Immediate Replantation of Maxillary Incisors in Rats: Effects of Tooth Immersion in Sodium Fluoride and Subsequent Removal of the Periodontal Ligament
 
 

Tetuo OKAMOTO1
Walter NICCOLI-FILHO2
Celso Koogi SONODA3
Ana Paula MARTINS1
Ricardo de SOUZA1

1Disciplina de Cirurgia e Traumatologia Buco-Maxilo-Facial, Faculdade de Odontologia, UNESP, Araçatuba, SP, Brasil
2Disciplina de Estomatologia, Faculdade de Odontologia, UNESP, São José dos Campos, SP, Brasil
3Disciplina de Clínica Integrada, Faculdade de Odontologia, UNESP, Araçatuba, SP, Brasil


Braz Dent J (1999) 10(2): 73-79  ISSN 0103-6440

| Introduction | Material and Methods | Results | Discussion | Conclusion | Acknowledgments | References |


The advantages and disadvantages of maintaining the periodontal ligament (PDL) in immediate replantation as well as chemical treatment of the root surface have been a matter of discussion because the vitality of such tissue in surgery is always questioned. This study evaluated the effects of conserving the tooth in sodium fluoride and the removal of the PDL before replantation of incisors in rats. There was more cementum-dentin resorption in the group with the PDL. The group without the PDL showed more discreet resorption, repair occurred through the newly formed bone tissue in the PDL space and ankylosis was more extensive than in the group with the PDL.


Key Words: incisor replantation, sodium fluoride, periodontal ligament.


Introduction

The study of the biological response after dental replantation is still a broad field of research due to the percentage of loss of teeth in such cases. Cementum-dentin resorption is the most important alteration observed not only because of its frequency but also because it is one of the causes of failure in dental replantation (Andreasen and Hjorting-Hansen, 1966; Okamoto, 1976; Barbakow et al., 1978; Andreasen, 1981a; Okamoto et al., 1987; Saad-Neto et al., 1987). Resorption seems to be particularly related to the amount of time the tooth is out of the socket (Barbakow et al., 1978) and the medium in which it is kept during this period. Of all the dental structures directly involved in replants, the PDL has been a target of interest since important alterations derived from it can lead to failure of replantation (Hammer, 1955; Flanagan and Myers, 1958; Loe and Waerhaug, 1961; Andreasen, 1981b; Okamoto et al., 1987).
The PDL that remains attached to the cementum after accidental tooth avulsion has drawn attention because its vitality at the moment of replantation is always uncertain. However, the advantages and disadvantages of maintaining this connective tissue at the moment of replantation has been a matter of discussion. According to Andreasen (1981b), small areas without the PDL show resorption and/or ankylosis. However, maintaining it without vitality on the root surface does not prevent cementum-dentin resorption (Andreasen and Kristerson, 1981) and some authors recommend its removal in such conditions (Coccia, 1980; Mahajan and Sidhu, 1981; Murad, 1982). On the other hand, several chemical treatments, varying from minor to major aggressiveness, have been utilized in immediate replants to avoid root resorption with varying degrees of clinical and experimental success.
According to Schulman et al. (1968), the immersion of the tooth in fluoride is a practical method to avoid resorption. This would be due to the fact that fluoride acts upon the dentin creating fluorapatite that is more resistant to resorption. In light of these controversies, this study evaluates the effects of tooth immersion in sodium fluoride and the subsequent removal of the PDL in immediate replantation of upper incisors in rats.


