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Experimental intra-oral caries models in fluoride research.

The use of experimental intra-oral caries models has increased in fluoride research. This paper focuses on the pre-clinical intra-oral models, the in situ and in vivo models, the various types, their benefits and disadvantages. Both preparation and sterilization of the hard tissue substrates can affect the substrates and therefore the results. Care needs to be taken that dentine samples are not exposed to drying and consequently shrinking during preparation and evaluation. Sterilization by gamma-radiation is at present the least tissue-damaging method. The most realistic experimental model is the in vivo model, followed by the in situ model using specimens with natural surfaces. The most accurate and direct evaluation technique for demineralization and remineralization studies is quantitative transversal microradiography, whereas confocal laser scanning microscopy (CLSM) is the most sensitive qualitative evaluation technique. Other evaluation techniques discussed are microhardness testing and the iodine permeability test. In light of the present skewed caries situation in western countries we suggest that fluoride research focuses on experimental caries models that can mimic severe cariogenic challenge. Testing of fluoride combinations and dosages that can prevent lesion development rather than promote remineralization would then be a practical consequence.

Animals↗

Keeping the pulp alive: the pharmacology and toxicology of agents applied to dentine.

Materials applied to teeth release chemicals which may diffuse through dentine to either harm or help the dental pulp. Chemical threats are minimized by material choice or by using relatively impermeable lining or base materials which are themselves of low chemical toxicity. The most probable long-term threat to the pulp in the restored tooth is bacterial, through leakage around restorations. This may be prevented by the use of treatment sequences designed to develop an effective seal. Pulpal inflammation is treated primarily by removal of the irritant cause; inflammation may also be suppressed in the very short term by corticosteroids, and its resolution aided in the longer term by zinc oxide-eugenol preparations.

Demeclocycline↗

Functional characterization of connexin43 mutations found in patients with oculodentodigital dysplasia.

Specific mutations in GJA1, the gene encoding the gap junction protein connexin43 (Cx43), cause an autosomal dominant disorder called oculodentodigital dysplasia (ODDD). Here, we characterize the effects of 8 of these mutations on Cx43 function. Immunochemical studies have shown that most of the mutant proteins formed gap junction plaques at the sites of cell-cell apposition. However, 2 of the mutations (a codon duplication in the first extracellular loop, F52dup, and a missense mutation in the second extracellular loop, R202H, produced full-length connexins that failed to properly form gap junction plaques. Cx43 proteins containing ODDD mutations found in the N-terminus (Y17S), first transmembrane domain (G21R, A40V), second transmembrane domain (L90V), and cytoplasmic loop (I130T, K134E) do form gap junction plaques but show compromised channel function. L90V, I130T, and K134E demonstrated a significant decrease in junctional conductance relative to Cx43WT. Mutations Y17S, G21R, and A40V demonstrated a complete lack of functional electrical coupling even in the presence of significant plaque formation between paired cells. Heterologous channels formed by coexpression of Cx43WT and mutation R202H resulted in electrically functional gap junctions that were not permeable to Lucifer yellow. Therefore, the mutations found in ODDD not only cause phenotypic variability, but also result in various functional consequences. Overall, our data show an extensive range of molecular phenotypes, consistent with the pleiotropic nature of the clinical syndrome as a whole.

Abnormalities, Multiple↗

Effect of bonded amalgam restorations on microleakage.

The purpose of this study was to evaluate quantitatively the effectiveness of adhesive dentin bonding systems in decreasing microleakage at the tooth-amalgam restoration interface. The results indicated that microleakage was significantly reduced when Amalgambond Plus or All-Bond 2 was used as liners in comparison to either Copalite varnish or no linear under amalgam restorations. No significant difference was found between the two dentin bonding systems at all time periods studied.

Analysis of Variance↗

Oxalate desensitising treatment of dentinal surface.

It is well known that a typical painful feeling is caused by impact of different agents and by thermodynamic conditions upon the dentine layer of the tooth. Therefore the action by artificial solutions should be tested to study how the induced modifications might inhibit the pain. The aim of the present study is to evaluate by scanning electron microscopy (SEM) the morphology of dentine surface after different chemical treatments. Oxalate solutions are able to produce a layer of large crystals, while acid solutions remove the smear layer and open the dentinal tubules.

Dentin↗

Tight junctions in differentiating ameloblasts and odontoblasts differentially express ZO-1, occludin, and claudin-1 in early odontogenesis of rat molars.

