MicroCT study on the enamel mineral density of primary molars.

AIM: The aim of this study is to report on the mineral density of the enamel of primary molars related to the age of the child and to compare the mineral density of sound and carious enamel in those molars. MATERIALS AND METHODS: This study included 23 children and 41 extracted primary molars. The primary molars of 21 children met all of the inclusion criteria, and these were studied and scanned using microCT. The teeth were embedded in Impregum (3M ESPE) and stored in a solution of tap water with thymol crystals. Sixteen primary molars from 7 children were used to compare the mineral density in sound and carious areas, and 13 primary molars from 11 children were used for the comparison between mineral density and time in situ. RESULTS: A statistically significant difference (31%) was found between the mineral density in carious enamel and sound enamel (p = 0.0006). In addition, a significant relationship was observed between the mineral density of sound enamel and the time the teeth had been in situ (r = 0.698). We also found two teeth with radiolucencies in the dentin with the enamel clinically showing only a non-cavitated carious lesion in the enamel. No significant differences were found between the mean mineral density in sound enamel surfaces and unaffected areas in surfaces of molars with enamel caries (p = 0.4373). CONCLUSION: Local and general differences in enamel mineralisation are presented. Post-eruptive maturation seems to be present not only in permanent teeth but also in primary molars. Carious enamel has significantly less mineral density than clinically sound enamel.

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.

Fluoride toothpaste containing 1.5% arginine and insoluble calcium as a new standard of care in caries prevention.

In spite of obvious achievements in prevention, caries remains a prevalent disease. Fluorides are effective by inhibiting enamel and dentin demineralization and enhancing remineralization, but have little or no influence on bacterial processes in dental plaque. Dental caries is a continuum of stages from reversible, early lesions to irreversible, pre-cavitated lesions and, ultimately, to cavities. Prevention should focus on strengthening protective and reducing pathological factors, and careful monitoring of the disease state. While fluoride and the mineral aspects of caries have been in focus for decades, new insights into the etiology of caries have generated novel concepts and approaches to its prevention and treatment. The observation that some plaque bacteria can produce alkali metabolites and, thus, raise pH or neutralize acid formed in plaque has long been known. Such pH rise factors are related to caries susceptibility. Nourishing the plaque with substrates that encourage alkali-producing reactions is a protective factor in the caries continuum. This article reviews the results of clinical studies with a novel toothpaste containing 1.5% arginine, an insoluble calcium compound, and fluoride which have demonstrated superior remineralization of white spot enamel lesions and rehardening of root surface lesions, favorable effects on the de-/remineralization balance, as well as superior cavity prevention efficacy compared to toothpaste with fluoride alone. Studies have also confirmed formation of ammonia and elevated pH levels in subjects using the arginine-containing toothpaste. This novel toothpaste effectively combines the established effects of fluoride on de- and remineralization with reduction of caries-inducing pathological factors resulting from plaque metabolism.

Child dental anxiety, parental rearing style and referral status of children.

OBJECTIVE: Treating children can be difficult for both dentist and child. In some cases treatment fails and those children are referred to a specialist paediatric dentist. Different factors can be put forward for referral of children, such as factors relating to the child, dentist and parent. Possible child-related factors can be dental anxiety and the child's temperament. A possible parental factor is the parental rearing style. The objective of this study was to assess the possible associations between dental anxiety, parental rearing style and referral status of children. METHODS: Parents of 120 non-referred and 335 referred paediatric dental patients were asked to fill out the Child Rearing Practices Report (CRPR) and the Child Fear Survey Schedule Dental Subscale (CFSS-DS) on behalf of their children. RESULTS: The questionnaires were filled out by 115 (96%) parents of primary schoolchildren and by 331 (99%) parents of referred children. Referred children were younger than non-referred children, t(442) = 6.9, p < 0.01, and had significantly more dental anxiety, t(430) = -8.7, p < 0.01. No differences existed between parents of referred children and parents of non-referred children on parental rearing-style. No differences existed between fearful and non-fearful children on parental rearing-style and also no correlation existed between children's dental anxiety and their parent's rearing style. However, non-referred children with parents using an authoritarian parenting style were more anxious than the other non-referred children. CONCLUSIONS: In the present study, referral status and dental anxiety of 4-12 year old children were not associated with parental rearing style.

