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Introduction: Visual imaging of subsurface caries lesions is of vital interest in dentistry, which can be obtained by invasive radiography technique as well as by available non-destructive imaging approaches. Thus, as a first step toward the development of a new innovative approach, Spectral-domain optical coherence tomography (SD-OCT) was applied to detect the lesion depth in comparison to the established reference technique (transverse microradiography [TMR])., Methods: Bovine enamel specimens were demineralized for 5 days, following previous studies. For OCT, the resulting artificial lesions were scanned three-dimensionally (SD-OCT) and semi-automated measured (CarLQuant). For TMR, specimens were sectioned and the lesion depth was manually determined (Inspektor Research System)., Results: The range of lesion depth detected with OCT was 24.0-174.0 μm (mouth rinse study), 18.0-178.0 μm (toothpastes study) and with TMR 59.2-198.0 μm (mouth rinse study), 33.2-133.4 μm (toothpastes study). We found a strong correlation between both methods in terms of lesion depth (Spearman rankwith outlierp < 0.001, Rho = 0.75, Spearman rankwithout outlierp = 0.001, Rho = 0.79). The two methods produce similar results (Passing-Bablok regression, 1.16). As deeper is the lesion, the smallest is the difference between both methods as indicated by Bland-Altman-plots., Conclusion: Especially in the case of deep lesions, the values obtained by both methods are in agreement, and OCT can potentially substitute TMR to detect and assess lesion depth with the benefit of being non-destructive., (© 2024 S. Karger AG, Basel.)

Assessment of enamel subsurface lesion remineralisation is essential for the evaluation of novel remineralisation technologies. The gold standard to assess subsurface mineral gain of enamel lesions is transverse microradiography (TMR). However, some studies have utilised surface microhardness (SMH) to evaluate efficacy of remineralisation agents. The aim of this study was to assess remineralisation of enamel subsurface lesions using TMR and SMH after in vitro treatment with calcium-containing technologies, and to test correlation between the TMR and SMH measurements. The parameters obtained from the TMR and SMH analyses of enamel subsurface remineralisation were not significantly correlated. Furthermore, the enamel subsurface remineralisation as measured by TMR was significantly correlated with the water-soluble calcium concentration of the remineralisation products. Scanning electron microscopy revealed surface precipitates formed by specific remineralisation treatments obfuscated accurate assessment of remineralisation by SMH. It was concluded that TMR is a more appropriate method for analysis of enamel subsurface remineralisation, and that SMH values of remineralised enamel should be interpreted with caution. Using TMR the level of remineralisation (%R) by the different technologies was CPP-ACP/F (31.3 ± 1.4%); CPP-ACP (24.2 ± 1.4%); CaSO 4 /K 2 HPO 4 /F (21.3 ± 1.4%); f-TCP/F (20.9 ± 1.0%); Nano-HA/F (16.3 ± 0.3%); Nano-HA (15.3 ± 0.6%) and F alone control (15.4 ± 1.3%)., (© 2022. The Author(s).)

Introduction: The acquired pellicle formation is the first step in dental biofilm formation. It distinguishes dental biofilms from other biofilm types., Objective: To explore the influence of salivary pellicle formation before biofilm formation on enamel demineralization., Methodology: Saliva collection was approved by Indiana University IRB. Three donors provided wax-stimulated saliva as the microcosm bacterial inoculum source. Acquired pellicle was formed on bovine enamel samples. Two groups (0.5% and 1% sucrose-supplemented growth media) with three subgroups (surface conditioning using filtered/pasteurized saliva; filtered saliva; and deionized water (DIW)) were included (n=9/subgroup). Biofilm was then allowed to grow for 48 h using Brain Heart Infusion media supplemented with 5 g/l yeast extract, 1 mM CaCl2.2H2O, 5% vitamin K and hemin (v/v), and sucrose. Enamel samples were analyzed for Vickers surface microhardness change (VHNchange), and transverse microradiography measuring lesion depth (L) and mineral loss (∆Z). Data were analyzed using two-way ANOVA., Results: The two-way interaction of sucrose concentration × surface conditioning was not significant for VHNchange (p=0.872), ∆Z (p=0.662) or L (p=0.436). Surface conditioning affected VHNchange (p=0.0079), while sucrose concentration impacted ∆Z (p<0.0001) and L (p<0.0001). Surface conditioning with filtered/pasteurized saliva resulted in the lowest VHNchange values for both sucrose concentrations. The differences between filtered/pasteurized subgroups and the two other surface conditionings were significant (filtered saliva p=0.006; DIW p=0.0075). Growing the biofilm in 1% sucrose resulted in lesions with higher ∆Z and L values when compared with 0.5% sucrose. The differences in ∆Z and L between sucrose concentration subgroups was significant, regardless of surface conditioning (both p<0.0001)., Conclusion: Within the study limitations, surface conditioning using human saliva does not influence biofilm-mediated enamel caries lesion formation as measured by transverse microradiography, while differences were observed using surface microhardness, indicating a complex interaction between pellicle proteins and biofilm-mediated demineralization of the enamel surface.

Aims: To evaluate the diagnostic abilities of near-infrared light transillumination (using the DIAGNOcam) and bitewing radiographs in detecting cavitated proximal carious lesions in primary molars., Subjects and Methods: The study was a cross-sectional analytical, clinical study. The proximal surfaces of primary molars of healthy 5- to 8-year-old children were radiographically screened for the presence of carious lesions in the enamel or outer third of dentin (D1). Two trained and calibrated examiners evaluated the depth of caries in bitewing radiographs and DIAGNOcam images and then verified the presence of cavitation by direct visual examination using the "International Caries Detection and Assessment System" after temporary tooth separation., Results: A total of 236 proximal lesions were included in the study. Most of the clinically cavitated lesions (51.9%) were D1 radiographically and in outer dentin lesions (scores 3 and 4) by the DIAGNOcam (37% and 48.1%, respectively). Although DIAGNOcam showed higher sensitivity (0.852) compared to the radiographs (0.519), it showed slightly less specificity (0.569) compared to the radiographs (0.579). However, DIAGNOcam showed higher value of the area under the curve (AUC = 0.722; P < 0.001) compared to the radiographic method (AUC = 0.561; P = 0.308)., Conclusions: The DIAGNOcam showed higher sensitivity and better accuracy than bitewing radiographs in diagnosing cavitated proximal lesions in primary molars and can be generally considered as an alternative to radiographs to detect cavitation without the hazards of ionizing radiation in children., Competing Interests: None

There is growing evidence that C. albicans is associated with dental caries, but its role on caries development needs to be better clarified. Label="OBJECTIVE">To evaluate at the hard tissue level the effect of C. albicans on the cariogenic potential of S. mutans biofilms focusing on the mineral profile of induced carious lesions. This study also aimed to evaluate the effect of C. albicans on the acidogenic potential of S. mutans biofilms. METHODOLOGY Dual-species (CA+SM) and single-species biofilms (CA or SM) were grown on the surface of enamel slabs in the presence of glucose/sucrose supplemented culture medium for 24, 48 and 72 hours. Demineralization was evaluated through percentage of surface microhardness change (%SMC) and transversal microradiography analysis (ILM and LD) and pH of the spent medium was recorded daily. Data were analyzed by two-way ANOVA followed by Bonferroni correction. RESULTS%SMC was statistically different among the biofilms at each time point being the highest for SM biofilms and the lowest for CA biofilms which also differed from CA+SM biofilms [SM (24 h: 47.0±7.3; 48 h: 66.3±8.3; 72 h: 75.4±3.9); CA (24 h: 7.3±3.3; 48 h: 7.1±6.4; 72 h: 6.6±3.6); CA+SM (24 h: 35.9±7.39.1; 48 h: 47.2±9.5; 72 h: 47.6±9.5)]. pH of spent medium was statistically lower for SM biofilms compared to the other biofilms at each time point and remained constant over time while pH values increased from 24 to 72 h for both CA and CA+SM biofilms [SM (24 h: 4.4±0.1; 48 h: 4.4±0.1; 72 h: 4.5±0.1); CA (24 h: 6.9±0.3; 48 h: 7.2±0.2; 72 h: 7.5±0.2); CA+MS (24 h: 4.7±0.2; 48 h: 5.1±0.1; 72 h: 6.1±0.6)]. IML and LD for SM biofilms increased over time while no difference was observed from 24 to 72 h for the other biofilms. CONCLUSIONS The present data suggest that C. albicans has low enamel demineralization potential and the presence of C. albicans can reduce both the cariogenic and acidogenic potentials of S. mutans biofilms.

