Fluoride release from restorative materials and its effects on dentin demineralization.

As the use of adhesive restorative materials has increased during the last several years, interest in adhesive materials that release fluoride has also grown. The purpose of this study was to measure fluoride release from several adhesive restorative materials and to evaluate its effect on dentin resistance to demineralization and on bacterial metabolism in a modified in vitro system. Standardized cavities (1.8 mm in diameter) were prepared in bovine teeth that had been ground to dentin. One cavity in each tooth was restored with one of the following restorative systems: (a) Single Bond and Z100; (b) Single Bond and Tetric Ceram; (c) Fuji Bond LC and Z100; (d) Fuji Bond LC and Tetric Ceram; (e) Fuji II LC; or (f) Fuji IX GP. The other cavity in each tooth was "restored" with wax as a control. For each restorative system, 12 specimens were evaluated for fluoride release during the first 24 hrs after restoration placement. Dentin adjacent to the restored sites was subjected to lactic acid challenge (pH 4.3) for 3 hrs, after which calcium release was measured. Another 12 specimens in each group were stored for 24 hrs in de-ionized water, and were exposed to an S. mutans suspension (1:1 THB/de-ionized water and 50 mM glucose, A660 = 0.2) for 6 hrs, followed by calcium release and pH measurement. Bulk specimens of each material were also made and stored in water. Fluoride released from Fuji Bond LC, Fuji IX GP, and Fuji II LC in bulk was significantly greater than from the other materials. In the restored dentin specimens, increased resistance to demineralization from a lactic acid challenge was directly related to fluoride release. The same effects were seen as a result of the S. mutans challenge. While fluoride release from restorative materials increased the resistance of dentin to demineralization in this system, the clinical relevance of the findings is not known.

Dental wear and growth performance in steers fed sweetpotato cannery waste.

OBJECTIVE: To determine whether feeding sweetpotato cannery waste (SPCW) to cattle had adverse effects on dental wear, growth performance, or ruminal tissues. DESIGN: Clinical trial. ANIMALS: 36 Holstein steers. PROCEDURE: Steers were assigned to 1 of 3 groups. All steers received ryegrass hay ad libitum. In addition, steers in group 1 were fed 3.2 kg of corn and soybean meal/steer/d, steers in group 2 were fed 0.45 kg of soybean meal/steer/d and SPCW ad libitum, and steers in group 3 were fed a mixture of SPCW and broiler litter ad libitum. Samples of rumen fluid were collected on day 56. Steers were slaughtered on day 84, and samples of rumen were submitted for histologic examination. Teeth from control steers were removed, and calcium ion loss in response to etching with 2.28% lactic acid solutions buffered to pH of 3.75, 4.0, 4.25, 4.5, and 4.75 was determined. RESULTS: Average daily gain was lower for steers fed SPCW than for steers in the other 2 groups. Steers fed the SPCW-broiler litter mixture had only mild increases in tooth wear and tooth color scores, compared with control steers, whereas steers fed unbuffered SPCW had substantial increases in tooth wear and tooth color scores. Histologic abnormalities were detected in rumens from steers fed diets containing SPCW. Calcium ion loss decreased as pH of the etching solution increased. CLINICAL IMPLICATIONS: Results indicate that feeding cattle unbuffered SPCW can cause dental erosion, ruminal epithelial changes, and poor growth; however, SPCW buffered with broiler litter can be used as a cattle feed.

In vitro effects of an acidic by-product feed on bovine teeth.

