Lead in Enamel and Saliva, Dental Caries and the Use of Enamel Biopsies for Measuring Past Exposure to Lead F. BRUDEVOLD, R. AASENDEN, B. N. SRINIVASIAN, and Y. BAKHOS Forsyth Dental Center, 140 Fenway, Boston, Massachusetts 20115, USA Enamel biopsies taken from schoolchildren in a comnmunity where exposure to lead was a health hazard were analyzed for lead and fluoride. The children with high enamel lead had significantly higher caries scores than the children with low enamel lead, in spite of the fact that the high lead group also was higher in enamel fluoride. There was no increase in enamel lead with age. The lead in saliva was only a fraction of that in blood. Infants with lead poisoning showed higher saliva lead than a normal infant. The use of the lead in enamel biopsies and in saliva for measuring exposure to lead is discussed. J Dent Res 56(10):1165-1171 October 1977. In a pilot enamel biopsy study, we recently found several cases of surprisingly high concentrations of lead among school children in Cambridge, Massachusetts.' The range was from 200 to 3,550 ppm, and the mean (S.D.) was 1,790 (960) ppm. We then learned that there had been several cases of lead poisoning in Cambridge and that ongoing screening indicated 11% of the child population had above normal concentrations of lead in the blood.2 The purpose of this study was to find out if this unusual exposure to lead affected the caries experience of the children. It was also considered that the lead in enamel biopsies and in saliva may be a useful indication of past and present exposures to lead. Several studies have been concerned with lead and dental caries. Wisotsky and Hein3 reported that consumption of drinking water containing 0.5 mEq of lead increased caries incidence in hamster males, but not in hamster females. Barnes,4 in a study on natives in New Guinea, found that high levels of dietary lead Received for publication November 16, 1976. Accepted for publication January 19, 1977. This study was supported by U.S.P.H.S. Grant DE02183.
were associated with increased caries experience. Ludvig, Adkins, and Losee,5 who compared caries in school populations in different eastern states in the United States, found relatively high levels of lead in the drinking water of the populations with the highest caries experience, but they were reluctant to postulate a causal relationship. Several studies indicate that posteruptive exposure to lead is without effect on caries. Aston6 reported that workers in lead plants had the same score of DMF teeth as other industrial workers. The lead in topically applied fluoride has failed to show caries inhibition. Studies by Bibby, DeRoche, and Wilkins7 and Gallagher and Knutson8 gave no reduction, and a third study by Klinkenberg and Bibby9 showed the same inhibition as was obtained from sodium fluoride. It seems from this review that ingestion of lead may possibly increase caries, but that topical effects are unlikely. In methods of detecting lead poisoning, the concentration of lead in the blood is generally considered the most reliable index, although the transitory nature of the lead in blood is a shortcoming.10 Shed deciduous teeth have been used to measure the body burden of lead.'1 Needleman et al.'2 found that a greater proportion of seriously exposed children was detected by determining lead in the dentin of deciduous teeth than by blood screening. They analyzed the secondary dentin adjacent to the pulp because, being formed continuously, this dentin gives a cumulative measure of posteruptive exposures to lead. One obvious disadvantage of using deciduous teeth is that they are difficult to obtain. The enamel biopsy procedure would be more suitable for large scale screening provided, of course, that the lead in enamel is a sensitive measure of past exposure to lead. Several investigators have shown that lead readily accumulates in the enamel, but there is uncertainty about whether the accumulation is continuous or occurs only during the pre-erup1165
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tive period. Brudevold and Steadman13 showed high concentrations in the surface enamel of unerupted teeth. They also found increased concentration in teeth from older persons as compared to teeth from younger persons, but were unable to conclude whether this reflected posteruptive acquisition or greater pre-eruptive exposure to lead in the older group. Although there is considerable current interest in the metabolic effects of lead, DiGregario et a114 seem to be the only workers who have been concemed with lead in saliva. They found that stimulated parotid saliva and blood contained nearly the same concentrations, but they analyzed only one sample of each fluid. In the present study, enamel biopsies and samples of saliva were taken from school children in Cambridge. The biopsies were analyzed for lead and fluoride, and the saliva for lead. A dental examination was also done, and the caries scores of the children with high and low biopsy lead were compared. The posteruptive uptake of lead was assessed by relating the concentrations of lead in the biopsies to age. A few samples of saliva and blood from both normal persons and patients suffering from lead poisoning were also analyzed in order to compare the concentrations of lead in the two fluids.
