Journal of Clinica! Periodontology 1979: 6: 186-193 Key words; Experimental gingivitis — juvenite dogs. Accepted for publication; June 29, 1978.
Development of experimental gingivitis in the juvenile and adult beagle dog Test of a model for comparative studies LARS MATSSON AKD ROLF ATTSTROM
Department of Pedodonties, University of Lund, School of Dentistry, Malmo, Sweden Department of Periodontology, Royal Dental College, Aarbus, Denmark Abstract. In a previous paper a lower tendency to develop gingivitis was reported among preschool children compared to adults. The aim of the present study was to clarify if the same age-dependent difference exists in dogs. Six beagle dogs from tbe same litter were used. Two periods of discontinued oral hygiene were studied, one period when the dogs were 3 months old and the other at 12 montbs. Prior to and between the experimental periods the animals were subjected to meticulous toothcleaning. During the experimental periods the Gingival and Plaque Index scores as well as the amounts of gingival exudate and crevicular leukocytes were determined on days 0, 4, 7, 14, 21 and 28. During the juvenile period the development of dental plaque was lower compared with the adult period. The frequency of gingival units with high Gingival Index scores was lower at every registration interval during tbe juvenile period. The amounts of gingival exudate and crevicular leukocytes increased to bigh values during the adult stage but only small increments were found in the juvenile stage. An intraindividual comparison of the increase in the amount of gingival exudate from day 0 to day 28 in areas with the same amount of dental plaque accumulation showed a lower increase in gingival exudate during the juvenile stage in all dogs.
Previous studies have shown a difference between preschool children and adults in the propensity to develop gingivitis (Mackler & Crawford 1973, Matsson 1978). By comparing tooth regions which showed a similar plaque accumulation, a statistically significant lower tendency to develop gingivitis was demonstrated in preschool children compared to adults (Matsson 1978). Longhurst et al. (1977) found that the density of plasma cells was greater in inflamed gingival tissues from adults than
from young children. These findings indicate that the cellular inflammatory response of the gingiva towards dental plaque differs between young and adult individuals. To elucidate the possible causes of the reported differences in the propensity to develop gingivitis between adults and young children, knowledge is needed about the histological characteristics of the developing gingival lesion in young individuals, including sequential gingival biopsying during a period of plaque and gingivitis develop-
0303-6979/79/030186-08$02.50/0 © 1979 Munksgaard, Copenhagen
DEVELOPMENT OF GINGIVITIS IN THE BEAGLE DOG ment. As such a histological study cannot be defended in human beings, it is desirable to establish a suitable animal model. Adult dogs have frequently been used in studies of experimental gingivitis. Morphological and functional changes in the gingiva during plaque development are well investigated (Attstrom & Egelberg 1971, Lindhe et al. 1973, Lindhe & Rylander 1975, Schroeder et al. 1975). Similar knowledge is scarce for juvenile dogs. In a recently published study. Hock (1978) reported a lower degree of gingivitis in dogs during the period of deciduous dentition compared to that of permanent dentition. However, in that study the deciduous teeth were exposed to a plaque-inducing diet for a shorter period than the succadaneous permanent teeth, which may have influenced the results. The aim of the present stndy was to clarify whether the difference which has been demonstrated between preschool children and adults in the propensity to develop gingivitis is present In dogs as well. Material and Methods
Six beagle dogs from the same Utter (two females and four males) were used. The animals were fed on gruel (Findus, Sweden) from the time of weaning. This diet was supplemented with minced meat. Three weeks prior to the start of the experiment the animals were given a hard diet according to Egelberg (1965). From the time of weaning to the start of the experiment the animals were subjected to tooth cleaning with a toothbrush and pumice once a day. During the last 3 weeks of the preexperimental period the teeth were also polished with pumice and a rotating rubber cup once a week. When the dogs had reached the age of 3 months the first part of the experiment was started. The gingiva was then in excellent health, i.e. less than 10 % of the gingival
units to be studied showed traces of gingival exudate as accepted by Attstrom et al. (1975). Al! cleaning of teeth was then discontinued for a period of 28 days, during which the animals were fed a soft diet (Egelberg 1965). The amount of dental plaque and the gingival state were determined on days 0, 4, 7, 14, 21 and 28 of the experiment. Thereafter the diet was changed to a hard one again, and the tooth cleaning was resumed. During the following months, the animals were subjected to toothcleaning with toothbrush and pumice three times a week. The teeth were also polished with pumice and a rotating rubber cup once a month. During the last 3 weeks of this period the dogs were subjected to toothcleaning with toothbrush and pumice once a day, and the teeth were polished with pumice and a rotation rubber cup once a week. When the dogs were 12 months old, the gingiva was in excellent health and toothcleaning was again discontinued and the second part of the experiment was started. The amount of dental plaque and the gingival state were determined at the same intervals as during the first part of the experiment. The outline of the experiment is given in Fig, 1, The degree of gingival inflammation was studied by determining: a) the amount of gingival exudate (Loe & Holm-Pedersen 1965) b) the amount of crevicular leukocytes (Attstrom & Egelberg 1971) c) the Gingival Index score (Loe & Silness 1963). The amount of bacterial plaque was assessed according to the criteria of the Plaque Index score (Silness & Loe 1964). One quadrant of the jaws in each dog was chosen by lot for the clinical registrations. The remaining quadrants were used for gingival biopsies. The results from the histological study will be presented in a subsequent paper.