OBJECTIVE: To evaluate in vitro erosive effects of sweet potato cannery waste (SPCW) on bovine incisor enamel. SAMPLE POPULATION: 20 bovine mandibles. PROCEDURE: Mandibles were collected and incisors were classified into 3 categories: lacking observable wear, advanced normal wear, or abnormal wear associated with feeding SPCW. Intact mandibles were radiographed. Contralateral normal teeth from the same jaw were used to compare Ca2+ loss (etching) with SPCW, lactic acid (pH 3.2), or SPCW neutralized with NaOH to pH 5.0 or 5.5. Scanning electron microscopy was performed to compare etched and unetched specimens. Two abnormally worn teeth were evaluated histologically. Knoop hardness testing was conducted on unexposed areas of surface enamel and enamel exposed to SPCW. RESULTS: Radiography revealed large periapical abscesses in the mandibles exposed to SPCW. Nearly identical amounts of Ca2+ were removed by SPCW and lactic acid solution at the same pH. Scanning electron microscopy did not indicate consistent differences between etch patterns resulting from exposure to SPCW or lactic acid. Mean rate of calcium removal was 56% higher in deciduous than permanent teeth. Knoop hardness data suggested that softening occurred in enamel exposed to SPCW. Neutralizing SPCW to pH 5.5 eliminated calcium removal. Histologic examination of sections indicated that SPCW degraded and removed some dentin matrix proteins. CONCLUSIONS: Exposure to SPCW results in enamel erosion in vitro; low pH is the most likely cause of erosion. Neutralizing SPCW to pH 5.5 eliminated erosive effects. CLINICAL RELEVANCE: Confirmation of SPCW's erosive effects on enamel in vitro supported the field diagnosis.

Immunohistochemical localization of signal transduction pathways during amelogenesis: an initial exploration.

This study was undertaken to map signal transduction pathway (STP) components uniquely associated with the four major receptor groups and their related STPs in association with the events involved in amelogenesis in the rat. Whole-head, freeze-dried sagittal sections were obtained at the level of the maxillary first molars and picked up on transparent adhesive tape. The sections were not decalcified or fixed, providing optimum conditions for immunohistochemical (IHC) localization. Antibodies to pathway components Gs alpha, Gi alpha, Gq alpha, Sos-1, Grb-2, p125Fak, Jak2, and Vav were localized. The respective patterns of localization indicate that the Gq alpha-linked, the receptor tyrosine kinase-initiated, and the integrin receptor-initiated pathways are involved in the proliferating pre-ameloblast cells. In the differentiating and differentiated ameloblasts, the Gs alpha-linked cAMP pathway is involved, apparently reading a factor(s) released by the dentin matrix. The Gq alpha-linked, the receptor tyrosine kinase-initiated, the integrin receptor-initiated, and the cytokine receptor-initiated pathways are also up-regulated in the proximal ends of the ameloblasts. These observations indicate that all four of the major receptor groups are involved in amelogenesis and that the role of classes of ligands not previously implicated in enamel formation must now be considered. It seems that the cells of the enamel organ respond to the appearance and disappearance of autocrine and paracrine growth factors, but they also up-regulate specific STPs to enable them to respond to circulating hormones and growth factors whose concentrations in the extracellular fluids remain relatively constant.

Structural and compositional alteration of tooth enamel in hereditary epidermolysis bullosa.

Epidermolysis bullosa (EB) is a group of conditions characterized by basement membrane and cellular defects that result in skin fragility and variable extra-cutaneous involvement. The teeth can be severely affected with marked enamel malformations. The purpose of this study was to characterize the structure and composition of teeth from individuals representing the major EB groups (EB simplex, dystrophic EB and junctional EB). Teeth were examined from 28 individuals with EB and 10 healthy people unaffected by EB. Teeth from individuals with junctional EB had marked enamel hypoplasia with varying abnormalities in the enamel structure. Minor structural defects of enamel, including areas of surface pitting, were seen in the other EB types. Although there was a slight reduction (approximately 10%) in the enamel mineral content in several dystrophic EB and junctional EB teeth, the mean mineral content was similar for all EB enamel types and normal enamel. This study shows that while individuals with junctional EB have marked alteration of the enamel structure, the composition may be normal to only mildly altered. Laminin-5, the molecular defect in junctional EB, is associated primarily with alteration in the amount and/or structure of enamel while the mineralization process appears relatively intact. The marked enamel hypoplasia in this EB type suggests that laminin-5 plays an important role in the secretory phase of enamel development.