Soy beverages are promoted as healthy alternatives to bovine milk even though they can contain added sugar., Objectives: To compare enamel mineral content after consumption of bovine milk or a soy beverage in a double-blind, randomized, cross-over in situ clinical study., Materials and Methods: Human enamel slabs with subsurface lesions were prepared and inserted into intra-oral appliances worn by volunteers who consumed 200 ml of either bovine milk or a soy beverage over a 60 s period once a day for 15 days. Enamel lesion depth and mineral content were measured using transverse microradiography. Saliva samples were collected immediately after consuming the beverages and calcium, inorganic phosphate and fluoride levels analysed. Data were statistically analysed using a linear mixed model., Results: Depth of the enamel subsurface lesions increased by 7.1 ± 2.0 μm and mineral content decreased by 47 ± 22 vol% min.μm after consumption of the soy beverage indicating demineralization. However, after consumption of bovine milk the depth of the lesions decreased by 7.6 ± 3.5 μm and mineral content increased by 202 ± 43 vol% min.μm indicating remineralization. The changes were significantly different (p < 0.001) between the two beverages. Fluoride levels were similar in the saliva samples for both beverages, however the calcium and inorganic phosphate levels for the bovine milk group were significantly higher (p < 0.02) than those for the soy beverage group., Conclusions: In this randomized, double-blind in situ clinical trial consumption of a soy beverage demineralized enamel whereas bovine milk produced remineralization., Clinical Significance: Although soy beverages are promoted as healthy alternatives to bovine milk the added sugar and low calcium bioavailability of the soy drink makes frequent consumption a caries risk. (Trial registration no. ISRCTN19137849)., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

Objectives: The aim of this study was to assess the effect of Myracrodruon urundeuva All. and Qualea grandiflora Mart. leaves hydroalcoholic extracts on viability and metabolism of a microcosm biofilm and on enamel demineralization prevention., Methodology: Microcosm biofilm was produced on bovine enamel using inoculum from pooled human saliva mixed with McBain saliva, under 0.2% sucrose exposure, for 14 days. The biofilm was daily-treated with the extracts for 1 min. At the end, it was analyzed with respect to viability by fluorescence, CFU counting and extracellular polysaccharides (phenol-sulphuric acid colorimetric assay) and lactic acid (enzymatic assay) production. The demineralization was measured by TMR. The data were compared using ANOVA or Kruskal-Wallis (p<0.05)., Results: M. urundeuva All. at 100, 10 and 0.1 μg/mL and Q. grandiflora Mart. at 100 and 0.1 μg/mL reduced biofilm viability similarly to positive control (chlorhexidine) and significantly more than the negative-vehicle control (35% ethanol). M. urundeuva at 1000, 100 and 0.1 μg/mL were able to reduce both lactobacilli and mutans streptococci CFU counting, while Q. grandiflora (1000 and 1.0 μg/mL) significantly reduced mutans streptococci CFU counting. On the other hand, the natural extracts were unable to significantly reduce extracellular polysaccharides and lactic acid productions neither the development of enamel carious lesions., Conclusions: The extracts showed antimicrobial properties on microcosm biofilm, however, they had no effect on biofilm metabolism and caries protection.

This chapter describes methods for preparing samples of bone and bone cells for scanning electron microscopy (SEM). Backscattered electron (BSE) imaging is by far the most useful in the bone field, followed by secondary electrons (SE) and the energy dispersive X-ray (EDX) analytical modes. Samples may have 3D detail in a 3D surface, or be topography-free, polished or micromilled, resin-embedded block surfaces, or resin casts of space compartments surrounded by bone matrix. Methods for cells include fixation, drying, looking at undersides of bone cells, and metallic conductive coating. Maceration with alkaline bacterial pronase, hypochlorite, hydrogen peroxide, and sodium or potassium hydroxide to remove cells and unmineralized matrix is described in detail. Attention is given especially to methods for 3D BSE SEM imaging of bone samples. Recommendations are made for the types of resin embedding for BSE SEM imaging. Correlated confocal and SEM imaging of PMMA embedded bone requires the use of glycerol to coverslip. Cathodoluminescence (CL) mode SEM imaging is an alternative for visualizing fluorescent mineralizing front labels such as calcein and tetracyclines. Making spatial casts from PMMA or other resin-embedded samples is an important use of this material. Correlation with other imaging means, including microradiography and microtomography is important. Shipping wet bone samples between labs is best done in glycerol. Control of the vacuum pressure in the SEM sample chamber (now generally available) can be used to eliminate "charging" problems which were common, for example, with large, complex, cancellous bone samples.

This chapter describes the use of point projection digital microradiography for rapid imaging and quantitation of bone mineral content in mice.

The purpose of this study was to compare optical properties of root caries under two observing conditions using swept-source optical coherence tomography (SS-OCT). In vitro and natural root caries were observed by SS-OCT under wet and dry conditions, followed by confocal laser scanning microscope (CLSM) and transverse microradiography (TMR). Signal intensity (SI), distance between SI peaks (SI-distance) and optical lesion depth were obtained from OCT. Lesion depth was measured from CLSM; lesion depth (LD TMR ) and mineral loss (ML) were obtained from TMR. In vitro root caries under wet and dry conditions showed different OCT images and SI patterns. Lesion depth of OCT and that of CLSM, SI-distance and LD TMR , LD TMR and ML significantly correlated. Under dry conditions, half natural root caries showed similar OCT images and SI patterns as in vitro root caries. The base of demineralized dentin could be detected more clearly under dry conditions than under wet conditions.

Objectives: To compare and explore the dose-response of phytate-containing 1150 ppm fluoride toothpastes on model caries lesions and to determine the impact of zinc ions., Methods: This was a single-centre, randomised, blinded (examiner/laboratory analyst), six-treatment, four-period crossover, in situ study in adults with a removable bilateral maxillary partial denture. Study treatments were toothpastes containing: 0.425% phytate/F; 0.85% phytate/F; 0.85% phytate/Zn/F; F-only; Zn/F and a 0% F placebo. Where present, F was 1150 ppm as NaF; Zn was 0.3% as ZnCl 2 . Human enamel specimens containing early-stage, surface-softened (A-lesions) or more advanced, subsurface (B-lesions) caries lesions were placed into the buccal flanges of participants' modified partial denture (one of each lesion type per side). A-lesions were removed after 14 days of twice-daily treatment use; B-lesions were removed after a further 14 days. A-lesions were analysed for surface microhardness recovery. Both lesion types were analysed by transverse microradiography and for enamel fluoride uptake, with B-lesions additionally analysed by quantitative light-induced fluorescence. Comparison was carried out using an analysis of covariance model., Results: Statistically significant differences between 1150 ppm F and the placebo toothpastes (p < 0.05) were shown for all measures, validating the model. No differences between fluoride toothpastes were observed for any measure with little evidence of a dose-response for phytate. Study treatments were generally well-tolerated., Conclusions: Results suggest phytate has little impact on fluoride's ability to promote early-stage lesion remineralisation or prevent more advanced lesion demineralisation in this in situ caries model. Similarly, results suggest zinc ions do not impair fluoride efficacy., Clinical Significance: Toothpastes may contain therapeutic or cosmetic agents that could interfere with fluoride's caries prevention efficacy. The present in situ caries study has demonstrated that phytate, added to provide enhanced extrinsic stain removal/prevention, and zinc, added to inhibit malodour, do not impair fluoride efficacy., (Copyright © 2018 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Objectives: To assess the accuracy of near-infrared-light transillumination (DIAGNO) compared to visual-tactile (VT) and radiographic (RA) evaluation of proximal carious lesions adjacent to composite restorations in vitro., Methods: Two hundred extracted posterior permanent human teeth with occluso-proximal composite restorations were allocated to 50 groups of four posterior teeth, and mounted in a pilot-tested diagnostic model in a dummy head. The teeth were independently assessed by two examiners. Transverse microradiography and visual assessment served as reference tests to detect any lesions (prevalence 24%) and cavitated lesions (18%), respectively, adjacent to restorations. Sensitivity, specificity, positive and negative predictive values and the area under the receiver-operating-characteristics curve (AUC) were calculated., Results: To detect any proximal carious lesions adjacent to composite, the mean sensitivity/specificity were 0.63/0.95 for DIAGNO, 0.70/0.88 for RA when lesions radiographically extending into enamel and dentin were considered, 0.26/0.98 for RA when only lesions extending into dentin were considered, and 0.31/0.96 for VT. For cavitated lesions adjacent to proximal composite restorations, these values were RA (enamel and dentin) 0.84/0.88, RA (dentin) 0.34/0.99, DIAGNO 0.69/0.94 and VT 0.40/0.97. AUC did not differ significantly between RA and DIAGNO, while VT showed significantly lower values (p < 0.05)., Conclusion: Within the limitations of this study, DIAGNO seems useful for detecting proximal carious lesions adjacent to restorations., Clinical Relevance: Near-infrared-light transillumination could be used as a radiation-free adjunct or alternative to RA for detecting carious lesions adjacent to composite restorations., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