The numerous microbial species in oral biofilms: how could antibacterial therapy be effective?

Hundreds of bacterial species inhabit the oral cavity. Many of these have never been cultivated and can be assessed only with DNA-based techniques. This new understanding has changed the paradigm of the etiology of oral disease from that associated with 'traditional pathogens' as being primarily responsible for all diseases. Increasingly, associations between oral bacteria and systemic diseases are being reported. The emergence of antibiotic resistance is alarming and calls for in-depth studies of biofilms, bacterial physiology, and a body-wide approach to infectious diseases. We propose that the borderline between commensal bacteria and pathogens is no longer discrete. In a field of science where so many of the established paradigms are being undermined, a thorough analysis of threats and opportunities is required. This article addresses some of the questions that can be raised and serves to identify research opportunities and needs to leverage the prevention of oral diseases through novel antimicrobial strategies.

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.

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.

Effects of casein phosphopeptide amorphous calcium fluoride phosphate paste on white spot lesions and dental plaque after orthodontic treatment: a 3-month follow-up.

The effects of casein phosphopeptide amorphous calcium fluoride phosphate (CPP-ACFP) paste vs. control paste on the remineralization of white spot caries lesions and on plaque composition were tested in a double-blind prospective randomized clinical trial. Fifty-four orthodontic patients, with multiple white spot lesions observed upon the removal of fixed appliances, were followed up for 3 months. Subjects were included and randomly assigned to either CPP-ACFP paste or control paste, for use supplementary to their normal oral hygiene. Caries regression was assessed on quantitative light-induced fluorescence (QLF) images captured directly after debonding and 6 and 12 wk thereafter. The total counts and proportions of aciduric bacteria, Streptococcus mutans, and Lactobacillus spp. were measured in plaque samples obtained just before debonding, and 6 and 12 wk afterwards. A significant decrease in fluorescence loss was found with respect to baseline for both groups and no difference was found between groups. The size of the lesion area did not change significantly over time or between the groups. The percentages of aciduric bacteria and of S. mutans decreased from 47.4 to 38.1% and from 9.6 to 6.6%, respectively. No differences were found between groups. We observed no clinical advantage for use of the CPP-ACFP paste supplementary to normal oral hygiene over the time span of 12 wk.

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.

Relationship between gap size and dentine secondary caries formation assessed in a microcosm biofilm model.

Secondary caries can develop at the tooth-restoration interface, depending on the presence of a gap and its size, but this process could be inhibited by fluoride. The aim of this study was to assess the relationship between gap size and dentine secondary caries adjacent to composite resin (CR) or glass ionomer (GI) restorations, using a microcosm biofilm model in a constant depth film fermentor (CDFF). Dentine discs restored with CR (Z250) or GI (Vitremer) with gap sizes of 0, 50, 100, 180 or 250 microm were mounted on the CDFF. Microcosm biofilms were formed on the restored discs and daily subjected to 8 pulses of 10% sucrose solution. On the 18th day, dentine mineral loss and lesion depth around the restorations were determined by transverse microradiography. The effect of gap size was overall not statistically significant either with regard to mineral loss (p = 0.449) or lesion depth (p = 0.328), but greater mineral loss and lesion depth were found adjacent to CR than to GI (p < 0.001). However, Spearman correlation showed that mineral loss and lesion depth increased with gap size for CR (p < 0.001) but not for GI (p > 0.05). The findings support the conclusion that fluoride released from GI inhibits dentine demineralization adjacent to restorations, irrespective of gap width.