Material and Methods

Thirty-six male Wistar rats (Rattus norvegicus albinus), weighing 180-200 g, were used. They were placed on a sodium feeding before and during the study (except for the first 24 hours post-operative), and water as needed. The rats were anesthetized intraperitoneally with sodium thionembutal (approximately 40 mg/kg of body weight, Abbott, São Paulo, SP). Before removal of the maxillary right incisor, a groove was made in the labial surface of the maxillary left incisor close to the cervical area using a carborundum disc mounted at slow speed with constant cooling. The maxillary right incisor of each animal was removed using instruments that were previously adapted for this procedure (Okamoto and Russo, 1973). The animals were randomly divided into two groups of 18 each. Group I: after extraction, the teeth were kept in 2% sodium fluoride at room temperature and humidity. After two hours, they were washed with NaCl. The teeth were handled by the crown only with gauze embedded in NaCl. The pulp tissue was removed in a retrograde fashion using a slightly curved #15 Kerr file. The pulp chamber was filled with calcium hydroxide paste (Herpe, Rio de Janeiro, RJ) and distilled water using a spiral lentulo. The tooth was then replanted in the socket using a pair of pliers and was splinted with 4-0 black cotton thread (Sutupack-Ethicon, Johnson & Johnson, São Paulo, SP) that passed through the groove made in the maxillary left incisor. Group II: the teeth were also kept in 2% sodium fluoride for two hours post-extraction. The PDL was removed carefully using a #11 blade. The pulp removal, canal filling and replantation were done in the same manner as described for group I. All animals received 20,000 IU of penicillin G benzatine (Benzatacil, São Paulo, SP) in a single dose after tooth replantation.
The splint was removed 48 hours later. Ten, 30 and 60 days post-surgery, 6 animals from each group were sacrificed by excessive inhalation of sulfuric ether. The maxilla was separated in the middle and the part which contained the replanted central incisors was obtained by cutting with scissors at the distal surface of the last molar. The specimens were fixed in 10% formalin for 24 h and then calcified with a solution of equal parts of sodium citrate and formic acid (Morse, 1945). They were then embedded in paraffin for slicing of the teeth and sockets longitudinally and buccal-lingually. The slices were 6-µm thick and were stained with hematoxylin-eosin for histomorphologic analysis. The root and the socket were divided into apical, middle and cervical thirds for the purpose of analysis. The extent of the cementum-dentin resorption and ankylosis were measured in all slices with a grid ocular eye-piece.


Results

In this section, we will discuss the findings at the gingival level of the lingual side of the socket, cementum and dentin, and the connective tissue located in the PDL space.

10 days

In group I (with PDL), the gingival epithelium showed a slight proliferation and the subjacent connective tissue with a small number lymphocytes. The PDL was well-vascularized with a moderate number of fibroblasts, some lymphocytes and macrophages. There were delicate collagen fibers parallel to the long axis of the tooth. At the cervical and middle thirds, the cementum and dentin showed moderate resorption and a degenerated PDL adhered to the cementum. The socket wall showed areas of discreet resorption in the cervical third and small newly formed bone spicules at the middle third.
In group II (without PDL), the gingival epithelium grew towards the apical area of the socket without reaching the cementum. In the PDL space close to the cementum, there was well-vascularized connective tissue with a moderate number of fibroblasts, some lymphocytes and macrophages. Some delicate collagen fibers parallel to the long axis of the tooth were also observed. In the bone wall, the remaining PDL was well-vascularized and rich in fibroblasts. The cementum and dentin did not show any alteration. The socket wall at the cervical third showed discreet resorption with osteoclasts. At the middle and apical thirds the bone neoformation in the alveolar wall was more intense than in group I.

30 days

In group I, the gingival epithelium of the lingual side was adhered to the cementum, slightly below the cementoenamel junction (CEJ) and the adjacent connective tissue showed a discreet number of lymphocytes. The presence of a well-vascularized connective tissue rich in fibroblasts was seen in some parts of the PDL. In other areas, the PDL was partially replaced by newly formed bone and few collagen fibers. At the cervical and middle thirds, the cementum and dentin displayed areas of extensive deep resorption, which in some places led to a communication between the pulp chamber and the PDL space (Figure 1). The alveolar wall showed areas of bone apposition which caused the narrowing of the PDL space.
In group II, the gingival epithelium adhered to the cementum surface well below the CEJ. The PDL space close to the middle and apical thirds was practically filled by newly formed bone with many osteoblasts at its edges (Figure 2). There was still a small quantity of connective tissue and a moderate number of fibroblasts and blood vessels. The cementum and dentin showed moderate superficial resorption repaired by newly formed bone.