This study aimed to test the hypotheses that (i) a parameter related to permeability, α d (ratio of squared water volume by the nonmineral volume) is, among all major component volumes (mineral, water, and organic volumes) the best predictor of quinoline infiltration in natural enamel caries (NEC), and (ii) the pore volume fraction infiltrated by quinoline (V qui ) in NEC is much lower than previous estimates that neglected water and organic enamel volumes. Mineral and nonmineral volumes and α d were measured at 341 histological points (from 20 approximal NEC lesions), and transport of quinoline was tracked by orientation-independent polarizing microscopy. R 2 values of V qui were 0.596 (α d ), 0.033 (mineral volume), 0.474 (water volume), and 0.011 (organic volume). V qui values were 23% (body of the lesion), 7% (dark zone), and 9% (translucent zone), lower than previous estimates (with high effect size). Transport of quinoline occurred both parallelly and perpendicularly to prism paths, and dark zones were seen where only transport parallel to prisms occurred. In conclusion, α d was the main predictor of quinoline infiltration, but it differed from the water volume with a small effect size, and the pore volume fraction with quinoline was much lower than previous estimates., (© 2017 Wiley Periodicals, Inc.)

Detection of occlusal caries with visual examination using ICDAS correlates strongly with histology under stereomicroscopy (SM), but dentin aspects under SM are ambiguous regarding mineral content. Thus, our aim was to test two null hypotheses: SM and microradiography result in similar correlations between ICDAS and histology; SM and microradiography result in similar positive (PPV) and negative predictive values (NPV) of ICDAS cut-off 1-2 (scores 0-2 as sound) with histological threshold D3 (demineralization in the inner third of dentin). Occlusal surfaces of extracted permanent teeth (n = 115) were scored using ICDAS. Undemineralized ground sections were histologically scored using both SM without contrast solution and microradiography after immersion in Thoulet's solution 1.47 for 24 h (MRC). Correlation between ICDAS and histology differed from SM (0.782) to MRC (0.511) (p = 0.0002), with a large effect size "q" of 0.49 (95% CI: 0.638/0.338). For ICDAS cut-off 1-2 and D3, PPV from MRC (0.56) was higher than that from SM (0.28) (p< 0.00001; effect size h = 0.81), and NPV from MRC (0.72) was lower than that from SM (1,00) (p < 0.00001; effect size h = 1.58). In conclusion, SM overestimated the correlation between ICDAS and lesion depth, and underestimated the number of occlusal surfaces with ICDAS cut-off 1-2 and deep dentin demineralization.

Objectives: Secondary caries limits the longevity of restorations and is thought to be associated with faulty restorations, e.g. dentin-restoration interfacial gaps. Recent evidence indicates that loading of restorations might aggravate the effects of gaps on interfacial mineral loss. It is unclear if this effect of loading is dose-dependent or not, and if restoration material properties like elasticity moderate the association between load and mineral loss. We hypothesized that mineral loss of secondary lesions increases with increasing load, and that this association is moderated by the elastic modulus of the placed restoration material., Methods: Dentin-restoration specimens with simulated interfacial gaps were submitted to cariogenic Lactobacillus-rhamnosus-biofilms for 10days, and concurrently loaded with different loads (0/42/84/126g per specimen, n=12/group). Two different composites (LEC: low elastic-modulus composite, HEC: high elastic-modulus composite) were employed. Transversal microradiography was used to evaluate the superficial and interfacial (wall) lesion mineral loss. Generalized linear modeling (GLM) was used to evaluate the association between loading, material and their interaction on mineral loss., Results: Surface mineral loss was not significantly associated with loading, material, or their interaction (p-values ranged between p=0.062 and 0.526). For deep interfacial (wall) lesions, the applied load (p=0.023) but not the material (p=0.382) showed a significant effect. The interaction between both significantly affected mineral loss (p=0.01). Loads of ≥84g per specimens were associated with higher wall lesion mineral loss., Conclusions: Loads above a certain threshold significantly increased interfacial (wall) lesion mineral loss. This association was moderated by the elasticity of the placed restoration materials., Clinical Significance: The clinical relevance of our findings remains unclear, as future studies are needed to understand how exactly both load and material elasticity affect secondary lesion induction., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

Purpose: The aim of this study was to analyze cone beam computed tomography (CBCT) densitometries of maxillary sinuses augmented with human bone allograft. In addition, previously obtained microradiographic specimens were used to verify the diagnostic potential of CBCT., Materials and Methods: A two-stage protocol was conducted in 21 consecutive patients, all with a crestal bone height < 2 mm. Mineralized human bone allograft particles were used to augment sinuses using a lateral window approach. A succession of CBCT scans of the maxilla were taken before surgery, immediately after sinus augmentation, and 6, 10, and 18 months after implant placement. Using virtual probes, CBCT images taken at 6, 8, and 10 mm from the crestal surface were processed with medical imaging software and expressed as gray level (GL)., Results: A total of 24 sinus augmentation procedures were performed in 21 patients. The average values of CBCT-GL ranged from 571 to 654, presenting the maximum value at 8 mm immediately after implant placement and the minimum value at 6 mm after 10 months. Furthermore, it was found that the graft mineral content decreased over time, completely disappearing between 10 and 11 months., Conclusion: CBCT and the medical imaging software employed for imaging visualization are reliable tools to study biomaterial behavior after sinus augmentation procedures. In addition, results from this study demonstrate that a complete resorption of human bone allograft is possible. Due to the limited sample size, further clinical and morphometric studies are needed.

Adult human mesenchymal stem cells show structural rearrangements of their cytoskeletal network during mechanically induced differentiation toward various cell types. In particular, the alignment of acto-myosin fibers is cell fate-dependent and can serve as an early morphological marker of differentiation. Quantification of such nanostructures on a mesoscopic scale requires high-resolution imaging techniques. Here, we use small- angle x-ray scattering with a spot size in the micro- and submicrometer range as a high-resolution and label-free imaging technique to reveal structural details of stem cells and differentiated cell types. We include principal component analysis into an automated empirical analysis scheme that allows the local characterization of oriented structures. Results on freeze-dried samples lead to quantitative structural information for all cell lines tested: differentiated cells reveal pronounced structural orientation and a relatively intense overall diffraction signal, whereas naive human mesenchymal stem cells lack these features. Our data support the hypothesis of stem cells establishing ordered structures along their differentiation process., (Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.)

Objective: Interpolation flaps are commonly used in plastic surgery to cover wide and deep defects. The need to, wait for 2 to 3 weeks until the division of the pedicle still, however, poses a serious challenge, not only extending treatment and hospital stay, but also increasing hospital expenses. To solve this problem, we have aimed to use the angiogenic potential of stem cells to selectively accelerate neovascularization with a view to increasing the viability of interpolation flaps and achieving early pedicle removal., Materials and Methods: A total of 32 rats were allocated to 2 groups as control (N = 16) and experiment (N = 16). The cranial flaps 6 × 5 cm in size located on the back of the rats were raised. Then, a total suspension containing 3 × 10(6) adipose-derived mesenchymal stem cells (ADSC) tagged with a green fluorescent protein (GFP) was injected diffusely into the distal part of the flap, receiving bed, and wound edges. In the control group, only a medium solution was injected into the same sites. After covering the 3 × 5 cm region in the proximal part of the area where the flap was removed, the distal part of the flap was adapted to the uncovered distal area. The pedicles of 4 rats in each group were divided on postoperative days 5, 8, 11, and 14. The areas were photographed 7 days after the pedicles were released. The photographs were processed using Adobe Acrobat 9 Pro software (San Jose, CA) to measure the flap survival area in millimeters and to compare groups. Seven days after the flap pedicle was divided, the rats were injected with 250 mCi Tc-99 mm (methoxy-isobutyl-isonitrie) from the penile vein, and scintigraphic images were obtained. The images obtained from each group were subjected to a numerical evaluation, which was then used in the comparison between groups. The flaps were then examined by histology to numerically compare the number of newly formed vessels. Neovascularization was also assessed by microangiography. In addition, radiographic images were obtained by mammography and evaluated quantitatively., Results: An evaluation of statistical results revealed a significant increase in the flap survival area of the group on stem cell treatment in comparison to the control group. In scintigraphic examinations, the rate of radioactive substance retention was significantly higher in the stem cell group, relative to the control group. Histopathologic examination showed that the capillary density in the stem cell group was higher than that in the control group. Green fluorescent protein had been used to label ADSC in the experiment and it was found by immunofluorescence staining that endothelial samples of control animals did not have GFP (+) cells, whereas all the animals in the experiment group had GFP (+) cells. The comparison of microangiographic images of the experiment and control groups demonstrated significantly elevated vascularity in the former, relative to the latter., Discussion: It has been established in the current study that ADSC injection worked well in speeding up the neovascularization of interpolated flaps and reducing the time of pedicle division. It seems possible to minimize the morbidity of interpolated skin flaps with mesenchymal stem cell therapy at an appropriate dose and for an appropriate length of time.

