[What is the functioning of tin in stannous fluoride?] / Wat is de werking van tin in tinfluoride?

Stannous fluoride is one of the first fluoride compounds that were added to dentifrices. Besides the well-known effect of fluoride, the presence of tin could also have an effect on dental health by its anti-microbial activity and the ability to form insoluble metal salts. The functioning of stannous fluoride has been studied extensively in many scientific publications. On the basis of the available literature, the use of stannous fluoride instead of sodium fluoride could be advantageous in case of gingivitis, halitosis, dentine hypersensitivity, or erosion. The effects that were found are statistically significant, albeit rather small, which makes it harder to predict the actual gain in dental health or the clinical relevance for an individual patient.

Comparison of Composition and Anticaries Effect of Galla Chinensis Extracts with Different Isolation Methods.

OBJECTIVES: Galla chinensis water extract (GCE) has been demonstrated to inhibit dental caries by favorably shifting the demineralization/remineralization balance of enamel and inhibiting the biomass and acid formation of dental biofilm. The present study focused on the comparison of composition and anticaries effect of Galla chinensis extracts with different isolation methods, aiming to improve the efficacy of caries prevention. METHODS: The composition of water extract (GCE), ethanol extract (eGCE) and commercial tannic acid was compared. High performance liquid chromatography coupled to electrospray ionization-time of flight-mass spectrometry (HPLC-ESI-TOF-MS) analysis was used to analyze the main ingredients. In vitro pH-cycling regime and polymicrobial biofilms model were used to assess the ability of different Galla chinensis extracts to inhibit enamel demineralization, acid formation and biofilm formation. RESULTS: All the GCE, eGCE and tannic acid contained a high level of total phenolics. HPLC-ESI-TOF-MS analysis showed that the main ingredients of GCE were gallic acid (GA), while eGCE mainly contained 4-7 galloylglucopyranoses (GGs) and tannic acid mainly contained 5-10 GGs. Furthermore, eGCE and tannic acid showed a better effect on inhibiting enamel demineralization, acid formation and biofilm formation compared to GCE. CONCLUSIONS: Galla chinensis extracts with higher tannin content were suggested to have higher potential to prevent dental caries.

A reproducible microcosm biofilm model of subgingival microbial communities.

OBJECTIVE: To develop a reproducible subgingival microcosm biofilm model. MATERIAL AND METHODS: Subgingival plaque samples were collected from four deep pockets (probing pocket depth ≥6 mm) in each of seven patients with periodontitis and from shallow pockets (probing pocket depth ≤3 mm) in two periodontally healthy donors. An active attachment model and a peptone medium (Thompson et. al., Appl Environ Microbiol 2015;81:8307-8314) supplemented with 30% serum was used. Biofilms were harvested at 2 and 4 weeks. DNA of dead cells was blocked for amplification by propidium monoazide treatment. Composition was analyzed using 16S rRNA gene amplicon pyrosequencing. Similarities between the biofilm samples were assessed by non-metric multidimensional scaling using the Bray-Curtis similarity index and similarity percentage analysis. Data from duplicate experiments, different biofilm sources and different biofilm age were compared. RESULTS: The non-metric multidimensional scaling revealed a strong clustering by the inoculum source, the donor and their periodontal status. Statistically significant differences were found between the sources of inoculum (P=.0001) and biofilm age (P=.0016). Furthermore, periodontitis biofilms (P) were distinct in composition from health-derived biofilms (H) by genera: Porphyromonas (P=19%; H=0%), Filifactor (P=10%; H=0%), Anaeroglobus (P=3%; H=0%), Phocaeicola (P=1.5%; H=0%), Parvimonas (P=19%; H=14%), Fusobacterium (P=2%; H=26%), Peptostreptococcus (P=20%; H=30%), Veillonella (P=7%; H=8%) and 57 other genera. Similarity distances (Bray-Curtis) (mean 0.73, SD 0.15) and the Shannon diversity index (mean 2, SD 0.2) revealed no differences between duplicate experiments (P=.121). CONCLUSION: This biofilm model allows reproducible production of complex subgingival microbial communities.

Noninvasive dentistry: a dream or reality?

Various caries prevention and repair strategies are reviewed in this article ranging from the use of fluoride to nanohydroxyapatite particles. Several of the strategies which combine fluoride and calcium and phosphate treatments have both in vitro and in vivo data showing them to be efficacious if the surface integrity of the lesion is not breached. Once this has occurred, the rationale for cutting off the nutrient supplies to the pathogenic bacteria without the removal of the infected dentine, a noninvasive restorative technique, is discussed using existing clinical studies as examples. Finally two novel noninvasive restorative techniques using fluorohydroxyapatite crystals are described. The need for clinical data in support of emerging caries-preventive and restorative strategies is emphasized.