Materials and Methods
STUDY SUBJECTS.-The study included 9to 12-year-old children from four schools in Cambridge, Mass. Of the 270 signed up for the study, only 251 (123 boys and 128 girls) were included in the comparisons involving all parameters studied, i.e. lead and fluoride concentrations of surface enamel, lead concentrations of saliva, and caries experience. The water in Cambridge has been fluoridated since August 1972. A number of children had received fluoride supplements for shorter or longer periods, and topical fluoride application is given regularly in these schools. Two adults and five infants, four of whom were hospitalized for lead poisoning, were included in an additional project concerned with a comparison of the concentrations of lead in saliva and blood. ENAMEL BIOPSIES, SALIVA AND BLOOD SAMPLES. An enamel biopsy was taken from a maxillary central incisor in each subject. Usually, the right central was biopsied, but when this tooth was not suitable, the left tooth was used. In ten subjects. tNvo consecutive biopsies
I Dent Res October 1977 wsere taken from the same area of the contralateral tooth. A shorter etching time (20 versus 35 seconds) was used for these biopsies. The sampling procedure was as described by Brudevold et al.1 Briefly, the procedure involved the use of an adhesive tape window technic, and etching the exposed enamel for 35 seconds with 10 ,1 of 1.6 N HCl in 70% glycerol. The depth of the etch was calculated from the sampling area (20.1 mm2) and the amount of calcium present in the sample, assuming that the enamel contained 37% calcium and had a density of 2.95 gm/cm3. About 1 ml of unstimulated whole saliva was collected from each subject in a plastic tube. Smaller amounts of whole saliva were collected from the infants by means of a plastic syringe. Stimulated parotid saliva was obtained utilizing lemon sours and a modified Carlson-Crittenden device.15 Blood samples were collected by collaborating medical personnel. The biopsies were analyzed for calcium, lead, and fluoride as described by Brudevold et al,' except that the addition method was used for lead in order to account for matrix effects. In the case of saliva, 0.5 ml was mixed with 0.5 ml 0.1 N HCl, and 10-#1 quantities were analyzed for lead as before. Nonlead adsorption was accounted for by employing D, arc corrector and subtracting the absorption obtained at X = 280.7 ttm (nonlead signal) from that at X 283.3 ,um. Triplicate analyses of several samples showed good reproducibility. The blood analyses were done at the laboratory of the Massachusetts Department of Public Health Childhood Lead Poisoning Prevention Program. CARIES EXAMINATION-.Without prior cleaning, the teeth were isolated with cotton rolls and dried with compressed air. The examinations were performed with a plane surface mirror and a sharp Starlite MG5 doubleended explorer. No radiographs were taken. The examinations included only the permanent teeth. DATA TREATMENT. The biopsy data were grouped according to biopsy depth in order to explore the variation with depth of the lead concentrations. These groupings of the data, and the results of the two consecutive biopsies indicated that the lead gradient in the surface enamel is extremely steep. Therefore, in the evaluation of the data, the biopsy depth was taken into consideration as follows: the biopsy data were grouped by depth using 0.3 amm as class interval. WVithin each class, biopsies with
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Vrol. 56 No. 10
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LEAD IN ENAMEL & SALIVA
1167
TABLE 1 LEAD CONCENTRATIONS OF ENAMEL BIOPSIES GROUPED BY DEPTH Biopsy depth (,im)
No. of samples Mean depth (,um) Mean Pb (ppm) S.D.
0.6-
were associated with increased caries experience. Ludvig, Adkins, and Losee,5 who compared caries in school populations in different eastern states in the United States, found relatively high levels of lead in the drinking water of the populations with the highest caries experience, but they were reluctant to postulate a causal relationship. Several studies indicate that posteruptive exposure to lead is without effect on caries. Aston6 reported that workers in lead plants had the same score of DMF teeth as other industrial workers. The lead in topically applied fluoride has failed to show caries inhibition. Studies by Bibby, DeRoche, and Wilkins7 and Gallagher and Knutson8 gave no reduction, and a third study by Klinkenberg and Bibby9 showed the same inhibition as was obtained from sodium fluoride. It seems from this review that ingestion of lead may possibly increase caries, but that topical effects are unlikely. In methods of detecting lead poisoning, the concentration of lead in the blood is generally considered the most reliable index, although the transitory nature of the lead in blood is a shortcoming.10 Shed deciduous teeth have been used to measure the body burden of lead.'1 Needleman et al.'2 found that a greater proportion of seriously exposed children was detected by determining lead in the dentin of deciduous teeth than by blood screening. They analyzed the secondary dentin adjacent to the pulp because, being formed continuously, this dentin gives a cumulative measure of posteruptive exposures to lead. One obvious disadvantage of using deciduous teeth is that they are difficult to obtain. The enamel biopsy procedure would be more suitable for large scale screening provided, of course, that the lead in enamel is a sensitive measure of past exposure to lead. Several investigators have shown that lead readily accumulates in the enamel, but there is uncertainty about whether the accumulation is continuous or occurs only during the pre-erup1165
Downloaded from jdr.sagepub.com at MICHIGAN STATE UNIV LIBRARIES on January 11, 2015 For personal use only. No other uses without permission.
