginal bone loss after 1 year was 1.31 mm. Thirty-four per cent of the implants lost at least 2 mm bone, and 14% more than 3 mm. The univariate analysis could not depict any parameter influencing marginal bone loss. Probing depth, Clinical Attachment Level, Bleeding and Plaque Index decreased over 1 year. CONCLUSIONS: The cumulative survival rate of the presented ceramic implant was comparable to the reported survival rate of titanium implants when immediately restored. However, the frequency of increased radiographic bone loss (>2 mm) after 1 year was considerably higher as compared to conventional two-piece titanium implants. The presented zirconia implant can therefore not be recommended for clinical usage.

Titanate biomaterials with enhanced antiinflammatory properties.

While titanium implants are generally recognized as having excellent biocompatibility, the mechanistic basis for this has yet to be established. We previously demonstrated that TiO2, found on surfaces of titanium, has antioxidant properties that degrade the reactive oxygen species (ROS) which mediate the inflammatory response. We hypothesized that the antioxidant mechanism was similar to that known to mediate photocatalysis by titanium oxides. Specifically, we investigated whether the electronic or valence state of the surface titanium atoms mediates the catalytic degradation of ROS. Surface Ti(IV) atoms in TiO2 and SrTiO3 single crystal substrates were converted into Ti(III) while maintaining the bulk crystalline structure by vacuum annealing or Niobium doping. The degradation of both chemically-induced and neutrophil-derived ROS were significantly increased by changing the valence state of surface titanium. These results suggest that titanium-mediated degradation of ROS is through a catalytic mechanism. Furthermore, we describe a series of novel biomaterials that have antioxidant properties superior to those of titanium.

Anti-inflammatory properties of micropatterned titanium coatings.

Prolonged inflammation and reactive oxygen species (ROS) generated around an implanted biosensor are the primary causes of the foreign body response, including encapsulation of biosensor membranes. We have previously demonstrated that TiO2 surfaces reduce ROS. Here we investigated the potential of using the anti-inflammatory properties of TiO2 in the design of biosensor membranes with improved long-term in vivo transport properties. Micropatterned Ti films were sputtered onto quartz surfaces in a series of hexagonally distributed dots with identical coverage area of 23% and dot size ranging from 5 to 100 microm. The antioxidant effect of the surfaces was investigated using a cell-free peroxynitrite donor assay and assays of superoxide released from stimulated surface-adhering neutrophils and macrophages. In all three assays, the amount of ROS was monitored using luminol-amplified chemiluminescence. Patterned surfaces in all experimental models significantly decreased ROS compared to the etched surfaces. In the cell-free experiment, the ROS reduction was only dependent on fractional surface coverage. In the cell experiments, however, a dot-size-dependent ROS reduction was seen, with the largest reduction at the smallest dot-size surfaces. These results indicate that micropatterned surfaces with small dots covering only 23% of the surface area exhibit similar antioxidative effect as fully covered surfaces.

Cell- rather than antibody-mediated immunity leads to the development of profound thrombocytopenia during experimental Plasmodium berghei malaria.

Experimental malarial thrombocytopenia can reach life-threatening levels and is believed to be due to Abs targeting platelets for destruction by the reticuloendothelial system. However, we report that Abs account for at most 15% of platelet destruction as Plasmodium berghei-infected B cell-deficient mice exhibited profound thrombocytopenia (83%) as did C57BL/6 controls (98%). Further, no significant increase in Abs bound to intact platelets was observed during infection. P. berghei infection can enhance the activity of anti-platelet Abs as indicated by a significantly (p < 0.005) increased thrombocytopenia on day 4 of infection in mice that were administered a low dose anti-CD41 mAb compared with rat IgG1-injected controls. RAG1-/- and CD4- plus CD8-deficient mice were markedly protected from thrombocytopenia (p < 0.005) and malarial pathogenesis. CD8- or TCRgammadelta-deficient mice were not protected from thrombocytopenia and CD4-deficient mice were modestly protected. RAG1-/- mice exhibited significantly (p < 0.05) lower levels of plasma TNF, IFN-gamma, and IL-12 during infection. IFNgamma-/- and IL-12-/- mice exhibited increased survival but similar thrombocytopenia to C57BL/6 controls. Collectively, these data indicate that thrombocytopenia is necessary but not sufficient for malarial pathogenesis and Abs are not the major contributors to malarial thrombocytopenia. Rather, we propose that both CD4+ and CD8+ T cell populations play key roles in malarial thrombocytopenia; a complex bidirectional interaction between cell-mediated immunity and platelets exists during experimental severe malaria that regulates both responses.

Contact activation and down regulation of neutrophils by cellulose wound dressings can be prevented by radical scavengers.

Neutrophil interaction with wound dressing materials was studied. A meshed non-woven cellulose was chosen as model dressing. Neutrophils isolated from human blood was added to the cellulose, and the production of reactive oxygen species was measured by luminol-amplified chemiluminescence. The respiratory burst response of the neutrophils was found to be activated upon contact with cellulose. The contact activation of the cells increased when the cellulose was oxidised with periodate, and decreased when the cellulose was reduced with cyanoborohydride, indicating that the activation of the respiratory burst response was due to carbonyl-induced stress. The contact activation of the respiratory burst response resulted in an inability of the neutrophils to respond to a secondary stimulation with zymosan. When radical scavenger enzymes were covalently bound to the cellulose, the contact activation was decreased and the ability to respond to stimuli was increased. Addition of the molecular scavenger glutathione (GSH) did not decrease the cell activation upon cellulose contact, but the cell showed an intact ability to respond to secondary stimuli after cellulose contact. In conclusion, the results show that the environmental redox potential effects neutrophils in a situations of clinical interest and that the addition of radical scavengers protects the neutrophils against material-induced damage resulting in preserved cell function.

Development of a wound dressing targeting neutrophil function.

Earlier studies show that neutrophils are virtually unable to kill Staphylococcus aureus in vitro. However, upon addition of 10 mM N-acetylcysteine (NAC) or reduced glutathione (GSH) the neutrophil bacterial killing ability becomes excellent. We want to exploit this phenomenon to develop a wound dressing material that will improve neutrophil function. To study the mechanisms behind the downregulation of neutrophil elimination of bacteria, we used different markers for neutrophil function on surface-adhering neutrophils in contact with S. aureus with or without addition of the antioxidants NAC or GSH. Analysis by scanning electron microscopy showed cell shrinkage and numerous cytoplasmic processes on surface-adhering neutrophils exposed to S. aureus. In cells exposed to S. aureus and GSH, the cells were of normal size and the cytoplasm was spread as in normal attachment. Staining for intracellular GSH, a hallmark of oxidative stress, showed little difference between the experimental groups, indicating that the cells were not damaged by traditional oxidative stress. The H(2)O(2) production of neutrophils, measured by Amplex red, was correlated to bacterial exposure and was not affected by the addition of scavengers. The intracellular and extracellular production of ROS was measured by luminol-amplified chemiluminescence. The apparent ROS-production was mostly intracellular and decreased in the presence of scavengers. However, extracellular production of ROS was not affected by the addition of NAC. The production of nitric oxide (NO) was measured spectrophotometrically as the production of nitrate apparently decreased in the presence of scavengers, probably as a result of interference with the reagents in the test system. In conclusion, differences between leukocytes that were able to eliminate S. aureus and those that were not were mainly seen in the morphology of the cells and in cell viability. The morphological findings point to a difference in NO signaling in the absence and presence of ROS scavengers.