Biofilm layers affect the treatment outcomes of NaF and Nano-hydroxyapatite.

During caries formation, dental biofilms function not only as acid producers but also as reservoirs and diffusion barriers for active caries-preventive components. The aim of this study was to investigate the influence of biofilms as a stagnant layer on the efficacy of NaF and nano-hydroxyapatite (nHA). Biofilms of Streptococcus mutans C180-2 were formed on the surfaces of artificially demineralized enamel in an active attachment biofilm model. After 2 days of biofilm formation, the model was subjected to a pH-cycling schedule, together with a control group without biofilms. Specimens were treated for 5 min twice daily with water, a 10% nHA slurry, or 18.4 mM NaF. At the end of the pH-cycling period, the biofilms were removed for the determination of the viable counts, the lactic acid production, and the calcium content. The mineral changes in the demineralized enamel blocks were analyzed by transversal microradiography. No differences in the biofilm viable counts and lactic acid production were found in the different treatment groups. The mean calcium content of the biofilms in the nHA group was 60.7 ± 15.3 mmol/g wet weight, which was approximately 8-fold higher than in the other 2 groups. The application of NaF resulted in net remineralization, but in the presence of a biofilm, net demineralization was observed. In contrast, nHA treatment reduced further demineralization compared with the water treatment, but the presence of a biofilm enhanced this effect. In conclusion, the presence of biofilms clearly influenced the treatment outcomes of anticaries products. Biofilms could either enhance or impede their efficacy. This result implies that biofilms should be included in the in vitro tests for the preclinical screening of caries-protective agents.

The effect of propidium monoazide treatment on the measured bacterial composition of clinical samples after the use of a mouthwash.

OBJECTIVES: The use of an anti-microbial mouthwash results not only in a reduction of the number of viable cells in dental plaque but potentially also in a shift in the oral microbiome. DNA-based techniques may be appropriate to monitor these shifts, but these techniques amplify DNA from both dead and living cells. Propidium monoazide (PMA) has been used to overcome this problem, by preventing the amplification of DNA from membrane-damaged cells. The aim of this study was to evaluate the use of PMA when measuring compositional shifts in clinical samples after mouthwash use. MATERIALS AND METHODS: On two consecutive days, baseline samples from buccal surfaces, tongue, and saliva were obtained from six volunteers, after which they used a mouthwash (Meridol, GABA, Switzerland) twice daily for 14 days. Subsequently similar samples were obtained on two consecutive days. The microbial composition of the samples, with or without ex vivo PMA treatment, was assessed with 16S rRNA gene amplicon sequencing. RESULTS: Data showed a clear effect of mouthwash usage on the tongue and saliva samples. PMA treatment enhanced the observed differences only for the saliva samples. Mouthwash treatments did not affect the composition of the plaque samples irrespective of the use of PMA. CONCLUSION: The necessity to use a PMA treatment to block the DNA from dead cells in clinical studies aimed at measuring compositional shifts after the use of a mouthwash is limited to salivary samples. CLINICAL RELEVANCE: Measuring shifts in the oral microbiome could be hampered by the presence of DNA from dead cells.

The effects of propidium monoazide treatment on the measured composition of polymicrobial biofilms after treatment with chlorhexidine.

BACKGROUND: The treatment of polymicrobial biofilms with antimicrobial compounds results in not only an overall loss of viability, but also compositional shifts. While DNA-based technologies may be more appropriate for the assessment of bacterial composition than culturing, these techniques amplify DNA from both live and dead cells. Propidium monoazide (PMA) has been used to discriminate between live and dead cells by blocking the DNA from membrane-damaged cells from being amplified. AIM: This study evaluated the use of PMA in a saliva-derived polymicrobial biofilm model subjected to a single chlorhexidine (CHX) treatment. MATERIALS AND METHODS: The effects of PMA on viable cells were tested using both untreated and PMA-treated saliva as an inoculum. Viability was determined by plate counts, metabolic activity was determined by lactic acid production, and biofilm composition was assessed by 16S rRNA gene amplicon sequencing. RESULTS: Exposure to a 0.2% CHX rinse (meridol® perio) reduced the viability and metabolic activity of 48-hour biofilms. The shift in biofilm composition observed after the CHX exposure was enhanced after a post-rinse PMA treatment. PMA treatment had a small effect on the measured composition of water-rinsed biofilms. Treating saliva with PMA reduced bacterial viability and shifted the bacterial composition of saliva and saliva-derived biofilms. CONCLUSION: The removal of DNA from non-viable cells with PMA treatment was shown to elicit an improvement in the detection of shifts in in vitro polymicrobial biofilms after antimicrobial treatment. However, PMA also influenced the ability of cells to grow, indicating that PMA should be used with caution.