1166
BRUDEVOLD ET AL
tive period. Brudevold and Steadman13 showed high concentrations in the surface enamel of unerupted teeth. They also found increased concentration in teeth from older persons as compared to teeth from younger persons, but were unable to conclude whether this reflected posteruptive acquisition or greater pre-eruptive exposure to lead in the older group. Although there is considerable current interest in the metabolic effects of lead, DiGregario et a114 seem to be the only workers who have been concemed with lead in saliva. They found that stimulated parotid saliva and blood contained nearly the same concentrations, but they analyzed only one sample of each fluid. In the present study, enamel biopsies and samples of saliva were taken from school children in Cambridge. The biopsies were analyzed for lead and fluoride, and the saliva for lead. A dental examination was also done, and the caries scores of the children with high and low biopsy lead were compared. The posteruptive uptake of lead was assessed by relating the concentrations of lead in the biopsies to age. A few samples of saliva and blood from both normal persons and patients suffering from lead poisoning were also analyzed in order to compare the concentrations of lead in the two fluids.
Materials and Methods
STUDY SUBJECTS.-The study included 9to 12-year-old children from four schools in Cambridge, Mass. Of the 270 signed up for the study, only 251 (123 boys and 128 girls) were included in the comparisons involving all parameters studied, i.e. lead and fluoride concentrations of surface enamel, lead concentrations of saliva, and caries experience. The water in Cambridge has been fluoridated since August 1972. A number of children had received fluoride supplements for shorter or longer periods, and topical fluoride application is given regularly in these schools. Two adults and five infants, four of whom were hospitalized for lead poisoning, were included in an additional project concerned with a comparison of the concentrations of lead in saliva and blood. ENAMEL BIOPSIES, SALIVA AND BLOOD SAMPLES. An enamel biopsy was taken from a maxillary central incisor in each subject. Usually, the right central was biopsied, but when this tooth was not suitable, the left tooth was used. In ten subjects. tNvo consecutive biopsies
I Dent Res October 1977 wsere taken from the same area of the contralateral tooth. A shorter etching time (20 versus 35 seconds) was used for these biopsies. The sampling procedure was as described by Brudevold et al.1 Briefly, the procedure involved the use of an adhesive tape window technic, and etching the exposed enamel for 35 seconds with 10 ,1 of 1.6 N HCl in 70% glycerol. The depth of the etch was calculated from the sampling area (20.1 mm2) and the amount of calcium present in the sample, assuming that the enamel contained 37% calcium and had a density of 2.95 gm/cm3. About 1 ml of unstimulated whole saliva was collected from each subject in a plastic tube. Smaller amounts of whole saliva were collected from the infants by means of a plastic syringe. Stimulated parotid saliva was obtained utilizing lemon sours and a modified Carlson-Crittenden device.15 Blood samples were collected by collaborating medical personnel. The biopsies were analyzed for calcium, lead, and fluoride as described by Brudevold et al,' except that the addition method was used for lead in order to account for matrix effects. In the case of saliva, 0.5 ml was mixed with 0.5 ml 0.1 N HCl, and 10-#1 quantities were analyzed for lead as before. Nonlead adsorption was accounted for by employing D, arc corrector and subtracting the absorption obtained at X = 280.7 ttm (nonlead signal) from that at X 283.3 ,um. Triplicate analyses of several samples showed good reproducibility. The blood analyses were done at the laboratory of the Massachusetts Department of Public Health Childhood Lead Poisoning Prevention Program. CARIES EXAMINATION-.Without prior cleaning, the teeth were isolated with cotton rolls and dried with compressed air. The examinations were performed with a plane surface mirror and a sharp Starlite MG5 doubleended explorer. No radiographs were taken. The examinations included only the permanent teeth. DATA TREATMENT. The biopsy data were grouped according to biopsy depth in order to explore the variation with depth of the lead concentrations. These groupings of the data, and the results of the two consecutive biopsies indicated that the lead gradient in the surface enamel is extremely steep. Therefore, in the evaluation of the data, the biopsy depth was taken into consideration as follows: the biopsy data were grouped by depth using 0.3 amm as class interval. WVithin each class, biopsies with
Downloaded from jdr.sagepub.com at MICHIGAN STATE UNIV LIBRARIES on January 11, 2015 For personal use only. No other uses without permission.
Vrol. 56 No. 10
-
LEAD IN ENAMEL & SALIVA
1167
TABLE 1 LEAD CONCENTRATIONS OF ENAMEL BIOPSIES GROUPED BY DEPTH Biopsy depth (,im)
No. of samples Mean depth (,um) Mean Pb (ppm) S.D.
0.6-