188
MATSSON AND ATTSTR5M
Age (months)
12
13
Plaque development Plaque control Fig. 1. Outline of tbe experiment. Die Versuchsplanung. (Age = Alter, plaque development = Plaqueentwicklung, plaque control = Plaquekontrolle). Plan de I'experience. (age en mois, periodes de formation (development) ou de maitrise (control) de la plaque).
In the respective quadrant of the primary dentition, gingival exudate and crevicular leukocytes were sampled mesiobuccally and distobnccally at the second and third molar and midbuccally at the first molar and the canine. Furthermore, in the upper jaw the midbuccal gingiva at the third incisor was also included. The amount of dental plaque and the Gingival Index were scored buccally at all these teeth. In the permanent dentition the registrations were performed in the same quadrant of each dog as in the primary dentition. Gingival exudate was sampled in the upper jaw midbuccally at tbe third incisor and first premolar and mesiobuccally and distobuccally at the canine, second and third premolars and first molar; in the lower jaw mesiobuccally and distobuccally at the canine, second to fourth premolars and first and second molars. Crevicular leukocytes were sampled mesiobuccally at the teeth used for the gingival exudate measurements. The amount of dental plaque and the Gingival Index scores were determined buccally at these teeth. All operative procedures in the dogs, except toothbrushing, were performed with the animals anesthetized by intravenous injection of Pentothal-Sodium® (Abbot, Belgium). A statistical comparison between the juvenile and adult stage was made for each dog with regard to the increase in the amount of gingival exudate from day 0 to day 28. The values for the gingival units
wliich had an adjacent tooth area with a Plaque Index score of 0 on day 0 were grouped in Plaque Index classes according to the Plaque Index score on day 28, Due to the small sample and to the fact that the values among the juvenile dogs may not be normally distributed, a distribution-free test, Wilcoxon's rank sum test, was applied. Differences at the 5 % level of probability were said to be statistically significant.
Results
At the start of each experimental period the amount of bacterial plaque was negligible. The amount of bacterial plaque increased continuously during both experimental periods. However, a difference in the frequency of teeth in different Plaque Index classes was found, more teeth with a high Plaque Index score being noted during the adult compared to the juvenile period (Table 1). On day 28 of the juvenile period 65 % of the gingival units showed a Gingival Index score of 0 and 35 % showed a score of 1. During the adult period no gingival units shovi'ed a Gingival Index score of 0 on day 28, 21 % showed a score of 1 and 79 % a score of 2 (Table 1). The amounts of gingival exudate and crevicular leukocytes increased during the juvenile as well as during the adult stage but the rise was slight during the juvenile and high during the adult period (Figs. 2 and 3). In areas with the same amount of dental plaque on day 28, the increase in gingival
DEVELOPMENT OF GINGIVITIS IN THE BEAGLE DOG
189
Table 1. Percentage distribution of registered units among Plaque Index classes and Gingival Index classes in dogs at juvenile and adult stages during 28 days of plaque development Verteilung der untersuchten Einheiten in Prozent auf Plaqueindex- und GlngivaUndexklassen wahrend 28-tdgiger Plaqueentwieklung bei Hunden im jugendlichen und im erwachsenen Alter Distribution des pourcentages d'unites enregistrees parmi les classes de l'indice de Plaque et les classes de l'indice Gingival chez les chiens au stade juvenile et au stade adulte pendant 28 jours de formation de la plaque Day
0
4
7
14
21
0 1 2 3
0 1 2 3
0 1 2 3
0 1 2 3
0 1 2 3
100 0 0 0 95 5 0 0
93 7 0 0 15 31 54 0
52 33 15 0 5 21 74 0
41 33 26 0 3 15 77 5
41 33 26 0 0 41 54 5
50 15 35 0 8 26 61 5
100 0 0 0 95 5 0 0
93 7 0 0 3 92 5 0
63 33 4 0 15 64 21 0
67 33 0 0 0 38 62 0
74 26 0 0 0 36 64 0
65 35 0 0 0 21 79 0
Scores Plaque Index Juvenile stage Adult stage Gingival Index Juvenile stage Adult stage
28 0 1 2 3
Day (Tag, jour), scores (Bewertungseinheiten, scores), juvenile stage (jugendliche Altersgruppe, stade juvenile), Plaque Index (Plaqueindex, Indice de Plaque), adult stage (erwachsene Altersgruppe, stade adulte), Gingival Index (Gingivalindex, Indice Gingival).