Little is known about the expression of associated proteins during the assembly of tight junctions (TJs). We studied the distribution of ZO-1, occludin, and claudin-1 between differentiating ameloblasts and odontoblasts in molar tooth germs from 1- to 3-day-old rats by confocal laser scanning microscopy. Immunoreactivity for ZO-1 was strong at proximal and distal junctional complexes of differentiating ameloblasts, while it was weak and punctuate at the distal region of differentiating odontoblasts. Occludin was immunoreactive at distal and proximal complexes of early differentiating ameloblasts and at distal regions of differentiating odontoblasts. However, in more advanced stages, occludin was only evident at the proximal complex of ameloblasts. Claudin-1 was strongly detected at the proximal complex but it was weak at distal complex of late differentiating ameloblasts. Thus, our results showed that ZO-1, occludin, and claudin-1 are differentially expressed as TJs assemble for regulating polarity and/or paracellular permeability in differentiating ameloblasts and odontoblasts.

Ameloblasts↗

Diffusion of Ca(OH)2 associated with different vehicles: chromatographic study (high-performance liquid chromatography).

Using high-performance liquid chromatography (HPLC), small amounts of liquid samples in which 25 premolar human teeth were immersed were evaluated. Each tooth was immersed separately in 800-ml flasks with distilled ultra-pure deionized water and remained there for 1678 h after the filling of their canals with Ca(OH)2 associated with different vehicles: group 1: polyethylene glycol and colophon (Calen); group 2: glycerin and camphorated paramonochlorophenol; group 3: camphorated paramonochlorophenol; group 4: glycerin and tricresol formol; and group 5: anesthetic solution (Citanest). Five polyethylene tubes were filled with each of these pastes and placed unsealed in similar flasks. At the end of this period, HPLC analyses of the aqueous medium related to each group were performed to detect other substances that had diffused from the pastes used in the canals of the teeth other than calcium and hydroxyl ions. Although the groups presented different maximum peaks when there was no barrier, they all showed higher values than when the tooth was present. At least 15 substances other than Ca2+ and OH- were detected in the aqueous medium of group 4. Analyzing the HPLC graphs, we concluded that not only Ca2+ and OH-, but also a considerable quantity of other components of the pastes diffused through the dentine and reached the external root surface.

Anesthetics, Local↗

Phospholipids in amelogenesis and dentinogenesis.

Phospholipids have been identified in enamel and dentin. Before demineralization, a group of phospholipids extracted by lipid solvents was associated with cell membranes and is therefore closely related to cell growth and intracellular regulations. After demineralization, a second group of phospholipids, associated with the extracellular matrix, was extracted; this group is probably linked to the mineralized phase. Using imidazole-osmium tetroxide fixation of rat incisors, we stained cellular unsaturated fatty acids, so that we could visualize the membrane domains, coated pits, and endocytic inclusions. Filipin, a probe for cholesterol, varied in density along the plasma membrane of secretory ameloblasts, and allowed us to visualize membrane remnants inside the forming enamel. With respect to phospholipids located in the extracellular matrix, the malachite-green-glutaraldehyde (MGA) method or iodoplatinate (IP) reaction retains and visualizes enamel and dentin phospholipids. In predentin, aggregates appearing as granules and filaments, or liposome-like structures, were located in the spaces between collagen fibrils. In dentin, organic envelopes coating the crystals, also named "crystal-ghost" structures, outlined groups of collagen fibrils. Histochemical data provided evidence that phospholipids are co-distributed or interact with proteoglycans. Radioautography after IP reaction established that [3H] choline was detected in dentin as early as 30 min after the intravenous injection of the labeled precursor, before any labeling was seen in odontoblasts and predentin. This suggests that blood-serum-labeled phospholipids pass between odontoblasts, cross the distal permeable junctional complex, and diffuse in dentin prior to any cellular uptake and phospholipid synthesis. Pharmacologically and genetically induced pathology also supports the suggestion that phospholipids play an important role in the formation and mineralization of dental tissues.

Ameloblasts↗

Intercellular junctions between human odontoblasts. A freeze-fracture study after demineralization.

Intercellular junctions in the odontoblastic layer have been studied with a freeze-fracture technique. Children's tooth germs were fixed, sliced and demineralized. Samples of the pulpodentinal border were routinely prepared for freeze-fracture. Three kinds of intercellular junctions were detected between human odontoblast cell bodies: gap junctions, desmosomes and tight junctions. Numerous gap junctions are responsible for intercellular communication at different levels of the cell bodies. Focal tight junctions, parallel to the axis of the cell, and desmosomes are sites of cell-to-cell adhesion between lateral plasma membranes. At the distal end of the cell bodies, junctional complexes consist of zonular tight junctions and gap junctions. These zonular tight junctions, never before described between odontoblasts, contribute to the pseudo-epithelial organization of the odontoblastic layer. They constitute a predentin-pulp barrier, the permeability of which must be studied to establish their role in relation to dentin formation.

Adolescent↗

The long term durability of bond strengths to dentin.