Objectives: The aim of the current study was to evaluate the application of two advanced noise-reduction algorithms for dental micro-CT images and to implement a comparative analysis of the performance of new and current denoising algorithms., Methods: Denoising was performed using gaussian and median filters as the current filtering approaches and the block-matching and three-dimensional (BM3D) method and total variation method as the proposed new filtering techniques. The performance of the denoising methods was evaluated quantitatively using contrast-to-noise ratio (CNR), edge preserving index (EPI) and blurring indexes, as well as qualitatively using the double-stimulus continuous quality scale procedure., Results: The BM3D method had the best performance with regard to preservation of fine textural features (CNREdge), non-blurring of the whole image (blurring index), the clinical visual score in images with very fine features and the overall visual score for all types of images. On the other hand, the total variation method provided the best results with regard to smoothing of images in texture-free areas (CNRTex-free) and in preserving the edges and borders of image features (EPI)., Conclusions: The BM3D method is the most reliable technique for denoising dental micro-CT images with very fine textural details, such as shallow enamel lesions, in which the preservation of the texture and fine features is of the greatest importance. On the other hand, the total variation method is the technique of choice for denoising images without very fine textural details in which the clinician or researcher is interested mainly in anatomical features and structural measurements.

Objective: In this study we give a preliminary study of a rationally designed small peptide, which is based on the enamel matrix protein amelogenin, to investigate its effect on remineralization of initial enamel caries lesions., Design: A novel peptide was designed and synthesized to investigate its effects on the remineralization of initial enamel carious lesions during pH cycling that simulates intra-oral conditions. Initial lesions were created in bovine enamel blocks, which were then pH-cycled for 12 days in the presence of 25μM peptide, 1g/L NaF (positive control), 50mM HEPES buffer(negative control). Before and after pH cycling, enamel blocks were analyzed by surface microhardness testing, polarized light microscopy and transverse microradiography., Results: Percentage of surface microhardness recovery (SMHR%) after pH cycling was significantly higher in peptide group than HEPES group. Lower lesion depth and less mineral mineral loss were found in peptide or NaF treatment groups after the cycling, and were significantly different to HEPES group. No significant differences were observed between the blocks treated with peptide and those treated with NaF., Conclussion: This study provides in vitro evidence that this amelogenin based peptide promotes enamel caries remineralization, offering a promising remineralizing biomaterial in initial enamel carious treatment., (Copyright © 2015 Elsevier Ltd. All rights reserved.)

A compact laboratory proximity soft X-ray microscope providing submicrometer spatial resolution and digital image registration is described. The microscope consists of a laser-plasma soft X-ray radiation source, a Schwarzschild objective to illuminate the test sample, and a two-coordinate detector for image registration. Radiation, which passes through the sample under study, generates an absorption image on the front surface of the detector. Optical ceramic YAG:Ce was used to convert the X-rays into visible light. An image was transferred from the scintillator to a charge-coupled device camera with a Mitutoyo Plan Apo series lens. The detector's design allows the use of lenses with numerical apertures of NA = 0.14, 0.28, and 0.55 without changing the dimensions and arrangement of the elements of the device. This design allows one to change the magnification, spatial resolution, and field of view of the X-ray microscope. A spatial resolution better than 0.7 μm and an energy conversion efficiency of the X-ray radiation with a wavelength of 13.5 nm into visible light collected by the detector of 7.2% were achieved with the largest aperture lens.

Objectives: To describe the tissue reactions at the bone-titanium interface of orthodontic miniplates in humans., Materials and Methods: Forty-two samples, consisting of tissue fragments attached or not to miniplates or their fixation screws, were collected from 24 orthodontic patients treated with miniplate anchorage, at the time of removal of their miniplates. The samples were embedded in methylmethacrylate and cut into undecalcified sections which were submitted to microradiographic analysis. The sections were also stained and examined under ordinary light., Results: Three types of reactions were observed both on the histological sections and on the microradiographs. 1. The majority of the stable miniplates were easy to remove (34/42). The tissue samples collected consisted mainly in mature lamellar bone with some medullary spaces containing blood vessels, 2. two screws were highly osseointegrated and required the surgeon to remove them by trephining (2/42). They were surrounded by bone tissue which extended to the miniplate. The histological features were similar to the previous group, though the bone-screw contact was higher, and 3. in six samples obtained after unstable miniplate removal during the treatment, we observed either some woven bone trabeculae or loose connective tissue, without any histological sign of inflammation., Limitations and Conclusion: For evident ethical reasons, our data were limited by the size of the tissue fragments and the limited number of patients and variety of clinical presentations. The healing reactions consisted mainly in mature lamellar bone tissue sparsely in contact with the screw or the miniplate, with signs of a moderate remodelling activity., (© The Author 2014. Published by Oxford University Press on behalf of the European Orthodontic Society. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

This in vitro study investigated whether restoration materials and adhesives influence secondary caries formation in gaps using a short-term in vitro biofilm model. Sixty enamel-dentin blocks were restored with 6 different restoration materials with or without adhesives (n = 10 per group) with a gap: 1) Clearfil AP-X composite, 2) Clearfil AP-X composite + SE Bond, 3) Clearfil AP-X composite + ProtectBond, 4) Filtek Silorane composite, 5) Filtek Silorane composite + Silorane System adhesive, or 6) Tytin amalgam. Specimens were subjected to an intermittent 1% sucrose biofilm model for 20 days to create artificial caries lesions. Lesion progression in the enamel-dentin next to the different materials was measured in lesion depth (LD) and mineral loss (ML) using transversal wavelength independent microradiography (T-WIM). A regression analysis was used to compare the LD and ML of the different restoration materials at 4 measurement locations: 1 location at the surface of the enamel, 1 location at the wall of the enamel, and 2 locations at the wall of the dentin. A statistically significant effect of AP-X composite with Protect Bond was found for LD and ML at the WallDentin1 location, leading to less advanced wall lesions. An additional finding was that gap size was also statistically significant at the 2 wall locations in dentin, leading to increasing lesion progression with wider gaps. In conclusion, adhesives can influence wall lesion development in gaps. Protect Bond showed significantly less caries progression compared to bare restoration materials or other adhesives in this short-term in vitro biofilm model., (© International & American Associations for Dental Research 2014.)

In order to simplify bone mineralization measurements, a system using radiographic films has been updated with a digital detector. The objective of this paper was to validate this new device. Technologies and physical phenomena involved in both systems (radiographic films and digital detector) are different. The methodology used to compare the two systems was based on image quality and assessed on two main parameters: contrast to noise ratio and spatial resolution. Results showed that the contrast to noise ratio was similar between the two systems, provided that acquisition parameters were optimized. With regard to spatial resolution, a magnification factor of at least 4 or more was required to achieve the same resolution as films. A final validation was also shown on a real image of a bone sample. The results showed that both systems have similar image quality performances, and the system using digital detector has several advantages (easier to use than films, no consumables and faster acquisition time).

Nuclear factor kappa B (NF-κB) signaling plays critical roles in many physiological and pathological processes, including regulating organogenesis. Down-regulation of NF-κB signaling during development results in hypohidrotic ectodermal dysplasia. The roles of NF-κB signaling in tooth development, however, are not fully understood. We examined mice overexpressing IKKβ, an essential component of the NF-κB pathway, under keratin 5 promoter (K5-Ikkβ). K5-Ikkβ mice showed supernumerary incisors whose formation was accompanied by up-regulation of canonical Wnt signaling. Apoptosis that is normally observed in wild-type incisor epithelium was reduced in K5-Ikkβ mice. The supernumerary incisors in K5-Ikkβ mice were found to phenocopy extra incisors in mice with mutations of Wnt inhibitor, Wise. Excess NF-κB activity thus induces an ectopic odontogenesis program that is usually suppressed under physiological conditions., (© International & American Associations for Dental Research 2014.)