Factors associated with alkali production from arginine in dental biofilms.

Alkali production by oral bacteria in the oral cavity has been linked to protection against dental caries. The current study assessed various parameters associated with ammonium produced during arginine catabolism in dental biofilms. Polymicrobial biofilms were formed with saliva as the inoculum. The NH(3) level and the pH of the spent medium were used to monitor and quantitate the bacterial reactions. The presence of sucrose, a low buffer capacity, and a low pH (≤ pH 4.5) were found to hamper alkali production from arginine. The rate of alkali production exhibited an optimum around pH 5.5. Biofilms were found to produce NH(3) also from polypeptides and proteins in the medium. The biofilm age affected these processes. The experimental model proved valuable for the assessment of the collective bacterial reactions determining the overall pH outcome. This experimental approach could bridge the gap in our knowledge between pH-rise phenomena and caries susceptibility from clinical observations and studies performed on alkali-producing bacteria in well- controlled, though simplified, in vitro models. Analysis of our data supports the hypothesis that the initiation and progression of dental caries may be influenced by the relative rates of acid and base formation, which critically depend on the aforementioned parameters.

Effect of pH on Galla chinensis extract's stability and anti-caries properties in vitro.

OBJECTIVES: Considering that Galla chinensis extract (GCE) solution has a low pH, which might dissolve dental enamel, we investigated the effects of elevation of pH on GCE stability, and on its anti-caries properties. DESIGNS: Stability of GCE solutions, either in H(2)O (pH less than 4.0) or when buffered at pH 5.5, 7.0 and 10.0, was assessed from UV-VIS spectra. Inhibition of enamel demineralization was determined in a pH-cycling set up, comprising treatments with either GCE solutions or negative control buffers and acid and neutral buffer immersions. Demineralization was assessed by calcium in the acetate buffers. To determine antimicrobial properties, polymicrobial biofilms were formed after saliva inoculation on glass surfaces which were treated after 48 h. Treatment output parameters were lactic acid formation and viability, the latter by colony forming unit (CFU) counts. RESULTS: At pH 7.0 and higher GCE solutions changed colour and absorption spectra in UV-VIS, indicative of chemical changes. Regarding enamel demineralization, significant inhibitions (P<0.05) were found for all GCE treatments when compared with corresponding controls. In polymicrobial biofilms, GCE reduced the acid production, compared with the negative controls (P<0.05). However, this difference was only significant at the lower pH values. CONCLUSIONS: GCE solutions were unstable under neutral and alkaline conditions. pH did not significantly influence the inhibiting effect of GCE on enamel demineralization. However, GCE was not effective on polymicrobial biofilms at alkaline pH (8.5). To avoid enamel damage due to acidic treatment, GCE solutions should be used at about pH 5.5.

Evaluation of different fluoride concentrations supplemented with trimetaphosphate on enamel de- and remineralization in vitro.

Trimetaphosphate (TMP) effects on demineralized bovine enamel were studied after 15 days of pH cycling. Treatments included 30 wt% (weight percent) dilutions of 0, 500, 1,500 or 3,000 µg F/g aqueous NaF solutions with or without 3% TMP. Treated specimens were assessed by transverse microradiography. With the exception of the 3,000 µg F/g case, 3% TMP addition provided significant additional overall remineralization compared with F alone. Mineral content profiles differed significantly between corresponding F and F + TMP groups. Fluoride alone resulted in more remineralization in the original demineralized zone, whereas F + TMP caused less demineralization in the underlying, originally sound enamel.

The effects of hyperosmosis or high pH on a dual-species biofilm of Enterococcus faecalis and Pseudomonas aeruginosa: an in vitro study.