Table 2. Distribution of the increase in the mean amount of gingival exudate from day 0 to day 28 in plaque index classes based on the plaque index score on day 28. Only the values of those gingival units which showed a plaque index score of 0 on day 0 are included. Wilcoxon's rank sum test applied Verteilung des Anstieges der Mittelwerte des gingivalen Exsudates vom Tage 0 bis zum Tage 28 auf Plaqueindexkiassen, die von den Bewertungseinheiten der Plaqueindizes des Tages 28 ausgehen. Hier werden nur die Werte soicher gingivaler Einheiten aufgefUhrt, bei denen am Tage 0 die Plaqueindex-Bewertungseinheit 0 festgestellt worden ist. Rangordnungs-Summentest nach Wilcoxon Distribution de l'augmentation de la quantite moyenne de l'exsudat gingival du jour 0 au jour 28 parmi les classes de l'indice de Plaque, en se basant sur les valeurs de l'indice de Plaque au jour 28. Seules les valeurs eoncernant les unites gingivales presetitant un Indice de Plaque de score 0 au jour 0 sont considerees. Application du test de Wilcoxon par la somme des rangs Pill Juvenile stage n Dogl 2
3 4, •5
6
0 1 0 4 0 2
0 -
-0.5 „_
5.5
PII 2
Adult stage n
^(liff
1 2 0
4.0 34,5 40.0 52.6 22.0
2 8 6
Juvenile stage P NS NS NS
Adult stage
n
^diff
n
^diff
4 4 4 0 4 2
6.5 5.3 7.0 8.0 4.5
8 8 10 4 3
43.0 36.5 56.9 21.0 53.3 57.3
6
P
Development of experimental gingivitis in the juvenile and adult beagle dog Test of a model for comparative studies LARS MATSSON AKD ROLF ATTSTROM
Department of Pedodonties, University of Lund, School of Dentistry, Malmo, Sweden Department of Periodontology, Royal Dental College, Aarbus, Denmark Abstract. In a previous paper a lower tendency to develop gingivitis was reported among preschool children compared to adults. The aim of the present study was to clarify if the same age-dependent difference exists in dogs. Six beagle dogs from tbe same litter were used. Two periods of discontinued oral hygiene were studied, one period when the dogs were 3 months old and the other at 12 montbs. Prior to and between the experimental periods the animals were subjected to meticulous toothcleaning. During the experimental periods the Gingival and Plaque Index scores as well as the amounts of gingival exudate and crevicular leukocytes were determined on days 0, 4, 7, 14, 21 and 28. During the juvenile period the development of dental plaque was lower compared with the adult period. The frequency of gingival units with high Gingival Index scores was lower at every registration interval during tbe juvenile period. The amounts of gingival exudate and crevicular leukocytes increased to bigh values during the adult stage but only small increments were found in the juvenile stage. An intraindividual comparison of the increase in the amount of gingival exudate from day 0 to day 28 in areas with the same amount of dental plaque accumulation showed a lower increase in gingival exudate during the juvenile stage in all dogs.