Problems associated with estimation of net calcium uptake during enamel formation using 45Ca.

45Ca uptake in mineralizing tissues may occur by net Ca uptake or by isotopic exchange. It is rarely possible to differentiate between these effects, making interpretation of the findings difficult. Unfortunately, this problem is not often considered, and 45Ca uptake is usually regarded as representative of only net calcium uptake. The study reported here was undertaken to estimate the extent to which 45Ca uptake in mineralizing enamel is due to net Ca deposition or to isotopic exchange, and to consider the implications. The enamel surfaces of the lower incisors of adult rats were notched at the gingival line, and the eruption distance over 16 hours was measured. This distance was used to establish the position of a 0.3-mm-wide increment of enamel at the beginning and end of the 16-hour period, during which it passed through the early-maturation stage of enamel formation. The rate of Ca uptake was determined by chemical assay. Other rats were injected with 45Ca, mean plasma specific activity values for the experimental period determined, and the rate of Ca uptake through the same area of enamel formation was estimated. The estimates were from two- to nearly ten-fold greater than those established by chemical assay, indicating that from 50 to 90% of the 45Ca uptake occurred by isotopic exchange. 45Ca uptake may indicate more about the labile state of Ca in mineralizing enamel than about the rate of mineral deposition.

The mineral and protein content of enamel in amelogenesis imperfecta.

The purpose of this investigation was to characterize the enamel composition of teeth affected with the hereditary enamel disorders known as amelogenesis imperfecta. Teeth from 10 individuals representing all 3 major AI types (hypocalcified, n = 3; hypomaturation, n = 3; hypoplastic, n = 4) and 10 normal teeth were studied. Half of each tooth was used for histological and biochemical studies. The enamel protein content was estimated by amino acid analysis. The enamel mineral content (volume %) was determined from the calcium and/or phosphorus content. Calcium was measured using atomic absorption and phosphorus was determined colorimetrically. The mean enamel mineral content was reduced for all hypomaturation and hypocalcified AI teeth while hypoplastic AI enamel varied from normal to reduced compared with normal enamel. The enamel protein content was increased in all but one AI case (7 cases were examined for protein) compared with the normal enamel. The mineral and protein content in AI enamel showed a significant inverse relationship (R = -0.939, P = 0.001). This study shows that all three of the major AI groups can have subtypes associated with substantial decreases in the enamel mineral content, although hypomineralization appears most severe in the hypomaturation and hypocalcified AI types. The decreased mineral content was associated with an increased protein content in AI enamel. These findings provide further evidence that altered enamel mineralization in AI teeth likely involves abnormal post-secretory processing of the enamel proteins.

Cytogenetic studies of sodium fluoride in mice.

The cytogenetic effects of sodium fluoride (NaF) were measured in mice following administration in the drinking water for 6 weeks. Bone fluoride levels were determined and showed a dose-related incorporation of fluoride. Micronuclei were measured in peripheral blood erythrocytes following 1 and 6 weeks of NaF administration. Bone marrow cell preparations were examined for the presence of chromosome aberrations following 6 weeks of treatment; metaphase and anaphase cells were examined. Anaphase cells were scored in three independent laboratories, two of which also scored metaphase cells from the same slides. No increases in micronuclei were seen in peripheral erythrocytes at either time point, and no increases in chromosome aberrations were seen in bone marrow cells when metaphase or anaphase cells were examined. A concurrent positive control, cyclophosphamide, produced significant increases in peripheral blood cell micronuclei and in chromosome aberrations in bone marrow cells in metaphase. No increases in aberrations were seen in the same cyclophosphamide-treated mice when anaphase cells were examined.

Fluoride uptake in hard tissues of fetal guinea pigs in response to various dose regimens.

