Animal Studies © 1990 S. Karger A G, Basel 0008-6568/90/0242-0117 $ 2.75/0

Caries Res 1990;24:117-120

Distribution of Fluoride across Cementum, Dentine and Alveolar Bone in Rats K. Katoa, H. Nakagaki\ C. Robinsonh, J.A. Weatherellb •' Department of Preventive Dentistry and Dental Public Health, School of Dentistry, Aichi-Gakuin University, Nagoya, Japan; h Department of Oral Biology, School of Dentistry, University of Leeds, UK

Key Words. Alveolar bone ■Cementum ■Dentine • Fluoride • Rat Abstract. This study was undertaken to determine the fluoride distribution in cementum and neighboring hard tissues of the rat after different levels of fluoride administration via the drinking water. Specimens of ce­ mentum with underlying dentine and adjacent bone were removed from the distal roots of the first lower mo­ lars. The fluoride distribution in each specimen was determined in samples removed sequentially using an ab­ rasive microsampling technique. Fluoride concentrations were highest at or near the surface and decreased towards the interior of cementum, dentine and alveolar bone in both control and experimental groups. With increasing fluoride intake, concentrations increased throughout the tissue. The distribution patterns of fluoride in cementum of contralateral teeth from the same animal were similar. Fluoride concentrations in cementum were higher than those of dentine and alveolar bone.

It was reported that in human cementum, fluoride concentrations were higher at or near the tissue sur­ face and decreased towards the cementum-dentine junction (CDJ), with an occasional peak of fluoride concentration in some regions of the fluoride gradient [Nakagaki et al., 1985]. Variations in fluoride concen­ tration across the tissue often produced a recogniz­ able profile. This generally appeared to be similar in the same type of tooth [Nakagaki et al., 1985], and tended to be characteristic of the patient from which the tooth had been extracted [Murakami et al., 1987], Nakagaki et al. [1988] also demonstrated a relation­ ship between the fluoride distribution and the histo­ logical structure of the cementum. In the rat cementum, however, there is no analo­ gous information and it was considered worthwhile to attempt a similar study on this very useful experimen­ tal animal. The present study describes the fluoride distribution in the cementum and neighboring hard tissues of rats receiving drinking water containing dif­ ferent concentrations of fluoride.

Materials and Methods Animals and Fluoride Administration Twenty-eight 5-week-old male Wistar rats, average weight 145 g, were divided into 4 equal groups. One group, given distilled water, served as a control. The 3 experimental groups received drinking water ad libitum containing 25, 50 and 100 ppm fluoride, respectively, for 10 weeks. All animals were given ad libitum a lab­ oratory diet (CE2, CLEA Japan Inc., Tokyo, Japan), containing an additional 32 ppm of fluoride. The weight of each rat was recorded weekly. Specimens The animals were killed by decapitation under chloroform anaesthesia. Longitudinal sections were cut through the lower mandible, including the first molars. These sections were ground to a thickness of about 500 urn and a specimen, which included den­ tine, cementum and the adjacent bone, was removed from the distal root of each section, as indicated in figure I. Removal and Recovery of Samples The specimen was attached to a brass rod and inserted in the sleeve of Mikrokator (C.E. Johansson, Eskilstuna, Sweden). Sam­ ple layers, 20 ,um thick, were serially abraded from the surface through the specimen with an abrasive microsampling technique as previously described, manipulating the specimen’s orientation in order to sample the cementum, dentine or bone, as required

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Introduction

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Kato/Nakagaki/Robinson/Weatherell x 1 O'* p p m F

[Weatherell et al., 1985]. After each layer had been removed, the specimen was carefully examined under a dissecting microscope. The powder from each layer was dissolved in 3 pi of 1 M HC104 and the solution was transferred to a 0.1-ml polythene capsule. Twelve microliters of acetate buffer were added to give a total solu­ tion volume of about 15 pi and a final pH of 5.2. Determination o f Phosphorus and Fluoride and Calculation of Results The concentration of phosphorus was determined on 4 pi of this 15-pl solution using the colorimetric method of Chen et al. [1956], Fluoride was determined on 3 pi of the same 15-pl solution, using the Orion ion-specific electrode (94-09, 90-01, Orion Res., Mass., USA) according to the microprocedure of Hallsworth et al. [1976], The fluoride concentrations in the original samples were calcu­ lated from the F:P ratios obtained. The results are expressed as parts/106 F per weight of cementum, dentine and alveolar bone cal­ culating these from the P estimation, on the assumption that the P concentration of cementum was 12.2%, dentine 13.5% and alveolar bone 10.3% per dry weight tissue [Lazzari, 1968], An electron mi­ croanalysis showed that the phosphorus concentration in the ce­ mentum was relatively constant. The fluoride concentrations esti­ mated from F : P ratios were therefore compared to each other with­ out a correction for the phosphorus concentration in the tissue. The statistical significance of differences between fluoride con­ centrations of the different groups was evaluated in the superficial and innermost layers of cementum using Student’s t test.

