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RESEARCH REPORT |
1 Departments of Endodontics and
2 Pathology, School of Dentistry, Aichi-Gakuin University, 2-11 Suemori-dori, Chikusa-ku, Nagoya 464-8651, Aichi, Japan; and
3 Department of Stomatology, University of California, San Francisco, CA 94143, USA;
*corresponding author, akihiro10{at}aol.com
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ABSTRACT |
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 TOP ABSTRACT INTRODUCTIONRESULTSDISCUSSIONREFERENCES |
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KEY WORDS: periradicular lesion • type 2 diabetes • diabetic rats
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INTRODUCTION |
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TOPABSTRACTINTRODUCTIONRESULTSDISCUSSIONREFERENCES |
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Rats with spontaneous or type 2 diabetes have been generated by selective breeding of normal rats with impaired glucose tolerance (Goto et al., 1976). Experimental periradicular lesions have been produced in many species, including rats (Waterman et al., 1998), cats (Maguire et al., 1998), dogs (Witherspoon and Gutmann, 2000), and monkeys (Walton and Garnick, 1986). The rat is considered a good model because the lesions—produced by exposure of the pulp of the molar to the oral cavity—are reproducible and because the histology has been well-described (Page and Schroeder, 1982; Yamasaki et al., 1994; Waterman et al., 1998).
We carried out the present study to investigate the relationship between type 2 diabetes mellitus and the development of periradicular lesions in rats.
MATERIALS & METHODS
Twenty normal Wistar rats and 20 GK rats with spontaneous type 2 diabetes mellitus (Goto et al., 1976) were studied. All rats were males weighing 230 g each and were conventionally housed. The rats were divided equally into two groups of normal rats (groups A and B) and two groups of diabetic rats (groups C and D). All rats were fed a normal laboratory diet, but the rats in groups A and C were given tap water (200 mL/kg/day), whereas those in groups B and D were given a 30% sucrose solution instead of tap water (200 mL/kg/day). The study design was approved by the Ethics Committee of the School of Dentistry of Aichi-Gakuin University.
Periradicular Lesions
The rats were anesthetized with ether, and the pulp of the left mandibular first molar was exposed through the occlusal surface with a size 1/2 round bur (Maillfer, Zürich, Switzerland).
Glucose Tolerance Test
Glucose tolerance tests were performed at the time of the pulp exposure and 2 and 4 wks thereafter. Blood samples were collected from a tail vein after a 20-hour fast, and the stomach was intubated. A 20% glucose solution (1 mL/kg body weight) was then infused, and blood samples were obtained 30, 60, and 120 min later. The blood glucose concentration was measured by a photospectroscopic method (Refloluxs, Boehringer Mannheim, Mannheim, Germany).
Measurement of Body Weight and Leukocyte Number
Body weight was measured immediately after the pulp exposure and at 2 and 4 wks. At the same time, peripheral blood was taken into capillary tubes containing an anti-coagulant, EDTA-2K (Yoneyama Co., Osaka, Japan), and the leukocyte count was determined with a hematology analyzer (STKS, Beckman Coulter, Fullerton, CA, USA).
Histology
At 2 or 4 wks after pulp exposure, the rats were killed with sodium pentobarbital. The left mandible was removed, immediately fixed in 10% neutral buffered formalin, decalcified in 0.5 M EDTA, embedded in paraffin, and sectioned serially at 6 µm in the mesiodistal plane. The sections were stained with hematoxylin-eosin, and the lesions of the periradicular and pulpal tissues in the mesial root of the mandibular first molar were investigated histologically.
Histometry
On 5 serial histologic sections from each rat, the central portion of the periradicular lesion was measured with an image-processing system, which consisted of a light microscope (BH-2, Olympus, Tokyo, Japan), a color camera (ICD-740, Olympus-Ikegami Co., Tokyo, Japan), a color image processor (Nexus 644, Kasiwagi Research, Tokyo, Japan), and a personal computer (Power Macintosh 8500, Apple, Cupertino, CA, USA) (Imaizumi, 1997). The average area was determined for each animal, and the average value (mean ± SD) was calculated for each experimental period. The histometric data were analyzed with the Mann-Whitney U test.