Objective: To determine the relative ability of various F-containing products to protect enamel against the initiation and progression of tooth surface loss due to erosive acid challenges., Methods: Cores of enamel were prepared from extracted human teeth, soaked in pooled human saliva (pellicle formation), and then treated in a 1:3 slurry (product:saliva) of either OTC level (1100 ppm F) or prescription level (5000 ppm F) products, followed by a standardized erosion cycling procedure (five days of cycling) that included 10-minute challenges with an erosive dietary acid (1% citric acid at pH 2.3) applied 60 minutes after each dentifrice treatment (repeated four times per day). Enamel surface loss was measured using transverse microradiography. Two studies were conducted. Study 1 included: A) 1100 ppm F as NaF; B) 1100 ppm F as stabilized SnF; C) 5000 ppm F as NaF; and D) 5000 ppm F as NaF + acidulated phosphate. Study 2 included: 1) 1100 ppm F as stabilized SnF; 2) 5000 ppm F as NaF + tricalcium phosphate; and 3) 1100 ppm F as NaF., Results: Study 1: Treatment B (1100 ppm F as SnF), where specimens lost only 8.0 μm of the enamel surface, was significantly more effective than Treatments A, C, and D at protecting enamel against the initiation and progression of erosive acid damage (p < 0.05). Specimens treated with product A exhibited 22.8 (1.25) μm (mean ± sem) of enamel loss; 20.0 (0.71) μm of enamel loss with treatment C and 24.0 (1.4) μm of enamel loss with Treatment D. Study 2 also demonstrated significantly greater erosion protection with the stabilized SnF2 dentifrice (p < 0.05), with only 5.8 (1.93) μm of tooth surface loss, while groups 2 and 3 lost 19.8 (0.75) μm and 18.0 (2.16) μm, respectively., Conclusion: Results from both studies demonstrated the OTC dentifrice formulated with stabilized SnF2 provides significantly greater protection against erosive acid attack compared to some of the most popular prescription level (5000 ppm F) fluoride treatments available.

Artificially inducing dentinal lesions mimicking those remaining after selective excavation should allow to investigate the effects and limits of such selective excavation, for example regarding the mechanical properties of treated teeth or the remineralisation of sealed residual lesions. Such analyses might otherwise be limited by the variability of natural lesions or ethical and practical concerns. This study compared different demineralisation protocols for their suitability to induce lesions similar to natural residual caries. Twelve natural deep lesions were excavated until leathery dentin remained, and analysed for their mineral loss (ΔZ), lesion depth (LD), mineral loss ratio (R), the slope of the mineral gradient and their nano-hardness profile. Artificial lesions were induced using four different demineralisation protocols (acetic acid pH = 4.95; 0.1 M lactic acid gel pH = 5.0; 0.5 M ethylenediaminetetraacetic acid pH = 7.2; Streptococcus mutans biofilms) and their depths monitored over different demineralisation times. Lesions with depths most according to those of natural lesions were analysed using transversal microradiography. Lesions induced by acetic acid solution did not significantly differ with regards to LD, ΔZ, R and mineral profile. Seven dentin specimens were subsequently submitted to a moderately acidic (pH = 5.3) methylhydroxydiphosphonate-buffered acetate solution for 12 weeks. Natural and artificial residual lesions were similarly deep (mean ± SD: LD = 626 ± 212 and 563 ± 88 µm), demineralised (R = 19.5 ± 4.7 and 29.8 ± 4.1%), showed a flat and continuous mineral gradient (slope = 0.10 ± 0.05 and 0.13 ± 0.06 vol%/µm) and did not significantly differ regarding their nano-hardness profile. The described protocol induces lesions with mineral content and mechanical properties similar to natural residual lesions.

The aim of this double-blind, randomized, cross-over in situ study was to compare the remineralizing effects induced by the application of casein phosphopeptide-stabilized amorphous calcium phosphate complexes (CPP-ACP)-containing cream (without fluoride) after the use of fluoride toothpaste with the prolonged use of fluoride toothpaste on enamel caries lesions in situ. During each of three experimental legs of 4 weeks, 13 participants wore intra-oral mandibular appliances with 8 pre-demineralized bovine enamel specimens in the vestibular flanges mimicking either 'easily cleanable' or 'proximal' surfaces (n = 312). The three randomly allocated treatments were as follows: (1) application of CPP-ACP-containing cream (GC Tooth Mouse, non-fluoride) after the use of fluoride toothpaste (1,400 ppm NaF; TM), (2) prolonged application of fluoride toothpaste (1,400 ppm NaF; positive control, PC) and (3) prolonged application of fluoride-free toothpaste (negative control, NC). Additionally, one of each of the two flanges was brushed twice daily with the respective toothpaste. The differences in integrated mineral loss as assessed by transversal microradiography were calculated between values before and after the in situ period. Changes in mineral loss were analysed for those pairs of subgroups differing in only one of the three factors (intervention, brushing and position). The PC treatment induced a significantly higher mineral gain compared with the TM and NC treatments. No significant differences between TM and NC for both positions were observed. In conclusion, the additional use of a CPP-ACP-containing cream seems to be less efficacious in remineralizing caries lesions than the prolonged application of fluoride toothpaste., (© 2014 S. Karger AG, Basel.)

Micro-computed tomography (micro-CT)-a version of X-ray CT operating at high spatial resolution-has had a considerable success for the investigation of trabecular bone micro-architecture. Currently, there is a lot of interest in exploiting CT techniques at even higher spatial resolutions to assess bone tissue at the cellular scale. After recalling the basic principles of micro-CT, we review the different existing system, based on either standard X-ray tubes or synchrotron sources. Then, we present recent applications of micro- and nano-CT for the analysis of osteocyte lacunae and the lacunar-canalicular network. We also address the question of the quantification of bone ultrastructure to go beyond the sole visualization.

Objectives: Selective caries removal involves sealing of carious dentine beneath restorations, which might decrease their marginal integrity and increase the susceptibility for secondary caries and microleakage. The present study compared these marginal characteristics of restorations in selectively and completely excavated teeth., Methods: In 32 premolars, shallow and deep artificial lesions were created on pulpo-axial walls of mesial-distal-occlusal cavities, with mesial and distal margins located in enamel and dentine, respectively. Demineralised dentine was either removed or left before adhesively restoring the teeth (n=8), which were then submitted to thermo-mechanical cycling. The integrity of gingivo-cervical margins was assessed using scanning electron microscopy. In half of each margin, caries was induced adjacent to restorations using a continuous-culture biofilm model, and resulting lesions were evaluated using transversal microradiography. The other half of each margin was used to assess microleakage., Results: Integrity or microleakage of margins located in enamel did not differ significantly between groups, and bacterial biofilms did not induce distinct caries lesions in enamel. Dentinal margins in teeth with deep compared with shallow lesions showed a significantly higher proportion of marginal imperfections, gaps and microleakage (p≤0.05, Mann-Whitney/χ(2)-test). In contrast, neither marginal integrity nor microleakage differed significantly between completely and selectively excavated teeth (p>0.05). Dentinal mineral loss adjacent to restorations did not differ significantly between groups (p>0.80)., Conclusions: The marginal characteristics of restorations were affected by the depth of sealed or excavated lesions, but not by the performed caries excavation. This study did not find selective excavation detrimental for restoration integrity in vitro., Clinical Significance: Selective excavation of deep lesions was shown to reduce pulpal risks, whilst leaving caries beneath restorations is feared to compromise the marginal characteristics of the subsequently placed restoration. Based on the present in vitro study, such assumptions cannot be supported., (Copyright © 2014 Elsevier Ltd. All rights reserved.)

Aim: This study aimed to evaluate the healing of peri-implant defects grafted with microparticles (MPs)., Material and Methods: Six domestic pigs received nine standardized defects at the calvaria, and an implant was inserted in the middle of each defect. The space between the implant and lateral bone portion was filled with MP pellets (n = 18) or MP supernatant (n = 18) or left unfilled (n = 18). After 14 and 28 days, three animals were sacrificed and specimens removed for further processing. Samples were microradiographically and histologically analysed. In addition, we immunohistochemically stained for anti-vWF as a marker of angiogenesis., Results: In the case of bone regeneration and vessel formation, the null hypothesis can be partially rejected. After 14 and 28 days, no significant difference was observed within groups regarding de novo bone formation, bone density and osseointegration. However, superior vessel formation was found at both time points., Conclusion: Microparticles represent a promising treatment option to accelerate peri-implant vessel formation. Further studies are needed to investigate the regenerative properties of MPs more precisely., (© 2014 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Introduction: Epinephrine potentiates and prolongs the efficacy of local anesthetics by reducing blood flow. We investigated the effect of epinephrine on the pharmacokinetics of lidocaine and the pulpal blood volume after maxillary infiltration anesthesia in rats., Methods: We measured the (14)C-radioactivity and (14)C-distribution in the maxilla and the dental pulp after the injection of 2% (14)C-lidocaine with or without 10 μg/mL epinephrine (n = 7) into the palatine mucosa proximal to the first molar. The blood volume in the pulp was measured using (99m)Tc-pertechnetate (n = 5)., Results: When lidocaine was injected together with epinephrine, the lidocaine became widely distributed throughout the maxilla and was observed mainly in the first molar pulp. The lidocaine amount in the dental pulp at 10-60 minutes was more than 2 times higher than that after the injection of lidocaine alone. The relative pulpal blood volume after 20 minutes decreased to 63.1% of the value after the injection of lidocaine alone., Conclusions: We found that lidocaine had infiltrated into the molar pulp after infiltration anesthesia. Furthermore, our results suggested that epinephrine augmented the retention of lidocaine in the pulp., (Copyright © 2014 American Association of Endodontists. Published by Elsevier Inc. All rights reserved.)