Fig. 2. Fluoride distribution, from surface to CDJ, across the cementum of rats which received water containing 0, 25, 50 and 100 ppm fluoride.

Results Fluoride Distribution from Surface to Interior o f Cementum Fluoride distributions from the surface to the CDJ are presented in figure 2. Fluoride concentrations were generally highest at the surface of cementum and decreased towards the interior of the tissue. Both con­ trol and the experimental groups exhibited a fluoridegradient curve. The concentration was not always maximal in the first layer, however. In some speci­ mens maximum concentrations occurred in the subsurface. With increasing levels of fluoride in the drinking water, the concentration of fluoride increased significantly in both the outer and inner cementum. Profiles o f Fluoride Distribution in Cementum o f Right versus Left Molars The profiles of fluoride distribution in the cemen­ tum of right and left molars are compared in figure 3. It can be seen that the fluoride-gradients of each pair of contralateral teeth were fairly similar. The fluoride

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.

Fig. 1 Change in fluoride concentration from root dentine towards alveolar bone of the rat.

Distribution of Fluoride in the Root of Rat Molars

Fluoride Distribution across Dentine, Cementum and Alveolar Bone The profiles of fluoride distribution across root dentine, cementum and alveolar bone are shown in figure 1. In each group, fluoride concentrations were higher at the pulpal surface of the dentine, decreased to the CDJ and rose again towards the root surface of the cementum. In the surface of alveolar bone, fluo­ ride concentrations were maximum, decreasing again towards the interior of the tissue. In every tissue, fluo­ ride concentrations were therefore highest in the sur­ face regions, decreasing towards the tissue interior. Fluoride concentrations increased markedly in each tissue as the intake of fluoride increased. The fluoride profile from the outer to the inner root surface thus exhibited a deep, symmetrical trough, with lowest concentrations coinciding with the CDJ. The highest average concentration of fluoride occurred in cemen­ tum, although this reflected the fact that cementum is of limited width compared, e.g. to bone.

Discussion Fluoride profiles have been described previously in human dental tissues [Nakagaki et al., 1987], in rat enamel [Kato et al., 1988] and rat bone [Narita et al., 1989], The present results provide analogous detailed information about fluoride distribution in rat cemen­ tum, dentine and bone. In mature human enamel, the concentration of flu­ oride is invariably highest at the surface and then usually decreased exponentially towards the interior [Nakagaki et al., 1984; Chow et al., 1985]. In the cementum, on the other hand, the concentration is higher near the root surface, although not always in the most superficial layer. There were also occasional peaks of higher concentration which frequently gave the profile a recognizable shape, characteristic of the patient’s dentition [Nakagaki et al., 1985; Murakami et al., 1987], These differences between the fluoride profiles of enamel and cementum probably reflect differences in their development and in their ultimate structure.

x 1 03ppmF

Fig. 3. Fluoride distribution, from surface to CDJ, across the ce­ mentum of right (...) and left (-) teeth from the same animal.

Kato et al. [1988] suggested that the distribution of fluoride in developing enamel of rats resulted from the pattern of protein withdrawal and the subsequent mineralization of enamel matrix. There was no subse­ quent enamel apposition and no evidence that the flu­ oride content of sound enamel changed significantly with age, although the surface enamel could be worn or demineralized, this leading to a loss or enhance­ ment of fluoride concentration [Nakagaki et al., 1987], In contrast, the deposition of the human cemen­ tum occurs continually throughout life [Zander and Hiirzeler, 1958; Hiirzeler and Zander, 1959]. The fluo­ ride concentration also increases with age [Nakagaki et al., 1985]. Sognnaes et al. [1955] showed the cemen­ tum was more permeable than other mineralized tis­ sues. Yoon et al. [1960] and Nakagaki et al. [1987] ob­ served the fluoride uptake from the tissue fluid in the periodontal space into the cementum. This and the overall shape of the fluoride gradients (fig. 1) suggest that the fluoride penetrates into the inner tissue from the exterior root surface. This is in accordance with the recent suggestion of Nopakun et al. [1988] that the fluoride released during the resorption o f alveolar bone might be absorbed by the cementum. Variations in the rate of apposition of cementum and the diffusion of fluoride into the tissue from tis­

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levels in the surface layers of both teeth from the con­ trol, 25- and 50-ppm F groups were also similar. The peak concentrations in the 100-ppm F group occurred approximately at the same point within the cementum of the right and left molars.