Antimicrobial activity of vanadium chloroperoxidase on planktonic Streptococcus mutans cells and Streptococcus mutans biofilms.

The aim of this study was to investigate the antimicrobial activity of vanadium chloroperoxidase (VCPO) reaction products on planktonic and biofilm cells of Streptococcus mutans C180-2. Planktonic and biofilm cells were incubated in a buffered reaction mixture containing VCPO, halide (either chloride or bromide) and hydrogen peroxide, and the killing efficacy was assessed by CFU counts. The enzymatic products formed by VCPO significantly reduced the viability of planktonic and biofilm cells compared to their negative controls and the effect on the biofilm cells was more effective than a 0.2% chlorhexidine digluconate treatment. We conclude that VCPO and its reaction products form a potent antimicrobial system against S. mutans.

Effect of various rinsing protocols after use of amine fluoride/stannous fluoride toothpaste on the bacterial composition of dental plaque.

This clinical study evaluated the effect of different oral hygiene protocols on the bacterial composition of dental plaque. After a 2-week period of using fluoride-free toothpaste, 30 participants followed three 1-week experimental protocols, each followed by 2-week fluoride-free washout periods in a randomized crossover examiner-blind controlled trial. The 1-week experimental protocols comprised the use of AmF/SnF(2) toothpaste twice daily, after which participants either (1) rinsed with tap water, (2) did not rinse but only spat out the toothpaste, or (3) rinsed with an AmF/SnF(2) mouthwash. In the fluoride-free washout periods, the participants brushed their teeth with fluoride-free toothpaste without further instructions. Six hours after the last brushing (+/- rinsing) of each period, buccal plaque samples in the upper molar region were taken. The microbiota composition of the plaque samples was analyzed by checkerboard DNA:DNA hybridization. A statistically significant reduction was found in the total amount of DNA of the 39 major plaque species measured, and in the proportions of some acid-producing bacterial strains after the period having used the AmF/SnF(2) toothpaste + AmF/SnF(2) mouthrinsing. The results indicate that using the AmF/SnF(2) toothpaste and rinse combination could result in plaque of lower cariogenicity.

Caries prevalence in Suriname schoolchildren.

OBJECTIVES: The aim of this study was to establish the oral health status of children living throughout the Interior of Suriname in order to define needs for dental care in line with WHO goals and guidelines. BASIC RESEARCH DESIGN: In this cross sectional study, dental caries was recorded according to the criteria of the WHO. Decayed, missing and filled (DMF)-teeth (T) and surfaces (S) indices for caries prevalence were used. A total of 951 children from four different regions and between 5-15 years of age, were examined. There was an approximately equal distribution of boys and girls. The children were divided into three age categories. RESULTS: The mean dmfs in the youngest children (5-7.5 yrs) was 11.81 (SD 11.19) and the mean dmft 5.16 (SD 3.93). 17.2% of the children was caries free. Statisticaly significant regional, racial and gender differences were found The mean dmfs of children in the middle age category (7.5-10 yrs) was 5.37 (SD 6.42) and the mean DMFS was 0.84 (SD 1.30). A mean DMFS of 2.31 (SD 4.97) was recorded in the oldest children. No regional, racial or gender differences were found in the last two categories. CONCLUSIONS: The results indicate that caries prevalence in young children in the Interior of Suriname is high according to the criteria of the WHO. In contrast, children in older age groups were found to experience low to moderate caries levels. This finding has consequences for the organisation and planning of future oral health care which should be focused on young children.

Fluoride Mode of Action: Once There Was an Observant Dentist . . .