Objectives: The effect of direct restorative materials on caries lesion formation was investigated with an 8-week in situ study with split-mouth design, testing the hypothesis that no difference in mineral loss next to a restoration would be found between different composite-based-materials and amalgam., Methods: Six groups (n=18) of restored dentine samples were prepared using amalgam, a microhybrid, a nanohybrid and a silorane composite. The composites were adhesively bonded with systems with or without an antibacterial monomer (Clearfil-SE-Protect, Clearfil-SE-bond, respectively), except for the silorane group (Silorane-System-Adhesive). Non-restored dentine samples were used as control (primary caries). Samples were inserted into slots, in lower prosthesis especially made for the experiment. Subjects were instructed to dip the lower prosthesis in a sucrose solution 4 times per day. At baseline and 8 weeks, samples were radiographed extra-orally and the integrated mineral loss was calculated. Data were statistically analyzed using multiple linear regression with a multilevel model (p=0.05)., Results: Nine subjects were selected, and only outer lesions were observed. The hypothesis was partially rejected, as the microhybrid composite bonded with the antibacterial system and the nanohybrid composite presented statistically significant lower mineral loss compared to amalgam. Also, no significant differences were seen for these groups compared to control., Conclusion: Within the limits of this study, the restorative material may influence outer lesion progression. Amalgam was not found to be related to lower secondary caries progression in dentine compared to composite-based materials after 8 weeks in situ., Clinical Significance: Although patient factors play a major role in caries progression, the restorative material may affect outer secondary lesion progression., (Copyright © 2014 Elsevier Ltd. All rights reserved.)

Purpose: Cone beam computed tomography (CBCT) and microradiographic analyses were comparatively performed in maxillary sinus augmentation to preliminarily verify the diagnostic potential of CBCT on the evaluation of bone regeneration., Materials and Methods: A two-stage protocol was conducted in 19 consenting patients, all having the crestal bone ≤2 mm, in private dental office. Mineralized human bone allograft particles were used to augment sinus using lateral window approach. A succession of CBCT scans of the maxilla was taken before surgery, after sinus augmentation, and immediately after implant insertion. Using virtual probes, CBCT data were processed by medical imaging software and expressed as gray level (GL). A bone core biopsy was taken at implant placement, 6 months after surgery. Microradiography of transverse sections, taken 6, 8, and 10 mm from the crestal surface, of methacrylate-embedded biopsies was performed to analyze and to evaluate the mineralized material amount (MM%)., Results: A total of 21 sinus augmentations were performed. CBCT (mean GL: 646-693) data were not statistically different when comparing 6-, 8-, and 10-mm sites to after grafting/implant-insertion values. Furthermore, microradiographic (mean MM%: 45.3-48.3) data were not statistically different comparing 6-, 8-, and 10-mm sites, due to variation of values among patients. A GL and MM% parallelism was identified considering each patient, instead. A significant correlation (p < .001) between GL and MM% was found after both Wilcoxon test for paired data and simple linear regression analysis., Conclusions: The preliminary result clearly demonstrated the predictability of the CBCT analysis. Due to the limited sample and great variations of the MM% recorded in patients, further clinical and morphometric studies are needed to fulfill diagnostic expectations., (© 2012 Wiley Periodicals, Inc.)

This in situ study investigated whether there is a relationship between gap size and wall lesion development in dentin next to 2 composite materials, and whether a clinically relevant threshold for the gap size could be established. For 21 days, 14 volunteers wore a modified occlusal splint containing human dentin samples with 5 different interfaces: 4 gaps of 50 µm, 100 µm, 200 µm, or 400 µm and 1 non-bonded interface without a gap. Eight times a day, the splint with samples was dipped in a 20% sucrose solution for 10 minutes. Before and after caries development, specimens were imaged with transversal wavelength-independent microradiography (T-WIM), and lesion depth (LD) and mineral loss (ML) were calculated at the 5 different interfaces. After correction for the confounder location (more mesial or distal), a paired t test clustered within volunteers was performed for comparison of gap widths. Results showed no trend for a relationship between the corrected lesion depth and the gap size. None of the differences in lesion depth for the different gap sizes was statistically significant. Also, the composite material (AP-X or Filtek Supreme) gave no statistically significant differences in lesion depth and mineral loss. A minimum gap size could not be established, although, in a non-bonded interface without a measurable gap, wall lesion development was never observed., (© International & American Associations for Dental Research.)

Objective: The aims of the present laboratory study were twofold: a) to investigate the suitability of Knoop and Vickers surface microhardness (SMH) in comparison to transverse microradiography (TMR) to investigate early enamel caries lesion formation; b) to compare the kinetics of caries lesion initiation and progression between human and bovine enamel., Design: Specimens (90×bovine and 90×human enamel) were divided into six groups (demineralization times of 8/16/24/32/40/48h) of 15 per enamel type and demineralized using a partially saturated lactic acid solution. SMH was measured before and after demineralization and changes in indentation length (ΔIL) calculated. Lesions were characterized using TMR. Data were analyzed (two-way ANOVA) and Pearson correlation coefficients calculated., Results: ΔIL increased with increasing demineralization times but plateaued after 40h, whereas lesion depth (L) and integrated mineral loss (ΔZ) increased almost linearly throughout. No differences between Knoop and Vickers SMH in their ability to measure enamel demineralization were observed as both correlated strongly. Overall, ΔIL correlated strongly with ΔZ and L but only moderately with the degree of surface zone mineralization, whereas ΔZ and L correlated strongly. Bovine demineralized faster than human enamel (all techniques)., Conclusions: Lesions in bovine formed faster than in human enamel, although the resulting lesions were almost indistinguishable in their mineral distribution characteristics. Early caries lesion demineralization can be sufficiently studied by SMH, but its limitations on the assessment of the mineral status of more demineralized lesions must be considered. Ideally, complementary techniques to assess changes in both physical and chemical lesion characteristics would be employed., (Copyright © 2014 Elsevier Ltd. All rights reserved.)

One-step incomplete excavation seals caries-affected dentin under a restoration and appears to be advantageous in the treatment of deep lesions. However, it is impossible to discriminate radiographically between intentionally left, arrested lesions and overlooked or active lesions. This diagnostic uncertainty decreases the acceptance of minimally invasive excavation and might lead to unnecessary re-treatment of incompletely excavated teeth. Radiopaque tagging of sealed lesions might mask arrested lesions and assist in discrimination from progressing lesions. Therefore, we microradiographically screened 4 substances (SnCl2, AgNO3, CsF, CsCH3COO) for their effect on artificial lesions. Since water-dissolved tin chloride (SnCl2×Aq) was found to stably mask artificial lesions, we then investigated its radiographic effects on progressing lesions. Natural lesions were incompletely excavated and radiopaque tagging performed. Grey-value differences (△GV) between sound and carious dentin were determined and radiographs assessed by 20 dentists. While radiographic effects of SnCl2×Aq were stable for non-progressing lesions, they significantly decreased during a second demineralization (p < .001, t test). For natural lesions, tagging with SnCl2×Aq significantly reduced △GV (p < .001, Wilcoxon). Tagged lesions were detected significantly less often than untagged lesions (p < .001). SnCl2×Aq was suitable to mask caries-affected dentin and discriminate between arrested and progressing lesions in vitro. Radiopaque tagging could resolve diagnostic uncertainties associated with incomplete excavation., (© International & American Associations for Dental Research.)

Transverse microradiography (TMR) and electron probe microanalysis (EPMA) are commonly used for characterizing dental tissues. TMR utilizes an approximately monochromatic X-ray beam to determine the mass attenuation of the sample, which is converted to volume percent mineral (vol%min). An EPMA stimulates the emission of characteristic X-rays from a variable volume of sample (dependent on density) to provide compositional information. The aim of this study was to compare the assessment of sound, demineralized, and remineralized enamel using both techniques. Human enamel samples were demineralized and a part of each was subsequently remineralized. The same line profile through each demineralized lesion was analyzed using TMR and EPMA to determine vol%min and wt% elemental composition and atomic concentration ratio information, respectively. The vol%min and wt% values determined by each technique were significantly correlated but the absolute values were not similar. This was attributable to the complex ultrastructural composition, the variable density of the samples analyzed, and the nonlinear interaction of the EPMA-generated X-rays. EPMA remains an important technique for obtaining atomic ratio information, but its limitations in determining absolute mineral content indicate that it should not be used in place of TMR for determining the mineral density of dental hard tissues.