This study was designed to explore various aspects of maternal-fetal fluoride metabolism. In its first phase, 57 female guinea pigs were bred and randomly assigned to a control group (I) or one of three experimental groups: (II) 3 parts/10(6) fluoride in drinking water, (III) a single daily oral dose and (IV) 3 parts/10(6) fluoride in water and a single daily oral fluoride dose. The total mean doses received by groups II and III were similar. The total mean dose received by group IV was approximately double that for groups II and III. Samples of maternal plasma, and fetal bone and enamel were collected on the 57th day of gestation. In its second phase, 53 pregnant guinea pigs were given drinking water containing 0, 2, 4, 6 or 8 parts/10(6) fluoride during gestation. On the 57th day of gestation samples of maternal and fetal enamel were collected. All samples in both phases of the study were assayed for fluoride using the microdiffusion, ion-selective electrode method. In the first part of the study, mean fetal enamel fluoride by group was: I, 21.6 parts/10(6); II, 38.6 parts/10(6); III, 33.5 parts/10(6); IV, 54.9 parts/10(6). In the second phase, maternal enamel was linearly related to the fluoride dose. The same was true for fetal enamel except at the 8 parts/10(6) fluoride level in the water where there was no increase over the 6 parts/10(6) group. At all dose levels, fetal enamel fluoride uptake was approximately an order of magnitude less than maternal enamel fluoride uptake.(ABSTRACT TRUNCATED AT 250 WORDS)

Fluoride and calcium content of enamel organ, muscle, liver and plasma in rats.

Nine-day-old rats were given oral fluoride doses (0.5 micrograms F/g body weight). Plasma, enamel organ, muscle and liver samples were collected from nondosed pups (baseline) and 15, 30, 60 and 120 min after the dose. Samples were assayed for fluoride and calcium concentrations. Adult rats were given 20 ppm F water for 6 weeks, and the same tissues were sampled and assayed for fluoride concentrations only. In the first phase of the study, enamel organ had significantly higher fluoride and calcium content than liver and muscle. In adult rats the fluoride content of enamel organ was again significantly higher than for liver and muscle.

Fluoride concentrations in human and rat bone.

The most recent report on fluoride concentrations ([F]) in human bone was published over a decade ago. Such data are of interest in the context of changing patterns in systemic fluoride exposure. In the study reported here, bone samples were collected from 24 human subjects who underwent orthopedic surgery. Medical histories and the best possible life-time systemic fluoride exposure information were obtained from each subject. Bone samples were assayed for fluoride concentration using the acid diffusion, ion selective electrode method. For ash from whole bone, the lowest value was 378 ppm in a 16-year-old subject, and the highest value was 3,708 ppm in a 79-year-old person. Fluoride concentrations in bone were significantly correlated with age (r = .62). The regression line intercept at birth was 442 ppm, and the slope was 22 ppm per year. When measured separately, trabecular bone ash fluoride concentrations were significantly higher than the corresponding cortical bone values. Trabecular and cortical bone samples from rats' drinking water containing 75 ppm F were assayed for F. The mean trabecular bone fluoride concentration was significantly higher than the mean cortical bone concentration. There was close agreement between F assay results using a modification of the acid diffusion method and the method originally reported by Singer and Armstrong. The human bone ash [F] values reported in this study are similar to those reported from other North American subjects over the last three decades. These findings are of interest in the context of evidence indicating increased systemic fluoride exposure in the United States population.(ABSTRACT TRUNCATED AT 250 WORDS)

Effect of an acute maternal fluoride dose on fetal plasma fluoride levels and enamel fluoride uptake in guinea pigs.