The discovery and implementation of fluoride in the prevention of dental caries is often praised as one of the most important achievements in health care. In the early 20th century, it took 30 y to identify fluoride as the cause of enamel mottling but also of reduced caries prevalence in a population drinking water containing fluoride. Similarly, from 1960 to 1990, it took major efforts to unravel the working mode of fluoride in such detail that a rational scheme of caries prevention could be formulated. This article describes the scientific struggle leading to a consensus on the topic. For a historic purpose, the field, the actors, and their main research achievements are described. Ultimately it was generally agreed that the effect of fluoride is primarily topical by fluorides in the oral fluids rather than systemic by incorporation of fluoride in the enamel mineral crystals. Fluoride concentrations, even <1 mg/L, enhance the deposition of calcium phosphates during remineralization of enamel (and dentin). Similarly, such low levels of fluoride are effective in reducing the dissolution of the calcified tissues. This understanding has led to the development of fluoride-containing caries-preventive products that had an undisputed beneficial effect on the levels of dental caries.

Effect of Veillonella parvula on the antimicrobial resistance and gene expression of Streptococcus mutans grown in a dual-species biofilm.

INTRODUCTION: Our previous studies showed that Streptococcus mutans and Veillonella parvula dual-species biofilms have a different acid production profile and a higher resistance to chlorhexidine than their single-species counterparts. The aim of the current study was to test whether the susceptibility of S. mutans grown in the presence of V. parvula is also decreased when it is exposed to various other antimicrobials. Furthermore, the aim was to identify other changes in the physiology of S. mutans when V. parvula was present using transcriptomics. METHODS: Susceptibility to antimicrobials was assessed in killing experiments. Transcript levels in S. mutans were measured with the help of S. mutans microarrays. RESULTS: When V. parvula was present, S. mutans showed an increase in survival after exposure to various antimicrobials. Furthermore, this co-existence altered the physiology of S. mutans. The expression of genes coding for proteins involved in amino acid synthesis, the signal recognition particle-translocation pathway, purine metabolism, intracellular polysaccharide synthesis, and protein synthesis all changed. CONCLUSION: Growing in a biofilm together with a non-pathogenic bacterium like V. parvula changes the physiology of S. mutans, and gives this bacterium an advantage in surviving antimicrobial treatment. Thus, the study of pathogens implicated in polymicrobial diseases, such as caries and periodontitis, should be focused more on multispecies biofilms. In addition, the testing of susceptibility to currently used and new antimicrobials should be performed on a multispecies microbial community rather than with single pathogens.

Effects of different kinds of fluorides on enolase and ATPase activity of a fluoride-sensitive and fluoride-resistant Streptococcus mutans strain.

Enolase and ATPase are sensitive to fluoride. It is unclear whether this sensitivity differs for F-sensitive and F-resistant cells or for different types of fluoride. Permeabilized cells of the fluoride-sensitive strain Streptococcus mutans C180-2 and its fluoride-resistant mutant strain C180-2 FR were preincubated at pH 7 or 4 with NaF, the amine fluorides Olaflur and Dectaflur and amine chloride controls. After preincubations, enolase and ATPase activities of the cells were assessed. Enolase activity was more inhibited after preincubation at pH 7 with NaF than with Olaflur. Amine chloride stimulated, although not with statistical significance, the enolase activity of both strains. After preincubation at pH 4 the enolases were strongly inactivated, but the fluoride-resistant strain's enolase to a lesser extent. The results suggested that amine acts to protect enolase activity against the detrimental low pH effect. Gene sequencing showed that the enolase genes of the fluoride-resistant and fluoride-sensitive strain were identical. ATPase activity was not reduced after NaF preincubation at either pH 7 or pH 4. The amine fluorides and their chloride controls in the preincubation mixture reduced the ATPase activity significantly at both pH values. In conclusion, our results showed that preincubation with amine fluoride did not inhibit enolase activity more effectively than NaF. The amine part of the molecule may protect enolase activity against preincubations at low pH. ATPase activity was not inhibited by NaF preincubation but was significantly inhibited after preincubation with amine fluorides and amine chlorides.

Chlorhexidine efficacy in preventing lesion formation in enamel and dentine: an in situ study.