Microautoradiography (MAR) is a conventional imaging method based on the daguerreotype. The technique is used to visualize the distribution of radionuclide-labeled compounds within a tissue section. However, application of the classical MAR method to plant tissue sections is associated with several difficulties. In this study, we report an MAR method applicable to fresh-frozen plant sections. Our method had two features: (i) the sample was kept frozen from plant tissue collection to radioisotope detection, making it possible to fix solutes without solvent exchange; and (ii) 1.2 µm thick polyphenylene sulfide film was inserted between the fresh-frozen plant section and the photosensitive nuclear emulsion to separate the section from the emulsion before autoradiography was conducted, which significantly improved the quality of the section until microscopic detection, the quality of the MAR image and the success rate. Then, the passage of cadmium (Cd) through vegetative rice stem tissue after 24 h of (109)Cd absorption was described for the first time using the MAR method. MAR clearly revealed the distribution of (109)Cd at the tissue level with high resolution. The (109)Cd concentration in phloem cells was found to be particularly high, whereas the xylem cells contained only small amounts of (109)Cd. The MAR method was also applicable for detecting (109)Cd and [(33)P]phosphate in roots. The MAR method developed here is expected to provide distribution images for a variety of compounds and ions in plant tissue., (© The Author 2014. Published by Oxford University Press on behalf of Japanese Society of Plant Physiologists. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

Objectives: To determine the role of Msx2 in craniofacial morphology and growth, we used a mouse model and performed a quantitative morphological characterization of the Msx2 (-/-) and the Msx2 (+/-) phenotype using a 2D cephalometric analysis applied on micrographs., Materials and Methods: Forty-four three-and-a-half-month-old female CD1 mice were divided into the following three groups: Msx2 (+/+) (n = 16), Msx2 (+/-) (n = 16), and Msx2 (-/-) (n = 12). Profile radiographs were scanned. Modified cephalometric analysis was performed to compare the three groups., Results: Compared with the wild-type mice, the Msx2 (-/-) mutant mice presented an overall craniofacial size decrease and modifications of the shape of the different parts of the craniofacial skeleton, namely the neurocranium, the viscerocranium, the mandible, and the teeth. In particular, dysmorphologies were seen in the cochlear apparatus and the teeth (taurodontism, reduced incisor curvature). Finally contrary to previous published results, we were able to record a specific phenotype of the Msx2 (+/-) mice with this methodology. This Msx2 (+/-) mouse phenotype was not intermediate between the Msx2 (-/-) and the wild-type animals., Conclusion: Msx2 plays an important role in craniofacial morphogenesis and growth because almost all craniofacial structures were affected in the Msx2(-/-) mice including both intramembranous and endochondral bones, the cochlear apparatus, and the teeth. In addition, Msx2 haploinsufficiency involves a specific phenotype with subtle craniofacial structures modifications compared with human mutations., (© 2014 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Objectives: The aim of the present study was to measure the effects of fluoride concentration on the real-time in vitro demineralization of enamel during exposure to caries-simulating conditions using Scanning Microradiography (SMR)., Methods: Enamel blocks obtained from non-carious human molars were fixed in SMR environmental cells, through which acidic solutions (0.1M acetic acid, pH 4.0) were circulated for periods of 48h. SMR was used to quantitatively measure continuous mineral mass loss. Subsequently, the effects of sequentially increasing fluoride concentration (0.1-4500mg/L [F(-)]) in the acidic solutions were measured on the rate of enamel demineralization., Results: The data shows a log-linear relationship between [F(-)] and reduction in demineralization up to 135mg/L [F(-)]. Above 135mg/L, no further significant decrease in demineralization occurred., Conclusion: The optimum range of local fluoride concentration for reducing enamel demineralization was in the range 0.1-135mg/L [F(-)] under the conditions studied., Clinical Significance: Relatively low [F(-)] can exhibit near-optimum protection. Increasing the fluoride concentrations above 135mg/L may not necessarily give an increased cariostatic benefit. Improving the means of delivery of relatively low fluoride concentrations to the oral fluids through slow releasing mechanisms, such as the oral fluoride reservoirs, is the more appropriate way forward for sustaining long-term clinical efficacy., (Copyright © 2013 Elsevier Ltd. All rights reserved.)

Purpose: To investigate the relative erosion protection potential of marketed dentifrices formulated with either stabilised stannous fluoride (SnF2 ), sodium fluoride (NaF) and/or sodium monofluorophosphate (SMFP) using an established laboratory erosion cycling model., Methods: Sound enamel cores from extracted, human enamel were cleaned, ground and polished, soaked in pooled saliva (pellicle formation) and treated with a 1:3 slurry of dentifrice and saliva. Specimens were subjected to daily challenges with 1% citric acid, a potentially damaging acid found in common food and drinks. Marketed dentifrices compared were: (1) a stabilised stannous fluoride product formulated with 1,100 ppm F as SnF2 ; (2) a cavity protection product containing 1,100 ppm F as NaF; (3) a cavity protection product comprising a mixed active fluoride system with 1,000 ppm F as SMFP + 450 ppm F as NaF; and (4) a sensitivity product containing 1,450 ppm F as SMFP + 8% arginine bicarbonate., Results: Specimens from Group 1 demonstrated an average loss of 5.5 (±1.2) μm of tooth surface enamel; Groups 2, 3 and 4 lost an average of 18.3 (±0.9) μm, 16.0 (±2.0) μm and 17.1 (±1.1) μm, respectively, of tooth surface enamel. Group 1 provided a statistically significant difference in protection compared with the other products., Conclusions: These results suggest that the marketed dentifrice formulated with stabilised SnF2 may provide enhanced protection of exposed tooth surfaces against dietary acid attack compared with the other products tested., (© 2014 FDI World Dental Federation.)

Purpose: To assess the potential of a stabilised stannous (Sn)-containing NaF dentifrice (Oral B/blend-a-Med(®) Pro-Expert), in addition to a number of other marketed European dentifrices formulated with various fluoride actives and two control dentifrices, to protect enamel against erosive acid damage., Methods: Cores of human enamel (four per group) were soaked in pooled human saliva, and then treated with a 1:3 slurry (dentifrice:saliva) using a standardised in vitro erosion model (5-day cycling) that includes 10-minute challenges with 1% citric acid applied 60 minutes after each dentifrice treatment. Enamel surface loss was measured using transverse microradiography (TMR)., Results: Specimens treated with the Sn-containing NaF dentifrice showed 6.5 μm of surface loss ± 1.2 (SEM), which was not significantly different (P < 0.05, Fisher LSD) from that of a clinically proven, stabilised SnF2 positive control [Crest(®) Pro-Health, 1,100 ppm F as SnF2 : 3.0 μm of surface loss ± 1.1 (SEM)]. The Sn-containing NaF dentifrice and the clinically proven positive control both provided significantly greater protection (P < 0.05, Fisher LSD) compared with all of the other products tested. Enamel loss (SEM) values for other European products and the reference control (active agents) were: Meridol(®) : (1,400 ppm F as AmF + SnF2 ) 12.0 μm (1.47); Colgate(®) Cavity Protection: (1,450 ppm F as SMFP + NaF) 12.9 μm (1.66); Odol med 3(®) (1,400 ppm F as NaF) 14.2 μm (1.49); Elmex(®) (1,400 ppm F as AmF) 14.5 μm (1.76); Colgate(®) Enamel Protect: (1,450 ppm F as NaF + KNO3 ) 16.3 μm (2.02); Lacalut(®) aktiv: (1,400 ppm F as AlF3 ) 18.5 μm (1.71); Sensodyne(®) ProNamel(™) : (1,450 ppm F as NaF + KNO3 ) 20.5 μm (1.26); Crest Cavity Protection (1,100 ppm F as NaF, reference control) 22.00 μm (2.04); and Mentadent(®) : (1,450 ppm F as NaF + Zn citrate) 22.3 μm (0.63)., Conclusion: These results support the potential for the stabilised, Sn-containing NaF dentifrice to provide erosion protection benefits that are not significantly different from the positive control benchmark for erosion protection (stabilised SnF2 ), and are significantly better than a broad range of dentifrice formulations available on the European market., (© 2014 FDI World Dental Federation.)