We conducted this study to measure maternal plasma, fetal plasma, and fetal enamel fluoride concentrations for four hours following an oral F dose to near-term pregnant guinea pigs. We placed female guinea pigs on de-ionized (Group I) or 3-ppm-F (Group II) drinking water prior to breeding and during gestation. On the 57th day of gestation, we administered a maternal dose of NaF solution (0.6 mg F/kg) by stomach tube. We collected samples of maternal plasma, fetal plasma, and fetal enamel at baseline, at 15 and 30 min, and at one, two, and four h after administration of the dose. We assayed samples for F using a modification of the micro-diffusion and ion-specific electrode method. Group I mean baseline F values were: maternal plasma, 0.016; fetal plasma, 0.002; and fetal enamel, 7.0 ppm. Group II mean values were: 0.055, 0.004, and 19.0 ppm. After the maternal fluoride dose, the mean maternal plasma [F] rose sharply for 30 to 60 min and declined to about 50% of peak values by four h. Fetal plasma [F] changed less in absolute values, but similarly to maternal changes relative to baseline. Fetal enamel mean [F] rose more in Group II than in Group I. Baseline F status had an important effect on F uptake in fetal enamel following an acute maternal fluoride dose.

The Ca, Pi, F, and proline content of developing bovine enamel under GBHA-stained and unstained bands.

The surface enamel of fetal bovine teeth was stained with GBHA to indicate the position of bands of smooth-ended and ruffle-ended ameloblasts relative to the developing enamel. The boundaries of the bands were scored, under a dissecting microscope, and the bulk enamel under each band was collected. The enamel samples were assayed for Ca, Pi, F, and proline. The amount of Ca and Pi in the enamel increased in successive bands and seemed unrelated to the overlying ameloblast cell type. The loss of proline seemed unrelated to cell type. The fluoride content of enamel increased by approximately 50% in the first stained band immediately adjacent to the secretory zone. The F level returned to secretory values in the succeeding unstained band. Thus, only changes in the F level of developing enamel appeared to be related to GBHA staining patterns.

The short-term uptake and retention of fluoride in developing enamel and bone.

Eight- and 12-day-old rat pups were injected intraperitoneally with fluoride. Plasma, molar enamel, and bone samples were collected at observation times up to six hr after injection. In a second series, adult rats maintained for six weeks on water containing 5 ppm F were injected with fluoride. Plasma, incisor enamel, and bone samples were collected at the same observation times as those used in the first series. Fluoride assays were conducted by means of the microdiffusion, ion-selective-electrode method. In the suckling rats, plasma [F] levels peaked at 15 min and returned nearly to baseline in one hr. Significant increases in the [F] of developing enamel and bone were observed. No significant decline from the peak [F] seen in the hard tissues was observed over the six-hour period. Similar results were seen in the developing enamel of the adult rats. The data gave no evidence of a short-term reversible component of fluoride uptake in developing enamel. Apparent increases in F uptake in enamel and bone beyond peak plasma values suggest the presence of a diffusion-limiting membrane for fluoride from the extracellular fluids into the mineralizing matrix.

Diffusion of fluoride through the rat enamel organ in vitro.

This study investigated the diffusion of fluoride through the enamel organ in vitro. The rat molar explants used were entirely in the secretory stage or predominantly in the maturation stage of enamel formation. The removal of the enamel organ or metabolic inhibition with iodoacetate caused significant increases in enamel fluoride uptake at both stages of enamel formation. Inhibition with dinitrophenol caused a significant increase only in the maturation phase. Uptake of fluoride in enamel was related to the fluoride concentration in the medium, except in the maturation stage explants, where increasing the medium fluoride concentration from 0.05 ppm to 0.08 ppm did not significantly increase fluoride uptake at any of the three observation times. The findings indicate that the enamel organ exists as a diffusion-limiting membrane to the movement of fluoride from the extracellular fluid compartment to the developing enamel.

A comparison of the effects of continuous and periodic fluoride delivery on fluoride levels in plasma, enamel, and bones of nursing rats.