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sue fluids may be responsible for the occasional peaks of higher concentration found in the profiles of fluo­ ride distribution. These patterns tended to be similar in certain aspects for the same tooth type from many subjects. The patterns also exhibited features charac­ teristic of the individual patient [Nakagaki et al., 1985; Murakami et al., 1987], The present findings re­ veal analogous variations in the patterns of fluoride distribution in rat molar cementum, although the pro­ files are less complex in rat molars than in human teeth. This is presumably due to the much greater pe­ riod of cement growth and tissue thickness in human cementum. Characteristic fluoride profiles in cementum of the right versus left rat molars (fig. 3), with similar con­ centrations and patterns of fluoride in the teeth from the same animal, suggest an overriding influence of systemic factors, e.g. fluoride exposure. In man, Mu­ rakami et al. [1987] reported that the fluoride profiles of cementum of the homologous upper and lower teeth could sometimes be more similar than those of homologous right and left teeth, suggesting that local factors such as mastication influence fluoride pro­ files. More specific studies should be undertaken to confirm these suggestions. In general, the present results suggest that fluoride concentrations in cementum were higher than those of dentine and alveolar bone. Previous reports of flu­ oride concentration in enamel [Kato et al., 1988] and bone [Narita et al., 1989] of rats under the same exper­ imental conditions indicated that the fluoride concen­ tration in the surface of cementum was approximately twice that of bone and 4-5 times that of enamel. It would appear that, as in human dental tissues, the av­ erage concentration of fluoride in the rat was highest in cementum and decreased, in descending order, in dentine and enamel [Nakagaki et al., 1987].

References Chen PS, Toribara TY, Warner H: Microdetermination of phos­ phorus. Anal Chem 1956;28:1756-1758. Chow LC, Beaudreau GM, Brown WE: Enamel fluoride profile construction from biopsy data. Caries Res 1985;19:103-112. Hallsworth AS, Weatherell JA, Deutsch D: Determination of sub­ nanogram amounts of fluoride with the fluoride electrode. Anal Chem 1976;48:1660-1664.

Hiirzeler B, Zander HA: Cementum apposition in periodontally diseased teeth. Helv Odontol Acta 1959;3:1-3. Kato K, Nakagaki H, Sakakibara Y, et al: Distribution of fluoride in the enamel of rat incisors examined by an abrasive micro­ sampling technique. Arch Oral Biol 1988;33:653-656. Lazzari EP: Dental Biochemistry. Philadelphia, Lea & Febiger, 1968. Murakami T, Nakagaki H, Sakakibara Y, et al: The distribution pattern of fluoride concentrations in human cementum. Arch Oral Biol 1987;32:567-571. Nakagaki H, Ishii T: An in vivo enamel solubility test; in Feanhead RW, Suga S (eds): Tooth Enamel. Amsterdam, Elsevier, 1984. Nakagaki H, Weatherell JA, Strong M, et al: Distribution of fluo­ ride in human cementum. Arch Oral Biol 1985;30:101-104. Nakagaki H, Koyama Y, Sakakibara Y, et al: Distribution of fluo­ ride across human dental enamel, dentine and cementum. Arch Oral Biol 1987;32:651-654. Nakagaki H, Kawai K, Murakami T, et al: Fluoride distribution and histological structure of human cementum. Arch Oral Biol 1988;33:257-264. Narita N, Nakagaki H, Kato K, et al: Distribution of fluoride con­ centration in the rat’s bone. Calcif Tissue Int, in press. Nopakun J, Guo MK, Messer HH, et al: Fluoride redeposition and retention during bone turnover in lactating rats. J Dent Res 1988:67:1213-1216. Sognnaes RF, Shaw JH, Bogoroch R: Radiotracer studies on bone, cementum, dentine and enamel of rhesus monkeys. Am J Phy­ siol 1955:180:408-420. Weatherell JA, Robinson C, Strong M, et al: Micro-sampling by abrasion. Caries Res 1985:19:97-102. Yoon SH, Brudevold F, Gardner DE, et al: Distribution of fluoride in teeth from areas with different levels of fluoride in the water supply. J Dent Res 1960;39:845-856. Zander HA, Hiirzeler B: Continuous cementum apposition. J Dent Res 1958;37:1035-1044.

Received: March 7, 1989 Accepted: July 5, 1989 K. Kato Department of Preventive Dentistry and Dental Public Health School of Dentistry Aichi-Gakuin University 1-100 Kusumoto-cho Chikusa-ku, Nagoya 464 (Japan)

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Distribution of fluoride across cementum, dentine and alveolar bone in rats.

This study was undertaken to determine the fluoride distribution in cementum and neighboring hard tissues of the rat after different levels of fluorid...
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