Probiotic bacteria have been suggested to inhibit Streptococcus mutans (SM) and thus prevent dental caries. However, supporting evidence is weak and probiotic species might be cariogenic themselves. Thus, we compared and combined the probiotic Lactobacillus rhamnosus GG (LGG) with SM and analysed the resulting mineral loss (ΔZ) in dental tissues. We simulated three biofilm compositions (SM, LGG, SM × LGG), two lesion sites (smooth enamel, dentin cavity) and two nutrition supply frequencies (twice/day, 6 times/day) in a multi-station, continuous-culture biofilm model. A total of 240 bovine enamel and dentin samples were cut, polished and embedded. All experimental procedures were performed in independent duplicates, with 10 samples being allocated to each group for each experiment (final sample size n = 20/group). Biofilms were cultured on the specimens and supplied with 2% sucrose medium and artificial saliva in consecutive pulses. After 10 days, ΔZ and bacterial numbers were assessed. SM × LGG biofilms caused significantly increased ΔZ compared with SM or LGG biofilms (p < 0.01, Mann-Whitney test), and ΔZ was significantly increased in dentin cavities compared with smooth enamel lesions (p < 0.01). Bacterial numbers did not significantly differ between biofilms of different species (p > 0.05, ANOVA). Frequent nutrition supply significantly increased bacterial numbers (p < 0.01). Biofilms in dentin cavities compared to smooth enamel harboured significantly more bacteria (p < 0.05). LGG induced mineral loss especially in dentin cavities and under highly cariogenic conditions. LGG did not have inhibitory effects on SM, but rather contributed to the caries process in vitro.

Gallium-doped phosphate-based glasses (Ga-PBG) were assessed for their impact on Streptococcus mutans and dental mineralisation, firstly by disc diffusion assays followed by biofilms grown on nitrocellulose filter membrane (NFM) and constant-depth film fermentor (CDFF). Short-time exposure (10 min) effects of Ga-PBG on S. mutans biofilm were compared with that of 0.2% chlorhexidine. The effects of Ga-PBG on bovine enamel (which was investigated under pH-cycling condition) and dentine were analysed using transverse microradiography (TMR), profilometry and inductively coupled plasma optical-emission spectrometry (ICP-OES). The disc diffusion assays showed inhibition zones of 24.5 ± 0.5 mm for Ga-PBG compared with controls (C-PBG). Ga-PBG showed statistically significant growth inhibition of S. mutans biofilms on NFM (p = 0.001) and CDFF (p < 0.046) compared with hydroxyapatite (HA) and C-PBG. The CDFF assay revealed a maximum of 2.11 log colony-forming unit (CFU) reduction at 48 h, but short-time exposure effects were comparable with that of 0.2% chlorhexidine only on older biofilms (maximum of 0.59 vs. 0.69 log CFU reduction at 120 h). TMR analyses of the enamel revealed non-significant mineral loss (p = 0.37) only in the case of Ga-PBG samples compared with controls including sodium fluoride. ICP-OES analyses indicated transient gallium adsorption into dentine by calcium displacement. The results confirmed that gallium inhibited S. mutans growth and appears to have the potential to protect the enamel surface under conditions representative of the oral environment. Further work is needed to establish whether it has an application in daily oral hygiene procedures to prevent or reduce caries.

Incipient caries lesions on smooth surfaces may be subjected to toothbrushing, potentially leading to remineralization and/or abrasive wear. The interplay of dentifrice abrasivity and fluoride on this process is largely unknown and was investigated on three artificially created lesions with different mineral content/distribution. 120 bovine enamel specimens were randomly allocated to 12 groups (n = 10), resulting from the association of (1) lesion type [methylcellulose acid gel (MeC); carboxymethylcellulose solution (CMC); hydroxyethylcellulose gel (HEC)], (2) slurry abrasive level [low (REA 4/ RDA 69); high (REA 7/RDA 208)], and (3) fluoride concentration [0/275 ppm (14.5 mM) F as NaF]. After lesion creation, specimens were brushed in an automated brushing machine with the test slurries (50 strokes 2×/day). Specimens were kept in artificial saliva in between brushings and overnight. Enamel surface loss (SL) was determined by optical profilometry after lesion creation, 1, 3 and 5 days. Two enamel sections (from baseline and post-brushing areas) were obtained and analyzed microradiographically. Data were analyzed by analysis of variance and Tukey's tests (α = 5%). Brushing with high-abrasive slurry caused more SL than brushing with low-abrasive slurry. For MeC and CMC lesions, fluoride had a protective effect on SL from day 3 on. Furthermore, for MeC and CMC, there was a significant mineral gain in the remaining lesions except when brushed with high-abrasive slurries and 0 ppm F. For HEC, a significant mineral gain took place when low-abrasive slurry was used with fluoride. The tested lesions responded differently to the toothbrushing procedures. Both slurry fluoride content and abrasivity directly impacted SL and mineral gain of enamel caries lesions.

Plants have efficient water-transporting vascular networks with a self-recovery function from embolism, which causes fatal discontinuity in sap flow. However, the embolism-refilling process in xylem vessel is still unclear. The water-refilling processes in the individual xylem vessels of excised Arabidopsis roots were visualized in this study using synchrotron X-ray micro-imaging technique with high spatial resolution up to 1 μm per pixel and temporal resolution up to 24 fps. In normal continuous water-refilling process, we could observe various flow patterns affected by the morphological structures of the xylem vessels, especially when water passed through perforation plates. A simple criterion based on the variation in dynamic pressure was suggested to evaluate the contribution of individual perforation plates to the water-refilling process. Meanwhile, the water-refilling embolized sections of xylem vessels through radial pathways were also observed. Separated water columns were formed from this discontinuous water-refilling process and the water influx rates through the radial pathways were estimated to be 478 and 928 μm(3) s(-1). The dynamic behavior of the separated water columns were quantitatively analyzed from the stoppage of volume growth to the translational phase. These water-refilling processes in excised roots of Arabidopsis may shed light on understanding the water refilling in the embolism vessels of intact plants and the interconnectivity of xylem vessel networks in vascular plants.

Objective: This study aimed at identifying the ideal concentration of a biofunctional surface coating of dental implants with a synthetic peptide (P-15). In a previous study, P-15 was shown to enhance osseointegration parameters., Material and Methods: Implants (modified ANKYLOS(®) A8; FRIADENT Plus(®) surface) with five different concentrations (0-400 μg/ml) of a P-15 coating as well as uncoated controls were inserted in the frontal bone of 45 adult domestic pigs. The histomorphometric and microradiographic findings for the coated implants were compared to those for the uncoated ones after 7, 14, and 30 days., Results: No significant differences were observed comparing the peri-implant bone density between the coated and uncoated implants The bone-to-implant contact, as the primary histological parameter for osseointegration, showed high rates for all surfaces investigated (between 73.3 ± 17.9% for the control and 81.9 ± 15.2% for P15 20 μg/ml after 30 days)., Conclusions: No significant benefit on osseointegration of a biofunctional P-15 coating of dental implants could be displayed in the present study., (© 2012 John Wiley & Sons A/S.)

Complete profiling would substantially facilitate the fundamental understanding of tumor angiogenesis and of possible anti-angiogenesis cancer treatments. We developed an integrated synchrotron-based methodology with excellent performances: detection of very small vessels by high spatial resolution (~1 μm) and nanoparticle contrast enhancement, in vivo dynamics investigations with high temporal resolution (~1 ms), and three-dimensional quantitative morphology parametrization by computer tracing. The smallest (3-10 μm) microvessels were found to constitute >80% of the tumor vasculature and exhibit many structural anomalies. Practical applications are presented, including vessel microanalysis in xenografted tumors, monitoring the effects of anti-angiogenetic agents and in vivo detection of tumor vascular rheological properties., (Copyright © 2011 Elsevier Inc. All rights reserved.)

Our experimental results demonstrate that full-field hard-X-ray microscopy is finally able to investigate the internal structure of cells in tissues. This result was made possible by three main factors: the use of a coherent (synchrotron) source of X-rays, the exploitation of contrast mechanisms based on the real part of the refractive index and the magnification provided by high-resolution Fresnel zone-plate objectives. We specifically obtained high-quality microradiographs of human and mouse cells with 29 nm Rayleigh spatial resolution and verified that tomographic reconstruction could be implemented with a final resolution level suitable for subcellular features. We also demonstrated that a phase retrieval method based on a wave propagation algorithm could yield good subcellular images starting from a series of defocused microradiographs. The concluding discussion compares cellular and subcellular hard-X-ray microradiology with other techniques and evaluates its potential impact on biomedical research., (Copyright © 2012 Elsevier Inc. All rights reserved.)