AIM: To investigate the effect of hyperosmotic hyperosmosis or alkaline stress on a dual-species biofilm of Enterococcus faecalis and Pseudomonas aeruginosa. METHODOLOGY: Biofilms were grown on glass cover slips suspended in bacterial inoculate for 96 h, after which the cover slips with attached biofilms were immersed in brain heart infusion broth (BHI-broth) with 6 mol L(-1) sodium chloride (NaCl) representing the hyperosmotic group or Ca(OH)(2), pH 12.1, representing the alkaline group. Two per cent sodium hypochlorite and BHI- broth served as positive and negative controls, respectively. After treatment, the biofilms were washed, harvested and plated on blood-agar plates after serial dilution. The bactericidal effect was assessed by determining the colony-forming units (CFU). The effect on the biofilm mass was imaged with confocal laser scanning microscopy (CLSM). RESULTS: Hyperosmosis reduced the CFU of both species significantly after 72 h (P < 0.0001). After 168 h, P. aeruginosa was eradicated and the E. faecalis reduction was more than 99%. High pH could not induce a significant bacterial reduction. CLSM revealed dense flocculation of the biofilms incubated in alkaline broth. CONCLUSION: Hyperosmosis effectively reduced a dual-species biofilm of E. faecalis and P. aeruginosa, whilst high pH had limited bactericidal effect in this model.

Effect of Galla chinensis on growth and metabolism of microcosm biofilms.

Galla chinensis extract (GCE) interferes with de- and remineralization of dental enamel and the growth and metabolism in planktonic bacteria. However, no information is available on GCE effects on biofilms formed with saliva as inoculum. The aim of the current experiments was to investigate the effects of GCE at different stages of salivary microcosm biofilm formation. Biofilms formed on glass or enamel surfaces were treated with GCE solutions at different concentrations and at different time points. Effects were assessed by lactic acid formation and colony-forming unit (CFU) counts of the biofilms. The results showed that GCE treatments inhibited growth and acid metabolism of both nascent and mature microcosm biofilms. Pretreatment of the substratum with GCE solutions inhibited growth and lactic acid production of biofilms grown on enamel, but had little effects on biofilms formed on glass surfaces. A maximum GCE effect was found when biofilms, on either surface type, were treated after 8 h of formation with 40 h of subsequent growth. In medium with sucrose-fermenting biofilms, low concentrations of GCE (0.2 and 0.1 mg/ml) inhibited acid production without killing bacteria of the biofilm. Differences were found in GCE effects on biofilms formed with saliva from different donors, with reductions in acid formation and CFU values ranging between 0 and 78%. In conclusion, bioactive components in GCE reduce or inhibit both growth and lactic acid formation in biofilms.

Different response to amine fluoride by Streptococcus mutans and polymicrobial biofilms in a novel high-throughput active attachment model.

BACKGROUND/AIMS: The antimicrobial resistance of microorganisms in biofilms and the polymicrobial interactions in these biofilms that modulate resistance require novel strategies to evaluate the efficacy of caries-preventive compounds. The current study aimed to evaluate the effects of a caries-preventive agent in Streptococcus mutans and polymicrobial biofilms. METHODS: We developed a novel high-throughput active attachment model. The model consisted of a custom-designed lid containing glass discs that fit on top of standard 24-well plates. Biofilms were formed using either S. mutans C180-2 or saliva. At the end of biofilm formation (up to 96 h) the biofilms were treated with amine fluoride (AmF) solutions. The viability of the biofilms was determined by CFU counts, and metabolic activity was measured via lactate production. RESULTS: The effect of AmF on the viability of the polymicrobial biofilms was significantly less than that on the S. mutans biofilms, indicating a higher resistance in the complex biofilms. Both types of biofilms became more resistant to AmF with age. The higher resistance of the polymicrobial biofilms was not reflected in metabolic activity; in dose-response experiments AmF reduced lactate production in both types of biofilms to the same extent. Moreover, the age-induced increased resistance in the polymicrobial biofilms was less pronounced in terms of the inhibition of metabolic activity. CONCLUSIONS: This study clearly shows that when evaluating the efficacy of caries-preventive compounds it is essential to use appropriate polymicrobial biofilm models, and more importantly that efficacy needs to be judged based on the reduction of acid formation (i.e. cariogenic potential) as well as on bacterial viability.

Elevated fluoride products enhance remineralization of advanced enamel lesions.

Caries prevention might benefit from the use of toothpastes containing over 1500 ppm F. With few clinical studies available, the aim of this pH-cycling study was to investigate the dose response between 0 and 5000 ppm F of de- and remineralization of advanced (> 150 microm) enamel lesions. Treatments included sodium and amine fluoride, and a fluoride-free control. Mineral uptake and loss were assessed from solution calcium changes and microradiographs. Treatments with 5000 ppm F both significantly enhanced remineralization and inhibited demineralization when compared with treatments with 1500 ppm F. Slight differences in favor of amine fluoride over sodium fluoride were observed. The ratio of de- over remineralization rates decreased from 13.8 to 2.1 in the range 0 to 5000 ppm F. As much as 71 (6)% of the remineralized mineral was calculated to be resistant to dissolution during subsequent demineralization periods. With 5000-ppm-F treatments, more demineralizing episodes per day (10 vs. 2 for placebo) would still be repaired by remineralization.