BACKGROUND: Clinical studies on the caries-preventive properties of chlorhexidine mouthrinses are limited and the results are inconclusive. AIM: The aim of this study was to elucidate the contribution of a 0.2% chlorhexidine mouthrinse to the protection of enamel and dentine against demineralization. METHODS: In this randomized two-treatment, two-leg study 14 individuals wearing partial prostheses were enrolled. Sound enamel and dentine specimens were placed in situ for 4 weeks. Twice daily, a mouthrinse was performed with either chlorhexidine or saline (control) depending on the experimental group the participant was allocated to. After the experimental period, plaque samples were collected from the surface of the specimens and from natural tooth surfaces to assess the organic acid production upon a sucrose challenge. The specimens were analyzed for mineral loss by transversal microradiography. RESULTS: This study could not demonstrate a significantly better protection of enamel and dentine against demineralization by the chlorhexidine treatment compared to saline. No differences in acid production of plaque samples from the chlorhexidine-treated and control groups were observed. This result was also found for plaque samples originating from the natural tooth surfaces. CONCLUSION: Mouth rinsing with 0.2% chlorhexidine did not prevent demineralization of dentine and enamel in situ.

Effect of Psidium cattleianum leaf extract on Streptococcus mutans viability, protein expression and acid production.

Plants naturally produce secondary metabolites that can be used as antimicrobials. The aim of this study was to assess the effects of Psidium cattleianum leaf extract on Streptococcus mutans. The extract (100%) was obtained by decoction of 100 g of leaves in 600 ml of deionized water. To assess killing, S. mutans biofilms were treated with water (negative control) or various extract dilutions [100, 50, 25% (v/v) in water] for 5 or 60 min. To evaluate the effect on protein expression, biofilms were exposed to water or 1.6% (v/v) extract for 120 min, proteins were extracted and submitted to 2-dimensional difference gel electrophoresis. Differentially expressed proteins were identified by mass spectrometry. The effect of 1.6% (v/v) extract on acid production was determined by pH measurements and compared to a water control. Viability was similar after 5 min of treatment with the 100% extract or 60 min with the 50% extract (about 0.03% survival). There were no differences in viability between the biofilms exposed to the 25 or 50% extract after 60 min of treatment (about 0.02% survival). Treatment with the 1.6% extract significantly changed protein expression. The abundance of 24 spots was decreased compared to water (p < 0.05). The extract significantly inhibited acid production (p < 0.05). It is concluded that P. cattleianum leaf extract kills S. mutans grown in biofilms when applied at high concentrations. At low concentrations it inhibits S. mutans acid production and reduces the expression of proteins involved in general metabolism, glycolysis and lactic acid production.

Effect of microleakage and fluoride on enamel-dentine demineralization around restorations.

There is no consensus about an association between microleakage and secondary caries, especially considering the presence of fluoride (F) at the tooth/restoration interface. Thus, a randomized, double-blind, crossover study was carried out to evaluate in situ the effect of microleakage on caries around enamel-dentine restorations in the presence of F from dental materials or dentifrice, either alone or in combination. In 4 phases of 14 days each, 14 volunteers wore palatal devices containing dental slabs restored with composite resin (CR) or resin-modified glass ionomer cement (GI). Restorations were made without leakage (L-), following the recommended adhesive procedures, or with leakage (L+), in the absence of adhesive procedures. Plaque-like biofilm (PLB) was left to accumulate on the restored slabs, which were exposed extraorally to a 20% sucrose solution 10x/day. The volunteers used a non-F (NF) or an F (FD) dentifrice 3x/day, depending on the experimental phase. No differences were found between L+ or L- restorations (p > 0.05). Higher demineralization in both enamel and dentine around CR restorations was observed under NF (p < 0.05). F concentration was higher in the fluid of PLB exposed to FD or formed onto GI restoration (p < 0.05). These results suggest that while microleakage does not affect caries development, GI or FD may maintain increased F levels in the PLB, thereby decreasing caries progression.