The aim of this study was to investigate the durability, throughout one year, of tensile bond strengths (TBS) to bovine dentin using various commercial and experimental bonding systems. Specimens were stored in a controlled solution of ion-exchanged water containing plaster chips and sodium azide. From the results it was concluded that the changes in TBS were not uniform over time, but a significant decrease was usually observed. For Super Bond D-liner and KB-100, the TBS were the highest and exhibited remarkable stability over the test period. The mode of fracture was noted to vary depending on the treatment system used, and was independent of TBS. Generally, the fracture mode tended to show increases in adhesive/cohesive failures within the resin over time. Super Bond D-liner always exhibited adhesive type failure at the tooth interface, and later involved failure in the hybrid layer. KB-100 showed very little change in failure over one year, being usually adhesive between bonding resin and resin composite. The results from this study indicate the need to carry out durability studies for the basic evaluation of all bonding systems. It was shown that the use of a controlled storage solution is important.

Acid Etching, Dental↗

Effects of fluoride on the calcium phosphate precipitation method for dentinal tubule occlusion.

Use of the calcium phosphate precipitation (CPP) method makes possible the occlusion of dentinal tubules to approximately 10 to 15 microns from the dentinal surface, and thus shows good potential for the treatment of dentin hypersensitivity. The precipitate formed in the dentinal tubules by the CPP method is, however, not apatite [HAP; Ca10(PO4)6(OH)2], a component of tooth and bone, but dicalcium phosphate dihydrate (DCPD; CaHPO4.2H2O). Since fluoride enhances the conversion of DCPD to HAP, we evaluated the effects of fluoride on the texture of the precipitate formed by the CPP method and on its capacity to occlude dentinal tubules in this in vitro study. CPP solution (1.0 mol/L CaHPO4.2H2O dissolved in 2.0 mol/L H3PO4) was applied to a dentin disk and was subsequently neutralized with a post-treatment solution (1 mol/L NaOH, from 0 to 0.1 mol/L NaF). Scanning electron microscopy revealed that the precipitate occluded dentinal tubules to a depth of approximately 10 to 15 microns from the dentinal surface, regardless of the NaF concentration (from 0 to 0.1 mol/L) in the post-treatment solution. Also, dentin permeability was reduced to 15% by the CPP treatment regardless of the NaF concentration. The Ca/P molar ratio of the precipitate, measured by x-ray micro-analysis, was higher (1.25 +/- 0.04) in the presence of NaF than in its absence (1.03 +/- 0.01). For further identification of the precipitate formed in the dentinal tubules, the same procedure was used in glass tubes (diameter, 1 mm), so that a larger amount of precipitate would be obtained.(ABSTRACT TRUNCATED AT 250 WORDS)

Calcium Phosphates↗

The movement of radioactive molecules across dentine in vivo in the dog.

Using a technique for measuring the movement of circulating molecules across the odontoblast-dentinal complex, the movement of water and alpha-aminoisobutyric acid through dentine and into a flow cell prepared on the tooth is described. The movement of water depended on dentine thickness, and varied from 0.40 microliter/min per cm2 in a flow cell overlying an average thickness of 0.74 mm of dentine, to 1.23 microliter/min per cm2 in a flow cell with a dentinal floor of 0.40 mm average thickness. The data suggest that alpha-aminoisobutyric-acid concentrations were lower in dentinal fluid than in blood plasma.

Aminoisobutyric Acids↗

Ascorbic acid and aging in the rat. Uptake of ascorbic acid by teeth and concentration of various forms of ascorbic acid in different organs.

1. The uptake of ascorbic acid in vitro by the teeth of rats showed a gradual decrease with age, indicating that the uptake may be related to collagen synthesis as in bone. 2. The concentration of total free ascorbic acid in various organs declined with age, but the rate of decline was different in different organs. In the spleen, however, it increased until maturity and then declined. 3. This decrease may be due to one or both of the following reasons: (a) the permeability of different tissues may decrease at different rates for ascorbic acid, or (b) the requirement for ascorbic acid may decrease at different rates. 4. The bound ascorbic acid declined with age in the skin, kidney, liver and brain after the age of 10-12 weeks, and in the spleen after the age of 26 weeks. 5. The concentration of dehydroascorbic acid and dioxogulonic acid declined with age in the skin.

Aging↗

Scanning electron microscopic study of the apical dentine surfaces lased with ND:YAG laser following apicectomy and retrofill.