Previous studies have shown a difference between preschool children and adults in the propensity to develop gingivitis (Mackler & Crawford 1973, Matsson 1978). By comparing tooth regions which showed a similar plaque accumulation, a statistically significant lower tendency to develop gingivitis was demonstrated in preschool children compared to adults (Matsson 1978). Longhurst et al. (1977) found that the density of plasma cells was greater in inflamed gingival tissues from adults than
from young children. These findings indicate that the cellular inflammatory response of the gingiva towards dental plaque differs between young and adult individuals. To elucidate the possible causes of the reported differences in the propensity to develop gingivitis between adults and young children, knowledge is needed about the histological characteristics of the developing gingival lesion in young individuals, including sequential gingival biopsying during a period of plaque and gingivitis develop-
0303-6979/79/030186-08$02.50/0 © 1979 Munksgaard, Copenhagen
DEVELOPMENT OF GINGIVITIS IN THE BEAGLE DOG ment. As such a histological study cannot be defended in human beings, it is desirable to establish a suitable animal model. Adult dogs have frequently been used in studies of experimental gingivitis. Morphological and functional changes in the gingiva during plaque development are well investigated (Attstrom & Egelberg 1971, Lindhe et al. 1973, Lindhe & Rylander 1975, Schroeder et al. 1975). Similar knowledge is scarce for juvenile dogs. In a recently published study. Hock (1978) reported a lower degree of gingivitis in dogs during the period of deciduous dentition compared to that of permanent dentition. However, in that study the deciduous teeth were exposed to a plaque-inducing diet for a shorter period than the succadaneous permanent teeth, which may have influenced the results. The aim of the present stndy was to clarify whether the difference which has been demonstrated between preschool children and adults in the propensity to develop gingivitis is present In dogs as well. Material and Methods
Six beagle dogs from the same Utter (two females and four males) were used. The animals were fed on gruel (Findus, Sweden) from the time of weaning. This diet was supplemented with minced meat. Three weeks prior to the start of the experiment the animals were given a hard diet according to Egelberg (1965). From the time of weaning to the start of the experiment the animals were subjected to tooth cleaning with a toothbrush and pumice once a day. During the last 3 weeks of the preexperimental period the teeth were also polished with pumice and a rotating rubber cup once a week. When the dogs had reached the age of 3 months the first part of the experiment was started. The gingiva was then in excellent health, i.e. less than 10 % of the gingival
units to be studied showed traces of gingival exudate as accepted by Attstrom et al. (1975). Al! cleaning of teeth was then discontinued for a period of 28 days, during which the animals were fed a soft diet (Egelberg 1965). The amount of dental plaque and the gingival state were determined on days 0, 4, 7, 14, 21 and 28 of the experiment. Thereafter the diet was changed to a hard one again, and the tooth cleaning was resumed. During the following months, the animals were subjected to toothcleaning with toothbrush and pumice three times a week. The teeth were also polished with pumice and a rotating rubber cup once a month. During the last 3 weeks of this period the dogs were subjected to toothcleaning with toothbrush and pumice once a day, and the teeth were polished with pumice and a rotation rubber cup once a week. When the dogs were 12 months old, the gingiva was in excellent health and toothcleaning was again discontinued and the second part of the experiment was started. The amount of dental plaque and the gingival state were determined at the same intervals as during the first part of the experiment. The outline of the experiment is given in Fig, 1, The degree of gingival inflammation was studied by determining: a) the amount of gingival exudate (Loe & Holm-Pedersen 1965) b) the amount of crevicular leukocytes (Attstrom & Egelberg 1971) c) the Gingival Index score (Loe & Silness 1963). The amount of bacterial plaque was assessed according to the criteria of the Plaque Index score (Silness & Loe 1964). One quadrant of the jaws in each dog was chosen by lot for the clinical registrations. The remaining quadrants were used for gingival biopsies. The results from the histological study will be presented in a subsequent paper.
188
MATSSON AND ATTSTR5M
Age (months)
12
13
Plaque development Plaque control Fig. 1. Outline of tbe experiment. Die Versuchsplanung. (Age = Alter, plaque development = Plaqueentwicklung, plaque control = Plaquekontrolle). Plan de I'experience. (age en mois, periodes de formation (development) ou de maitrise (control) de la plaque).
In the respective quadrant of the primary dentition, gingival exudate and crevicular leukocytes were sampled mesiobuccally and distobnccally at the second and third molar and midbuccally at the first molar and the canine. Furthermore, in the upper jaw the midbuccal gingiva at the third incisor was also included. The amount of dental plaque and the Gingival Index were scored buccally at all these teeth. In the permanent dentition the registrations were performed in the same quadrant of each dog as in the primary dentition. Gingival exudate was sampled in the upper jaw midbuccally at tbe third incisor and first premolar and mesiobuccally and distobuccally at the canine, second and third premolars and first molar; in the lower jaw mesiobuccally and distobuccally at the canine, second to fourth premolars and first and second molars. Crevicular leukocytes were sampled mesiobuccally at the teeth used for the gingival exudate measurements. The amount of dental plaque and the Gingival Index scores were determined buccally at these teeth. All operative procedures in the dogs, except toothbrushing, were performed with the animals anesthetized by intravenous injection of Pentothal-Sodium® (Abbot, Belgium). A statistical comparison between the juvenile and adult stage was made for each dog with regard to the increase in the amount of gingival exudate from day 0 to day 28. The values for the gingival units
wliich had an adjacent tooth area with a Plaque Index score of 0 on day 0 were grouped in Plaque Index classes according to the Plaque Index score on day 28, Due to the small sample and to the fact that the values among the juvenile dogs may not be normally distributed, a distribution-free test, Wilcoxon's rank sum test, was applied. Differences at the 5 % level of probability were said to be statistically significant.