The purpose of this study was to determine and compare the F levels in plasma, enamel, and bones of nursing rat pups that received the same daily dose of F by continuous or periodic delivery during enamel development. The hypothesis was that F delivered continuously would result in enamel F levels higher than those attained when F was delivered periodically. For continuous delivery, copolymer devices (Southern Research Institute) that provide slow release of F were implanted in the backs of four-day-old rat pups. For periodic delivery, rat pups received F by intraperitoneal injection or gastric intubation. The doses were 0.01, 0.02, or 0.04 mg F/day. The rats were killed at 13 days of age, 24 hours after the last periodic delivery. Plasma was collected, femur and calvaria bones were removed, and enamel was scraped from developing first molars. Fluoride assay was by the microdiffusion method of Taves, with a F electrode. For the 0.02 mg F/day dose, plasma levels in control, implanted, injected, and gastric-intubated rats were 0.004, 0.020, 0.011, and 0.009 ppm, respectively. Enamel F levels were 1.1, 61.9, 54.0, and 42.3 ppm, respectively. Femur F levels were 2.2, 81.2, 84.8, and 68.1 ppm, respectively. Calvaria F levels were 2.5, 79.3, 80.1, and 67.9 ppm, respectively. The results showed that there was no significant difference in the enamel F levels or in the bone F levels in rat pups that received continuous or periodic, by injection, delivery of F at the same daily dose.(ABSTRACT TRUNCATED AT 250 WORDS)

Fluoride uptake and retention at various stages of rat molar enamel development.

Suckling rat pups were given intraperitoneal fluoride injections at selected ages so that we could study fluoride uptake in the enamel of the maxillary first molar at various stages of enamel development. Plasma fluoride levels in six-day-old and 11-day-old pups were monitored following the intraperitoneal injection of fluoride. The findings indicate that: (1) fluoride was more easily taken up and retained during the early stages of enamel formation, but fluoride uptake can occur during all stages of enamel formation; (2) when injections were started early in enamel formation, more fluoride was contained in the enamel of the maxillary first molar at 13 days of age; and (3) the same dose of fluoride per gram body weight resulted in greater exposure to elevated plasma fluoride levels in six-day-old pups than in 11-day-old pups.

Enamel fluoride in nursing rats with mothers drinking water with high fluoride concentrations.

The purpose of this study was to determine the F levels in plasma and molar enamel from rat pups whose mothers had received various levels of F during pregnancy and/or lactation. Rats were started on water containing 0 (Group I), 50 (Group II), or 100 (Group III) ppm F at the beginning of pregnancy or on the day of delivery. The mothers and pups were killed 13 days after delivery, and plasma F levels, milk F levels, and pup molar enamel F levels were determined. The mean maternal plasma F concentrations were 0.02 +/- 0.005 ppm in Group I, 0.10 +/- 0.031 ppm in Group II, and 0.21 +/- 0.057 ppm in Group III. The milk F values were about twice as high as the respective plasma concentrations. The plasma F concentration in control pups was 0.003 +/- 0.0002 ppm, and there was a rise to 0.006 +/- 0.0002 ppm in Group III. Enamel F concentrations were 0.62 +/- 0.13 ppm, 4.72 +/- 0.79 ppm, and 8.80 +/- 1.74 ppm, respectively. The plasma and enamel F values obtained from pups were not significantly different between the pre-natal/post-natal, and the post-natal-only groups. It was concluded that: fluoride levels in the plasma and enamel of control rat pups were much lower than those found in adult rats, such values could be increased only slightly when high doses of F were given to the mother, and unlike values reported for other species, rat milk fluoride concentrations were higher than the respective plasma values.

The effects of parathyroid hormone, calcitonin, and vitamin D metabolites on calcium transport in the secretory rat enamel organ.

The effects of parathyroid hormone (PTH), calcitonin (CT), 1,25(OH)2D3, and 24,25(OH)2D3 on calcium transport through the secretory stage enamel organ were studied on developing rat molars in vitro. 24,25(OH)2D3 increased 45Ca uptake by the explants. 24,25(OH)2D3 plus PTH further enhanced 45Ca uptake and resulted in an increase in net calcium uptake by the developing enamel.