The purpose of this investigation was to study the effects of a Nd:YAG laser on the cut surface of teeth using scanning electron microscopy (SEM). Eighteen single-rooted teeth were cleaned, shaped, and obturated with gutta-percha and root canal sealer. The apical 3 mm of each tooth were resected with a diamond fissure bur, and the teeth were randomly divided into two groups of nine teeth each. The resected surface of each root in one group was lased twice. The duration of lasing and the number of pulses were recorded for each tooth. The teeth were air dried, mounted on stubs, sputter-coated with gold-palladium and examined under SEM. Application of the Nd:YAG laser caused melting of apical dentine surfaces. The melted material resembled the appearance of glazed interconnected droplets. Resolidification and recrystalization of the melted areas appeared to be incomplete and discontinuous. Some areas between the glazed regions appeared similar to those of non-lased apical dentine resected root surfaces.

Apicoectomy↗

In vitro investigation of the liberation of fluoride ions from toothpaste compounds in a permeation model.

The most important requirement for effective topical fluoride prophylaxis from toothpaste containing fluoride is that the active fluoride agent must be chemically free, and the rapid spread of the dissolved fluoride ions over the tooth surface. Abrasive compounds in the toothpastes and the brief residence time at the site of action, the oral cavity and tooth surface must not prevent the liberation. Using a two-chamber diffusion cell and an ion-selective fluoride electrode, the content of fluoride ions in five different fluoride-containing toothpastes was determined by direct potentiometry as a function of time and the different abrasive compounds employed. The investigation has demonstrated that a reduction of the release rate of fluoride ions by nearly 50% is seen when calcium carbonate and calcium hydrogen phosphate dihydrate are used as abrasive compounds and combined with sodium fluoride.

Fluorides↗

Fate of horseradish peroxidase in the secretion zone of the rat incisor enamel organ.

Adult CDF albino rats were killed from 10 min to 6 hr after a single intravenous dose of HRP. Experimental and control tissues were reacted for peroxidase activity and processed for light and electron microscopy. At 10 min, all extracellular spaces of the secretion zone showed reaction product. A reaction was also seen around Tomes' processes and in a layer of enamel spaces in the region of thin enamel. At later times intervals, reactions around Tomes' processes were also seen in regions of thicker enamel. Tracer was located preferentially at the growth fronts of rod and interrod enamel, and also diffused for some distance into enamel. From 2 to 6 hr, the enamel over the transition zone became heavily labeled. The tracer penetrated for more than 90 micrometer into the enamel and was localized mainly in the interrod enamel. Droplets of dense stippled material in the extracellular spaces between Tomes' processes did not mix with tracer, but sites which contain a light stippled material in the controls (extracellular spaces, vesicles within ameloblasts) showed a reaction. It is concluded that (1) the basal terminal bars of secretory ameloblasts do not impede the flow of large molecules, (2) the apical terminal bars are permeable in early secretion, become increasingly tight as secretion progresses, and are again permeable in the transition zone, (3) ameloblasts can shuttle large extracellular molecules towards the enamel growth fronts, (4) large molecules can diffuse into enamel; rod and interrod enamel differ with regard to the diffusion of large molecules, (5) ameloblasts phagocytose significant amounts of light stippled material. The possibility is considered that extracellular enamel precursor molecules move preferentially towards the enamel growth fronts, perhaps by a mechanism involving membrane flow, and diffuse through enamel in similar fashion as HRP.

Ameloblasts↗

Ionic transport through bis-glycidyl methacrylate films.

Specific ion meter studies have shown that ionic transport through bis-GMA fissure sealant films is selective and that dissolved fluoride ions disappear from the solution around tooth sections at a more rapid rate than do calcium and chloride ions. This would indicate that topical fluoridation of pits and fissures can still be effected after placement of a fissure sealant. A causative mechanism for this phenomenon has been postulated. Calcium exchange studies indicate that calcium is removed from the enamel and that the microsolubility of fissure sealed teeth in CaCl2 solution is increased by the preparative and sealing procedure used for the enamel.

Acrylates↗

Effect of hydrostatic pressure on the diffusion of monomers through dentin in vitro.

In previous work, the diffusion of monomers from composite and bonding resins through dentin was demonstrated in vitro. The monomers triethylene glycol dimethacrylate (TEGDMA) and 2-hydroxyethyl methacrylate (HEMA) were identified in samples from the pulp space. In the current study, we examined the effects of two levels of positive hydrostatic pressure on the passage of resin monomers through dentin in vitro from a composite-resin/bonding-resin combination to test the hypothesis that monomer diffusion is prevented by such pressure. An occlusal cavity prepared in the tooth crown was restored with the resins. Distilled water samples from the pulpal space were removed over time and analyzed for monomer content by high-performance liquid chromatography and mass spectrometry. Positive pulpal pressure reduced but did not prevent pulpward movement of diluent monomers that leach from bonding agents and from resin composites through dentin in vitro. The degree of reduction of diffusion was greater with TEGDMA than with the lower-molecular-weight monomer HEMA.

Adolescent↗