Results
At the start of each experimental period the amount of bacterial plaque was negligible. The amount of bacterial plaque increased continuously during both experimental periods. However, a difference in the frequency of teeth in different Plaque Index classes was found, more teeth with a high Plaque Index score being noted during the adult compared to the juvenile period (Table 1). On day 28 of the juvenile period 65 % of the gingival units showed a Gingival Index score of 0 and 35 % showed a score of 1. During the adult period no gingival units shovi'ed a Gingival Index score of 0 on day 28, 21 % showed a score of 1 and 79 % a score of 2 (Table 1). The amounts of gingival exudate and crevicular leukocytes increased during the juvenile as well as during the adult stage but the rise was slight during the juvenile and high during the adult period (Figs. 2 and 3). In areas with the same amount of dental plaque on day 28, the increase in gingival
DEVELOPMENT OF GINGIVITIS IN THE BEAGLE DOG
189
Table 1. Percentage distribution of registered units among Plaque Index classes and Gingival Index classes in dogs at juvenile and adult stages during 28 days of plaque development Verteilung der untersuchten Einheiten in Prozent auf Plaqueindex- und GlngivaUndexklassen wahrend 28-tdgiger Plaqueentwieklung bei Hunden im jugendlichen und im erwachsenen Alter Distribution des pourcentages d'unites enregistrees parmi les classes de l'indice de Plaque et les classes de l'indice Gingival chez les chiens au stade juvenile et au stade adulte pendant 28 jours de formation de la plaque Day
0
4
7
14
21
0 1 2 3
0 1 2 3
0 1 2 3
0 1 2 3
0 1 2 3
100 0 0 0 95 5 0 0
93 7 0 0 15 31 54 0
52 33 15 0 5 21 74 0
41 33 26 0 3 15 77 5
41 33 26 0 0 41 54 5
50 15 35 0 8 26 61 5
100 0 0 0 95 5 0 0
93 7 0 0 3 92 5 0
63 33 4 0 15 64 21 0
67 33 0 0 0 38 62 0
74 26 0 0 0 36 64 0
65 35 0 0 0 21 79 0
Scores Plaque Index Juvenile stage Adult stage Gingival Index Juvenile stage Adult stage
28 0 1 2 3
Day (Tag, jour), scores (Bewertungseinheiten, scores), juvenile stage (jugendliche Altersgruppe, stade juvenile), Plaque Index (Plaqueindex, Indice de Plaque), adult stage (erwachsene Altersgruppe, stade adulte), Gingival Index (Gingivalindex, Indice Gingival).
Table 2. Distribution of the increase in the mean amount of gingival exudate from day 0 to day 28 in plaque index classes based on the plaque index score on day 28. Only the values of those gingival units which showed a plaque index score of 0 on day 0 are included. Wilcoxon's rank sum test applied Verteilung des Anstieges der Mittelwerte des gingivalen Exsudates vom Tage 0 bis zum Tage 28 auf Plaqueindexkiassen, die von den Bewertungseinheiten der Plaqueindizes des Tages 28 ausgehen. Hier werden nur die Werte soicher gingivaler Einheiten aufgefUhrt, bei denen am Tage 0 die Plaqueindex-Bewertungseinheit 0 festgestellt worden ist. Rangordnungs-Summentest nach Wilcoxon Distribution de l'augmentation de la quantite moyenne de l'exsudat gingival du jour 0 au jour 28 parmi les classes de l'indice de Plaque, en se basant sur les valeurs de l'indice de Plaque au jour 28. Seules les valeurs eoncernant les unites gingivales presetitant un Indice de Plaque de score 0 au jour 0 sont considerees. Application du test de Wilcoxon par la somme des rangs Pill Juvenile stage n Dogl 2
3 4, •5
6
0 1 0 4 0 2
0 -
-0.5 „_
5.5
PII 2
Adult stage n
^(liff
1 2 0
4.0 34,5 40.0 52.6 22.0
2 8 6
Juvenile stage P NS NS NS
Adult stage
n
^diff
n
^diff
4 4 4 0 4 2
6.5 5.3 7.0 8.0 4.5
8 8 10 4 3
43.0 36.5 56.9 21.0 53.3 57.3
6
P