60 days

In group I, the gingival epithelium presented the same characteristics observed at 30 days. There was still a moderate quantity of connective tissue throughout the PDL. In other areas, it was replaced by bone tissue which at some points filled the space left by resorption (Figure 3). The cementum and dentin were present in small quantities and in some areas there was active resorption with multinuclear cells.
In group II, the gingival epithelium and the subcutaneous connective tissue showed the same morphologic characteristics observed at 30 days. In the PDL, there was more newly formed bone compared to group I. At the middle and apical thirds, the bone tissue filled almost all of the PDL space (Figure 4). In some places this bone tissue occupied moderate cementum-dentin resorption areas and in other areas it was in direct contact with the cementum.


Discussion

Many procedures of dental replantation have been studied in order to find ways to achieve periodontal reconstitution, even if not biologically acceptable but at least acceptable regarding cementum-dentin resorption. Thus, replantation in animals is a relatively controllable and reproducible experimental model. In this study, besides using the fluoride solution which decreases the degree of cementum-dentin resorption after replantation (Shulman et al., 1968; Barbakow et al., 1978), we removed the PDL in one of the groups because advantages of maintaining this connective tissue remains controversial.
At the gingival epithelium level, PDL removal led to relatively deep periodontal pockets. This may have occurred due to the absence of the PDL which would facilitate the loss of the blood clot next to the alveolar crest. This loss can also be caused by the mobility of the gingival mucosa. On the other hand, there were significant differences between the two groups at the PDL level.
The teeth in which the PDL was retained showed an early and more accentuated cementum-dentin resorption leading to an almost total loss of the replanted tooth. The neoformation of bone next to the PDL space was more discreet, although it was possible to observe areas of repair through the bone tissue.
Group II showed a more discreet cementum-dentin resorption and an intense bone neoformation between the socket wall and the root. These results indicate that there is a relationship between the presence of a degenerated PDL and the likelihood of neoformation in the periodontal space, particularly next to the root surface. Thus, the presence of this degenerated tissue adhered to the cementum makes the osseous differentiation more difficult, favoring the initiation and progression of cementum-dentin resorption.
The results obtained in this study are in agreement with the observations of Mahajan and Sidhu (1981), who conducted a clinical study in humans using topical application of 2% sodium fluoride for 4 min after extraction. The PDL was removed prior to replantation in some of the teeth in their study and after replantation, they observed clinically and radiographically better results in the group in which the PDL was removed and the root surface was treated with fluoride.
Hammarstrom et al. (1986) pointed out that when the PDL is completely destroyed at the moment of replantation, ankylosis is most likely to occur, especially when there is no inflammatory resorption. From a clinical standpoint, this suggests that the PDL should be removed before replantation if it was manipulated and/or degenerated. Studies have shown the positive effect of fluoride as a factor in decreasing cementum-dentin resorption (Schulman et al., 1968; Björvatn and Massler, 1981; Mahajan and Sidhu, 1981). Its mechanism of action in such cases is not clear. However, it is possible that the fluoride acts on the dentin and cementum by transforming hydroxyapatite into fluorapatite, which could be more resistant to resorption or even inhibit the formation of clast cells (Schulman et al., 1968; Björvatn and Massler, 1981). In the present study, using fluoride on the root surface did not reduce the cementum-dentin resorption significantly. It is possible that the time the teeth were kept in the solution (2 h) was short. Therefore, new studies are needed to evaluate the length of time most suitable to maintain the teeth in the solution.


Conclusions

In the group without the PDL, the cementum-dentin resorption was less noticeable, the formation of bone tissue in the PDL space was more rapid and intense, and the ankylosis area was larger compared to the group with the PDL.


Acknowledgments

This study was supported by FAPESP.


References

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Correspondence: Prof. Tetuo Okamoto, Rua José Bonifácio, 1193, Vila Mendonça, 16015-050 Araçatuba, SP, Brasil. E-mail: tokamoto@foa.unesp.br


Accepted June 22, 1999
Eletronic publication: April, 2000


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