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RESULTS |
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TOPABSTRACTINTRODUCTIONRESULTSDISCUSSIONREFERENCES |
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Blood Glucose Level
Throughout the study, the blood glucose levels were significantly higher in groups C and D than in groups A and B (100 vs. 400 mg/dL; p < 0.01). In particular, group D showed a significant time-dependent increase (p < 0.01) in blood glucose value (Figs. 1a, 1b
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Histologic Findings
Two weeks after pulp exposure, necrosis was found in the upper half of the pulpal tissue in group A. Inflammatory cell infiltration was observed in the lower half of the pulpal tissue and in the periradicular periodontal tissue. Bone resorption was seen in the periradicular alveolar tissue. In groups B and C, the pulp and periradicular lesions were similar to those in group A. In group D, however, the pulpal tissue was almost completely necrotic, and the inflammatory cell infiltration and alveolar bone resorption were more severe than in the three other groups (Figs. 2a, 2b, 2c).
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DISCUSSION |
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TOPABSTRACTINTRODUCTIONRESULTSDISCUSSIONREFERENCES |
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Several factors might explain why the lesions were most severe in the diabetic rats given the sucrose solution. One reason may be related to impaired leukocyte function. In several studies, the increased susceptibility to infection associated with diabetes mellitus has been ascribed to impaired function of polymorphonuclear leukocytes (Saiki et al., 1980; Gustke et al., 1998). Consistent with this idea, the diabetic rats in our study had significantly fewer leukocytes than the control rats at both 2 and 4 wks. Thus, the diabetic rats may have had an insufficient number of leukocytes to defend against the continuous penetration of bacteria, resulting in more severe periradicular lesions.
Another possible explanation for the greater severity of lesions in the diabetic rats relates to the regulation of insulin levels. It has been reported that sucrose exacerbates the diabetic condition of rats with type 2 diabetes (Nakamura et al., 2001). This worsening appears to be caused by reduced insulin secretion resulting from damage to the pancreatic B-cells (Goto and Kakizaki, 1984). In fact, Goto and Kakizaki (1982) reported that rats with type 2 diabetes given a 30% sucrose solution had a remarkably decreased insulin content in the pancreas and in the plasma. Since our diabetic rat model is the same as that examined in these other studies, it stands to reason that our animals, which were also given a 30% sucrose solution, also had reduced insulin secretion. Furthermore, our rats had high blood glucose levels—approximately 400 mg/dL— similar to those in diabetic rats in other studies. For example, the mean blood glucose level was over 300 mg/dL in rats with alloxan-induced diabetes (Bissada et al., 1966) and ranged from 388 (Kohsaka et al., 1996) to 843 mg/dL (Johnson, 1985) in rats with streptozotocin-induced diabetes. Therefore, the diabetic rats in our study likely had reduced insulin levels, and thus a reduced level of diabetic control that made them susceptible to the development of more severe periradicular lesions.
Finally, diabetes in humans has been consistently associated with altered calcium homeostasis and bone loss, and the same appears to be the case in experimental rat models of diabetes (Ward et al., 2001). It has been reported that a lack of insulin in diabetic rats correlates with a negative calcium balance in the body (Schneider and Schedl, 1972), which leads to reduced bone turnover and to growth arrest (Hough et al., 1981). This calcium imbalance and reduced bone turnover may, in part, also explain why the periradicular lesions were most severe in the diabetic rats given the sucrose solution.
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ACKNOWLEDGMENTS |
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Received March 13, 2001; Last revision December 20, 2002; Accepted January 28, 2003
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REFERENCES |
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0.01). (c) The rats in groups A and B had significantly more leukocytes than those in groups C and D at all experimental periods (n = 5, p 