The relative efficacy of fluoride toothpastes assessed with pH cycling.

This study addressed the dose response between fluoride toothpastes and in vitro de- and remineralization, to predict the efficacy of toothpastes and understand the mode of action in the range 0-3,000 ppm F. Enamel lesions were pH-cycled with calcium uptake and loss being assessed daily. Both 'shallow' (about 50 microm deep) and 'deep' (about 200 microm deep) lesions were studied. F treatments were given in 30 (w/v)% toothpaste dilutions for up to 5 min daily. Calcium loss during the demineralization periods showed a dose response, resulting in 72% reduction for 3,000 ppm F compared to 0 ppm F. Calcium uptake during remineralization was increased in the F compared to non-F groups, with F concentration being less important than its mere presence. Significant differences were observed in F response between shallow and deep lesions, suggesting that this parameter should be included when testing caries-preventive products. Microradiographic analysis showed that lesion depth and severity had increased significantly in the non-F groups. In the F groups, the original lesion was partly remineralized, while a new lesion had formed beyond the original lesion front. Mineral loss of this second lesion correlated inversely with the F concentration of the treatments. These data revealed that fluoride can drive demineralization further into enamel by making the surface tissue less soluble, hence by not neutralizing acids penetrating into the tissue. It is also concluded that depth analysis of mineral uptake and loss is important to understand the mode of action of different F products.

Effect of fluoride-releasing filling materials on underlying dentinal lesions in vitro.

Fluoride-releasing materials placed over carious tissue are assumed to enhance remineralisation of the underlying lesion. This remineralisation, however, also depends on the availability of calcium and phosphate, which may be supplied by the pulpal fluid. The aim of this study was to measure the fluoride release of glass ionomer cements (GICs) into underlying dentin and to measure the effect of the released fluoride on the remineralisation of the underlying dentinal lesions using transversal microradiography. Discs of fluoride-releasing GIC were placed on top of dentinal lesions in an in vitro model. The discs and the dentin slabs were covered completely by a protective layer of nail varnish, leaving only the pulpal side of the dentin slab open, and hence the dentinal tubules as the pathway for the incubation fluid to the GIC disc. Specimens were incubated in a remineralisation buffer. The materials tested were a conventional GIC, an experimental GIC that was designed to have a high fluoride release, and an inert material. Fluoride was found to penetrate through the dentin slab into the surrounding fluid. Fluoride uptake from the experimental GIC was higher than from the conventional GIC. Mineral content-depth profiles after 10 weeks' remineralisation revealed that in the outer 30 microm of the lesion a higher mineral deposition occurred for the experimental GIC than in both other groups. No differences in the overall change of integrated mineral loss were found for the tested materials. We conclude that high fluoride release from filling materials only results in superficially increased remineralisation of underlying demineralised dentin.

Light-induced fluorescence studies on dehydration of incipient enamel lesions.

Changes in the hydration state of enamel affect its optical qualities, such as light scattering and fluorescence. In this study, the rate of fluorescence loss was measured when incipient enamel lesions with different de-remineralization history were left to dehydrate. Four groups of lesions were studied. In groups A, B and C, the lesions were prepared in vitro in an acid-gel system. Group A was kept as control, and groups B and C were remineralized (4 weeks) without and with 1 ppm F in solution, respectively. Group D consisted of natural incipient lesions. Enamel fluorescence was measured for all lesions immediately after removal from water and subsequently at short intervals for 30 min. The change in fluorescence with dehydration varied between the groups. In lesions from groups A and B, it followed a double exponential decrease, while in lesions from groups C and D, it followed a mono-exponential decrease. In all groups, the fluorescence of sound surfaces declined mono-exponentially. The 'fractional fluorescence difference', defined as (L(sound) - L(carious) )/L(sound), became constant after periods of dehydration of about 5, 5, 20 and 5 min for groups A to D, respectively. The observation of the change of fluorescence with dehydration should be taken into consideration when planning studies that use fluorescence as an assessment method. However, it might also be used to gain insight into the properties for fluid transport inside the various lesions, relevant to de-remineralization or fluoride treatments.